Flavonols and dihydroflavonols inhibit the main protease activity of SARS-CoV-2 and the replication of human coronavirus 229E
}, volume={571}, ISSN={["1089-862X"]}, DOI={10.1016/j.virol.2022.04.005}, abstractNote={Since December 2019, the deadly novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the current COVID-19 pandemic. To date, vaccines are available in the developed countries to prevent the infection of this virus; however, medicines are necessary to help control COVID-19. Human coronavirus 229E (HCoV-229E) causes the common cold. The main protease (M pro ) is an essential enzyme required for the multiplication of these two viruses in the host cells, and thus is an appropriate candidate to screen potential medicinal compounds. Flavonols and dihydroflavonols are two groups of plant flavonoids. In this study, we report docking simulation with two M pro enzymes and five flavonols and three dihydroflavonols, in vitro inhibition of the SARS-CoV-2 M pro , and in vitro inhibition of the HCoV 229E replication. The docking simulation results predicted that (+)-dihydrokaempferol, (+)- dihydroquercetin, (+)-dihydromyricetin, kaempferol, quercetin, myricentin, isoquercitrin, and rutin could bind to at least two subsites (S1, S1', S2, and S4) in the binding pocket and inhibit the activity of SARS-CoV-2 M pro . Their affinity scores ranged from -8.8 to -7.4 (kcal/mol). Likewise, these compounds were predicted to bind and inhibit the HCoV-229E M pro activity with affinity scores ranging from -7.1 to -7.8 (kcal/mol). In vitro inhibition assays showed that seven available compounds effectively inhibited the SARS-CoV-2 M pro activity and their IC 50 values ranged from 0.125 to 12.9 μM. Five compounds inhibited the replication of HCoV-229E in Huh-7 cells. These findings indicate that these antioxidative flavonols and dihydroflavonols are promising candidates for curbing the two viruses.}, journal={VIROLOGY}, author={Zhu, Yue and Scholle, Frank and Kisthardt, Samantha C. and Xie, De-Yu}, year={2022}, month={Jun}, pages={21–33} } @article{zhu_yan_liu_xia_an_xu_zhao_liu_guo_zhang_et al._2022, title={Alternative splicing of CsJAZ1 negatively regulates flavan-3-ol biosynthesis in tea plants}, volume={1}, ISSN={["1365-313X"]}, url={http://dx.doi.org/10.1111/tpj.15670}, DOI={10.1111/tpj.15670}, abstractNote={SUMMARYFlavan‐3‐ols are abundant in the tea plant (Camellia sinensis) and confer tea with flavor and health benefits. We recently found that alternative splicing of genes is likely involved in the regulation of flavan‐3‐ol biosynthesis; however, the underlying regulatory mechanisms remain unknown. Here, we integrated metabolomics and transcriptomics to construct metabolite–gene networks in tea leaves, collected over five different months and from five spatial positions, and found positive correlations between endogenous jasmonic acid (JA), flavan‐3‐ols, and numerous transcripts. Transcriptome mining further identified CsJAZ1, which is negatively associated with flavan‐3‐ols formation and has three CsJAZ1 transcripts, one full‐length (CsJAZ1‐1), and two splice variants (CsJAZ1‐2 and ‐3) that lacked 3′ coding sequences, with CsJAZ1‐3 also lacking the coding region for the Jas domain. Confocal microscopy showed that CsJAZ1‐1 was localized to the nucleus, while CsJAZ1‐2 and CsJAZ1‐3 were present in both the nucleus and the cytosol. In the absence of JA, CsJAZ1‐1 was bound to CsMYC2, a positive regulator of flavan‐3‐ol biosynthesis; CsJAZ1‐2 functioned as an alternative enhancer of CsJAZ1‐1 and an antagonist of CsJAZ1‐1 in binding to CsMYC2; and CsJAZ1‐3 did not interact with CsMYC2. In the presence of JA, CsJAZ1‐3 interacted with CsJAZ1‐1 and CsJAZ1‐2 to form heterodimers that stabilized the CsJAZ1‐1–CsMYC2 and CsJAZ1‐2–CsMYC2 complexes, thereby repressing the transcription of four genes that act late in the flavan‐3‐ol biosynthetic pathway. These data indicate that the alternative splicing variants of CsJAZ1 coordinately regulate flavan‐3‐ol biosynthesis in the tea plant and improve our understanding of JA‐mediated flavan‐3‐ol biosynthesis.}, journal={PLANT JOURNAL}, publisher={Wiley}, author={Zhu, Junyan and Yan, Xiaomei and Liu, Shengrui and Xia, Xiaobo and An, Yanlin and Xu, Qingshan and Zhao, Shiqi and Liu, Lu and Guo, Rui and Zhang, Zhaoliang and et al.}, year={2022}, month={Mar} } @article{yuzuak_xie_2022, title={Anthocyanins from muscadine (Vitis rotundifolia) grape fruit}, volume={30}, ISSN={["2214-6628"]}, DOI={10.1016/j.cpb.2022.100243}, abstractNote={Muscadine grapes (Vitis rotundifolia) have multiple health benefits to human health. The high nutritional values of muscadine berries result from antioxidative anthocyanins and other phenolic compounds. Since the middle of the 18th century, muscadine grapes have been cropped in the southeastern United States. Early cultivars were selected from wild vines. To date, the breeding efforts have created more than 100 cultivars featured by different fruit pigmentations for wine, juice, or fresh market industries. Herein, we review features of anthocyanin profiles in muscadine berries and different final products. Main anthocyanidins include cyanidin, delphinidin, petunidin, peonidin, and malvidin. Pelargonidin has been also reported in certain types of varieties. Main anthocyanins are comprised of cyanin and delphinin, which derive from non-acylated 3,5-O-diglucosides of the main five anthocyanidins. In addition, minor pelargonin such as pelargonidin 3, 5-diglucoside and other minor anthocyanins have been identified in some cultivars. Moreover, we discussed biosynthesis of anthocyanins, color instability and intensity of anthocyanins, and effects of copigments such as proanthocyanidins on color stability and intensity of muscadine products.}, journal={CURRENT PLANT BIOLOGY}, author={Yuzuak, Seyit and Xie, De-Yu}, year={2022}, month={Apr} } @article{yao_liu_zhuang_zhao_dai_jiang_wang_jiang_zhang_qian_et al._2022, title={Insights into acylation mechanisms: co-expression of serine carboxypeptidase-like acyltransferases and their non-catalytic companion paralogs}, volume={5}, ISSN={["1365-313X"]}, DOI={10.1111/tpj.15782}, abstractNote={SUMMARYSerine carboxypeptidase‐like acyltransferases (SCPL‐ATs) play a vital role in the diversification of plant metabolites. Galloylated flavan‐3‐ols highly accumulate in tea (Camellia sinensis), grape (Vitis vinifera), and persimmon (Diospyros kaki). To date, the biosynthetic mechanism of these compounds remains unknown. Herein, we report that two SCPL‐AT paralogs are involved in galloylation of flavan‐3‐ols: CsSCPL4, which contains the conserved catalytic triad S‐D‐H, and CsSCPL5, which has the alternative triad T‐D‐Y. Integrated data from transgenic plants, recombinant enzymes, and gene mutations showed that CsSCPL4 is a catalytic acyltransferase, while CsSCPL5 is a non‐catalytic companion paralog (NCCP). Co‐expression of CsSCPL4 and CsSCPL5 is likely responsible for the galloylation. Furthermore, pull‐down and co‐immunoprecipitation assays showed that CsSCPL4 and CsSCPL5 interact, increasing protein stability and promoting post‐translational processing. Moreover, phylogenetic analyses revealed that their homologs co‐exist in galloylated flavan‐3‐ol‐ or hydrolyzable tannin‐rich plant species. Enzymatic assays further revealed the necessity of co‐expression of those homologs for acyltransferase activity. Evolution analysis revealed that the mutations of the CsSCPL5 catalytic residues may have taken place about 10 million years ago. These findings show that the co‐expression of SCPL‐ATs and their NCCPs contributes to the acylation of flavan‐3‐ols in the plant kingdom.}, journal={PLANT JOURNAL}, author={Yao, Shengbo and Liu, Yajun and Zhuang, Juhua and Zhao, Yue and Dai, Xinlong and Jiang, Changjuan and Wang, Zhihui and Jiang, Xiaolan and Zhang, Shuxiang and Qian, Yumei and et al.}, year={2022}, month={May} } @article{li_guo_wang_li_jiang_liu_xie_gao_xia_2022, title={Molecular and biochemical characterization of two 4-coumarate: CoA ligase genes in tea plant (Camellia sinensis)}, volume={109}, ISSN={["1573-5028"]}, url={https://doi.org/10.1007/s11103-022-01269-6}, DOI={10.1007/s11103-022-01269-6}, abstractNote={Two 4-coumarate: CoA ligase genes in tea plant involved in phenylpropanoids biosynthesis and response to environmental stresses. Tea plant is rich in flavonoids benefiting human health. Lignin is essential for tea plant growth. Both flavonoids and lignin defend plants from stresses. The biosynthesis of lignin and flavonoids shares a key intermediate, 4-coumaroyl-CoA, which is formed from 4-coumaric acid catalyzed by 4-coumaric acid: CoA ligase (4CL). Herein, we report two 4CL paralogs from tea plant, Cs4CL1 and Cs4CL2, which are a member of class I and II of this gene family, respectively. Cs4CL1 was mainly expressed in roots and stems, while Cs4CL2 was mainly expressed in leaves. The promoter of Cs4CL1 had AC, nine types of light sensitive (LSE), four types of stress-inducible (SIE), and two types of meristem-specific elements (MSE). The promoter of Cs4CL2 also had AC and nine types of LSEs, but only had two types of SIEs and did not have MSEs. In addition, the LSEs varied in the two promoters. Based on the different features of regulatory elements, three stress treatments were tested to understand their expression responses to different conditions. The resulting data indicated that the expression of Cs4CL1 was sensitive to mechanical wounding, while the expression of Cs4CL2 was UV-B-inducible. Enzymatic assays showed that both recombinant Cs4CL1 and Cs4CL2 transformed 4-coumaric acid (CM), ferulic acid (FR), and caffeic acid (CF) to their corresponding CoA ethers. Kinetic analysis indicated that the recombinant Cs4CL1 preferred to catalyze CF, while the recombinant Cs4CL2 favored to catalyze CM. The overexpression of both Cs4CL1 and Cs4CL2 increased the levels of chlorogenic acid and total lignin in transgenic tobacco seedlings. In addition, the overexpression of Cs4CL2 consistently increased the levels of three flavonoid compounds. These findings indicate the differences of Cs4CL1 and Cs4CL2 in the phenylpropanoid metabolism.}, number={4-5}, journal={PLANT MOLECULAR BIOLOGY}, publisher={Springer Science and Business Media LLC}, author={Li, Mingzhuo and Guo, Lili and Wang, Yeru and Li, Yanzhi and Jiang, Xiaolan and Liu, Yajun and Xie, De-Yu and Gao, Liping and Xia, Tao}, year={2022}, month={May} } @article{jie_ma_xie_jie_2022, title={Transcriptional and Metabolic Characterization of Feeding Ramie Growth Enhanced by a Combined Application of Gibberellin and Ethrel}, volume={23}, ISSN={["1422-0067"]}, url={https://doi.org/10.3390/ijms231912025}, DOI={10.3390/ijms231912025}, abstractNote={Feeding ramie cultivars (Boehmaria nivea L.) are an important feedstock for livestock. Increasing their biomass and improving their nutritional values are essential for animal feeding. Gibberellin (GA3) and ethylene (ETH) are two plant hormones that regulate the growth, development, and metabolism of plants. Herein, we report effects of the GA3 and ETH application on the growth and plant metabolism of feeding ramie in the field. A combination of GA3 and ETH was designed to spray new plants. The two hormones enhanced the growth of plants to produce more biomass. Meanwhile, the two hormones reduced the contents of lignin in leaves and stems, while increased the content of flavonoids in leaves. To understand the potential mechanisms behind these results, we used RNA-seq-based transcriptomics and UPLC-MS/MS-based metabolomics to characterize gene expression and metabolite profiles associated with the treatment of GA3 and ETH. 1562 and 2364 differentially expressed genes (DEGs) were obtained from leaves and stems (treated versus control), respectively. Meanwhile, 99 and 88 differentially accumulated metabolites (DAMs) were annotated from treated versus control leaves and treated versus control stems, respectively. Data mining revealed that both DEGs and DAMs were associated with multiple plant metabolisms, especially plant secondary metabolism. A specific focus on the plant phenylpropanoid pathway identified candidates of DEGs and DEMs that were associated with lignin and flavonoid biosynthesis. Shikimate hydroxycinnamoyl transferase (HCT) is a key enzyme that is involved in the lignin biosynthesis. The gene encoding B. nivea HCT was downregulated in the treated leaves and stems. In addition, genes encoding 4-coumaryl CoA ligase (4CL) and trans-cinnamate 4-monooxygenase (CYP73A), two lignin pathway enzymes, were downregulated in the treated stems. Meanwhile, the reduction in lignin in the treated leaves led to an increase in cinnamic acid and p-coumaryl CoA, two shared substrates of flavonoids that are enhanced in contents. Taken together, these findings indicated that an appropriate combination of GA3 and ETH is an effective strategy to enhance plant growth via altering gene expression and plant secondary metabolism for biomass-enhanced and value-improved feeding ramie.}, number={19}, journal={INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES}, author={Jie, Hongdong and Ma, Yushen and Xie, De-Yu and Jie, Yucheng}, year={2022}, month={Oct} } @article{li_he_la hovary_zhu_dong_liu_xing_liu_jie_ma_et al._2022, title={A de novo regulation design shows an effectiveness in altering plant secondary metabolism}, volume={37}, ISSN={["2090-1224"]}, url={http://dx.doi.org/10.1016/j.jare.2021.06.017}, DOI={10.1016/j.jare.2021.06.017}, abstractNote={Transcription factors (TFs) and}, journal={JOURNAL OF ADVANCED RESEARCH}, publisher={Elsevier BV}, author={Li, Mingzhuo and He, Xianzhi and La Hovary, Christophe and Zhu, Yue and Dong, Yilun and Liu, Shibiao and Xing, Hucheng and Liu, Yajun and Jie, Yucheng and Ma, Dongming and et al.}, year={2022}, month={Mar}, pages={43–60} } @article{zhu_scholle_kisthardt_xie_2021, title={Flavonols and dihydroflavonols inhibit the main protease activity of SARS-CoV-2 and the replication of human coronavirus 229E}, volume={7}, url={http://dx.doi.org/10.1101/2021.07.01.450756}, DOI={10.1101/2021.07.01.450756}, abstractNote={AbstractSince December 2019, the deadly novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the current COVID-19 pandemic. To date, vaccines are available in the developed countries to prevent the infection of this virus, however, medicines are necessary to help control COVID-19. Human coronavirus 229E (HCoV-229E) causes the common cold. The main protease (Mpro) is an essential enzyme required for the multiplication of these two viruses in the host cells, and thus is an appropriate candidate to screen potential medicinal compounds. Flavonols and dihydroflavonols are two groups of plant flavonoids. In this study, we report docking simulation with two Mpro enzymes and five flavonols and three dihydroflavonols, in vitro inhibition of the SARS-CoV-2 Mpro, and in vitro inhibition of the HCoV 229E replication. The docking simulation results predicted that (+)-dihydrokaempferol, (+)-dihydroquercetin, (+)-dihydromyricetin, kaempferol, quercetin, myricentin, isoquercetin, and rutin could bind to at least two subsites (S1, S1’, S2, and S4) in the binding pocket and inhibit the activity of SARS-CoV-2 Mpro. Their affinity scores ranged from −8.8 to −7.4. Likewise, these compounds were predicted to bind and inhibit the HCoV-229E Mpro activity with affinity scores ranging from −7.1 to −7.8. In vitro inhibition assays showed that seven available compounds effectively inhibited the SARS-CoV-2 Mpro activity and their IC50 values ranged from 0.125 to 12.9 µM. Five compounds inhibited the replication of HCoV-229E in Huh-7 cells. These findings indicate that these antioxidative flavonols and dihydroflavonols are promising candidates for curbing the two viruses.}, journal={[]}, publisher={Cold Spring Harbor Laboratory}, author={Zhu, Yue and Scholle, Frank and Kisthardt, Samantha C. and Xie, De-Yu}, year={2021}, month={Jul} } @article{li_guo_wang_li_jiang_liu_xie_gao_xia_2021, title={Molecular and Biochemical Characterization of Two 4-Coumarate: Coa Ligase Genes in Tea Plant (Camellia Sinensis)}, url={https://doi.org/10.21203/rs.3.rs-897959/v1}, DOI={10.21203/rs.3.rs-897959/v1}, abstractNote={Abstract Tea is rich in flavonoids benefiting human health. Lignin is essential for tea plant growth. Both flavonoids and lignin defend plants from stresses. The biosynthesis of lignin and flavonoids shares a key intermediate, p-coumaroyl-CoA, which is formed from p-coumaric acid catalyzed by p-coumaric acid: CoA ligase (4CL). Herein, we reported two 4CL paralogs from tea plant, Cs4CL1 and Cs4CL2, which were a member of class I and II, respectively. Cs4CL1 was mainly expressed in roots and stems, while Cs4CL2 was mainly expressed in leaves. The promoter of Cs4CL1 had AC, light and stress-inducible (LSI), and meristem-specific elements, while that of Cs4CL2 had AC and LSI elements only. Moreover, the promoter of Cs4CL1 had two more stress-inducible elements than Cs4CL2 had and the two promoters had six different light-inducible elements. These features suggested their differences in their responses to environmental conditions. Three stress treatments indicated that the expression of Cs4CL1 was sensitive to mechanical wounding, while the expression of Cs4CL2 was UV-B-inducible. Enzymatic assay showed that both recombinant Cs4CL1 and Cs4CL2 transformed p-coumaric acid, ferulic acid and caffeic acid to their corresponding CoA ethers. Kinetic analysis indicated that the recombinant Cs4CL1 preferred to catalyze caffeic acid, while the recombinant Cs4CL2 favored to catalyze p-coumaric acid. The overexpression of both Cs4CL1 and Cs4CL2 increased the levels of chlorogenic acid and total lignin in transgenic tobacco seedlings. In addition, the overexpression of Cs4CL2 increased the levels of three flavonoid compounds. These findings indicate the differences of Cs4CL1 and Cs4CL2 in the phenylpropanoid metabolism.