Plant biomass fermentation by the extreme thermophile Caldicellulosiruptor bescii for co-production of green hydrogen and acetone: Technoeconomic analysis
}, volume={348}, ISSN={["1873-2976"]}, url={http://europepmc.org/abstract/med/35093526}, DOI={10.1016/j.biortech.2022.126780}, abstractNote={A variety of chemical and biological processes have been proposed for conversion of sustainable low-cost feedstocks into industrial products. Here, a biorefinery concept is formulated, modeled, and analyzed in which a naturally (hemi)cellulolytic and extremely thermophilic bacterium, Caldicellulosiruptor bescii, is metabolically engineered to convert the carbohydrate content of lignocellulosic biomasses (i.e., soybean hulls, transgenic poplar) into green hydrogen and acetone. Experimental validation of C. bescii fermentative performance demonstrated 82% carbohydrate solubilization of soybean hulls and 55% for transgenic poplar. A detailed technical design, including equipment specifications, provides the basis for an economic analysis that establishes metabolic engineering targets. This robust industrial process leveraging metabolically engineered C. bescii yields 206 kg acetone and 25 kg H2 per metric ton of soybean hull, or 174 kg acetone and 21 kg H2 per metric ton transgenic poplar. Beyond this specific case, the model demonstrates industrial feasibility and economic advantages of thermophilic fermentation.}, journal={BIORESOURCE TECHNOLOGY}, author={Bing, Ryan G. and Straub, Christopher T. and Sulis, Daniel B. and Wang, Jack P. and Adams, Michel W. W. and Kelly, Robert M.}, year={2022}, month={Mar} } @article{liu_gao_sun_li_zhang_wang_zhou_sulis_wang_chiang_et al._2022, title={Dimerization of PtrMYB074 and PtrWRKY19 mediates transcriptional activation of PtrbHLH186 for secondary xylem development in Populus trichocarpa}, volume={5}, ISSN={["1469-8137"]}, url={http://europepmc.org/abstract/med/35152419}, DOI={10.1111/nph.18028}, abstractNote={Summary}, journal={NEW PHYTOLOGIST}, author={Liu, Huizi and Gao, Jinghui and Sun, Jiatong and Li, Shuang and Zhang, Baofeng and Wang, Zhuwen and Zhou, Chenguang and Sulis, Daniel Barletta and Wang, Jack P. and Chiang, Vincent L. and et al.}, year={2022}, month={Mar} } @article{chen_neubauer_wang_2022, title={Enhancing HR Frequency for Precise Genome Editing in Plants}, volume={13}, ISSN={["1664-462X"]}, url={http://europepmc.org/abstract/med/35592579}, DOI={10.3389/fpls.2022.883421}, abstractNote={Gene-editing tools, such as Zinc-fingers, TALENs, and CRISPR-Cas, have fostered a new frontier in the genetic improvement of plants across the tree of life. In eukaryotes, genome editing occurs primarily through two DNA repair pathways: non-homologous end joining (NHEJ) and homologous recombination (HR). NHEJ is the primary mechanism in higher plants, but it is unpredictable and often results in undesired mutations, frameshift insertions, and deletions. Homology-directed repair (HDR), which proceeds through HR, is typically the preferred editing method by genetic engineers. HR-mediated gene editing can enable error-free editing by incorporating a sequence provided by a donor template. However, the low frequency of native HR in plants is a barrier to attaining efficient plant genome engineering. This review summarizes various strategies implemented to increase the frequency of HDR in plant cells. Such strategies include methods for targeting double-strand DNA breaks, optimizing donor sequences, altering plant DNA repair machinery, and environmental factors shown to influence HR frequency in plants. Through the use and further refinement of these methods, HR-based gene editing may one day be commonplace in plants, as it is in other systems.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Chen, Hao and Neubauer, Matthew and Wang, Jack P.}, year={2022}, month={May} } @article{yu_zhou_li_li_lin_wang_chiang_li_2022, title={p A PtrLBD39-mediated transcriptional network regulates tension wood formation in Populus trichocarpa}, volume={3}, ISSN={["2590-3462"]}, url={http://europepmc.org/abstract/med/35059630}, DOI={10.1016/j.xplc.2021.100250}, abstractNote={Tension wood (TW) is a specialized xylem tissue formed in angiosperm trees under gravitational stimulus or mechanical stresses (e.g., bending). The genetic regulation that underlies this important mechanism remains poorly understood. Here, we used laser capture microdissection of stem xylem cells coupled with full transcriptome RNA-sequencing to analyze TW formation in Populus trichocarpa. After tree bending, PtrLBD39 was the most significantly induced transcription factor gene; it has a phylogenetically paired homolog, PtrLBD22. CRISPR-based knockout of PtrLBD39/22 severely inhibited TW formation, reducing cellulose and increasing lignin content. Transcriptomic analyses of CRISPR-based PtrLBD39/22 double mutants showed that these two genes regulate a set of TW-related genes. Chromatin immunoprecipitation sequencing (ChIP-seq) was used to identify direct targets of PtrLBD39. We integrated transcriptomic analyses and ChIP-seq assays to construct a transcriptional regulatory network (TRN) mediated by PtrLBD39. In this TRN, PtrLBD39 directly regulates 26 novel TW-responsive transcription factor genes. Our work suggests that PtrLBD39 and PtrLBD22 specifically control TW formation by mediating a TW-specific TRN in Populus.