}, author={Li, Mingzhuo and Guo, Lili and Wang, Yeru and Li, Yanzhi and Jiang, Xiaolan and Liu, Yajun and Xie, Deyu and Gao, Liping and Xia, Tao}, year={2021}, month={Sep} } @article{xie_li_jie_xie_yang_shi_zhong_2020, title={Comparative transcriptomics of stem bark reveals genes associated with bast fiber development in Boehmeria nivea L. gaud (ramie)}, volume={21}, ISSN={["1471-2164"]}, DOI={10.1186/s12864-020-6457-8}, abstractNote={Abstract Background Boehmeria nivea L. Gaud (Ramie) produces one of the longest natural fibers in nature. The bark of ramie mainly comprises of the phloem tissue of stem and is the raw material for fiber. Therefore, identifying the molecular regulation of phloem development is important for understanding of bast fiber biosynthesis and improvement of fiber quality in ramie. Results In this study, we collected top bud (TB), bark from internode elongating region (ER) and bark from internode fully elongated region (FER) from the ramie variety Zhongzhu No. 1. Histological study indicated that these samples contain phloem tissues at different developmental and maturation stages, with a higher degree of maturation of phloem tissue in FER. RNA sequencing (RNA-seq) was performed and de novo transcriptome was assembled. Unigenes and differentially expressed genes (DEGs) in these three samples were identified. The analysis of DEGs by using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed clear differences in gene expression between ER and FER. Some unigenes involved in secondary cell wall biosynthesis were up-regulated in both ER and FER, while unigenes for some cell wall components or cell wall modifications showed differential expression between ER and FER. In addition, the ethylene respond factors (ERFs) in the ethylene signaling pathway were up-regulated in FER, and ent-kaurenoic acid oxidase (KAO) and GA 20-oxidase (GA20ox) for gibberellins biosynthesis were up-regulated while GA 2-oxidase (GA2ox) for gibberellin inactivation was down-regulated in FER. Conclusions Both morphological study and gene expression analysis supported a burst of phloem and vascular developmental processes during the fiber maturation in the ramie stem, and ethylene and gibberellin are likely to be involved in this process. Our findings provide novel insights into the phloem development and fiber maturation in ramie, which could be useful for fiber improvement in ramie and other fiber crops. }, number={1}, journal={BMC GENOMICS}, author={Xie, Jiyong and Li, Jiaqi and Jie, Yucheng and Xie, Deyu and Yang, Di and Shi, Huazhong and Zhong, Yingli}, year={2020}, month={Jan} } @article{dai_liu_zhuang_yao_liu_jiang_zhou_wang_xie_bennetzen_et al._2020, title={Discovery and characterization of tannase genes in plants: roles in hydrolysis of tannins}, volume={226}, ISSN={["1469-8137"]}, url={https://doi.org/10.1111/nph.16425}, DOI={10.1111/nph.16425}, abstractNote={Summary Plant tannins, including condensed tannins (CTs) and hydrolyzable tannins (HTs), are widely distributed in the plant kingdom. To date, tannase (TA) – is a type of tannin acyl‐hydrolase hydrolyzing HTs, CT monomer gallates and depsides – has been reported in microbes only. Whether plants express TA remains unknown. Herein, we report plant TA genes. A native Camellia sinensis TA (CsTA) is identified from leaves. Six TAs are cloned from tea, strawberry (Fragaria × ananassa, Fa) and four other crops. Biochemical analysis shows that the native CsTA and six recombinant TAs hydrolyze tannin compounds, depsides and phenolic glycosides. Transcriptional and metabolic analyses reveal that the expression of CsTA is oppositely associated with the accumulation of galloylated catechins. Moreover, the transient overexpression and RNA interference of FaTA are positively associated with the accumulation of ellagitannins in strawberry fruit. Phylogenetic analysis across different kingdoms shows that 29 plant TA homologs are clustered as a plant‐specific TA clade in class I carboxylesterases. Further analysis across the angiosperms reveals that these TA genes are dispersed in tannin‐rich plants, which share a single phylogenetic origin c. 120 million yr ago. Plant TA is discovered for the first time in the plant kingdom and is shown to be valuable to improve tannin compositions in plants. }, number={4}, journal={NEW PHYTOLOGIST}, publisher={Wiley}, author={Dai, Xinlong and Liu, Yajun and Zhuang, Juhua and Yao, Shengbo and Liu, Li and Jiang, Xiaolan and Zhou, Kang and Wang, Yunsheng and Xie, Deyu and Bennetzen, Jeffrey L. and et al.}, year={2020}, month={May}, pages={1104–1116} } @article{zhu_xie_2020, title={Docking Characterization and in vitro Inhibitory Activity of Flavan-3-ols and Dimeric Proanthocyanidins Against the Main Protease Activity of SARS-Cov-2}, volume={11}, ISSN={1664-462X}, url={http://dx.doi.org/10.3389/fpls.2020.601316}, DOI={10.3389/fpls.2020.601316}, abstractNote={We report to use the main protease (Mpro) of SARS-Cov-2 to screen plant flavan-3-ols and proanthocyanidins. Twelve compounds, (–)-afzelechin (AF), (–)-epiafzelechin (EAF), (+)-catechin (CA), (–)-epicatechin (EC), (+)-gallocatechin (GC), (–)-epigallocatechin (EGC), (+)-catechin-3-O-gallate (CAG), (–)-epicatechin-3-O-gallate (ECG), (–)-gallocatechin-3-O-gallate (GCG), (–)-epigallocatechin-3-O-gallate (EGCG), procyanidin A2 (PA2), and procyanidin B2 (PB2), were selected for docking simulation. The resulting data predicted that all 12 metabolites could bind to Mpro. The affinity scores of PA2 and PB2 were predicted to be −9.2, followed by ECG, GCG, EGCG, and CAG, −8.3 to −8.7, and then six flavan-3-ol aglycones, −7.0 to −7.7. Docking characterization predicted that these compounds bound to three or four subsites (S1, S1′, S2, and S4) in the binding pocket of Mpro via different spatial ways and various formation of one to four hydrogen bonds. In vitro analysis with 10 available compounds showed that CAG, ECG, GCG, EGCG, and PB2 inhibited the Mpro activity with an IC50 value, 2.98 ± 0.21, 5.21 ± 0.5, 6.38 ± 0.5, 7.51 ± 0.21, and 75.3 ± 1.29 μM, respectively, while CA, EC, EGC, GC, and PA2 did not have inhibitory activities. To further substantiate the inhibitory activities, extracts prepared from green tea (GT), two muscadine grapes (MG), cacao, and dark chocolate (DC), which are rich in CAG, ECG, GAG, EGCG, or/and PB2, were used for inhibitory assay. The resulting data showed that GT, two MG, cacao, and DC extracts inhibited the Mpro activity with an IC50 value, 2.84 ± 0.25, 29.54 ± 0.41, 29.93 ± 0.83, 153.3 ± 47.3, and 256.39 ± 66.3 μg/ml, respectively. These findings indicate that on the one hand, the structural features of flavan-3-ols are closely associated with the affinity scores; on the other hand, the galloylation and oligomeric types of flavan-3-ols are critical in creating the inhibitory activity against the Mpro activity.}, journal={Frontiers in Plant Science}, publisher={Frontiers Media SA}, author={Zhu, Yue and Xie, De-Yu}, year={2020}, month={Nov} } @article{peng_long_du_li_xie_2020, title={RNA-seq of aboveground sporophyte's transcriptome of Huperzia serrata and transcriptional understanding of early steps associated with huperzine biosynthesis in forest}, volume={24}, ISSN={["2214-6628"]}, DOI={10.1016/j.cpb.2020.100159}, abstractNote={Chinese toothed clubmoss (Huperzia serrata) is a primitive fern native in certain types of forests. It is severely endangered in China due to its difficult propagation and massive hunting for huperzine A to improve and prevent Alzheimer's disease. In this study, we completed RNA-seq for young leaves (HSYL), old leaves (HSOL), and stems (HSS) of H. serrata (HS) plants collected from a forest in 2016. From these tissues, we generated 77,430,786 trimmed paired reads (paired 32,418,231,517 pb). Sequence assembly obtained 621,023 contigs and 755,420 transcripts, which were annotated to be 49,923 unigenes. Of all unigenes, 40,612 were expressed in the three tissues, while 9311 were differentially expressed. 1158, 1675, and 1326 unigenes are specifically expressed in HSYL, HSOL, and HSS, respectively. Sequence mining obtained two unigenes encoding lysine decarboxylase (LDC1 and 2) and three unigenes encoding copper amine oxidase (CAO1, 2, and 3), which were involved in two early steps of the huperzine pathway. Quantitative RT-PCR was carried out to validate these early pathway genes using samples collected in 2017. RPKM values and qRT-PCR analysis characterized that the transcriptional level of LDC1 was the highest in old leaves followed by young leaves and stems, while the transcriptional level of LDC2 was similar in three tissues. Of three CAO genes, qRT-PCR validated the expression of CAO1 and CAO2 but not CAO3. Metabolite analysis showed the formation and differentiation of huperzine A in the three tissues collected in 2019, demonstrating the expression of the biosynthetic pathway of huperzine. Furthermore, the gene expression and huperzine A formation are discussed to understand the biosynthesis of huperzine in the forest. Taken together, this study provides a valuable genome-wide transcriptome of the aboveground sporophyte tissues and shows a dynamically transcriptional and metabolic feature of the huperzine biosynthesis in the forest.}, journal={CURRENT PLANT BIOLOGY}, author={Peng, Qing-Zhong and Long, Hua and Du, Ci and Li, Jing and Xie, De-Yu}, year={2020}, month={Dec} } @article{noar_thomas_xie_carter_ma_daub_2019, title={A polyketide synthase gene cluster associated with the sexual reproductive cycle of the banana pathogen, Pseudocercospora fijiensis}, volume={14}, ISSN={1932-6203}, url={http://dx.doi.org/10.1371/journal.pone.0220319}, DOI={10.1371/journal.pone.0220319}, abstractNote={Disease spread of Pseudocercospora fijiensis, causal agent of the black Sigatoka disease of banana, depends on ascospores produced through the sexual reproductive cycle. We used phylogenetic analysis to identify P. fijiensis homologs (PKS8-4 and Hybrid8-3) to the PKS4 polyketide synthases (PKS) from Neurospora crassa and Sordaria macrospora involved in sexual reproduction. These sequences also formed a clade with lovastatin, compactin, and betaenone-producing PKS sequences. Transcriptome analysis showed that both the P. fijiensis Hybrid8-3 and PKS8-4 genes have higher expression in infected leaf tissue compared to in culture. Domain analysis showed that PKS8-4 is more similar than Hybrid8-3 to PKS4. pPKS8-4:GFP transcriptional fusion transformants showed expression of GFP in flask-shaped structures in mycelial cultures as well as in crosses between compatible and incompatible mating types. Confocal microscopy confirmed expression in spermagonia in leaf substomatal cavities, consistent with a role in sexual reproduction. A disruption mutant of pks8-4 retained normal pathogenicity on banana, and no differences were observed in growth, conidial production, and spermagonia production. GC-MS profiling of the mutant and wild type did not identify differences in polyketide metabolites, but did identify changes in saturated fatty acid methyl esters and alkene and alkane derivatives. To our knowledge, this is the first report of a polyketide synthase pathway associated with spermagonia.}, number={7}, journal={PLOS ONE}, publisher={Public Library of Science (PLoS)}, author={Noar, Roslyn D. and Thomas, Elizabeth and Xie, De-Yu and Carter, Morgan E. and Ma, Dongming and Daub, Margaret E.}, editor={Lespinet, OlivierEditor}, year={2019}, month={Jul}, pages={e0220319} } @article{judd_bagley_li_zhu_lei_yuzuak_ekelöf_pu_zhao_muddiman_et al._2019, title={Artemisinin Biosynthesis in Non-glandular Trichome Cells of Artemisia annua}, volume={12}, ISSN={1674-2052}, url={http://dx.doi.org/10.1016/J.MOLP.2019.02.011}, DOI={10.1016/J.MOLP.2019.02.011}, abstractNote={Artemisinin-based combination therapy (ACT) forms the first line of malaria treatment. However, the yield fluctuation of artemisinin has remained an unsolved problem in meeting the global demand for ACT. This problem is mainly caused by the glandular trichome (GT)-specific biosynthesis of artemisinin in all currently used Artemisia annua cultivars. Here, we report that non-GT cells of self-pollinated inbred A. annua plants can express the artemisinin biosynthetic pathway. Gene expression analysis demonstrated the transcription of six known pathway genes in GT-free leaves and calli of inbred A. annua plants. LC-qTOF-MS/MS analysis showed that these two types of GT-free materials produce artemisinin, artemisinic acid, and arteannuin B. Detailed IR-MALDESI image profiling revealed that these three metabolites and dihydroartemisinin are localized in non-GT cells of leaves of inbred A. annua plants. Moreover, we employed all the above approaches to examine artemisinin biosynthesis in the reported A. annua glandless (gl) mutant. The resulting data demonstrated that leaves of regenerated gl plantlets biosynthesize artemisinin. Collectively, these findings not only add new knowledge leading to a revision of the current dogma of artemisinin biosynthesis in A. annua but also may expedite innovation of novel metabolic engineering approaches for high and stable production of artemisinin in the future.}, number={5}, journal={Molecular Plant}, publisher={Elsevier BV}, author={Judd, Rika and Bagley, M. Caleb and Li, Mingzhuo and Zhu, Yue and Lei, Caiyan and Yuzuak, Seyit and Ekelöf, Måns and Pu, Gaobin and Zhao, Xiting and Muddiman, David C. and et al.}, year={2019}, month={May}, pages={704–714} } @article{li_ji_xi_xie_su_2019, title={Creation of elite growth and development features in PAP1-programmed red Nicotiana tabacum Xanthi via overexpression of synthetic geranyl pyrophosphate synthase genes}, volume={39}, ISSN={1380-3743 1572-9788}, url={http://dx.doi.org/10.1007/s11032-019-0968-5}, DOI={10.1007/s11032-019-0968-5}, number={4}, journal={Molecular Breeding}, publisher={Springer Science and Business Media LLC}, author={Li, Gui and Ji, Xiaoming and Xi, Jing and Xie, De-Yu and Su, Xiaohua}, year={2019}, month={Apr} } @article{liu_zhang_xie_franks_xiang_2019, title={Functional characterization of Terminal Flower1 homolog in Cornus canadensis by genetic transformation}, volume={38}, ISSN={0721-7714 1432-203X}, url={http://dx.doi.org/10.1007/s00299-019-02369-2}, DOI={10.1007/s00299-019-02369-2}, abstractNote={TFL1homologCorcanTFL1suppresses the initiation of inflorescence development and regulates the inflorescence morphology inCornus canadensis. In flowering plants, there is a wide range of variation of inflorescence morphology. Despite the ecological and evolutionary importance, efforts devoted to the evolutionary study of the genetic basis of inflorescence morphology are far fewer compared to those on flower development. Our previous study on gene expression patterns suggested a CorTFL1-CorAP1 based model for the evolution of determinate umbels, heads, and mini dichasia from elongated inflorescences in Cornus. Here, we tested the function of CorcanTFL1 in regulating inflorescence development in Cornus canadensis through Agrobacterium-mediated transformation. We showed that transgenic plants overexpressing CorcanTFL1 displayed delayed or suppressed inflorescence initiation and development and extended periods of vegetative growth. Transgenic plants within which CorcanTFL1 had been down-regulated displayed earlier emergence of inflorescence and a reduction of bract and inflorescence sizes, conversions of leaves to bracts and axillary leaf buds to small inflorescences at the uppermost node bearing the inflorescence, or phyllotaxy changes of inflorescence branches and leaves from decussate opposite to spirally alternate. These observations support an important role of CorcanTFL1 in determining flowering time and the morphological destinies of leaves and buds at the node bearing the inflorescence. The evidence is in agreement with the predicted function of CorTFL1 from the gene expression model, supporting a key role of CorTFL1 in the evolutionary divergence of inflorescence forms in Cornus.}, number={3}, journal={Plant Cell Reports}, publisher={Springer Science and Business Media LLC}, author={Liu, Xiang and Zhang, Jian and Xie, Deyu and Franks, Robert G. and Xiang, Qiu-Yun (Jenny)}, year={2019}, month={Jan}, pages={333–343} } @article{wang_liu_zhang_wang_hou_zhao_jiang_yu_tan_wang_et al._2020, title={Functional demonstration of plant flavonoid carbocations proposed to be involved in the biosynthesis of proanthocyanidins}, volume={101}, ISSN={["1365-313X"]}, DOI={10.1111/tpj.14515}, abstractNote={SummaryThe plant flavonoid dogma proposes that labile plant flavonoid carbocations (PFCs) play vital roles in the biosynthesis of proanthocyanidins (PAs). However, whether PFCs exist in plants and how PFCs function remain unclear. Here, we report the use of an integrative strategy including enzymatic assays, mutant analysis, metabolic engineering, isotope labeling and metabolic profiling to capture PFCs and demonstrate their functions. In anthocyanidin reductase (ANR) assays, an (−)‐epicatechin conjugate was captured in protic polar nucleophilic methanol alone or methanol−HCl extracts. Tandem mass spectrum (MS/MS) analysis characterized this compound as an (−)‐epicatechin‐4‐O‐methyl (EOM) ether, which resulted from (−)‐epicatechin carbocation and the methyl group of methanol. Acid‐based catalysis of procyanidin B2 and B3 produced four compounds, which were annotated as two EOM and two (+)‐catechin‐4‐O‐methyl (COM) ethers. Metabolic profiling of seven PA pathway mutants showed an absence or reduction of two EOM ether isomers in seeds. Camellia sinensis ANRa (CsANRa), leucoanthocyanidin reductase c (CsLARc), and CsMYB5b (a transcription factor) were independently overexpressed for successful PA engineering in tobacco. The EOM ether was remarkably increased in CsANRa and CsMYB5b transgenic flowers. Further metabolic profiling for eight green tea tissues revealed two EOM and two COM ethers associated with PA biosynthesis. Moreover, an incubation of (−)‐epicatechin or (+)‐catechin with epicatechin carbocation in CsANRa transgenic flower extracts formed dimeric procyanidin B1 or B2, demonstrating the role of flavan‐3‐ol carbocation in the formation of PAs. Taken together, these findings indicated that flavan‐3‐ol carbocations exist in extracts and are involved in the biosynthesis of PAs of plants.}, number={1}, journal={PLANT JOURNAL}, author={Wang, Peiqiang and Liu, Yajun and Zhang, Lingjie and Wang, Wenzhao and Hou, Hua and Zhao, Yue and Jiang, Xiaolan and Yu, Jie and Tan, Huarong and Wang, Yunsheng and et al.}, year={2020}, month={Jan}, pages={18–36} } @article{ma_gandra_manoharlal_la hovary_xie_2019, title={Untargeted Metabolomics of Nicotiana tabacum Grown in United States and India Characterizes the Association of Plant Metabolomes With Natural Climate and Geography}, volume={10}, ISSN={1664-462X}, url={http://dx.doi.org/10.3389/fpls.2019.01370}, DOI={10.3389/fpls.2019.01370}, abstractNote={Climate change and geography affect all the living organisms. To date, the effects of climate and geographical factors on plant metabolome largely remain open for worldwide and local investigations. In this study, we designed field experiments with tobacco (Nicotiana tabacum) in India versus USA and used untargeted metabolomics to understand the association of two weather factors and two different continental locations with respect to tobacco metabolism. Field research stations in Oxford, North Carolina, USA, and Rajahmundry, Andhra Pradesh India were selected to grow a commercial tobacco genotype (K326) for 2 years. Plant growth, field management, and leaf curing followed protocols standardized for tobacco cultivation. Gas chromatography–mass spectrometry based unbiased profiling annotated 171 non-polar and 225 polar metabolites from cured tobacco leaves. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) showed that two growing years and two field locations played primary and secondary roles affecting metabolite profiles, respectively. PCA and Pearson analysis, which used nicotine, 11 other groups of metabolites, two locations, temperatures, and precipitation, revealed that in North Carolina, temperature changes were positively associated with the profiles of sesquiterpenes, diterpenes, and triterpenes, but negatively associated with the profiles of nicotine, organic acids of tricarboxylic acid, and sugars; in addition, precipitation was positively associated with the profiles of triterpenes. In India, temperature was positively associated with the profiles of benzenes and polycyclic aromatic hydrocarbons, but negatively associated with the profiles of amino acids and sugar. Further comparative analysis revealed that nicotine levels were affected by weather conditions, nevertheless, its trend in leaves was independent of two geographical locations and weather changes. All these findings suggested that climate and geographical variation significantly differentiated the tobacco metabolism.}, journal={Frontiers in Plant Science}, publisher={Frontiers Media SA}, author={Ma, Dong-Ming and Gandra, Saiprasad V. S. and Manoharlal, Raman and La Hovary, Christophe and Xie, De-Yu}, year={2019}, month={Oct} } @article{ekelöf_garrard_judd_rosen_xie_kashuba_muddiman_2018, title={Evaluation of Digital Image Recognition Methods for Mass Spectrometry Imaging Data Analysis}, volume={29}, ISSN={1044-0305 1879-1123}, url={http://dx.doi.org/10.1007/S13361-018-2073-0}, DOI={10.1007/S13361-018-2073-0}, abstractNote={Analyzing mass spectrometry imaging data can be laborious and time consuming, and as the size and complexity of datasets grow, so does the need for robust automated processing methods. We here present a method for comprehensive, semi-targeted discovery of molecular distributions of interest from mass spectrometry imaging data, using widely available image similarity scoring algorithms to rank images by spatial correlation. A fast and powerful batch search method using a MATLAB implementation of structural similarity (SSIM) index scoring with a pre-selected reference distribution is demonstrated for two sample imaging datasets, a plant metabolite study using Artemisia annua leaf, and a drug distribution study using maraviroc-dosed macaque tissue. Graphical Abstract ᅟ.}, number={12}, journal={Journal of The American Society for Mass Spectrometry}, publisher={Springer Science and Business Media LLC}, author={Ekelöf, Måns and Garrard, Kenneth P. and Judd, Rika and Rosen, Elias P. and Xie, De-Yu and Kashuba, Angela D. M. and Muddiman, David C.}, year={2018}, month={Oct}, pages={2467–2470} } @article{yuzuak_ballington_xie_2018, title={HPLC-qTOE-MS/MS-Based Profiling of Flavan-3-ols and Dimeric Proanthocyanidins in Berries of Two Muscadine Grape Hybrids FLH 13-11 and FLH 17-66}, volume={8}, ISSN={["2218-1989"]}, url={http://www.mdpi.com/2218-1989/8/4/57}, DOI={10.3390/metabo8040057}, abstractNote={FLH 13-11 FL and FLH 17-66 FL are two interspecific hybrid varieties of muscadine grape resulting from the cross of Vitis munsoniana (Simpson) ex Munson and V. rotundifolia. However, profiles of flavan-3-ols and proanthocyanidins in these two hybrids have not been characterized. Herein, we report the use of high-performance liquid chromatography-quadrupole, time-of-flight, tandem mass spectrometry (HPLC-qTOF-MS/MS) to characterize these two groups of metabolites in berries. Ripe berries collected from two consecutive cropping years were used to extract metabolites. Metabolites were ionized using the negative mode. Collision-induced dissociation was performed to fragmentize ions to obtain feature fragment profiles. Based on standards, MS features, and fragments resulted from MS/MS, four flavan-3-ol aglycones, 18 gallated or glycosylated conjugates, and eight dimeric procyanidins, were annotated from berry extracts. Of these 30 metabolites, six are new methylated flavan-3-ol gallates. Furthermore, comparative profiling analysis showed obvious effects of each cultivar on the composition these 30 metabolites, indicating that genotypes control biosynthesis. In addition, cropping seasons altered profiles of these metabolites, showing effects of growing years on metabolic composition. These data are informative to enhance the application of the two cultivars in grape and wine industries in the future.}, number={4}, journal={METABOLITES}, publisher={MDPI AG}, author={Yuzuak, Seyit and Ballington, James and Xie, De-Yu}, year={2018}, month={Dec} } @article{zhu_peng_li_xie_2018, title={Molecular Cloning and Functional Characterization of a Dihydroflavonol 4-Reductase from Vitis bellula}, volume={23}, ISSN={["1420-3049"]}, url={http://www.mdpi.com/1420-3049/23/4/861}, DOI={10.3390/molecules23040861}, abstractNote={Vitis bellula is a new grape crop in southern China. Berries of this species are rich in antioxidative anthocyanins and proanthocyanidins. This study reports cloning and functional characterization of a cDNA encoding a V. bellula dihydroflavonol reductase (VbDFR) involved in the biosynthesis of anthocyanins and proanthocyanidins. A cDNA including 1014 bp was cloned from young leaves and its open reading frame (ORF) was deduced encoding 337 amino acids, highly similar to V. vinifera DFR (VvDFR). Green florescence protein fusion and confocal microscopy analysis determined the cytosolic localization of VbDFR in plant cells. A soluble recombinant VbDFR was induced and purified from E. coli for enzyme assay. In the presence of NADPH, the recombinant enzyme catalyzed dihydrokaempferol (DHK) and dihydroquercetin (DHQ) to their corresponding leucoanthocyanidins. The VbDFR cDNA was introduced into tobacco plants via Agrobacterium-mediated transformation. The overexpression of VbDFR increased anthocyanin production in flowers. Anthocyanin hydrolysis and chromatographic analysis revealed that transgenic flowers produced pelargonidin and delphinidin, which were not detected in control flowers. These data demonstrated that the overexpression of VbDFR produced new tobacco anthocyanidins. In summary, all data demonstrate that VbDFR is a useful gene to provide three types of substrates for metabolic engineering of anthocyanins and proanthocyanidins in grape crops and other crops.}, number={4}, journal={MOLECULES}, publisher={MDPI AG}, author={Zhu, Yue and Peng, Qingzhong and Li, Kegang and Xie, De-Yu}, year={2018}, month={Apr} } @article{ma_xu_alejos-gonzalez_wang_yang_judd_xie_2018, title={Overexpression of Artemisia annua Cinnamyl Alcohol Dehydrogenase Increases Lignin and Coumarin and Reduces Artemisinin and Other Sesquiterpenes}, volume={9}, ISSN={["1664-462X"]}, DOI={10.3389/fpls.2018.00828}, abstractNote={Artemisia annua is the only medicinal crop that produces artemisinin for malarial treatment. Herein, we describe the cloning of a cinnamyl alcohol dehydrogenase (AaCAD) from an inbred self-pollinating (SP) A. annua cultivar and its effects on lignin and artemisinin production. A recombinant AaCAD was purified via heterogeneous expression. Enzyme assays showed that the recombinant AaCAD converted p-coumaryl, coniferyl, and sinapyl aldehydes to their corresponding alcohols, which are key intermediates involved in the biosynthesis of lignin. Km, Vmax, and Vmax/Km values were calculated for all three substrates. To characterize its function in planta, AaCAD was overexpressed in SP plants. Quantification using acetyl bromide (AcBr) showed significantly higher lignin contents in transgenics compared with wild-type (WT) plants. Moreover, GC-MS-based profiling revealed a significant increase in coumarin contents in transgenic plants. By contrast, HPLC-MS analysis showed significantly reduced artemisinin contents in transgenics compared with WT plants. Furthermore, GC-MS analysis revealed a decrease in the contents of arteannuin B and six other sesquiterpenes in transgenic plants. Confocal microscopy analysis showed the cytosolic localization of AaCAD. These data demonstrate that AaCAD plays a dual pathway function in the cytosol, in which it positively enhances lignin formation but negatively controls artemisinin formation. Based on these data, crosstalk between these two pathways mediated by AaCAD catalysis is discussed to understand the metabolic control of artemisinin biosynthesis in plants for high production.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Ma, Dongming and Xu, Chong and Alejos-Gonzalez, Fatima and Wang, Hong and Yang, Jinfen and Judd, Rika and Xie, De-Yu}, year={2018}, month={Jun} } @article{borghi_xie_2018, title={Cloning and characterization of a monoterpene synthase gene from flowers of Camelina sativa}, volume={247}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-017-2801-x}, abstractNote={CsTPS1 encodes for a monoterpene synthase that contributes to the emission of a blend of volatile compounds emitted from flowers of Camelina sativa. The work describes the in vitro characterization of a monoterpene synthase and its regulatory region that we cloned from Camelina sativa (Camelina). Here, we named this gene as C. sativa terpene synthase 1 (CsTPS1). In vitro experiments performed with the CsTPS1 protein after expression and purification from Escherichia coli (E. coli) showed production of a blend of monoterpene volatile organic compounds, of which the emission was also detected in the floral bouquet of wild-type Camelina plants. Quantitative-PCR measurements revealed a high abundance of CsTPS1 transcripts in flowers and experiments performed with the GUS reporter showed high CsTPS1 expression in the pistil, in the cells of the wall of the ovary and in the stigma. Subcellular localization of the CsTPS1 protein was investigated with a GFP reporter construct that showed expression in plastids. The CsTPS1 gene identified in this study belongs to a mid-size family of 60 genes putatively codifying for TPS enzymes. This enlarged family of TPS genes suggests that Camelina has the structural framework for the production of terpenes and other secondary metabolites of relevance for the consumers.}, number={2}, journal={PLANTA}, author={Borghi, Monica and Xie, De-Yu}, year={2018}, month={Feb}, pages={443–457} } @article{borghi_xie_2018, title={Cloning and characterization of a monoterpene synthase gene from flowers of Camelina sativa (vol 247, pg 443, 2018)}, volume={247}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-017-2810-9}, abstractNote={In the original publication, the order of figures and citations was incorrect. The corrections are listed below.}, number={1}, journal={PLANTA}, author={Borghi, Monica and Xie, De-Yu}, year={2018}, month={Jan}, pages={287–288} } @article{li_ji_swantko_xie_2017, title={Compartmentalized Overexpression of a Synthetic Geranyl Pyrophosphate Synthase and Its Regulation on Plant Growth and Metabolism}, volume={53}, number={Suppl. 1}, journal={In Vitro Cellular & Developmental Biology - Animal}, author={Li, Gui and Ji, Xiaoming and Swantko, Sarah and Xie, De-Yu}, year={2017}, pages={54–55} } @article{li_li_guo_gong_pang_jiang_liu_jiang_zhao_wang_et al._2017, title={Functional Characterization of Tea (Camellia sinensis) MYB4a Transcription Factor Using an Integrative Approach}, volume={8}, ISSN={["1664-462X"]}, DOI={10.3389/fpls.2017.00943}, abstractNote={Green tea (Camellia sinensis, Cs) abundantly produces a diverse array of phenylpropanoid compounds benefiting human health. To date, the regulation of the phenylpropanoid biosynthesis in tea remains to be investigated. Here, we report a cDNA isolated from leaf tissues, which encodes a R2R3-MYB transcription factor. Amino acid sequence alignment and phylogenetic analysis indicate that it is a member of the MYB4-subgroup and named as CsMYB4a. Transcriptional and metabolic analyses show that the expression profile of CsMYB4a is negatively correlated to the accumulation of six flavan-3-ols and other phenolic acids. GFP fusion analysis shows CsMYB4a’s localization in the nucleus. Promoters of five tea phenylpropanoid pathway genes are isolated and characterized to contain four types of AC-elements, which are targets of MYB4 members. Interaction of CsMYB4a and five promoters shows that CsMYB4a decreases all five promoters’ activity. To further characterize its function, CsMYB4a is overexpressed in tobacco plants. The resulting transgenic plants show dwarf, shrinking and yellowish leaf, and early senescence phenotypes. A further genome-wide transcriptomic analysis reveals that the expression levels of 20 tobacco genes involved in the shikimate and the phenylpropanoid pathways are significantly downregulated in transgenic tobacco plants. UPLC-MS and HPLC based metabolic profiling reveals significant reduction of total lignin content, rutin, chlorogenic acid, and phenylalanine in CsMYB4a transgenic tobacco plants. Promoter sequence analysis of the 20 tobacco genes characterizes four types of AC-elements. Further CsMYB4a-AC element and CsMYB4a-promoter interaction analyses indicate that the negative regulation of CsMYB4a on the shikimate and phenylpropanoid pathways in tobacco is via reducing promoter activity. Taken together, all data indicate that CsMYB4a negatively regulates the phenylpropanoid and shikimate pathways. Highlight: A tea (Camellia sinensis) MYB4a is characterized to encode a R2R3-MYB transcription factor. It is shown to repressively control the phenylpropanoid and shikimate pathway.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Li, Mingzhuo and Li, Yanzhi and Guo, Lili and Gong, Niandi and Pang, Yongzheng and Jiang, Wenbo and Liu, Yajun and Jiang, Xiaolan and Zhao, Lei and Wang, Yunsheng and et al.}, year={2017}, month={Jun} } @article{zhao_jiang_qian_wang_xie_gao_xia_2017, title={Metabolic Characterization of the Anthocyanidin Reductase Pathway Involved in the Biosynthesis of Flavan-3-ols in Elite Shuchazao Tea (Camellia sinensis) Cultivar in the Field}, volume={22}, url={http://www.mdpi.com/1420-3049/22/12/2241}, DOI={10.3390/molecules22122241}, abstractNote={Anthocyanidin reductase (ANR) is a key enzyme in the ANR biosynthetic pathway of flavan-3-ols and proanthocyanidins (PAs) in plants. Herein, we report characterization of the ANR pathway of flavan-3-ols in Shuchazao tea (Camellia sinesis), which is an elite and widely grown cultivar in China and is rich in flavan-3-ols providing with high nutritional value to human health. In our study, metabolic profiling was preformed to identify two conjugates and four aglycones of flavan-3-ols: (−)-epigallocatechin-gallate [(−)-EGCG], (−)-epicatechin-gallate [(−)-ECG], (−)-epigallocatechin [(−)-EGC], (−)-epicatechin [(−)-EC], (+)-catechin [(+)-Ca], and (+)-gallocatechin [(+)-GC], of which (−)-EGCG, (−)-ECG, (−)-EGC, and (−)-EC accounted for 70–85% of total flavan-3-ols in different tissues. Crude ANR enzyme was extracted from young leaves. Enzymatic assays showed that crude ANR extracts catalyzed cyanidin and delphinidin to (−)-EC and (−)-Ca and (−)-EGC and (−)-GC, respectively, in which (−)-EC and (−)-EGC were major products. Moreover, two ANR cDNAs were cloned from leaves, namely CssANRa and CssANRb. His-Tag fused recombinant CssANRa and CssANRb converted cyanidin and delphinidin to (−)-EC and (−)-Ca and (−)-EGC and (−)-GC, respectively. In addition, (+)-EC was observed from the catalysis of recombinant CssANRa and CssANRb. Further overexpression of the two genes in tobacco led to the formation of PAs in flowers and the reduction of anthocyanins. Taken together, these data indicate that the majority of leaf flavan-3-ols in Shuchazao’s leaves were produced from the ANR pathway.}, number={12}, journal={Molecules}, publisher={MDPI AG}, author={Zhao, Lei and Jiang, Xiao-Lan and Qian, Yu-Mei and Wang, Pei-Qiang and Xie, De-Yu and Gao, Li-Ping and Xia, Tao}, year={2017}, month={Dec}, pages={2241} } @article{zhao_jiang_qian_wang_xie_gao_xia_2017, title={Metabolic characterization of the anthocyanidin reductase pathway involved in the biosynthesis of flavan-3-ols in Elite Shuchazao Tea (Camellia sinensis) cultivar in the field}, volume={22}, number={12}, journal={Molecules}, author={Zhao, L. and Jiang, X. L. and Qian, Y. M. and Wang, P. Q. and Xie, D. Y. and Gao, L. P. and Xia, T.}, year={2017} } @article{li_xi_ji_li_xie_2018, title={Non-plastidial expression of a synthetic insect geranyl pyrophosphate synthase effectively increases tobacco plant biomass}, volume={221}, ISSN={["1618-1328"]}, DOI={10.1016/j.jplph.2017.12.014}, abstractNote={Designing effective synthetic genes of interest is a fundamental step in plant synthetic biology for biomass. Geranyl pyrophosphate (diphosphate) synthase (GPPS) catalyzes a bottleneck step toward terpenoid metabolism. We previously designed and synthesized a plant (Arabidopsis thaliana)-insect (Myzus persicae, Mp) GPPS- human influenza hemagglutinin (HA) cDNA, namely PTP-MpGPPS-HA (or PTP-sMpGPPS-HA, s: synthetic), to localize the protein in plastids and improve plant biomass. To better understand the effects of different subcellular localizations on plant performance, herein we report PTP-sMpGPPS-HA re-design to synthesize a new MpGPPS-HA cDNA, namely sMpGPPS-HA, to express a non-plastidial sMpGPPS-HA protein. The sMpGPPS-HA cDNA driven by a 2 × S 35S promoter was introduced into Nicotiana tabacum Xanthi. PTP-MpGPPS-HA and PMDC84 vector transgenic plants were also generated as positive and negative controls, respectively. Eighteen to twenty transgenic T0 lines were generated for each sMpGPPS-HA, PTP-sMpGPPS-HA, and PMDC84. Transcriptional genotyping analysis demonstrated the expression of sMpGPPS-HA in transgenic plants. Confocal microscopy analysis of transgenic progeny demonstrated the non-plastidial localization of sMpGPPS-HA. Growth of T1 transgenic and wild-type control plants showed that the expression of sMpGPPS-HA effectively increased plant height by 50-80%, leaf numbers and sizes, and dry biomass by 60-80%. Calculation of the vegetative growth rates showed that the expression of sMpGPPS-HA increased plant height each week. Moreover, sMpGPPS-HA expression promoted early flowering and reduced leaf carotenoid levels. In conclusion, non-plastidial expression of the novel sMpGPPS-HA was effective for improving tobacco growth and biomass. Our data indicate that research examining different subcellular localizations facilitates a better understanding of in planta functions of proteins encoded by synthetic cDNAs.