}, number={1}, journal={PLANT COMMUNICATIONS}, author={Yu, Jing and Zhou, Chenguang and Li, Danning and Li, Shuang and Lin, Ying-Chung Jimmy and Wang, Jack P. and Chiang, Vincent L. and Li, Wei}, year={2022}, month={Jan} } @article{matthews_wang_sederoff_chiang_williams_2021, title={A multiscale model of lignin biosynthesis for predicting bioenergy traits in Populus trichocarpa}, volume={19}, ISSN={["2001-0370"]}, url={http://europepmc.org/abstract/med/33425249}, DOI={10.1016/j.csbj.2020.11.046}, abstractNote={Understanding the mechanisms behind lignin formation is an important research area with significant implications for the bioenergy and biomaterial industries. Computational models are indispensable tools for understanding this complex process. Models of the monolignol pathway in Populus trichocarpa and other plants have been developed to explore how transgenic modifications affect important bioenergy traits. Many of these models, however, only capture one level of biological organization and are unable to capture regulation across multiple biological scales. This limits their ability to predict how gene modification strategies will impact lignin and other wood properties. While the first multiscale model of lignin biosynthesis in P. trichocarpa spanned the transcript, protein, metabolic, and phenotypic layers, it did not account for cross-regulatory influences that could impact abundances of untargeted monolignol transcripts and proteins. Here, we present a multiscale model incorporating these cross-regulatory influences for predicting lignin and wood traits from transgenic knockdowns of the monolignol genes. The three main components of this multiscale model are (1) a transcript-protein model capturing cross-regulatory influences, (2) a kinetic-based metabolic model, and (3) random forest models relating the steady state metabolic fluxes to 25 physical traits. We demonstrate that including the cross-regulatory behavior results in smaller predictive error for 23 of the 25 traits. We use this multiscale model to explore the predicted impact of novel combinatorial knockdowns on key bioenergy traits, and identify the perturbation of PtrC3H3 and PtrCAld5H1&2 monolignol genes as a candidate strategy for increasing saccharification efficiencies while reducing negative impacts on wood density and height.}, journal={COMPUTATIONAL AND STRUCTURAL BIOTECHNOLOGY JOURNAL}, author={Matthews, Megan L. and Wang, Jack P. and Sederoff, Ronald and Chiang, Vincent L. and Williams, Cranos M.}, year={2021}, pages={168–182} } @article{vries_brouckaert_chanoca_kim_regner_timokhin_sun_de meester_van doorsselaere_goeminne_et al._2021, title={CRISPR-Cas9 editing of CAFFEOYL SHIKIMATE ESTERASE 1 and 2 shows their importance and partial redundancy in lignification in Populus tremula x P. alba}, volume={11}, ISSN={["1467-7652"]}, url={http://europepmc.org/abstract/med/34160888}, DOI={10.1111/pbi.13651}, abstractNote={Summary}, journal={PLANT BIOTECHNOLOGY JOURNAL}, author={Vries, Lisanne and Brouckaert, Marlies and Chanoca, Alexandra and Kim, Hoon and Regner, Matthew R. and Timokhin, Vitaliy I. and Sun, Yi and De Meester, Barbara and Van Doorsselaere, Jan and Goeminne, Geert and et al.}, year={2021}, month={Aug} } @article{lin_sun_song_chen_shi_yang_liu_tunlaya-anukit_liu_loziuk_et al._2021, title={Enzyme Complexes of Ptr4CL and PtrHCT Modulate Co-enzyme A Ligation of Hydroxycinnamic Acids for Monolignol Biosynthesis in Populus trichocarpa}, volume={12}, ISSN={["1664-462X"]}, url={http://europepmc.org/abstract/med/34691108}, DOI={10.3389/fpls.2021.727932}, abstractNote={Co-enzyme A (CoA) ligation of hydroxycinnamic acids by 4-coumaric acid:CoA ligase (4CL) is a critical step in the biosynthesis of monolignols. Perturbation of 4CL activity significantly impacts the lignin content of diverse plant species. InPopulus trichocarpa, two well-studied xylem-specific Ptr4CLs (Ptr4CL3 and Ptr4CL5) catalyze the CoA ligation of 4-coumaric acid to 4-coumaroyl-CoA and caffeic acid to caffeoyl-CoA. Subsequently, two 4-hydroxycinnamoyl-CoA:shikimic acid hydroxycinnamoyl transferases (PtrHCT1 and PtrHCT6) mediate the conversion of 4-coumaroyl-CoA to caffeoyl-CoA. Here, we show that the CoA ligation of 4-coumaric and caffeic acids is modulated by Ptr4CL/PtrHCT protein complexes. Downregulation ofPtrHCTsreduced Ptr4CL activities in the stem-differentiating xylem (SDX) of transgenicP. trichocarpa. The Ptr4CL/PtrHCT interactions were then validatedin vivousing biomolecular fluorescence complementation (BiFC) and protein pull-down assays inP. trichocarpaSDX extracts. Enzyme activity assays using recombinant proteins of Ptr4CL and PtrHCT showed elevated CoA ligation activity for Ptr4CL when supplemented with PtrHCT. Numerical analyses based on an evolutionary computation of the CoA ligation activity estimated the stoichiometry of the protein complex to consist of one Ptr4CL and two PtrHCTs, which was experimentally confirmed by chemical cross-linking using SDX plant protein extracts and recombinant proteins. Based on these results, we propose that Ptr4CL/PtrHCT complexes modulate the metabolic flux of CoA ligation for monolignol biosynthesis during wood formation inP. trichocarpa.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Lin, Chien-Yuan and Sun, Yi and Song, Jina and Chen, Hsi-Chuan and Shi, Rui and Yang, Chenmin and Liu, Jie and Tunlaya-Anukit, Sermsawat and Liu, Baoguang and Loziuk, Philip L. and et al.}, year={2021}, month={Oct} } @article{bing_sulis_wang_adams_kelly_2021, title={Thermophilic microbial deconstruction and conversion of natural and transgenic lignocellulose}, volume={13}, ISSN={1758-2229 1758-2229}, url={http://dx.doi.org/10.1111/1758-2229.12943}, DOI={10.1111/1758-2229.12943}, abstractNote={Summary}, number={3}, journal={Environmental Microbiology Reports}, publisher={Wiley}, author={Bing, Ryan G. and Sulis, Daniel B. and Wang, Jack P. and Adams, Michael W. W. and Kelly, Robert M.