}, journal={JOURNAL OF PLANT PHYSIOLOGY}, author={Li, Gui and Xi, Jing and Ji, Xiaoming and Li, Ming-Zhuo and Xie, De-Yu}, year={2018}, month={Feb}, pages={144–155} } @article{ma_li_zhu_xie_2017, title={Overexpression and Suppression of Artemisia annua 4-Hydroxy-3-Methylbut-2-enyl Diphosphate Reductase 1 Gene (AaHDR1) Differentially Regulate Artemisinin and Terpenoid Biosynthesis}, volume={8}, ISSN={["1664-462X"]}, DOI={10.3389/fpls.2017.00077}, abstractNote={4-Hydroxy-3-methylbut-2-enyl diphosphate reductase (HDR) catalyzes the last step of the 2-C-methyl-D-erythritol 4- phosphate (MEP) pathway to synthesize isopentenyl pyrophosphate (IPP) and dimethylallyl diphosphate (DMAPP). To date, little is known regarding effects of an increase or a decrease of a HDR expression on terpenoid and other metabolite profiles in plants. In our study, an Artemisia annua HDR cDNA (namely AaHDR1) was cloned from leaves. Expression profiling showed that it was highly expressed in leaves, roots, stems, and flowers with different levels. Green florescence protein fusion and confocal microscope analyses showed that AaHDR1 was localized in chloroplasts. The overexpression of AaHDR1 increased contents of artemisinin, arteannuin B and other sesquiterpenes, and multiple monoterpenes. By contrast, the suppression of AaHDR1 by anti-sense led to opposite results. In addition, an untargeted metabolic profiling showed that the overexpression and suppression altered non-polar metabolite profiles. In conclusion, the overexpression and suppression of AaHDR1 protein level in plastids differentially affect artemisinin and other terpenoid biosynthesis, and alter non-polar metabolite profiles of A. annua. Particularly, its overexpression leading to the increase of artemisinin production is informative to future metabolic engineering of this antimalarial medicine.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Ma, Dongming and Li, Gui and Zhu, Yue and Xie, De-Yu}, year={2017}, month={Jan} } @article{rossi_borghi_yang_xie_2017, title={Overexpression of Populus×canescens isoprene synthase gene in Camelina sativa leads to alterations in its growth and metabolism}, volume={215}, ISSN={0176-1617}, url={http://dx.doi.org/10.1016/J.JPLPH.2017.06.005}, DOI={10.1016/J.JPLPH.2017.06.005}, abstractNote={Isoprene (2-methyl-1,3-butadiene) is a hemiterpene molecule. It has been estimated that the plant kingdom emits 500-750 million tons of isoprene in the environment, half of which results from tropical broadleaf trees and the remainder from shrubs. Camelina (Camelina sativa (L.) Crantz) is an emerging bioenergy plant for biodiesel. In this study, we characterized isoprene formation following a diurnal/nocturnal cycle in wild-type Camelina plants. To understand the potential effects of isoprene emission on this herbaceous plant, a gray poplar Populus×canescens isoprene synthase gene (PcISPS) was overexpressed in Camelina. Transgenic plants showed increased isoprene production, and the emissions were characterized by a diurnal/nocturnal cycle. Measurements of the expression of six genes of the plastidial 2-C-methyl-d-erythriol-4-phosphate (MEP) pathway revealed that the expression patterns of three key genes were associated with isoprene formation dynamics in the three genotypic plants. Conversely, dissimilar gene expression levels existed in different genotypes, indicating that dynamics and variations occurred among plants. Moreover, transgenic plants grew shorter and developed smaller leaves than the wild-type and empty vector control transgenic plants. Photosynthetic analysis showed that the CO 2 assimilation rate, intracellular CO 2 concentration, mesophyll conductance and contents of chlorophylls a and b were similar among PcISPS transgenic, empty-vector control transgenic, and wild-type plants, indicating that the transgene did not negatively affect photosynthesis. Based on these results, we suggest that the reduced biomass was likely a trade-off consequence of the increased isoprene emission.}, journal={Journal of Plant Physiology}, publisher={Elsevier BV}, author={Rossi, Lorenzo and Borghi, Monica and Yang, Jinfen and Xie, De-Yu}, year={2017}, month={Aug}, pages={122–131} } @article{ma_li_alejos-gonzalez_zhu_xue_wang_zhang_li_ye_wang_et al._2017, title={Overexpression of a type-I isopentenyl pyrophosphate isomerase of Artemisia annua in the cytosol leads to high arteannuinB production and artemisinin increase}, volume={91}, DOI={10.1111/tpj.13583}, abstractNote={SummaryWe recently characterized a gene–terpene network that is associated with artemisinin biosynthesis in self‐pollinated (SP) Artemisia annua, an effective antimalarial plant. We hypothesize that an alteration of gene expression in the network may improve the production of artemisinin and its precursors. In this study, we cloned an isopentenyl pyrophosphate isomerase (IPPI) cDNA, AaIPPI1, from Artemisia annua (Aa). The full‐length cDNA encodes a type‐I IPPI containing a plastid transit peptide (PTP) at its amino terminus. After the removal of the PTP, the recombinant truncated AaIPPI1 isomerized isopentenyl pyrophosphate (IPP) to dimethyl allyl pyrophosphate (DMAPP) and vice versa. The steady‐state equilibrium ratio of IPP/DMAPP in the enzymatic reactions was approximately 1:7. The truncated AaIPPI1 was overexpressed in the cytosol of the SP A. annua variety. The leaves of transgenic plants produced approximately 4% arteannuin B (g g−1, dry weight, dw) and 0.17–0.25% artemisinin (g g−1, dw), the levels of which were significantly higher than those in the leaves of wild‐type plants. In addition, transgenic plants showed an increase in artemisinic acid production of more than 1% (g g−1, dw). In contrast, isoprene formation was significantly reduced in transgenic plants. These results provide evidence that overexpression of AaIPPI1 in the cytosol can lead to metabolic alterations of terpenoid biosynthesis, and show that these transgenic plants have the potential to yield high production levels of arteannuin B as a new precursor source for artemisinin.}, number={3}, journal={Plant Journal}, author={Ma, D. M. and Li, G. and Alejos-Gonzalez, F. and Zhu, Y. and Xue, Z. and Wang, A. M. and Zhang, H. and Li, X. and Ye, H. C. and Wang, H. and et al.}, year={2017}, pages={466–479} } @article{zhu_xie_2017, title={Red and White PAP1-controlled Arabidopsis Cells Are Dependent Upon TT8}, volume={53}, number={Supplement 1}, journal={In Vitro Cellular & Developmental Biology - Animal}, author={Zhu, Yue and Xie, De-Yu}, year={2017}, pages={S51–S51} } @article{xie_2017, title={Regulation of Anthocyanin Biosynthesis in the WD40-bHLH-MYB Complex-Programmed Arabidopsis Cells}, volume={53}, number={Suppl 1}, journal={In Vitro Cellular & Developmental Biology - Animal}, author={Xie, De-Yu}, year={2017}, pages={13–13} } @article{liu_zhang_abuahmad_franks_xie_xiang_2016, title={Analysis of two TFL1 homologs of dogwood species (Cornus L.) indicates functional conservation in control of transition to flowering}, volume={243}, ISSN={0032-0935, 1432-2048}, url={http://link.springer.com/10.1007/s00425-016-2466-x}, DOI={10.1007/s00425-016-2466-x}, abstractNote={Two TFL1 -like genes, CorfloTFL1 and CorcanTFL1 cloned from Cornus florida and C. canadensis, function in regulating the transition to reproductive development in Arabidopsis. TERMINAL FLOWER 1 (TFL1) is known to regulate inflorescence development in Arabidopsis thaliana and to inhibit the transition from a vegetative to reproductive phase within the shoot apical meristem. Despite the importance, TFL1 homologs have been functionally characterized in only a handful eudicots. Here we report the role of TFL1 homologs of Cornus L. in asterid clade of eudicots. Two TFL1-like genes, CorfloTFL1 and CorcanTFL1, were cloned from Cornus florida (a tree) and C. canadensis (a subshrub), respectively. Both are deduced to encode proteins of 175 amino acids. The amino acid sequences of these two Cornus TFL1 homologs share a high similarity to Arabidopsis TFL1 and phylogenetically more close to TFL1 paralogous copy ATC (Arabidopsis thaliana CENTRORADIALIS homologue). Two genes are overexpressed in wild-type and tfl1 mutant plants of A. thaliana. The over-expression of each gene in wild-type Arabidopsis plants results in delaying flowering time, increase of plant height and cauline and rosette leaf numbers, excessive shoot buds, and secondary inflorescence branches. The over-expression of each gene in the tfl1 mutant rescued developmental defects, such as the early determinate inflorescence development, early flowering time, and other vegetative growth defects, to normal phenotypes of wild-type plants. These transgenic phenotypes are inherited in progenies. All data indicate that CorfloTFL1 and CorcanTFL1 have conserved the ancestral function of TFL1 and CEN regulating flowering time and inflorescence determinacy.}, number={5}, journal={Planta}, publisher={Springer Science and Business Media LLC}, author={Liu, Xiang and Zhang, Jian and Abuahmad, Ahmad and Franks, Robert G. and Xie, De-Yu and Xiang, Qiu-Yun}, year={2016}, month={May}, pages={1129–1141} } @article{xie_2016, title={Artemisia annua, artemisinin, and the Nobel Prize: beauty of natural products and educational significance}, volume={61}, ISSN={["2095-9281"]}, DOI={10.1007/s11434-015-0989-3}, abstractNote={In the beautiful evening on December 10, 2015 (local time in Sweden), the annual Nobel Prize ceremony was hosted by the Swedish Royal Family at Stockholm’s City Hall, Stockholm, Sweden. Congratulations to all ten new Nobel Prize Laureates and thanks them for using their brightest talents to make our world better! The 2015 Nobel Prize in Physiology or Medicine was awarded to Professors Youyou Tu, William C. Campbell, and Satoshi Omura for their inventions in novel therapies using natural product medicines that help humankind fight against diseases caused by two types of severe endemic parasites, and save millions of people’s lives [1]. The Nobel Prize awarded to Professor Tu was to recognize and appreciate her pioneering discovery of artemisinin from Artemisia annua and clinical innovation in fighting against malaria, one of the top three diseases leading to the loss of people’s life [2]. Since the announcement of the award in October 2015, hundreds of praising reports have been published in scientific journals and the media to introduce and appreciate Professor Tu’s scientific achievements and medicinal contributions to global health [1–3]. Particularly, people in China from the top administers of the central Chinese government to elementary school students have been excited to applaud, discuss, and comment on her greatest research achievements that benefit human health. Here, as a university’s teacher in Medicinal Plants, Phytochemistry, and Plant Natural Products, I would like to specifically highlight the beauty and the educational significance of the Nobel Prize awarded to Professor Tu. To me, the 2015 Nobel Prize in Medicines is the most magnificent award. Not only is it awarded to Professor Tu for her invention of the novel therapy treating malaria, but also it is awarded to her collaborative teams for their research efforts in the development of artemisinin for malaria treatment. After the Nobel Prize committee announced her as one of the winners of the award, Professor Tu has always humbly expressed that the honor should also belong to her entire team and collaborators. In her interviews and speeches, she always respectfully appreciated her team members and collaborators for their supportive collaboration 40 years ago. The discovery of artemisinin was carried out during the Cultural Revolution period in China [4]. The Chinese government funded a secret project coded as ‘‘Project 523’’ to treat severely lethal malaria associated with the Vietnam War. The code number means the date ‘‘the 23rd of May, 1967,’’ when the project was officially launched. In addition, my personal opinion is that the other main reason was the extremely severe endemic malaria (Da Bai Zi, in Chinese Pin Yin) in south China during 1960s and 1970s. For example, I was one of the malarial patients infected by Plasmodium falciparum in 1977. I also remember that many people in my village were victims of malaria. In 1967, a group of Chinese scientists were funded to develop effective medicines from Traditional Chinese Medicines to fight against lethal malaria. At that time, young Tu was one of the junior scientists in the entire national team. Tu and her team endeavored to screen hundreds of different Chinese medicinal plants. In October of 1971, Tu led her team for the first time to demonstrate the medicinal activity of A. annua in treating malaria [5]. By the end of 1972, she and SPECIAL TOPIC: Advances in Artemisinin Study}, number={1}, journal={SCIENCE BULLETIN}, author={Xie, De-Yu}, year={2016}, month={Jan}, pages={42–44} } @article{xie_ma_judd_jones_2016, title={Artemisinin biosynthesis in Artemisia annua and metabolic engineering: questions, challenges, and perspectives}, volume={15}, ISSN={["1572-980X"]}, url={https://doi.org/10.1007/s11101-016-9480-2}, DOI={10.1007/s11101-016-9480-2}, number={6}, journal={PHYTOCHEMISTRY REVIEWS}, publisher={Springer Nature}, author={Xie, De-Yu and Ma, Dong-Ming and Judd, Rika and Jones, Ashley Loray}, year={2016}, month={Dec}, pages={1093–1114} } @article{xie_2016, title={Metabolic Network Based Regulation of Artemisinin Biosynthesis for Anti-malarial Medicine}, volume={52}, number={Supplement}, journal={In Vitro Cellular & Developmental Biology - Plant}, author={Xie, De-Yu}, year={2016}, pages={S24} } @article{he_li_lawson_xie_2017, title={Metabolic engineering of anthocyanins in dark tobacco varieties}, volume={159}, ISSN={["1399-3054"]}, DOI={10.1111/ppl.12475}, abstractNote={In this study, we investigate the metabolic engineering of anthocyanins in two dark tobacco crops (Narrow Leaf Madole and KY171) and evaluate the effects on physiological features of plant photosynthesis. Arabidopsis PAP1 (production of anthocyanin pigment 1) gene (AtPAP1) encodes a R2R3‐type MYB transcript factor that is a master component of regulatory complexes controlling anthocyanin biosynthesis. AtPAP1 was introduced to Narrow Leaf Madole and KY171 plants. Multiple transgenic plants developed red/purple pigmentation in different tissues. Quantitative real‐time polymerase chain reaction (qRT‐PCR) analysis showed that the expression levels of six pathway genes were increased two‐ to eight‐fold in AtPAP1 transgenic plants compared with vector control plants. Dihydroflavonol reductase and anthocyanidin synthase genes that were not expressed in wild‐type plants were activated. Spectrophotometric measurement showed that the amount of anthocyanins in AtPAP1 transgenic plants were 400–800 µg g−1 fresh weight (FW). High‐performance liquid chromatography (HPLC) analysis showed that one main anthocyanin molecule accounted for approximately 98% of the total anthocyanins. Tandem MS/MS analysis using HPLC coupled to electrospray ionization and quadrupole time‐of‐flight mass spectrometry identified the main anthocyanin as cyanidin 3‐O‐rutinoside, an important medicinal anthocyanin. Analysis of photosynthesis rate, chlorophylls and carotenoids contents showed no differences between red/purple transgenic and control plants, indicating that this metabolic engineering did not alter photosynthetic physiological traits. This study shows that AtPAP1 is of significance for metabolic engineering of anthocyanins in crop plants for value‐added traits.}, number={1}, journal={PHYSIOLOGIA PLANTARUM}, author={He, Xianzhi and Li, Yong and Lawson, Darlene and Xie, De-Yu}, year={2017}, month={Jan}, pages={2–12} } @article{long_li_li_xie_peng_li_2016, title={Ontogenetic characterization of sporangium and spore of Huperzia serrata: an anti-aging disease fern}, volume={57}, ISSN={["1999-3110"]}, url={https://doi.org/10.1186/s40529-016-0151-9}, DOI={10.1186/s40529-016-0151-9}, abstractNote={Huperzia serrata is a medicinal plant used in Traditional Chinese Medicine, which has been used to prevent against aging diseases. It is mainly propagated by spores and grows extremely slowly. Due to severe harvest, it is a highly endangered species. In this report, we characterize ontogenesis of sporangia and spores that are associated with propagation. A wild population of H. serrata plants is localized in western Hunan province, China and protected by Chinese Government to study its development (e.g. sporangia and spores) and ecology. Both field and microscopic observations were conducted for a few of years. The development of sporangia from their initiation to maturation took nearly 1 year. Microscopic observations showed that the sporangial walls were developed from epidermal cells via initiation, cell division, and maturation. The structure of the mature sporangial wall is composed of one layer of epidermis, two middle layers of cells, and one layer of tapetum. Therefore, the sporangium is the eusporangium type. Spore development is characterized into six stages, initiation from epidermal cell and formation of sporogenous cells, primary sporogenous cell, secondary sporogenous cell, spore mother cell, tetrad, and maturation. The sporangial development of H. serrata belongs to the eusporangium type. The development takes approximately 1 year period from the initiation to the maturation. These data are useful for improving propagation of this medicinal plant in the future.