}, year={2021}, month={Mar}, pages={272–293} } @article{liu_liu_yu_wang_sun_li_lin_chiang_li_wang_2021, title={Transcriptional reprogramming of xylem cell wall biosynthesis in tension wood}, volume={186}, ISSN={["1532-2548"]}, url={https://doi.org/10.1093/plphys/kiab038}, DOI={10.1093/plphys/kiab038}, abstractNote={Abstract}, number={1}, journal={PLANT PHYSIOLOGY}, publisher={Oxford University Press (OUP)}, author={Liu, Baoguang and Liu, Juan and Yu, Jing and Wang, Zhifeng and Sun, Yi and Li, Shuang and Lin, Ying-Chung Jimmy and Chiang, Vincent L. and Li, Wei and Wang, Jack P.}, year={2021}, month={May}, pages={250–269} } @article{wang_mao_guo_gao_liu_li_lin_chen_wang_chiang_et al._2020, title={MYB Transcription Factor161 Mediates Feedback Regulation of Secondary wall-associated NAC-Domain1 Family Genes for Wood Formation}, volume={9}, url={https://doi.org/10.1104/pp.20.01033}, DOI={10.1104/pp.20.01033}, abstractNote={Wood formation is a complex process that involves cell differentiation, cell expansion, secondary wall deposition, and programmed cell death. We constructed a four-layer wood formation transcriptional regulatory network (TRN) in Populus trichocarpa (black cottonwood) that has four Secondary wall-associated NAC-Domain1 (PtrSND1) transcription factor (TF) family members as the top-layer regulators. We characterized the function of a MYB (PtrMYB161) TF in this PtrSND1-TRN, using transgenic P. trichocarpa cells and whole plants. PtrMYB161 is a third-layer regulator that directly transactivates five wood formation genes. Overexpression of PtrMYB161 in P. trichocarpa (OE-PtrMYB161) led to reduced wood, altered cell type proportions, and inhibited growth. Integrative analysis of wood cell-based chromatin-binding assays with OE-PtrMYB161 transcriptomics revealed a feedback regulation system in the PtrSND1-TRN, where PtrMYB161 represses all four top-layer regulators and one second-layer regulator, PtrMYB021, possibly affecting many downstream TFs in, and likely beyond, the TRN, to generate the observed phenotypic changes. Our data also suggested that the PtrMYB161's repressor function operates through interaction of the base PtrMYB161 target-binding system with gene-silencing cofactors. PtrMYB161 protein does not contain any known negative regulatory domains. CRISPR-based mutants of PtrMYB161 in P. trichocarpa exhibited phenotypes similar to the wild type, suggesting that PtrMYB161's activator functions are redundant among many TFs. Our work demonstrated that PtrMYB161 binds to multiple sets of target genes, a feature that allows it to function as an activator as well as a repressor. The balance of the two functions may be important to the establishment of regulatory homeostasis for normal growth and development.}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Wang, Zhifeng and Mao, Yuli and Guo, Yanjiao and Gao, Jinghui and Liu, Xinying and Li, Shuang and Lin, Ying-Chung Jimmy and Chen, Hao and Wang, Jack P. and Chiang, Vincent L. and et al.}, year={2020}, month={Nov}, pages={pp.01033.2020} } @article{matthews_wang_sederoff_chiang_williams_2020, title={Modeling cross-regulatory influences on monolignol transcripts and proteins under single and combinatorial gene knockdowns in Populus trichocarpa}, url={https://doi.org/10.1371/journal.pcbi.1007197}, DOI={10.1371/journal.pcbi.1007197}, abstractNote={Accurate manipulation of metabolites in monolignol biosynthesis is a key step for controlling lignin content, structure, and other wood properties important to the bioenergy and biomaterial industries. A crucial component of this strategy is predicting how single and combinatorial knockdowns of monolignol specific gene transcripts influence the abundance of monolignol proteins, which are the driving mechanisms of monolignol biosynthesis. Computational models have been developed to estimate protein abundances from transcript perturbations of monolignol specific genes. The accuracy of these models, however, is hindered by their inability to capture indirect regulatory influences on other pathway genes. Here, we examine the manifestation of these indirect influences on transgenic transcript and protein abundances, identifying putative indirect regulatory influences that occur when one or more specific monolignol pathway genes are perturbed. We created a computational model using sparse maximum likelihood to estimate the resulting monolignol transcript and protein abundances in transgenic Populus trichocarpa based on targeted knockdowns of specific monolignol genes. Using in-silico simulations of this model and root mean square error, we showed that our model more accurately estimated transcript and protein abundances, in comparison to previous models, when individual and families of monolignol genes were perturbed. We leveraged insight from the inferred network structure obtained from our model to identify potential genes, including PtrHCT, PtrCAD, and Ptr4CL, involved in post-transcriptional and/or post-translational regulation. Our model provides a useful computational tool for exploring the cascaded impact of single and combinatorial modifications of monolignol specific genes on lignin and other wood properties.}, journal={PLOS Computational Biology}, author={Matthews, Megan L. and Wang, Jack P. and Sederoff, Ronald and Chiang, Vincent L. and Williams, Cranos M.}, editor={Roy, SushmitaEditor}, year={2020}, month={Apr} } @article{kim_li_karlen_smith_shi_liu_yang_tunlaya-anukit_wang_chang_et al._2020, title={Monolignol Benzoates Incorporate into the Lignin of Transgenic Populus trichocarpa Depleted in C3H and C4H}, volume={8}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85082016107&partnerID=MN8TOARS}, DOI={10.1021/acssuschemeng.9b06389}, abstractNote={Lignin, an abundant renewable aromatic biopolymer, is an essential macromolecule in terrestrial vascular plants. Genetic modifications affecting monolignol biosynthesis, which produces lignin monom...}, number={9}, journal={ACS Sustainable Chemistry and Engineering}, author={Kim, H. and Li, Q. and Karlen, S.D. and Smith, R.A. and Shi, R. and Liu, J. and Yang, C. and Tunlaya-Anukit, S. and Wang, J.P. and Chang, H.-M. and et al.}, year={2020}, pages={3644–3654} } @misc{sulis_wang_2020, title={Regulation of Lignin Biosynthesis by Post-translational Protein Modifications}, volume={11}, ISSN={["1664-462X"]}, url={http://europepmc.org/abstract/med/32714349}, DOI={10.3389/fpls.2020.00914}, abstractNote={Post-translational modification of proteins exerts essential roles in many biological processes in plants. The function of these chemical modifications has been extensively characterized in many physiological processes, but how these modifications regulate lignin biosynthesis for wood formation remained largely unknown. Over the past decade, post-translational modification of several proteins has been associated with lignification. Phosphorylation, ubiquitination, glycosylation, and S-nitrosylation of transcription factors, monolignol enzymes, and peroxidases were shown to have primordial roles in the regulation of lignin biosynthesis. The main discoveries of post-translational modifications in lignin biosynthesis are discussed in this review.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Sulis, Daniel B. and Wang, Jack P.}, year={2020}, month={Jul} } @article{liu_wang_2020, title={Tracheid Associated Transcription Factors in Loblolly Pine.}, volume={2}, url={http://europepmc.org/abstract/med/32050028}, DOI={10.1093/treephys/tpaa014}, journal={Tree physiology}, author={Liu, B and Wang, JP}, year={2020}, month={Feb} } @article{straub_bing_wang_chiang_adams_kelly_2020, title={Use of the lignocellulose-degrading bacterium Caldicellulosiruptor bescii to assess recalcitrance and conversion of wild-type and transgenic poplar}, volume={13}, ISSN={["1754-6834"]}, url={http://europepmc.org/abstract/med/32180826}, DOI={10.1186/s13068-020-01675-2}, abstractNote={Abstract}, number={1}, journal={BIOTECHNOLOGY FOR BIOFUELS}, author={Straub, Christopher T. and Bing, Ryan G. and Wang, Jack P. and Chiang, Vincent L. and Adams, Michael W. W. and Kelly, Robert M.}, year={2020}, month={Mar} } @article{yeh_wang_miao_ma_kao_hsu_yu_hung_lin_kuan_et al._2019, title={A novel synthetic-genetic-array-based yeast one-hybrid system for high discovery rate and short processing time}, volume={29}, ISSN={["1549-5469"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85071056042&partnerID=MN8TOARS}, DOI={10.1101/gr.245951.118}, abstractNote={Eukaryotic gene expression is often tightly regulated by interactions between transcription factors (TFs) and their DNA cis targets. Yeast one-hybrid (Y1H) is one of the most extensively used methods to discover these interactions. We developed a high-throughput meiosis-directed yeast one-hybrid system using the Magic Markers of the synthetic genetic array analysis. The system has a transcription factor–DNA interaction discovery rate twice as high as the conventional diploid-mating approach and a processing time nearly one-tenth of the haploid-transformation method. The system also offers the highest accuracy in identifying TF–DNA interactions that can be authenticated in vivo by chromatin immunoprecipitation. With these unique features, this meiosis-directed Y1H system is particularly suited for constructing novel and comprehensive genome-scale gene regulatory networks for various organisms.}, number={8}, journal={GENOME RESEARCH}, author={Yeh, Chung-Shu and Wang, Zhifeng and Miao, Fang and Ma, Hongyan and Kao, Chung-Ting and Hsu, Tzu-Shu and Yu, Jhong-He and Hung, Er-Tsi and Lin, Chia-Chang and Kuan, Chen-Yu and et al.}, year={2019}, month={Aug}, pages={1343–1351} } @article{yan_liu_kim_liu_huang_yang_lin_chen_yang_wang_et al._2019, title={CAD1 and CCR2 protein complex formation in monolignol biosynthesis in Populus trichocarpa}, volume={222}, ISSN={["1469-8137"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85055556739&partnerID=MN8TOARS}, DOI={10.1111/nph.15505}, abstractNote={Summary}, number={1}, journal={NEW PHYTOLOGIST}, author={Yan, Xiaojing and Liu, Jie and Kim, Hoon and Liu, Baoguang and Huang, Xiong and Yang, Zhichang and Lin, Ying-Chung Jimmy and Chen, Hao and Yang, Chenmin and Wang, Jack P. and et al.}, year={2019}, month={Apr}, pages={244–260} } @misc{wang_liu_sun_chiang_sederoff_2019, title={Enzyme-Enzyme Interactions in Monolignol Biosynthesis}, volume={9}, ISSN={["1664-462X"]}, url={http://dx.doi.org/10.3389/fpls.2018.01942}, DOI={10.3389/fpls.2018.01942}, abstractNote={The enzymes that comprise the monolignol biosynthetic pathway have been studied intensively for more than half a century. A major interest has been the role of pathway in the biosynthesis of lignin and the role of lignin in the formation of wood. The pathway has been typically conceived as linear steps that convert phenylalanine into three major monolignols or as a network of enzymes in a metabolic grid. Potential interactions of enzymes have been investigated to test models of metabolic channeling or for higher order interactions. Evidence for enzymatic or physical interactions has been fragmentary and limited to a few enzymes studied in different species. Only recently the entire pathway has been studied comprehensively in any single plant species. Support for interactions comes from new studies of enzyme activity, co-immunoprecipitation, chemical crosslinking, bimolecular fluorescence complementation, yeast 2-hybrid functional screening, and cell type–specific gene expression based on light amplification by stimulated emission of radiation capture microdissection. The most extensive experiments have been done on differentiating xylem of Populus trichocarpa, where genomic, biochemical, chemical, and cellular experiments have been carried out. Interactions affect the rate, direction, and specificity of both 3 and 4-hydroxylation in the monolignol biosynthetic pathway. Three monolignol P450 mono-oxygenases form heterodimeric and heterotetrameric protein complexes that activate specific hydroxylation of cinnamic acid derivatives. Other interactions include regulatory kinetic control of 4-coumarate CoA ligases through subunit specificity and interactions between a cinnamyl alcohol dehydrogenase and a cinnamoyl-CoA reductase. Monolignol enzyme interactions with other pathway proteins have been associated with biotic and abiotic stress response. Evidence challenging or supporting metabolic channeling in this pathway will be discussed.