}, number={1}, journal={BOTANICAL STUDIES}, publisher={Springer Nature}, author={Long, Hua and Li, Jing and Li, You-You and Xie, De-Yu and Peng, Qing-Zhong and Li, Li}, year={2016}, month={Nov} } @article{overexpression of a synthetic insect-plant geranyl pyrophosphate synthase gene in camelina sativa alters plant growth and terpene biosynthesis_2016, volume={244}, number={1}, journal={Planta}, year={2016}, pages={215–230} } @article{xi_rossi_lin_xie_2016, title={Overexpression of a synthetic insect–plant geranyl pyrophosphate synthase gene in Camelina sativa alters plant growth and terpene biosynthesis}, volume={244}, ISSN={0032-0935 1432-2048}, url={http://dx.doi.org/10.1007/S00425-016-2504-8}, DOI={10.1007/S00425-016-2504-8}, abstractNote={A novel plastidial homodimeric insect-plant geranyl pyrophosphate synthase gene is synthesized from three different cDNA origins. Its overexpression in Camelina sativa effectively alters plant development and terpenoid metabolism. Geranyl pyrophosphate synthase (GPPS) converts one isopentenyl pyrophosphate and dimethylallyl pyrophosphate to GPP. Here, we report a synthetic insect-plant GPPS gene and effects of its overexpression on plant growth and terpenoid metabolism of Camelina sativa. We synthesized a 1353-bp cDNA, namely PTP-MpGPPS. This synthetic cDNA was composed of a 1086-bp cDNA fragment encoding a small GPPS isomer of the aphid Myzus persicae (Mp), 240-bp Arabidopsis thaliana cDNA fragment encoding a plastidial transit peptide (PTP), and a 27-bp short cDNA fragment encoding a human influenza hemagglutinin tag peptide. Structural modeling showed that the deduced protein was a homodimeric prenyltransferase. Confocal microscopy analysis demonstrated that the PTP-MpGPPS fused with green florescent protein was localized in the plastids. The synthetic PTP-MpGPPS cDNA driven by 2 × 35S promoters was introduced into Camelina (Camelina sativa) by Agrobacterium-mediated transformation and its overexpression in transgenic plants were demonstrated by western blot. T2 and T3 progeny of transgenic plants developed larger leaves, grew more and longer internodes, and flowered earlier than wild-type plants. Metabolic analysis showed that the levels of beta-amyrin and campesterol were higher in tissues of transgenic plants than in those of wild-type plants. Fast isoprene sensor analysis demonstrated that transgenic Camelina plants emitted significantly less isoprene than wild-type plants. In addition, transcriptional analyses revealed that the expression levels of gibberellic acid and brassinosteroids-responsive genes were higher in transgenic plants than in wild-type plants. Taken together, these data demonstrated that this novel synthetic insect-plant GPPS cDNA was effective to improve growth traits and alter terpenoid metabolism of Camelina.}, number={1}, journal={Planta}, publisher={Springer Science and Business Media LLC}, author={Xi, Jing and Rossi, Lorenzo and Lin, Xiuli and Xie, De-Yu}, year={2016}, month={Mar}, pages={215–230} } @article{rossi_borghi_francini_lin_xie_sebastiani_2016, title={Salt stress induces differential regulation of the phenylpropanoid pathway in Olea europaea cultivars Frantoio (salt-tolerant) and Leccino (salt-sensitive)}, volume={204}, ISSN={0176-1617}, url={http://dx.doi.org/10.1016/j.jplph.2016.07.014}, DOI={10.1016/j.jplph.2016.07.014}, abstractNote={Olive tree (Olea europaea L.) is an important crop in the Mediterranean Basin where drought and salinity are two of the main factors affecting plant productivity. Despite several studies have reported different responses of various olive tree cultivars to salt stress, the mechanisms that convey tolerance and sensitivity remain largely unknown. To investigate this issue, potted olive plants of Leccino (salt-sensitive) and Frantoio (salt-tolerant) cultivars were grown in a phytotron chamber and treated with 0, 60 and 120mM NaCl. After forty days of treatment, growth analysis was performed and the concentration of sodium in root, stem and leaves was measured by atomic absorption spectroscopy. Phenolic compounds were extracted using methanol, hydrolyzed with butanol-HCl, and quercetin and kaempferol quantified via high performance liquid-chromatography-electrospray-mass spectrometry (HPLC-ESI-MS) and HPLC-q-Time of Flight-MS analyses. In addition, the transcripts levels of five key genes of the phenylpropanoid pathway were measured by quantitative Real-Time PCR. The results of this study corroborate the previous observations, which showed that Frantoio and Leccino differ in allocating sodium in root and leaves. This study also revealed that phenolic compounds remain stable or are strongly depleted under long-time treatment with sodium in Leccino, despite a strong up-regulation of key genes of the phenylpropanoid pathway was observed. Frantoio instead, showed a less intense up-regulation of the phenylpropanoid genes but overall higher content of phenolic compounds. These data suggest that Frantoio copes with the toxicity imposed by elevated sodium not only with mechanisms of Na + exclusion, but also promptly allocating effective and adequate antioxidant compounds to more sensitive organs.}, journal={Journal of Plant Physiology}, publisher={Elsevier BV}, author={Rossi, Lorenzo and Borghi, Monica and Francini, Alessandra and Lin, Xiuli and Xie, De-Yu and Sebastiani, Luca}, year={2016}, month={Oct}, pages={8–15} } @article{ma_wang_wang_alejos-gonzales_sun_xie_2015, title={A Genome-Wide Scenario of Terpene Pathways in Self-pollinated Artemisia annua}, volume={8}, ISSN={["1752-9867"]}, DOI={10.1016/j.molp.2015.07.004}, abstractNote={Scenarios of genes to metabolites in Artemisia annua remain uninvestigated. Here, we report the use of an integrated approach combining metabolomics, transcriptomics, and gene function analyses to characterize gene-to-terpene and terpene pathway scenarios in a self-pollinating variety of this species. Eighty-eight metabolites including 22 sesquiterpenes (e.g., artemisinin), 26 monoterpenes, two triterpenes, one diterpene and 38 other non-polar metabolites were identified from 14 tissues. These metabolites were differentially produced by leaves and flowers at lower to higher positions. Sequences from cDNA libraries of six tissues were assembled into 18 871 contigs and genome-wide gene expression profiles in tissues were strongly associated with developmental stages and spatial specificities. Sequence mining identified 47 genes that mapped to the artemisinin, non-amorphadiene sesquiterpene, monoterpene, triterpene, 2-C-methyl-D-erythritol 4-phosphate and mevalonate pathways. Pearson correlation analysis resulted in network integration that characterized significant correlations of gene-to-gene expression patterns and gene expression-to-metabolite levels in six tissues simultaneously. More importantly, manipulations of amorpha-4,11-diene synthase gene expression not only affected the activity of this pathway toward artemisinin, artemisinic acid, and arteannuin b but also altered non-amorphadiene sesquiterpene and genome-wide volatile profiles. Such gene-to-terpene landscapes associated with different tissues are fundamental to the metabolic engineering of artemisinin.}, number={11}, journal={MOLECULAR PLANT}, author={Ma, Dong-Ming and Wang, Zhilong and Wang, Liangjiang and Alejos-Gonzales, Fatima and Sun, Ming-An and Xie, De-Yu}, year={2015}, month={Nov}, pages={1580–1598} } @article{dalal_lopez_vasani_hu_swift_yalamanchili_dvora_lin_xie_qu_et al._2015, title={A photorespiratory bypass increases plant growth and seed yield in biofuel crop Camelina sativa}, volume={8}, ISSN={["1754-6834"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84945972179&partnerID=MN8TOARS}, DOI={10.1186/s13068-015-0357-1}, abstractNote={Camelina sativa is an oilseed crop with great potential for biofuel production on marginal land. The seed oil from camelina has been converted to jet fuel and improved fuel efficiency in commercial and military test flights. Hydrogenation-derived renewable diesel from camelina is environmentally superior to that from canola due to lower agricultural inputs, and the seed meal is FDA approved for animal consumption. However, relatively low yield makes its farming less profitable. Our study is aimed at increasing camelina seed yield by reducing carbon loss from photorespiration via a photorespiratory bypass. Genes encoding three enzymes of the Escherichia coli glycolate catabolic pathway were introduced: glycolate dehydrogenase (GDH), glyoxylate carboxyligase (GCL) and tartronic semialdehyde reductase (TSR). These enzymes compete for the photorespiratory substrate, glycolate, convert it to glycerate within the chloroplasts, and reduce photorespiration. As a by-product of the reaction, CO2 is released in the chloroplast, which increases photosynthesis. Camelina plants were transformed with either partial bypass (GDH), or full bypass (GDH, GCL and TSR) genes. Transgenic plants were evaluated for physiological and metabolic traits.Expressing the photorespiratory bypass genes in camelina reduced photorespiration and increased photosynthesis in both partial and full bypass expressing lines. Expression of partial bypass increased seed yield by 50-57 %, while expression of full bypass increased seed yield by 57-73 %, with no loss in seed quality. The transgenic plants also showed increased vegetative biomass and faster development; they flowered, set seed and reached seed maturity about 1 week earlier than WT. At the transcriptional level, transgenic plants showed differential expression in categories such as respiration, amino acid biosynthesis and fatty acid metabolism. The increased growth of the bypass transgenics compared to WT was only observed in ambient or low CO2 conditions, but not in elevated CO2 conditions.The photorespiratory bypass is an effective approach to increase photosynthetic productivity in camelina. By reducing photorespiratory losses and increasing photosynthetic CO2 fixation rates, transgenic plants show dramatic increases in seed yield. Because photorespiration causes losses in productivity of most C3 plants, the bypass approach may have significant impact on increasing agricultural productivity for C3 crops.}, number={1}, journal={BIOTECHNOLOGY FOR BIOFUELS}, author={Dalal, Jyoti and Lopez, Harry and Vasani, Naresh B. and Hu, Zhaohui and Swift, Jennifer E. and Yalamanchili, Roopa and Dvora, Mia and Lin, Xiuli and Xie, Deyu and Qu, Rongda and et al.}, year={2015}, month={Oct} } @article{olarte_worthington_horn_moore_singh_monacell_dorner_stone_xie_carbone_2015, title={Enhanced diversity and aflatoxigenicity in interspecific hybrids ofAspergillus flavusandAspergillus parasiticus}, volume={24}, ISSN={0962-1083}, url={http://dx.doi.org/10.1111/mec.13153}, DOI={10.1111/mec.13153}, abstractNote={AbstractAspergillus flavus and A. parasiticus are the two most important aflatoxin‐producing fungi responsible for the contamination of agricultural commodities worldwide. Both species are heterothallic and undergo sexual reproduction in laboratory crosses. Here we examine the possibility of interspecific matings between A. flavus and A. parasiticus. These species can be distinguished morphologically and genetically, as well as by their mycotoxin profiles. Aspergillus flavus produces both B aflatoxins and cyclopiazonic acid (CPA), B aflatoxins or CPA alone, or neither mycotoxin; Aspergillus parasiticus produces B and G aflatoxins or the aflatoxin precursor O‐methylsterigmatocystin, but not CPA. Only four of forty‐five attempted interspecific crosses between opposite mating types of A. flavus and A. parasiticus were fertile and produced viable ascospores. Single ascospore strains from each cross were shown to be recombinant hybrids using multilocus genotyping and array comparative genome hybridization. Conidia of parents and their hybrid progeny were haploid and predominantly monokaryons and dikaryons based on flow cytometry. Multilocus phylogenetic inference showed that experimental hybrid progeny were grouped with naturally occurring A. flavus L strain and A. parasiticus. Higher total aflatoxin concentrations in some F1 progeny strains compared to midpoint parent aflatoxin levels indicate synergism in aflatoxin production; moreover, three progeny strains synthesized G aflatoxins that were not produced by the parents, and there was evidence of allopolyploidization in one strain. These results suggest that hybridization is an important diversifying force resulting in the genesis of novel toxin profiles in these agriculturally important fungi.}, number={8}, journal={Molecular Ecology}, publisher={Wiley}, author={Olarte, Rodrigo A. and Worthington, Carolyn J. and Horn, Bruce W. and Moore, Geromy G. and Singh, Rakhi and Monacell, James T. and Dorner, Joe W. and Stone, Eric A. and Xie, De-Yu and Carbone, Ignazio}, year={2015}, month={Apr}, pages={1889–1909} } @article{zhu_wang_peng_tang_xia_wu_xie_2015, title={Functional characterization of an anthocyanidin reductase gene from the fibers of upland cotton (Gossypium hirsutum)}, volume={241}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-014-2238-4}, abstractNote={Metabolic profiling, gene cloning, enzymatic analysis, ectopic expression, and gene silencing experiments demonstrate that the anthocyanidin reductase (ANR) pathway is involved in the biosynthesis of proanthocyanidins in upland cotton. Proanthocyanidins (PAs) are oligomeric or polymeric flavan-3-ols, however, the biosynthetic pathway of PAs in cotton remains to be elucidated. Here, we report on an anthocyanidin reductase (ANR) gene from cotton fibers and the ANR pathway of PAs. Phytochemical analysis demonstrated that leaves, stems, roots, and early developing fibers produced PAs and their monomers, including (-)-epicatechin, (-)-catechin, (-)-epigallocatechin, and (-)-gallocatechin. Crude PA extractions from different tissues were boiled in Butanol:HCl. Cyanidin, delphinidin, and pelargonidin were produced, indicating that cotton PAs include diverse extension unit structures. An ANR cDNA homolog (named GhANR1) was cloned from developing fibers. The open reading frame, composed of 1,011 bp nucleotides, was expressed in E. coli to obtain a recombinant protein. In the presence of NADPH, the recombinant enzyme catalyzed cyanidin, delphinidin, and pelargonidin to (-)-epicatechin and (-)-catechin, (-)-epigallocatechin and (-)-gallocatechin, and (-)-epiafzelechin and (-)-afzelechin, respectively. The ectopic expression of GhANR11 in an Arabidopsis ban mutant allowed for the reconstruction of the ANR pathway and PA biosynthesis in the seed coat. Virus-induced gene silencing (VIGS) of GhANR11 led to a significant increase in anthocyanins and a decrease in the PAs, (-)-epicatechin, and (-)-catechin in the stems and leaves of VIGS-infected plants. Taken together, these data demonstrate that the ANR pathway contributes to the biosynthesis of flavan-3-ols and PAs in cotton.}, number={5}, journal={PLANTA}, author={Zhu, Yue and Wang, Haiyun and Peng, Qingzhong and Tang, Yuntao and Xia, Guixian and Wu, Jiahe and Xie, De-Yu}, year={2015}, month={May}, pages={1075–1089} } @article{borghi_xie_2016, title={Tissue-specific production of limonene in Camelina sativa with the Arabidopsis promoters of genes BANYULS and FRUITFULL}, volume={243}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-015-2425-y}, abstractNote={Arabidopsis promoters of genes BANYULS and FRUITFULL are transcribed in Camelina. They triggered the transcription of limonene synthase and induced higher limonene production in seeds and fruits than CaMV 35S promoter. Camelina sativa (Camelina) is an oilseed crop of relevance for the production of biofuels and the plant has been target of a recent and intense program of genetic manipulation aimed to increase performance, seed yield and to modify the fatty acid composition of the oil. Here, we have explored the performance of two Arabidopsis thaliana (Arabidopsis) promoters in triggering transgene expression in Camelina. The promoters of two genes BANYULS (AtBAN pro ) and FRUITFULL (AtFUL pro ), which are expressed in seed coat and valves of Arabidopsis, respectively, have been chosen to induce the expression of limonene synthase (LS) from Citrus limon. In addition, the constitutive CaMV 35S promoter was utilized to overexpress LS in Camelina . The results of experiments revealed that AtBAN pro and AtFUL pro are actively transcribed in Camelina where they also retain specificity of expression in seeds and valves as previously observed in Arabidopsis. LS induced by AtBAN pro and AtFUL pro leads to higher limonene production in seeds and fruits than when the CaMV 35S was used to trigger the expression. In conclusion, the results of experiments indicate that AtBAN pro and AtFUL pro can be successfully utilized to induce the expression of the transgenes of interest in seeds and fruits of Camelina.}, number={2}, journal={PLANTA}, author={Borghi, Monica and Xie, De-Yu}, year={2016}, month={Feb}, pages={549–561} } @article{shi_xie_2014, title={Biosynthesis and Metabolic Engineering of Anthocyanins in Arabidopsis thaliana}, volume={8}, ISSN={1872-2083}, url={http://dx.doi.org/10.2174/1872208307666131218123538}, DOI={10.2174/1872208307666131218123538}, abstractNote={Arabidopsis thaliana is the first model plant, the genome of which has been sequenced. In general, intensive studies on this model plant over the past nearly 30 years have led to many new revolutionary understandings in every single aspect of plant biology. Here, we review the current understanding of anthocyanin biosynthesis in this model plant. Although the investigation of anthocyanin structures in this model plant was not performed until 2002, numerous studies over the past three decades have been conducted to understand the biosynthesis of anthocyanins. To date, it appears that all pathway genes of anthocyanins have been molecularly, genetically and biochemically characterized in this plant. These fundamental accomplishments have made Arabidopsis an ideal model to understand the regulatory mechanisms of anthocyanin pathway. Several studies have revealed that the biosynthesis of anthocyanins is controlled by WD40-bHLH-MYB (WBM) transcription factor complexes under lighting conditions. However, how different regulatory complexes coordinately and specifically regulate the pathway genes of anthocyanins remains unclear. In this review, we discuss current progresses and findings including structural diversity, regulatory properties and metabolic engineering of anthocyanins in Arabidopsis thaliana.}, number={1}, journal={Recent Patents on Biotechnology}, publisher={Bentham Science Publishers Ltd.}, author={Shi, Ming-Zhu and Xie, De-Yu}, year={2014}, month={Apr}, pages={47–60} } @article{zhu_peng_li_xie_2014, title={Molecular cloning and functional characterization of the anthocyanidin reductase gene from Vitis bellula}, volume={240}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-014-2094-2}, abstractNote={Anthocyanidin reductase (ANR) is an NADPH-/NADH-dependent enzyme that transfers two hydrides to anthocyanidins to produce three types of isomeric flavan-3-ols. This reductase forms the ANR pathway toward the biosynthesis of proanthocyanidins (PAs, which are also called condensed tannins). Here, we report cloning and functional characterization of an ANR (called VbANR) homolog from the leaves of Vitis bellula, a newly developed grape crop in southern China. The open reading frame (ORF) of VbANR is 1,017 bp in length and encodes 339 amino acids. A phylogenetic analysis and an alignment using 17 sequences revealed that VbANR is approximately 99.9 % identical to the ANR homolog from Vitis vinifera. The VbANR ORF is fused to the Trx gene containing a His-tag in the pET32a(+) vector to obtain a pET32a(+)-VbANR construct for expressing the recombinant VbANR. In vitro enzyme assays show that VbANR converts cyanidin, delphinidin, and pelargonidin to their corresponding flavan-3-ols. Enzymatic products include 2S,3R-trans- and 2R,3R-cis-flavan-3-ols isomers, such as (-)-catechin and (-)-epicatechin. In addition, the third compound that is observed from the enzymatic products is most likely a 2S,3S-cis-flavan-3-ol. To analyze the kinetics and optimize pH and temperature values, a UV spectrometry method was developed to quantify the concentrations of total enzymatic products. The optimum pH and temperature values are 4.0 and 40 °C, respectively. The K m , K cat, V max, and K cat/K m values for pelargonidin and delphinidin were similar. In comparison, VbANR exhibits a slightly lower affinity to cyanidin. VbANR uses both NADPH and NADH but prefers to employ NADPH. GFP fusion and confocal microscopy analyses revealed the cytosolic localization of VbANR. The overexpression of VbANR in ban mutants reconstructed the biosynthetic pathway of PAs in the seed coat. These data demonstrate that VbANR forms the ANR pathway, leading to the formation of three types of isomeric flavan-3-ols and PAs in the leaves of V. bellula.