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Wang, Jack P. and Liu, Baoguang and Sun, Yi and Chiang, Vincent L. and Sederoff, Ronald R.}, year={2019}, month={Jan} } @article{wang_matthews_naik_williams_ducoste_sederoff_chiang_2019, title={Flux modeling for monolignol biosynthesis}, volume={56}, ISSN={0958-1669}, url={http://dx.doi.org/10.1016/J.COPBIO.2018.12.003}, DOI={10.1016/j.copbio.2018.12.003}, abstractNote={The pathway of monolignol biosynthesis involves many components interacting in a metabolic grid to regulate the supply and ratios of monolignols for lignification. The complexity of the pathway challenges any intuitive prediction of the output without mathematical modeling. Several models have been presented to quantify the metabolic flux for monolignol biosynthesis and the regulation of lignin content, composition, and structure in plant cell walls. Constraint-based models using data from transgenic plants were formulated to describe steady-state flux distribution in the pathway. Kinetic-based models using enzyme reaction and inhibition constants were developed to predict flux dynamics for monolignol biosynthesis in wood-forming cells. This review summarizes the recent progress in flux modeling and its application to lignin engineering for improved plant development and utilization.}, journal={Current Opinion in Biotechnology}, publisher={Elsevier BV}, author={Wang, Jack P and Matthews, Megan L and Naik, Punith P and Williams, Cranos M and Ducoste, Joel J and Sederoff, Ronald R and Chiang, Vincent L}, year={2019}, month={Apr}, pages={187–192} } @article{chen_wang_liu_li_lin_shi_yang_gao_zhou_li_et al._2019, title={Hierarchical Transcription Factor and Chromatin Binding Network for Wood Formation in Populus trichocarpa}, volume={31}, url={https://doi.org/10.1105/tpc.18.00620}, DOI={10.1105/tpc.18.00620}, abstractNote={A hierarchical transcriptional regulatory network for wood formation in Populus contains 57 direct transcription factor (TF)–DNA interactions linking 17 TFs regulating 27 cell wall biosynthetic genes. Wood remains the world’s most abundant and renewable resource for timber and pulp and is an alternative to fossil fuels. Understanding the molecular regulation of wood formation can advance the engineering of wood for more efficient material and energy productions. We integrated a black cottonwood (Populus trichocarpa) wood-forming cell system with quantitative transcriptomics and chromatin binding assays to construct a transcriptional regulatory network (TRN) directed by a key transcription factor (TF), PtrSND1-B1 (secondary wall-associated NAC-domain protein). The network consists of four layers of TF–target gene interactions with quantitative regulatory effects, describing the specificity of how the regulation is transduced through these interactions to activate cell wall genes (effector genes) for wood formation. PtrSND1-B1 directs 57 TF–DNA interactions through 17 TFs transregulating 27 effector genes. Of the 57 interactions, 55 are novel. We tested 42 of these 57 interactions in 30 genotypes of transgenic P. trichocarpa and verified that ∼90% of the tested interactions function in vivo. The TRN reveals common transregulatory targets for distinct TFs, leading to the discovery of nine TF protein complexes (dimers and trimers) implicated in regulating the biosynthesis of specific types of lignin. Our work suggests that wood formation may involve regulatory homeostasis determined by combinations of TF–DNA and TF–TF (protein–protein) regulations.}, number={3}, journal={The Plant Cell}, publisher={American Society of Plant Biologists (ASPB)}, author={Chen, Hao and Wang, Jack P. and Liu, Huizi and Li, Huiyu and Lin, Ying-Chung Jimmy and Shi, Rui and Yang, Chenmin and Gao, Jinghui and Zhou, Chenguang and Li, Quanzi and et al.}, year={2019}, month={Mar}, pages={602–626} } @article{matthews_wang_sederoff_chiang_williams_2019, title={Modeling cross-regulatory influences on monolignol transcripts and proteins under single and combinatorial gene knockdowns in Populus trichocarpa}, volume={6}, url={https://doi.org/10.1101/677047}, DOI={10.1101/677047}, abstractNote={Abstract}, publisher={Cold Spring Harbor Laboratory}, author={Matthews, Megan L. and Wang, Jack P. and Sederoff, Ronald and Chiang, Vincent L. and Williams, Cranos M.}, year={2019}, month={Jun} } @article{straub_khatibi_wang_conway_williams-rhaesa_peszlen_chiang_adams_kelly_2019, title={Quantitative fermentation of unpretreated transgenic poplar by Caldicellulosiruptor bescii}, volume={10}, ISSN={["2041-1723"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85070390326&partnerID=MN8TOARS}, DOI={10.1038/s41467-019-11376-6}, abstractNote={Abstract}, number={1}, journal={NATURE COMMUNICATIONS}, publisher={Springer Science and Business Media LLC}, author={Straub, Christopher T. and Khatibi, Piyum A. and Wang, Jack P. and Conway, Jonathan M. and Williams-Rhaesa, Amanda M. and Peszlen, Ilona M. and Chiang, Vincent L. and Adams, Michael W. W. and Kelly, Robert M.