}, number={2}, journal={PLANTA}, author={Zhu, Yue and Peng, Qing-Zhong and Li, Ke-Gang and Xie, De-Yu}, year={2014}, month={Aug}, pages={381–398} } @article{zhu_peng_du_li_xie_2013, title={Characterization of Flavan-3-ols and Expression of MYB and Late Pathway Genes Involved in Proanthocyanidin Biosynthesis in Foliage of Vitis bellula}, volume={3}, ISSN={2218-1989}, url={http://dx.doi.org/10.3390/metabo3010185}, DOI={10.3390/metabo3010185}, abstractNote={Proanthocyanidins (PAs) are fundamental nutritional metabolites in different types of grape products consumed by human beings. Although the biosynthesis of PAs in berry of Vitis vinifera has gained intensive investigations, the understanding of PAs in other Vitis species is limited. In this study, we report PA formation and characterization of gene expression involved in PA biosynthesis in leaves of V. bellula, a wild edible grape species native to south and south-west China. Leaves are collected at five developmental stages defined by sizes ranging from 0.5 to 5 cm in length. Analyses of thin layer chromatography (TLC) and high performance liquid chromatography-photodiode array detector (HPLC-PAD) show the formation of (+)-catechin, (−)-epicatechin, (+)-gallocatechin and (−)-epigallocatechin during the entire development of leaves. Analyses of butanol-HCl boiling cleavage coupled with spectrometry measurement at 550 nm show a temporal trend of extractable PA levels, which is characterized by an increase from 0.5 cm to 1.5 cm long leaves followed by a decrease in late stages. TLC and HPLC-PAD analyses identify cyanidin, delphinidin and pelargonidin produced from the cleavage of PAs in the butanol-HCl boiling, showing that the foliage PAs of V. bellula include three different types of extension units. Four cDNAs, which encode VbANR, VbDFR, VbLAR1 and VbLAR2, respectively, are cloned from young leaves. The expression patterns of VbANR and VbLAR2 but not VbLAR1 and VbDFR follow a similar trend as the accumulation patterns of PAs. Two cDNAs encoding VbMYBPA1 and VbMYB5a, the homologs of which have been demonstrated to regulate the expression of both ANR and LAR in V. vinifera, are also cloned and their expression profiles are similar to those of VbANR and VbLAR2. In contrast, the expression profiles of MYBA1 and 2 homologs involved in anthocyanin biosynthesis are different from those of VbANR and VbLAR2. Our data show that both ANR and LAR branches are involved in PA biosynthesis in leaves of V. bellula.}, number={1}, journal={Metabolites}, publisher={MDPI AG}, author={Zhu, Yue and Peng, Qing-Zhong and Du, Ci and Li, Ke-Gang and Xie, De-Yu}, year={2013}, month={Mar}, pages={185–203} } @article{alejos-gonzalez_perkins_winston_xie_2013, title={Efficient Somatic Embryogenesis and Organogenesis of Self-Pollination Artemisia annua Progeny and Artemisinin Formation in Regenerated Plants}, volume={04}, ISSN={2158-2742 2158-2750}, url={http://dx.doi.org/10.4236/ajps.2013.411274}, DOI={10.4236/ajps.2013.411274}, abstractNote={To enhance the understanding of artemisinin biosynthesis, we have successfully bred self-pollination Artemisia annua plants. Here, we report efficient somatic embryogenesis and organogenesis of self-pollination plants and artemisinin formation in regenerated plants. The first through sixth nodal leaves of seedlings are used as explants. On agar-solidified MS basal medium supplemented with TDZ (0.6 mg/l) and IBA (0.1 mg/l), all explants after inoculation of less than 3 weeks start to form embryogenic calli, which further produce globular, torpedo, heart and early cotyledon embryos. In all six positional leaves, explants from the sixth leaf show the rapidest responses to induction of embryogenic calli and somatic embryos. On this medium, somatic embryos continuously develop into adventitious buds, which can form adventitious roots on a rooting medium containing NAA (0.5 mg/l). Meanwhile, on agar-solidified MS basal medium supplemented with BAP (1 mg/l) and NAA (0.05 mg/l), approximately 100% of explants from leaves #3-6 form calli in less than 3 weeks of inoculation and adventitious buds via organogenesis in 3-4 weeks. In all six positional leaves, explants from the sixth leaf exhibit the rapidest response to induction of calli and adventitious buds. Nearly 100% adventitious buds can form adventitious roots on the rooting medium. Regenerated plants from both somatic embryogenesis and organogenesis complete self-pollination to produce seeds in 80-90 days of growth in growth chamber. LC-ESI-MS analysis demonstrates that regenerated plants biosynthesize artemisinin. These results show the highly efficient regeneration capacity of self-pollination A. annua plants that can form a new platform to enhance the understanding of artemisinin biosynthesis and metabolic engineering.}, number={11}, journal={American Journal of Plant Sciences}, publisher={Scientific Research Publishing, Inc.}, author={Alejos-Gonzalez, Fatima and Perkins, Kelly and Winston, Malcolm Isaiah and Xie, De-Yu}, year={2013}, pages={2206–2217} } @article{xie_shi_liu_2013, title={Mechanisms of Metabolic Programming Toward Anthocyanin Biosynthesis in Cultured Red Arabidopsis Cells}, volume={49}, number={4}, journal={In Vitro Cellular & Developmental Biology - Plant}, author={Xie, De-Yu and Shi, M.Z. and Liu, Z.}, year={2013}, pages={484–485} } @article{liu_shi_xie_2014, title={Regulation of anthocyanin biosynthesis in Arabidopsis thaliana red pap1-D cells metabolically programmed by auxins}, volume={239}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-013-2011-0}, abstractNote={Red pap1-D cells of Arabidopsis thaliana have been cloned from production of anthocyanin pigmentation 1-Dominant (pap1-D) plants. The red cells are metabolically programmed to produce high levels of anthocyanins by a WD40-bHLH-MYB complex that is composed of the TTG1, TT8/GL3 and PAP1 transcription factors. Here, we report that indole 3-acetic acid (IAA), naphthaleneacetic acid (NAA) and 2,4-dichlorophenoxyacetic acid (2,4-D) regulate anthocyanin biosynthesis in these red cells. Seven concentrations (0, 0.2, 0.4, 2.2, 9, 18 and 27 μM) were tested for the three auxins. IAA and 2,4-D at 2.2-27 μM reduced anthocyanin levels. NAA at 0-0.2 μM or above 9 μM also decreased anthocyanin levels, but from 0.4 to 9 μM, it increased them. HPLC-ESI-MS analysis identified seven cyanin molecules that were produced in red pap1-D cells, and their levels were affected by auxins. The expression levels of ten genes, including six transcription factors (TTG1, EGL3, MYBL2, TT8, GL3 and PAP1) and four pathway genes (PAL1, CHS, DFR and ANS) involved in anthocyanin biosynthesis were analyzed upon various auxin treatments. The resulting data showed that 2,4-D, NAA and IAA control anthocyanin biosynthesis by regulating the expression of TT8, GL3 and PAP1 as well as genes in the anthocyanin biosynthetic pathway, such as DFR and ANS. In addition, the expression of MYBL2, PAL1 and CHS in red pap1-D and wild-type cells differentially respond to the three auxins. Our data demonstrate that the three auxins regulate anthocyanin biosynthesis in metabolically programmed red cells via altering the expression of transcription factor genes and pathway genes.}, number={4}, journal={PLANTA}, author={Liu, Zhong and Shi, Ming-Zhu and Xie, De-Yu}, year={2014}, month={Apr}, pages={765–781} } @article{peng_zhu_liu_du_li_xie_2012, title={An integrated approach to demonstrating the ANR pathway of proanthocyanidin biosynthesis in plants}, volume={236}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-012-1670-6}, abstractNote={Proanthocyanidins (PAs) are oligomers or polymers of plant flavan-3-ols and are important to plant adaptation in extreme environmental conditions. The characterization of anthocyanidin reductase (ANR) and leucoanthocyanidin reductase (LAR) has demonstrated the different biogenesis of four stereo-configurations of flavan-3-ols. It is important to understand whether ANR and the ANR pathway widely occur in the plant kingdom. Here, we report an integrated approach to demonstrate the ANR pathway in plants. This includes different methods to extract native ANR from different tissues of eight angiosperm plants (Lotus corniculatus, Desmodium uncinatum, Medicago sativa, Hordeum vulgare, Vitis vinifera, Vitis bellula, Parthenocissus heterophylla, and Cerasus serrulata) and one fern plant (Dryopteris pycnopteroides), a general enzymatic analysis approach to demonstrate the ANR activity, high-performance liquid chromatography-based fingerprinting to demonstrate (-)-epicatechin and other flavan-3-ol molecules, and phytochemical analysis of PAs. Results demonstrate that in addition to leaves of M. sativa, tissues of other eight plants contain an active ANR pathway. Particularly, the leaves, flowers and pods of D. uncinatum, which is a model plant to study LAR and the LAR pathways, are demonstrated to express an active ANR pathway. This finding suggests that the ANR pathway involves PA biosynthesis in D. uncinatum. In addition, a sequence BLAST analysis reveals that ANR homologs have been sequenced in plants from both gymnosperms and angiosperms. These data show that the ANR pathway to PA biosynthesis occurs in both seed and seedless vascular plants.}, number={3}, journal={PLANTA}, author={Peng, Qing-Zhong and Zhu, Yue and Liu, Zhong and Du, Ci and Li, Ke-Gang and Xie, De-Yu}, year={2012}, month={Sep}, pages={901–918} } @inbook{broeckling_li_xie_2012, place={Rijeka, Croatia}, title={Comparative Metabolomics of Transgenic Tobacco Plants (Nicotiana tabacum var. Xanthi) Reveals Differential Effects of Engineered Complete and Incomplete Flavonoid Pathways on the Metabolome}, ISBN={9789535101819}, url={http://dx.doi.org/10.5772/32872}, DOI={10.5772/32872}, abstractNote={Anthocyanins and proanthocyanidins (PAs) are two groups of end products of the plant flavonoid pathway (Fig. 1). Biochemical and genetic evidences have demonstrated that they share the same upstream pathway beginning with phenylalanine through a series of enzymatic reaction to anthocyanidins. Anthocyanidins are either modified by glycosylation, methylation, or other reactions to form diverse anthocyanins (Springob et al. 2003) or catalyzed into flavan-3-ols by an anthocyanidin reductase (ANR) (Xie et al. 2003; Xie et al. 2006). In addition, leucoanthocyanidin reductase (LAR) has been enzymatically demonstrated to catalyze leucoanthocyanidins into catechin (Tanner et al. 2003). To date, whether or not this branch catalyzed by LAR exists in plants still remains open to genetic studies.}, booktitle={Transgenic Plants - Advances and Limitations}, publisher={InTech}, author={Broeckling, Corey D. and Li, Ke-Gang and Xie, De-Yu}, editor={Çiftçi, Yelda OzdenEditor}, year={2012}, month={Mar} } @article{xie_shi_2012, title={Differentiation of programmed Arabidopsis cells}, volume={3}, ISSN={2165-5979 2165-5987}, url={http://dx.doi.org/10.4161/bbug.3.1.17786}, DOI={10.4161/bbug.3.1.17786}, abstractNote={Plants express genes that encode enzymes that catalyse reactions to form plant secondary metabolites in specific cell types. However, the mechanisms of how plants decide their cellular metabolic fate and how cells diversify and specialise their specific secondary metabolites remains largely unknown. Additionally, whether and how an established metabolic program impacts genome-wide reprogramming of plant gene expression is unclear. We recently isolated PAP1-programmed anthocyanin-producing (red) and -free (white) cells from Arabidopsis thaliana; our previous studies have indicated that the PAP1 expression level is similar between these two different cell types. Transcriptional analysis showed that the red cells contain the TTG1-GL3/TT8-PAP1 regulatory complex, which controls anthocyanin biosynthesis; in contrast, the white cells and the wild-type cells lack this entire complex. These data indicate that different regulatory programming underlies the different metabolic states of these cells. In addition, our previous transcriptomic comparison indicated that there is a clear difference in the gene expression profiles of the red and wild-type cells, which is probably a consequence of cell-specific reprogramming. Based on these observations, in this report we discuss the potential mechanisms that underlie the programming and reprogramming of gene expression involved in anthocyanin biosynthesis.}, number={1}, journal={Bioengineered}, publisher={Informa UK Limited}, author={Xie, De-Yu and Shi, Ming-Zhu}, year={2012}, month={Jan}, pages={54–59} } @article{feng_liu_yu_xie_franks_xiang_2012, title={Evolution of bract development and B‐class MADS box gene expression in petaloid bracts of Cornus s. l. (Cornaceae)}, volume={196}, ISSN={0028-646X 1469-8137}, url={http://dx.doi.org/10.1111/j.1469-8137.2012.04255.x}, DOI={10.1111/j.1469-8137.2012.04255.x}, abstractNote={Despite increasing interest in the molecular mechanisms of floral diversity, few studies have investigated the developmental and genetic bases of petaloid bracts. This study examined morphological patterns of bract initiation and expression patterns of B-class MADS-box genes in bracts of several Cornus species. We suggest that petaloid bracts in this genus may not share a single evolutionary origin. Developmental pathways of bracts and spatiotemporal expression of B-class genes in bracts and flowers were examined for four closely related dogwood species. Divergent morphological progressions and gene expression patterns were found in the two sister lineages with petaloid bracts, represented by Cornus florida and Cornus canadensis. Phylogeny-based analysis identified developmental and gene expression changes that are correlated with the evolution of petaloid bracts in C. florida and C. canadensis. Our data support the existence of independent evolutionary origins of petaloid bracts in C. canadensis and C. florida. Additionally, we suggest that functional transference within B-class gene families may have contributed to the origin of bract petaloidy in C. florida. However, the underlying mechanisms of petaloid bract development likely differ between C. florida and C. canadensis. In the future this hypothesis can be tested by functional analyses of Cornus B-class genes.}, number={2}, journal={New Phytologist}, publisher={Wiley}, author={Feng, Chun‐Miao and Liu, Xiang and Yu, Yi and Xie, Deyu and Franks, Robert G. and Xiang, Qiu‐Yun (Jenny)}, year={2012}, month={Aug}, pages={631–643} } @article{ma_gandra_sharma_xie_2012, title={Integration of GC-MS Based Non-Targeted Metabolic Profiling with Headspace Solid Phase Microextraction Enhances the Understanding of Volatile Differentiation in Tobacco Leaves from North Carolina, India and Brazil}, volume={03}, ISSN={2158-2742 2158-2750}, url={http://dx.doi.org/10.4236/ajps.2012.312215}, DOI={10.4236/ajps.2012.312215}, abstractNote={In this report, gas chromatography-mass spectrometry (GC-MS) based non-targeted metabolomics is used to develop appropriate headspace solid phase microextractions (HS-SPME) to enhance the understanding of volatile complexity of flue-cured tobacco leaves. Non-targeted metabolic profiling of GC-MS shows that the extraction condition of HS-SPME at 100?C for 30 min provides a better metabolite profile than other extraction conditions tested. GC-MS and principal component analyses (PCA) show that among five types of fibers tested, 100 μm polydimethylsiloxane (PMDS), 65 μm polydimethylsiloxane/divinylbenzene (PMDS/DVB) and 75 μm carboxen/polydimethylsiloxane (CAR/ PMS) provide a better reproducible metabolite profile. Based on an appropriate PDMS extraction condition optimized, we use GC-MS analysis and PCA to compare metabolite profiles in flue-cured leaves of tobacco plants grown in North Carolina, India and Brazil, respectively. The resulting data of PCA show that the global metabolic profiles in North Carolina samples are separated from those in Brazil and India samples, two groups of which are characterized by a partially overlapped pattern. Several peaks that were differentially accumulated in samples were annotated to known metabolites by deconvolution analysis, such as norsolanadione, solavetivone and rishtin. Norsolanadione is detected only in Brazil samples. Solavetivone is detected in samples of India and Brazil but not in those of North Carolina. Rishtin is detected in samples of North Carolina and India but not in Brazil samples. These data indicate that not only can a non-targeted metabolic profiling approach enhance the understanding of volatile complexity, but also can identify marker volatile metabolites in tobacco leaves produced in different growth regions.}, number={12}, journal={American Journal of Plant Sciences}, publisher={Scientific Research Publishing, Inc.