}, year={2019}, month={Aug} } @misc{myburg_hussey_wang_street_mizrachi_2019, title={Systems and Synthetic Biology of Forest Trees: A Bioengineering Paradigm for Woody Biomass Feedstocks}, volume={10}, ISSN={["1664-462X"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85068483814&partnerID=MN8TOARS}, DOI={10.3389/fpls.2019.00775}, abstractNote={Fast-growing forest plantations are sustainable feedstocks of plant biomass that can serve as alternatives to fossil carbon resources for materials, chemicals, and energy. Their ability to efficiently harvest light energy and carbon from the atmosphere and sequester this into metabolic precursors for lignocellulosic biopolymers and a wide range of plant specialized metabolites make them excellent biochemical production platforms and living biorefineries. Their large sizes have facilitated multi-omics analyses and systems modeling of key biological processes such as lignin biosynthesis in trees. High-throughput ‘omics’ approaches have also been applied in segregating tree populations where genetic variation creates abundant genetic perturbations of system components allowing construction of systems genetics models linking genes and pathways to complex trait variation. With this information in hand, it is now possible to start using synthetic biology and genome editing techniques in a bioengineering approach based on a deeper understanding and rational design of biological parts, devices, and integrated systems. However, the complexity of the biology and interacting components will require investment in big data informatics, machine learning, and intuitive visualization to fully explore multi-dimensional patterns and identify emergent properties of biological systems. Predictive systems models could be tested rapidly through high-throughput synthetic biology approaches and multigene editing. Such a bioengineering paradigm, together with accelerated genomic breeding, will be crucial for the development of a new generation of woody biorefinery crops.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Myburg, Alexander A. and Hussey, Steven G. and Wang, Jack P. and Street, Nathaniel R. and Mizrachi, Eshchar}, year={2019}, month={Jun} } @article{li_lin_wang_zhang_li_chen_shi_tunlaya-anukit_liu_wang_et al._2019, title={The AREB1 Transcription Factor Influences Histone Acetylation to Regulate Drought Responses and Tolerance in Populus trichocarpa}, volume={31}, url={http://dx.doi.org/10.1105/tpc.18.00437}, DOI={10.1105/tpc.18.00437}, abstractNote={AREB1 recruits the ADA2b-GCN5 complex to control H3K9ac and RNA polymerase II enrichment on drought-responsive genes, thereby driving high expression of these genes for drought tolerance. Plants develop tolerance to drought by activating genes with altered levels of epigenetic modifications. Specific transcription factors are involved in this activation, but the molecular connections within the regulatory system are unclear. Here, we analyzed genome-wide acetylated lysine residue 9 of histone H3 (H3K9ac) enrichment and examined its association with transcriptomes in Populus trichocarpa under drought stress. We revealed that abscisic acid-Responsive Element (ABRE) motifs in promoters of the drought-responsive genes PtrNAC006, PtrNAC007, and PtrNAC120 are involved in H3K9ac enhancement and activation of these genes. Overexpressing these PtrNAC genes in P. trichocarpa resulted in strong drought-tolerance phenotypes. We showed that the ABRE binding protein PtrAREB1-2 binds to ABRE motifs associated with these PtrNAC genes and recruits the histone acetyltransferase unit ADA2b-GCN5, forming AREB1-ADA2b-GCN5 ternary protein complexes. Moreover, this recruitment enables GCN5-mediated histone acetylation to enhance H3K9ac and enrich RNA polymerase II specifically at these PtrNAC genes for the development of drought tolerance. CRISPR editing or RNA interference-mediated downregulation of any of the ternary members results in highly drought-sensitive P. trichocarpa. Thus, the combinatorial function of the ternary proteins establishes a coordinated histone acetylation and transcription factor-mediated gene activation for drought response and tolerance in Populus species.}, number={3}, journal={Plant Cell}, author={Li, S. and Lin, Y.-C.J. and Wang, P. and Zhang, B. and Li, M. and Chen, S. and Shi, R. and Tunlaya-Anukit, S. and Liu, X. and Wang, Z. and et al.}, year={2019}, month={Mar}, pages={663–686} } @article{naik_wang_sederoff_chiang_williams_ducoste_2018, title={Assessing the impact of the 4CL enzyme complex on the robustness of monolignol biosynthesis using metabolic pathway analysis}, volume={13}, ISSN={1932-6203}, url={http://dx.doi.org/10.1371/journal.pone.0193896}, DOI={10.1371/journal.pone.0193896}, abstractNote={Lignin is a polymer present in the secondary cell walls of all vascular plants. It is a known barrier to pulping and the extraction of high-energy sugars from cellulosic biomass. The challenge faced with predicting outcomes of transgenic plants with reduced lignin is due in part to the presence of unique protein-protein interactions that influence the regulation and metabolic flux in the pathway. Yet, it is unclear why certain plants have evolved to create these protein complexes. In this study, we use mathematical models to investigate the role that the protein complex, formed specifically between Ptr4CL3 and Ptr4CL5 enzymes, have on the monolignol biosynthesis pathway. The role of this Ptr4CL3-Ptr4CL5 enzyme complex on the steady state flux distribution was quantified by performing Monte Carlo simulations. The effect of this complex on the robustness and the homeostatic properties of the pathway were identified by performing sensitivity and stability analyses, respectively. Results from these robustness and stability analyses suggest that the monolignol biosynthetic pathway is resilient to mild perturbations in the presence of the Ptr4CL3-Ptr4CL5 complex. Specifically, the presence of Ptr4CL3-Ptr4CL5 complex increased the stability of the pathway by 22%. The robustness in the pathway is maintained due to the presence of multiple enzyme isoforms as well as the presence of alternative pathways resulting from the presence of the Ptr4CL3-Ptr4CL5 complex.