}, author={Ma, Dong-Ming and Gandra, Saiprasad V. S. and Sharma, Navin and Xie, De-Yu}, year={2012}, pages={1759–1769} } @article{liu_feng_franks_qu_xie_xiang_2013, title={Plant regeneration and genetic transformation of C. canadensis: a non-model plant appropriate for investigation of flower development in Cornus (Cornaceae)}, volume={32}, ISSN={0721-7714, 1432-203X}, url={http://link.springer.com/10.1007/s00299-012-1341-x}, DOI={10.1007/s00299-012-1341-x}, abstractNote={KEY MESSAGE : Efficient Agrobacterium -mediated genetic transformation for investigation of genetic and molecular mechanisms involved in inflorescence architectures in Cornus species. Cornus canadensis is a subshrub species in Cornus, Cornaceae. It has recently become a favored non-model plant species to study genes involved in development and evolution of inflorescence architectures in Cornaceae. Here, we report an effective protocol of plant regeneration and genetic transformation of C. canadensis. We use young inflorescence buds as explants to efficiently induce calli and multiple adventitious shoots on an optimized induction medium consisting of basal MS medium supplemented with 1 mg/l of 6-benzylaminopurine and 0.1 mg/l of 1-naphthaleneacetic acid. On the same medium, primary adventitious shoots can produce a large number of secondary adventitious shoots. Using leaves of 8-week-old secondary shoots as explants, GFP as a reporter gene controlled by 35S promoter and hygromycin B as the selection antibiotic, a standard procedure including pre-culture of explants, infection, co-cultivation, resting and selection has been developed to transform C. canadensis via Agrobacterium strain EHA105-mediated transformation. Under a strict selection condition using 14 mg/l hygromycin B, approximately 5 % explants infected by Agrobacterium produce resistant calli, from which clusters of adventitious shoots are induced. On an optimized rooting medium consisting of basal MS medium supplemented with 0.1 mg/l of indole-3-butyric acid and 7 mg/l hygromycin B, most of the resistant shoots develop adventitious roots to form complete transgenic plantlets, which can grow normally in soil. RT-PCR analysis demonstrates the expression of GFP transgene. Green fluorescence emitted by GFP is observed in transgenic calli, roots and cells of transgenic leaves under both stereo fluorescence microscope and confocal microscope. The success of genetic transformation provides an appropriate platform to investigate the molecular mechanisms by which the various inflorescence forms are developed in Cornus plants.}, number={1}, journal={Plant Cell Reports}, publisher={Springer Science and Business Media LLC}, author={Liu, Xiang and Feng, Chun-Miao and Franks, Robert and Qu, Rongda and Xie, De-Yu and Xiang, Qiu-Yun Jenny}, year={2013}, month={Jan}, pages={77–87} } @inproceedings{xie_shi_2012, title={Programming and reprogramming of gene expression: Mechanisms of cellular specific metabolisms?}, volume={50}, number={11}, booktitle={Pharmaceutical Biology}, author={Xie, D.Y. and Shi, M.Z.}, year={2012}, pages={1356–1356} } @article{zhou_shi_xie_2012, title={Regulation of anthocyanin biosynthesis by nitrogen in TTG1-GL3/TT8-PAP1-programmed red cells of Arabidopsis thaliana}, volume={236}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-012-1674-2}, abstractNote={Nitrogen nutrients can regulate anthocyanin biosynthesis in Arabidopsis thaliana. In this investigation, we report the nitrogen regulation of anthocyanin biosynthesis activated by TTG1-GL3/TT8-PAP1 in red pap1-D cells. To understand the mechanisms of nitrogen regulation, we employed red pap1-D cells and wild-type cells (as a control) to examine the effects of different nitrogen treatments on anthocyanin biosynthesis. In general, the higher concentrations of ammonium and high total nitrogen tested (e.g., 58.8 and 29.8 mM total nitrogen consisting of NH(4)NO(3) and KNO(3)) reduced the levels and molecular diversity of anthocyanins; in contrast, the lower concentrations of ammonium and total nitrogen conditions (e.g., 9.4 mM KNO(3) and the depletion of nitrogen) increased the levels and molecular diversity of anthocyanins. An expression analysis of the main regulatory and pathway genes showed that at conditions of higher concentrations of ammonium and total nitrogen, the expression levels of PAP1 and TT8 decreased, but the expression levels of LBD37, 38 and 39, three negative regulators of anthocyanin biosynthesis, increased. In addition, the expression levels of the main pathway genes decreased. In contrast, at conditions of lower concentrations of ammonium and total nitrogen, the expression levels of PAP1, TT8 and the main pathway genes increased, whereas those of LBD37, 38 and 39 decreased. These results show that nitrogen regulation of anthocyanin biosynthesis in red cells undergoes a mechanism by which nitrogen controls the expression of genes encoding both main components of the TTG1-GL3/TT8-PAP1 complex and negative regulators. Based on these observations, we propose that the regulatory mechanism of nitrogen may occur via two pathways to control the expression of genes encoding positive and negative regulators in red pap1-D cells.}, number={3}, journal={PLANTA}, author={Zhou, Li-Li and Shi, Ming-Zhu and Xie, De-Yu}, year={2012}, month={Sep}, pages={825–837} } @article{xie_alejos-gonzalez_qu_2012, title={Self-pollinated Artemisia annua plants form a new platform to understand artemisinin biosynthesis}, volume={50}, number={5}, journal={Pharmaceutical Biology}, author={Xie, D.Y. and Alejos-Gonzalez, F. and Qu, G.}, year={2012}, pages={662–662} } @article{xie_melis_2012, title={Special Issue on Metabolic Plant Biology}, volume={236}, ISSN={["0032-0935"]}, DOI={10.1007/s00425-012-1700-4}, number={3}, journal={PLANTA}, author={Xie, De-Yu and Melis, Anastasios}, year={2012}, month={Sep}, pages={763–764} } @article{he_brumos_li_ji_ke_gong_zeng_li_zhang_an_et al._2011, title={A Small-Molecule Screen Identifies l-Kynurenine as a Competitive Inhibitor of TAA1/TAR Activity in Ethylene-Directed Auxin Biosynthesis and Root Growth in Arabidopsis}, volume={23}, ISSN={1040-4651 1532-298X}, url={http://dx.doi.org/10.1105/tpc.111.089029}, DOI={10.1105/tpc.111.089029}, abstractNote={AbstractThe interactions between phytohormones are crucial for plants to adapt to complex environmental changes. One example is the ethylene-regulated local auxin biosynthesis in roots, which partly contributes to ethylene-directed root development and gravitropism. Using a chemical biology approach, we identified a small molecule, l-kynurenine (Kyn), which effectively inhibited ethylene responses in Arabidopsis thaliana root tissues. Kyn application repressed nuclear accumulation of the ETHYLENE INSENSITIVE3 (EIN3) transcription factor. Moreover, Kyn application decreased ethylene-induced auxin biosynthesis in roots, and TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1/TRYPTOPHAN AMINOTRANSFERASE RELATEDs (TAA1/TARs), the key enzymes in the indole-3-pyruvic acid pathway of auxin biosynthesis, were identified as the molecular targets of Kyn. Further biochemical and phenotypic analyses revealed that Kyn, being an alternate substrate, competitively inhibits TAA1/TAR activity, and Kyn treatment mimicked the loss of TAA1/TAR functions. Molecular modeling and sequence alignments suggested that Kyn effectively and selectively binds to the substrate pocket of TAA1/TAR proteins but not those of other families of aminotransferases. To elucidate the destabilizing effect of Kyn on EIN3, we further found that auxin enhanced EIN3 nuclear accumulation in an EIN3 BINDING F-BOX PROTEIN1 (EBF1)/EBF2-dependent manner, suggesting the existence of a positive feedback loop between auxin biosynthesis and ethylene signaling. Thus, our study not only reveals a new level of interactions between ethylene and auxin pathways but also offers an efficient method to explore and exploit TAA1/TAR-dependent auxin biosynthesis.}, number={11}, journal={The Plant Cell}, publisher={American Society of Plant Biologists (ASPB)}, author={He, Wenrong and Brumos, Javier and Li, Hongjiang and Ji, Yusi and Ke, Meng and Gong, Xinqi and Zeng, Qinglong and Li, Wenyang and Zhang, Xinyan and An, Fengying and et al.}, year={2011}, month={Nov}, pages={3944–3960} } @article{alejos-gonzalez_qu_zhou_saravitz_shurtleff_xie_2011, title={Characterization of development and artemisinin biosynthesis in self-pollinated Artemisia annua plants}, volume={234}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-011-1430-z}, abstractNote={Artemisia annua L. is the only natural resource that produces artemisinin (Qinghaosu), an endoperoxide sesquiterpene lactone used in the artemisinin-combination therapy of malaria. The cross-hybridization properties of A. annua do not favor studying artemisinin biosynthesis. To overcome this problem, in this study, we report on selection of self-pollinated A. annua plants and characterize their development and artemisinin biosynthesis. Self-pollinated F2 plants selected were grown under optimized growth conditions, consisting of long day (16 h of light) and short day (9 h of light) exposures in a phytotron. The life cycles of these plants were approximately 3 months long, and final heights of 30-35 cm were achieved. The leaves on the main stems exhibited obvious morphological changes, from indented single leaves to odd, pinnately compound leaves. Leaves and flowers formed glandular and T-shaped trichomes on their surfaces. The glandular trichome densities increased from the bottom to the top leaves. High performance liquid chromatography-mass spectrometry-based metabolic profiling analyses showed that leaves, flowers, and young seedlings of F2 plants produced artemisinin. In leaves, the levels of artemisinin increased from the bottom to the top of the plants, showing a positive correlation to the density increase of glandular trichomes. RT-PCR analysis showed that progeny of self-pollinated plants expressed the amorpha-4, 11-diene synthase (ADS) and cytochrome P450 monooxygenase 71 AV1 (CYP71AV1) genes, which are involved in artemisinin biosynthesis in leaves and flowers. The use of self-pollinated A. annua plants will be a valuable approach to the study of artemisinin biosynthesis.}, number={4}, journal={PLANTA}, author={Alejos-Gonzalez, Fatima and Qu, Guosheng and Zhou, Li-Li and Saravitz, Carole H. and Shurtleff, Janet L. and Xie, De-Yu}, year={2011}, month={Oct}, pages={685–697} } @article{shi_xie_2011, title={Engineering of red cells of Arabidopsis thaliana and comparative genome-wide gene expression analysis of red cells versus wild-type cells}, volume={233}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-010-1335-2}, abstractNote={We report metabolic engineering of Arabidopsis red cells and genome-wide gene expression analysis associated with anthocyanin biosynthesis and other metabolic pathways between red cells and wild-type (WT) cells. Red cells of A. thaliana were engineered for the first time from the leaves of production of anthocyanin pigment 1-Dominant (pap1-D). These red cells produced seven anthocyanin molecules including a new one that was characterized by LC-MS analysis. Wild-type cells established as a control did not produce anthocyanins. A genome-wide microarray analysis revealed that nearly 66 and 65% of genes in the genome were expressed in the red cells and wild-type cells, respectively. In comparison with the WT cells, 3.2% of expressed genes in the red cells were differentially expressed. The expression levels of 14 genes involved in the biosynthetic pathway of anthocyanin were significantly higher in the red cells than in the WT cells. Microarray and RT-PCR analyses demonstrated that the TTG1-GL3/TT8-PAP1 complex regulated the biosynthesis of anthocyanins. Furthermore, most of the genes with significant differential expression levels in the red cells versus the WT cells were characterized with diverse biochemical functions, many of which were mapped to different metabolic pathways (e.g., ribosomal protein biosynthesis, photosynthesis, glycolysis, glyoxylate metabolism, and plant secondary metabolisms) or organelles (e.g., chloroplast). We suggest that the difference in gene expression profiles between the two cell lines likely results from cell types, the overexpression of PAP1, and the high metabolic flux toward anthocyanins.}, number={4}, journal={PLANTA}, author={Shi, Ming-Zhu and Xie, De-Yu}, year={2011}, month={Apr}, pages={787–805} } @article{shi_xie_2010, title={Features of anthocyanin biosynthesis in pap1-D and wild-type Arabidopsis thaliana plants grown in different light intensity and culture media conditions}, volume={231}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-010-1142-9}, abstractNote={The number of different anthocyanin molecules potentially produced by Arabidopsis thaliana and which anthocyanin molecule is the first product of anthocyanidin modification remain unknown. To accelerate the understanding of these questions, we investigated anthocyanin biosynthesis in rosette leaves of both pap1-D and wild-type (WT) A. thaliana plants grown in nine growth conditions, which were composed of three light intensities (low light, middle light, and high light) and three media derived from MS medium (medium-1, 2, and 3). These nine growth conditions differentially affected the levels of anthocyanins and pigmentation patterns of rosette leaves, which were closely related to the diversification levels of cyanin structures. The combined growth conditions of high light and either medium-2 or medium-1 induced the most molecular diversity of anthocyanin structures in rosette leaves of pap1-D plants. Twenty cyanin molecules, including five that were previously unknown, were characterized by HPLC-ESI-MS and HPLC-TOF-MS analyses. We detected that the A. thaliana anthocyanin molecule A11 was most likely the first cyanin derived from the multiple modification steps of cyanidin. In addition, in the same growth condition, rosette leaves of pap1-D plants produced much higher levels and more diverse molecular profiling of cyanins than those of WT plants. The transcript levels of PAP1, PAL1, CHS, DFR, and ANS cDNAs were much higher in pap1-D rosette leaves than in WT ones. Furthermore, on the same agar-solidified medium, an enhancement of light intensity increased levels and molecular diversity of cyanins in both pap1-D and WT rosette leaves. In the same light intensity condition, the responses of anthocyanin levels and profiling to medium alternation were different between pap1-D and WT plants.}, number={6}, journal={PLANTA}, author={Shi, Ming-Zhu and Xie, De-Yu}, year={2010}, month={May}, pages={1385–1400} } @article{almaarri_xie_2010, title={In vitro direct organogenesis and micropropagation of Artemisia annua}, volume={26}, journal={Journal of Biotechnologie Vegetale}, author={ALMaarri, Khalil and Xie, De-Yu}, year={2010}, pages={327–337} } @article{almaarri_alamir_junaid_xie_2010, title={Volatile compounds from leaf extracts of Juniperus excelsa growing in Syria via gas chromatography mass spectrometry}, volume={2}, ISSN={["1759-9679"]}, DOI={10.1039/b9ay00256a}, abstractNote={This study reports the use of gas chromatography mass spectrometry (GC-MS) to investigate volatile compounds in leaves of Juniperus excelsa native to Syria. Leaf samples were collected from both 100-year and 10-year old J. excelsa plants. Dried leaf samples were extracted with hexane to obtain essential oil metabolites and other non-polar compounds. GC-MS was used to profile and analyze metabolites in hexane extracts. Mass spectral deconvolution and identification and analysis of KI values allowed us to characterize sixty-nine metabolites, including twenty-four monoterpenes; twenty-nine sesquiterpenes; and sixteen other compounds including alkanes. Among these sixty-nine metabolites, germacrene B, cedrol, γ-elemene, and stenol were produced in leaf extracts of both 100-year and 10-year old trees. Interestingly, we observed that ten monoterpene and nineteen sesquiterpene compounds produced in leaves of the 100-year old trees were not detected in ones of the 10-year old trees investigated. In contrast, junipene was a dominant essential oil component in leaves of the 10-year old trees, but was either just at a detectable level or undetectable in leaves of the 100-year old trees.}, number={6}, journal={ANALYTICAL METHODS}, author={Almaarri, Khalil and Alamir, Lina and Junaid, Yasmin and Xie, De-Yu}, year={2010}, month={Jun}, pages={673–677} } @inproceedings{liu_liu_zhang_cai_chen_xie_2009, title={Extraction of Essential Oil and Anatomical Characterization of Oil Cell from Leaves of Litsea Euosma}, booktitle={Proceedings of 2009 International Conference of Natural Products and Traditional Medicine}, author={Liu, Shibiao and Liu, Zhu-Xiang and Zhang, Hui and Cai, Shi-Jian and Chen, Gong-Xi and Xie, De-Yu}, year={2009}, pages={737–744} } @article{feng_qu_zhou_xie_xiang_2009, title={Shoot regeneration of dwarf dogwood (Cornus canadensis L.) and morphological characterization of the regenerated plants}, volume={97}, ISSN={["1573-5044"]}, DOI={10.1007/s11240-009-9495-0}, number={1}, journal={PLANT CELL TISSUE AND ORGAN CULTURE}, author={Feng, Chun-Miao and Qu, Rongda and Zhou, Li-Li and Xie, De-Yu and Xiang, Qiu-Yun}, year={2009}, month={Apr}, pages={27–37} } @article{zhou_zeng_shi_xie_2008, title={Development of tobacco callus cultures over expressing Arabidopsis PAP1/MYB75 transcription factor and characterization of anthocyanin biosynthesis}, volume={229}, ISSN={0032-0935 1432-2048}, url={http://dx.doi.org/10.1007/s00425-008-0809-y}, DOI={10.1007/s00425-008-0809-y}, abstractNote={The Arabidopsis PAP1 gene (At1g56650) encodes the MYB75 transcription factor, which has been demonstrated to essentially regulate the biosynthesis of anthocyanins. Our previous study showed that ectopic expression of the PAP1 gene led to high pigmentation of anthocyanins in all tissues of transgenic tobacco plants. In order to understand the mechanisms of how PAP1 regulates anthocyanin biosynthesis and what can regulate the function of PAP1, we have established PAP1 transgenic tobacco callus cultures. Phenotypically different calli including anthocyanin-producing red and anthocyanin-free white calli lines were differentially induced from the same genotype of PAP1 transgenic plants. RT-PCR analysis showed that the expression of the PAP1 transgene was similar in the two types of calli, indicating that the transgenic red and white calli had differential responses to the regulation of PAP1. The growth of transgenic red calli followed a "sigmoid-like" curve in a 25-day callus culture period, during which the time course obviously impacted the profiles and the average levels of anthocyanins even though the expression of the PAP1 transgene was constitutive. A HPLC-UV-ESI-mass spectrum-based profiling characterized nine anthocyanin molecules (e.g., 595, 579 and 609 m/z) in the transgenic red calli over the course of the culture period. Cyanidin, pelargonidin, and peonidin were the major anthocyanidins identified by HPLC-mass spectrum analysis. We have demonstrated that dark, nitrogen nutrients, and auxin apparently affect the anthocyanin profiles in PAP1 transgenic callus cultures; and suggest that these cell cultures are an appropriate system to study the regulatory function of PAP1 on the anthocyanin biosynthesis at post-transcriptional level in vivo.}, number={1}, journal={Planta}, publisher={Springer Science and Business Media LLC}, author={Zhou, Li-Li and Zeng, Hai-Nian and Shi, Ming-Zhu and Xie, De-Yu}, year={2008}, month={Sep}, pages={37–51} } @article{bhatnagar_chandrasekharan_xie_hong_2008, title={In vitro studies of tropical woody species}, volume={44}, number={Supplement}, journal={In Vitro Cellular & Developmental Biology - Animal}, author={Bhatnagar, S. and Chandrasekharan, S. and Xie, D.Y. and Hong, Y.}, year={2008}, pages={S54} } @article{stepanova_robertson-hoyt_yun_benavente_xie_doležal_schlereth_jürgens_alonso_2008, title={TAA1-Mediated Auxin Biosynthesis Is Essential for Hormone Crosstalk and Plant Development}, volume={133}, ISSN={0092-8674}, url={http://dx.doi.org/10.1016/j.cell.2008.01.047}, DOI={10.1016/j.cell.2008.01.047}, abstractNote={Plants have evolved a tremendous ability to respond to environmental changes by adapting their growth and development. The interaction between hormonal and developmental signals is a critical mechanism in the generation of this enormous plasticity. A good example is the response to the hormone ethylene that depends on tissue type, developmental stage, and environmental conditions. By characterizing the Arabidopsis wei8 mutant, we have found that a small family of genes mediates tissue-specific responses to ethylene. Biochemical studies revealed that WEI8 encodes a long-anticipated tryptophan aminotransferase, TAA1, in the essential, yet genetically uncharacterized, indole-3-pyruvic acid (IPA) branch of the auxin biosynthetic pathway. Analysis of TAA1 and its paralogues revealed a link between local auxin production, tissue-specific ethylene effects, and organ development. Thus, the IPA route of auxin production is key to generating robust auxin gradients in response to environmental and developmental cues.}, number={1}, journal={Cell}, publisher={Elsevier BV}, author={Stepanova, Anna N. and Robertson-Hoyt, Joyce and Yun, Jeonga and Benavente, Larissa M. and Xie, De-Yu and Doležal, Karel and Schlereth, Alexandra and Jürgens, Gerd and Alonso, Jose M.}, year={2008}, month={Apr}, pages={177–191} } @article{xie_sharma_wright_wang_dixon_2006, title={Metabolic engineering of proanthocyanidins through co-expression of anthocyanidin reductase and the PAP1 MYB transcription factor}, volume={45}, ISSN={0960-7412 1365-313X}, url={http://dx.doi.org/10.1111/j.1365-313x.2006.02655.x}, DOI={10.1111/j.1365-313x.2006.02655.x}, abstractNote={SummaryProanthocyanidins (PAs) and their monomeric building blocks, the (epi)‐flavan‐3‐ols, are plant antioxidants that confer multiple human health benefits. The presence of PAs in forage crops is an important agronomic trait, preventing pasture bloat in ruminant animals. However, many consumed plant materials lack PAs, and there has been little success to date in introducing monomeric or polymeric flavan‐3‐ols de novo into plant tissues for disease prevention by dietary means or development of ‘bloat‐safe’ forages. We report the introduction of PAs into plants by combined expression of a MYB family transcription factor and anthocyanidin reductase for conversion of anthocyanidin into (epi)‐flavan‐3‐ol. Tobacco leaves expressing both transgenes accumulated epicatechin and gallocatechin monomers, and a series of dimers and oligomers consisting primarily of epicatechin units. The levels of PAs reached values that would confer bloat reduction in forage species. Expression of anthocyanidin reductase in anthocyanin‐containing leaves of the forage legume Medicago truncatula resulted in production of a specific subset of PA oligomers.}, number={6}, journal={The Plant Journal}, publisher={Wiley}, author={Xie, De-Yu and Sharma, Shashi B. and Wright, Elane and Wang, Zeng-Yu and Dixon, Richard A.}, year={2006}, month={Mar}, pages={895–907} } @article{xie_dixon_2005, title={Proanthocyanidin biosynthesis – still more questions than answers?}, volume={66}, ISSN={0031-9422}, url={http://dx.doi.org/10.1016/j.phytochem.2005.01.008}, DOI={10.1016/j.phytochem.2005.01.008}, abstractNote={Proanthocyanidins, also known as condensed tannins, are oligomers or polymers of flavan-3-ol units. In spite of important breakthroughs in our understanding of the biosynthesis of the major building blocks of proanthocyanidins, (+)-catechin and (-)-epicatechin, important questions still remain to be answered as to the exact nature of the molecular species that undergo polymerization, and the mechanisms of assembly. We review the structures of proanthocyanidins reported over the past 12 years in the context of biosynthesis, and summarize the outstanding questions concerning synthesis of proanthocyanidins from the chemical, biochemical and molecular genetic perspectives.}, number={18}, journal={Phytochemistry}, publisher={Elsevier BV}, author={Xie, De-Yu and Dixon, Richard A.}, year={2005}, month={Sep}, pages={2127–2144} } @article{xie_jackson_cooper_ferreira_paiva_2004, title={Molecular and Biochemical Analysis of Two cDNA Clones Encoding Dihydroflavonol-4-Reductase from Medicago truncatula}, volume={134}, ISSN={0032-0889 1532-2548}, url={http://dx.doi.org/10.1104/pp.103.030221}, DOI={10.1104/pp.103.030221}, abstractNote={AbstractDihydroflavonol-4-reductase (DFR; EC1.1.1.219) catalyzes a key step late in the biosynthesis of anthocyanins, condensed tannins (proanthocyanidins), and other flavonoids important to plant survival and human nutrition. Two DFR cDNA clones (MtDFR1 and MtDFR2) were isolated from the model legume Medicago truncatula cv Jemalong. Both clones were functionally expressed in Escherichia coli, confirming that both encode active DFR proteins that readily reduce taxifolin (dihydroquercetin) to leucocyanidin. M. truncatula leaf anthocyanins were shown to be cyanidin-glucoside derivatives, and the seed coat proanthocyanidins are known catechin and epicatechin derivatives, all biosynthesized from leucocyanidin. Despite high amino acid similarity (79% identical), the recombinant DFR proteins exhibited differing pH and temperature profiles and differing relative substrate preferences. Although no pelargonidin derivatives were identified in M. truncatula, MtDFR1 readily reduced dihydrokaempferol, consistent with the presence of an asparagine residue at a location known to determine substrate specificity in other DFRs, whereas MtDFR2 contained an aspartate residue at the same site and was only marginally active on dihydrokaempferol. Both recombinant DFR proteins very efficiently reduced 5-deoxydihydroflavonol substrates fustin and dihydrorobinetin, substances not previously reported as constituents of M. truncatula. Transcript accumulation for both genes was highest in young seeds and flowers, consistent with accumulation of condensed tannins and leucoanthocyanidins in these tissues. MtDFR1 transcript levels in developing leaves closely paralleled leaf anthocyanin accumulation. Overexpression of MtDFR1 in transgenic tobacco (Nicotiana tabacum) resulted in visible increases in anthocyanin accumulation in flowers, whereas MtDFR2 did not. The data reveal unexpected properties and differences in two DFR proteins from a single species.}, number={3}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Xie, De-Yu and Jackson, Lisa A. and Cooper, John D. and Ferreira, Daneel and Paiva, Nancy L.}, year={2004}, month={Feb}, pages={979–994} } @article{dixon_xie_sharma_2004, title={Proanthocyanidins - a final frontier in flavonoid research?}, volume={165}, ISSN={0028-646X 1469-8137}, url={http://dx.doi.org/10.1111/j.1469-8137.2004.01217.x}, DOI={10.1111/j.1469-8137.2004.01217.x}, abstractNote={SummaryProanthocyanidins are oligomeric and polymeric end products of the flavonoid biosynthetic pathway. They are present in the fruits, bark, leaves and seeds of many plants, where they provide protection against predation. At the same time they give flavor and astringency to beverages such as wine, fruit juices and teas, and are increasingly recognized as having beneficial effects on human health. The presence of proanthocyanidins is also a major quality factor for forage crops. The past 2 years have seen important breakthroughs in our understanding of the biosynthesis of the building blocks of proanthocyanidins, the flavan‐3‐ols (+)‐catechin and (–)‐epicatechin. However, virtually nothing is known about the ways in which these units are assembled into the corresponding oligomers in vivo. Molecular genetic approaches are leading to an understanding of the regulatory genes that control proanthocyanidin biosynthesis, and this information, together with increased knowledge of the enzymes specific for the pathway, will facilitate the genetic engineering of plants for introduction of value‐added nutraceutical and forage quality traits. Contents Summary 9 I. Introduction 9 II. Sources and structures of proanthocyanidins 10 III. Functions of proanthocyanidins in the plant 12 IV. Proanthocyanidins and plant quality traits 13 V. Flavanols, proanthocyanidins and human health 14 VI. Biosynthesis of proanthocyanidins 16 VII. Genetic manipulation of the proanthocyanidin pathway 23 VIII. Conclusions and future prospects 24 Acknowledgements 25 References 25 }, number={1}, journal={New Phytologist}, publisher={Wiley}, author={Dixon, Richard A. and Xie, De-Yu and Sharma, Shashi B.}, year={2004}, month={Nov}, pages={9–28} } @article{xie_sharma_dixon_2004, title={Anthocyanidin reductases from Medicago truncatula and Arabidopsis thaliana}, volume={422}, ISSN={0003-9861}, url={http://dx.doi.org/10.1016/j.abb.2003.12.011}, DOI={10.1016/j.abb.2003.12.011}, abstractNote={Anthocyanidin reductase (ANR), encoded by the BANYULS gene, is a newly discovered enzyme of the flavonoid pathway involved in the biosynthesis of condensed tannins. ANR functions immediately downstream of anthocyanidin synthase to convert anthocyanidins into the corresponding 2,3-cis-flavan-3-ols. We report the biochemical properties of ANRs from the model legume Medicago truncatula (MtANR) and the model crucifer Arabidopsis thaliana (AtANR). Both enzymes have high temperature optima. MtANR uses both NADPH and NADH as reductant with slight preference for NADPH over NADH. In contrast, AtANR only uses NADPH and exhibits positive cooperativity for the co-substrate. MtANR shows preference for potential anthocyanidin substrates in the order cyanidin>pelargonidin>delphinidin, with typical Michaelis-Menten kinetics for each substrate. In contrast, AtANR exhibits the reverse preference, with substrate inhibition at high concentrations of cyanidin and pelargonidin. (+)-Catechin and (+/-)-dihydroquercetin inhibit AtANR but not MtANR, whereas quercetin inhibits both enzymes. Possible catalytic reaction sequences for ANRs are discussed.}, number={1}, journal={Archives of Biochemistry and Biophysics}, publisher={Elsevier BV}, author={Xie, De-Yu and Sharma, Shashi B and Dixon, Richard A}, year={2004}, month={Feb}, pages={91–102} } @article{xie_2003, title={Role of Anthocyanidin Reductase, Encoded by BANYULS in Plant Flavonoid Biosynthesis}, volume={299}, ISSN={0036-8075 1095-9203}, url={http://dx.doi.org/10.1126/science.1078540}, DOI={10.1126/science.1078540}, abstractNote={ Condensed tannins (CTs) are flavonoid oligomers, many of which have beneficial effects on animal and human health. The flavanol (–)-epicatechin is a component of many CTs and contributes to flavor and astringency in tea and wine. We show that the BANYULS ( BAN ) genes from Arabidopsis thaliana and Medicago truncatula encode anthocyanidin reductase, which converts anthocyanidins to their corresponding 2,3- cis -flavan-3-ols. Ectopic expression of BAN in tobacco flower petals and Arabidopsis leaves results in loss of anthocyanins and accumulation of CTs. }, number={5605}, journal={Science}, publisher={American Association for the Advancement of Science (AAAS)}, author={Xie, D.-Y.}, year={2003}, month={Jan}, pages={396–399} } @article{xie_hong_2002, title={Agrobacterium-mediated genetic transformation of Acacia mangium}, volume={20}, ISSN={0721-7714 1432-203X}, url={http://dx.doi.org/10.1007/s00299-001-0397-9}, DOI={10.1007/s00299-001-0397-9}, number={10}, journal={Plant Cell Reports}, publisher={Springer Science and Business Media LLC}, author={Xie, D. and Hong, Y.}, year={2002}, month={Mar}, pages={917–922} } @article{xie_hong_2001, volume={66}, ISSN={0167-6857}, url={http://dx.doi.org/10.1023/a:1010632619342}, DOI={10.1023/a:1010632619342}, number={3}, journal={Plant Cell, Tissue and Organ Culture}, publisher={Springer Science and Business Media LLC}, author={Xie, Deyu and Hong, Yan}, year={2001}, pages={167–173} } @article{xie_hong_2001, title={Regeneration of Acacia mangium through somatic embryogenesis}, volume={20}, ISSN={0721-7714 1432-203X}, url={http://dx.doi.org/10.1007/s002990000288}, DOI={10.1007/s002990000288}, abstractNote={ Somatic embryogenesis and whole plant regeneration were achieved in callus cultures derived from immature zygotic embryos of Acacia mangium. Embryogenic callus was induced on MS medium containing combinations of TDZ (1-2 mg/l), IAA (0.25-2 mg/l) and a mixture of amino acids. Globular embryos developed on embryogenic callus cultured on the induction medium. Nearly 42% of embryogenic cultures with globular embryos produced torpedo- and cotyledonary-stage embryos by a two-step maturation phase. The first stage occurred on 1/2-strength MS basal medium containing 30 g/l sucrose and 5 mg/l GA 3 followed by the second stage on 1/2-strength MS basal medium containing 50 g/l sucrose. Of the cotyledonary-stage somatic embryos, 11% germinated into seedlings that could be successfully transferred to pots. Light- and scanning electron microscopy showed that the somatic embryos originated from single cells of the embryogenic callus. Further, a single cell layer could be detected beneath the developing somatic embryos that appeared to be a demarcation layer isolating the somatic proembryonic structure from the rest of the maternal callus. A suspensor-like structure connected the globular embryos to the demarcation layer. This is the first successful report of plant regeneration through somatic embryogenesis for this economically important tropical forest species.}, number={1}, journal={Plant Cell Reports}, publisher={Springer Science and Business Media LLC}, author={Xie, D. Y. and Hong, Y.}, year={2001}, month={Jan}, pages={34–40} } @article{xie_ye_li_guo_zou_guo_2001, title={Selection of hairy root clones of Artemisia annua L. for artemisinin production}, volume={49}, ISSN={0792-9978}, url={http://dx.doi.org/10.1560/n11n-6blg-er7c-xkct}, DOI={10.1560/n11n-6blg-er7c-xkct}, abstractNote={Hairy roots were induced from two kinds of explants and selection of hairy root clones was studied for artemisin in production. Leaf blade pieces and petiole segments of Artemisia annua plantlets were infected with Agrobacterium rhizogenesstrain 1601. The efficiency of leaf blade pieces forming hairy roots was higher than that of petiole segments. Light promoted hairy root induction and branching. Two hundred and twenty hairy root clones showed considerable variations in capacity of growth and branching. Six hairy root clones were established in suspension culture and showed obvious differences in their biomass and artemisin in content. Among six clones, clone 1601-L-3 produced the highest biomass, more than 70 times the inoculum, while clone 1601-L-1 gave the lowest biomass. The artemisin in content of clone 1601-L-1 was the highest, 1.195 mg/g DW, and this line yielded the highest artemisinin level of 9.08 mg/L.}, number={2}, journal={Israel Journal of Plant Sciences}, publisher={Laser Pages Publishing Ltd.}, author={Xie, Deyu and Ye, Hechun and Li, Guofeng and Guo, Zhongchen and Zou, Zhuorong and Guo, Zhongchen}, year={2001}, month={Jan}, pages={129–134} } @article{xie_wang_ye_li_2000, volume={63}, ISSN={0167-6857}, url={http://dx.doi.org/10.1023/a:1006438919841}, DOI={10.1023/a:1006438919841}, number={2}, journal={Plant Cell, Tissue and Organ Culture}, publisher={Springer Science and Business Media LLC}, author={Xie, Deyu and Wang, Lianhui and Ye, Hechun and Li, Guofeng}, year={2000}, pages={161–166} } @article{xie_guo_1999, title={ZYGOTIC EMBRYO CULTURE OF TAXUS CHINESIS VAR. MAIREI AND PLANT REGENERATION THROUGH ORGANOGENESIS}, volume={47}, ISSN={0792-9978 2223-8980}, url={http://dx.doi.org/10.1080/07929978.1999.10676786}, DOI={10.1080/07929978.1999.10676786}, abstractNote={When different media were compared for their effect on zygotic embryo culture, DCR was found more effective than MS medium. From these embryos, adventitious bud induction was achieved by optimizing formular conditions as described in this paper. Adventitious buds were formed from calli derived from hypocotyls of germinated embryos. Finally, adventitious roots were induced and complete plantlets were obtained.}, number={4}, journal={Israel Journal of Plant Sciences}, publisher={Brill}, author={Xie, Deyu and Guo, Zhongchen}, year={1999}, month={May}, pages={287–289} } @inproceedings{xie_ye_li_guo_1998, place={Fyshwick, Canberra}, title={Artemisia annua L. Transformation with different Agrobacterium rhizogenesis and large scale culture of hairy roots for Artemisinin (Qinghaosu) production}, booktitle={Agricultural biotechnology : laboratory, field and market : proceedings of the 4th Asia-Pacific Conference on Agricultural Biotechnology, 13-16 July 1998, Darwin, Australia}, publisher={Under the Counter Publishing}, author={Xie, Deyu and Ye, Hechun and Li, Guofeng and Guo, Zhongchen}, editor={Larkin, P.J.Editor}, year={1998}, pages={134–136} } @article{xie_kang_li_1995, title={Studies on the karyotype of Artemisia annua}, volume={12}, number={Supplement}, journal={Chinese Bulletin of Botany}, author={Xie, De-Yu and Kang, Ning-ling and Li, Guo-zhen}, year={1995}, pages={71–72} } @article{xie_ye_li_1995, title={The Progress of Artemisia annua research--the application of biotechnology and prospects}, volume={12}, number={4}, journal={Chinese Bulletin of Botany}, author={Xie, De-Yu and Ye, Hechun and Li, Guofeng}, year={1995}, pages={28–31} } @article{xie_li_ye_li_1993, title={Studies on the relation between the changes of endogenous IPA and the growth of cell of Arnebia euchroma during the culture}, volume={14}, number={6}, journal={Journal of Jishou University (Natural Science)}, author={Xie, De-Yu and Li, Guozhen and Ye, Hechun and Li, Guofeng}, year={1993}, pages={29–31} } @article{li_qi_kang_xie_ye_li_1992, title={Tissue culture and chromosome analysis of Arnebia euchroma}, volume={9}, number={1}, journal={Chinese Bulletin of Botany}, author={Li, Guozhen and Qi, Mingpo and Kang, Ningling and Xie, Deyu and Ye, Yechun and Li, Guofeng}, year={1992}, pages={37–41} } @inproceedings{dixon_xie_sharma_chen_ferreira, place={Washington, DC}, title={Biochemical and molecular genetic approaches to proanthocyanidin biosynthesis}, volume={228}, booktitle={Abstracts of papers : 228th ACS National Meeting, American Chemical Society, Philadelphia, PA, August 22-26, 2004}, publisher={American Chemical Society}, author={Dixon, R.A. and Xie, D.Y. and Sharma, S.B. and Chen, F. and Ferreira, D.}, pages={U247} }