}, number={3}, journal={PLOS ONE}, publisher={Public Library of Science (PLoS)}, author={Naik, Punith and Wang, Jack P. and Sederoff, Ronald and Chiang, Vincent and Williams, Cranos and Ducoste, Joel J.}, editor={Cullen, DanielEditor}, year={2018}, month={Mar}, pages={e0193896} } @article{wang_matthews_williams_shi_yang_tunlaya-anukit_chen_li_liu_lin_et al._2018, title={Improving wood properties for wood utilization through multi-omics integration in lignin biosynthesis}, volume={9}, ISSN={2041-1723}, url={http://dx.doi.org/10.1038/s41467-018-03863-z}, DOI={10.1038/s41467-018-03863-z}, abstractNote={Abstract}, number={1}, journal={Nature Communications}, publisher={Springer Science and Business Media LLC}, author={Wang, Jack P. and Matthews, Megan L. and Williams, Cranos M. and Shi, Rui and Yang, Chenmin and Tunlaya-Anukit, Sermsawat and Chen, Hsi-Chuan and Li, Quanzi and Liu, Jie and Lin, Chien-Yuan and et al.}, year={2018}, month={Apr}, pages={1579} } @article{lin_chen_li_li_wang_shi_tunlaya-anukit_shuai_wang_ma_et al._2017, title={Reciprocal cross-regulation of VND and SND multigene TF families for wood formation in Populus trichocarpa}, volume={114}, ISSN={["0027-8424"]}, url={http://europepmc.org/abstract/med/29078399}, DOI={10.1073/pnas.1714422114}, abstractNote={Significance}, number={45}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, author={Lin, Ying-Chung Jimmy and Chen, Hao and Li, Quanzi and Li, Wei and Wang, Jack P. and Shi, Rui and Tunlaya-Anukit, Sermsawat and Shuai, Peng and Wang, Zhifeng and Ma, Hongyan and et al.}, year={2017}, month={Nov}, pages={E9722–E9729} } @article{shi_wang_lin_li_sun_chen_sederoff_chiang_2017, title={Tissue and cell-type co-expression networks of transcription factors and wood component genes in Populus trichocarpa}, volume={245}, ISSN={["1432-2048"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85009291513&partnerID=MN8TOARS}, DOI={10.1007/s00425-016-2640-1}, abstractNote={Co-expression networks based on transcriptomes of Populus trichocarpa major tissues and specific cell types suggest redundant control of cell wall component biosynthetic genes by transcription factors in wood formation. We analyzed the transcriptomes of five tissues (xylem, phloem, shoot, leaf, and root) and two wood forming cell types (fiber and vessel) of Populus trichocarpa to assemble gene co-expression subnetworks associated with wood formation. We identified 165 transcription factors (TFs) that showed xylem-, fiber-, and vessel-specific expression. Of these 165 TFs, 101 co-expressed (correlation coefficient, r > 0.7) with the 45 secondary cell wall cellulose, hemicellulose, and lignin biosynthetic genes. Each cell wall component gene co-expressed on average with 34 TFs, suggesting redundant control of the cell wall component gene expression. Co-expression analysis showed that the 101 TFs and the 45 cell wall component genes each has two distinct groups (groups 1 and 2), based on their co-expression patterns. The group 1 TFs (44 members) are predominantly xylem and fiber specific, and are all highly positively co-expressed with the group 1 cell wall component genes (30 members), suggesting their roles as major wood formation regulators. Group 1 TFs include a lateral organ boundary domain gene (LBD) that has the highest number of positively correlated cell wall component genes (36) and TFs (47). The group 2 TFs have 57 members, including 14 vessel-specific TFs, and are generally less correlated with the cell wall component genes. An exception is a vessel-specific basic helix-loop-helix (bHLH) gene that negatively correlates with 20 cell wall component genes, and may function as a key transcriptional suppressor. The co-expression networks revealed here suggest a well-structured transcriptional homeostasis for cell wall component biosynthesis during wood formation.}, number={5}, journal={PLANTA}, author={Shi, Rui and Wang, Jack P. and Lin, Ying-Chung and Li, Quanzi and Sun, Ying-Hsuan and Chen, Hao and Sederoff, Ronald R. and Chiang, Vincent L.}, year={2017}, month={May}, pages={927–938} } @inbook{wang_tunlaya-anukit_shi_yeh_chuang_isik_yang_liu_li_loziuk_et al._2016, title={A Proteomic-Based Quantitative Analysis of the Relationship Between Monolignol Biosynthetic Protein Abundance and Lignin Content Using TransgenicPopulus trichocarpa}, volume={5}, ISBN={9781118883303 9781118883266}, url={http://dx.doi.org/10.1002/9781118883303.ch4}, DOI={10.1002/9781118883303.ch4}, abstractNote={As part of a long-term project to develop a predictive model of lignin biosynthesis in the stem differentiating xylem of Populus trichocarpa, we explored the quantitative relationships of gene-specific monolignol pathway proteins and the amount of the lignin polymer. We determined the absolute abundance of monolignol pathway proteins in wild-type (Nisqually-1) and 80 transgenic trees, downregulated for the expression of genes in the monolignol pathway. Total lignin content for wild-type and transgenics ranged from 9.4 to 24.2%. Comparison of protein variation with lignin content showed that the proteins are produced in several-fold excess, suggesting that the monolignol pathway is highly homeostatic. Strong downregulation of xylem-specific PtrPAL2, 4, and 5 showed the most direct relationship with lignin content. These results are consistent with the Predictive Kinetic Metabolic Flux (PKMF) model of Wang et al., based on differential equations of mass action kinetics. Functional redundancy of multiple genes also moderates gene-specific effects. In addition, targeted production of specific proteins may affect the concentration of nontarget monolignol proteins, suggesting a yet-to-be-described level of coordinated control.}, booktitle={Recent Advances in Polyphenol Research}, publisher={John Wiley & Sons, Ltd}, author={Wang, Jack P. and Tunlaya-Anukit, Sermsawat and Shi, Rui and Yeh, Ting-Feng and Chuang, Ling and Isik, Fikret and Yang, Chenmin and Liu, Jie and Li, Quanzi and Loziuk, Philip L. and et al.}, editor={Kumi Yoshida, Veronique Cheynier and Quideau, StephaneEditors}, year={2016}, month={Dec}, pages={89–107} } @article{lin_li_tunlaya-anukit_shi_sun_wang_liu_loziuk_edmunds_miller_et al._2016, title={A cell wall-bound anionic peroxidase, PtrPO21, is involved in lignin polymerization in Populus trichocarpa}, volume={12}, ISSN={1614-2942 1614-2950}, url={http://dx.doi.org/10.1007/S11295-016-0978-Y}, DOI={10.1007/s11295-016-0978-y}, number={2}, journal={Tree Genetics & Genomes}, publisher={Springer Science and Business Media LLC}, author={Lin, Chien-Yuan and Li, Quanzi and Tunlaya-Anukit, Sermsawat and Shi, Rui and Sun, Ying-Hsuan and Wang, Jack P. and Liu, Jie and Loziuk, Philip and Edmunds, Charles W. and Miller, Zachary D. and et al.}, year={2016}, month={Mar} } @article{lin_wang_li_chen_liu_loziuk_song_williams_muddiman_sederoff_et al._2015, title={4-Coumaroyl and Caffeoyl Shikimic Acids Inhibit 4-Coumaric Acid: Coenzyme A Ligases and Modulate Metabolic Flux for 3-Hydroxylation in Monolignol Biosynthesis of Populus trichocarpa}, volume={8}, ISSN={["1752-9867"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84925201417&partnerID=MN8TOARS}, DOI={10.1016/j.molp.2014.12.003}, abstractNote={Downregulation of 4-coumaric acid:coenzyme A ligase (4CL) can reduce lignin content in a number of plant species. In lignin precursor (monolignol) biosynthesis during stem wood formation in Populus trichocarpa, two enzymes, Ptr4CL3 and Ptr4CL5, catalyze the coenzyme A (CoA) ligation of 4-coumaric acid to 4-coumaroyl-CoA and caffeic acid to caffeoyl-CoA. CoA ligation of 4-coumaric acid is essential for the 3-hydroxylation of 4-coumaroyl shikimic acid. This hydroxylation results from sequential reactions of 4-hydroxycinnamoyl-CoA:shikimic acid hydroxycinnamoyl transferases (PtrHCT1 and PtrHCT6) and 4-coumaric acid 3-hydroxylase 3 (PtrC3H3). Alternatively, 3-hydroxylation of 4-coumaric acid to caffeic acid may occur through an enzyme complex of cinnamic acid 4-hydroxylase 1 and 2 (PtrC4H1 and PtrC4H2) and PtrC3H3. We found that 4-coumaroyl and caffeoyl shikimic acids are inhibitors of Ptr4CL3 and Ptr4CL5. 4-Coumaroyl shikimic acid strongly inhibits the formation of 4-coumaroyl-CoA and caffeoyl-CoA. Caffeoyl shikimic acid inhibits only the formation of 4-coumaroyl-CoA. 4-Coumaroyl and caffeoyl shikimic acids both act as competitive and uncompetitive inhibitors. Metabolic flux in wild-type and PtrC3H3 downregulated P. trichocarpa transgenics has been estimated by absolute protein and metabolite quantification based on liquid chromatography–tandem mass spectrometry, mass action kinetics, and inhibition equations. Inhibition by 4-coumaroyl and caffeoyl shikimic acids may play significant regulatory roles when these inhibitors accumulate.}, number={1}, journal={MOLECULAR PLANT}, author={Lin, Chien-Yuan and Wang, Jack P. and Li, Quanzi and Chen, Hsi-Chuan and Liu, Jie and Loziuk, Philip and Song, Jina and Williams, Cranos and Muddiman, David C. and Sederoff, Ronald R. and et al.}, year={2015}, month={Jan}, pages={176–187} } @article{liu_hai_wang_cao_xu_jia_yang_wang_dai_cheng_2015, title={Comparative proteomic analysis of Populus trichocarpa early stem from primary to secondary growth}, volume={126}, ISSN={["1876-7737"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84930933098&partnerID=MN8TOARS}, DOI={10.1016/j.jprot.2015.05.032}, abstractNote={Wood is derived from the secondary growth of tree stems. In this study, we investigated the global changes of protein abundance in Populus early stems using a proteomic approach. Morphological and histochemical analyses revealed three typical stages during Populus early stems, which were the primary growth stage, the transition stage from primary to secondary growth and the secondary growth stage. A total of 231 spots were differentially abundant during various growth stages of Populus early stems. During Populus early stem lignifications, 87 differential spots continuously increased, while 49 spots continuously decreased. These two categories encompass 58.9% of all differential spots, which suggests significant molecular changes from primary to secondary growth. Among 231 spots, 165 unique proteins were identified using LC–ESI-Q-TOF-MS, which were classified into 14 biological function groups. The proteomic characteristics indicated that carbohydrate metabolism, oxido-reduction, protein degradation and secondary cell wall metabolism were the dominantly occurring biochemical processes during Populus early stem development. This study helps in elucidating biochemical processes and identifies potential wood formation-related proteins during tree early stem development. It is a comprehensive proteomic investigation on tree early stem development that, for the first time, reveals the overall molecular networks that occur during Populus early stem lignifications.}, journal={JOURNAL OF PROTEOMICS}, author={Liu, Jinwen and Hai, Guanghui and Wang, Chong and Cao, Shenquan and Xu, Wenjing and Jia, Zhigang and Yang, Chuanping and Wang, Jack P. and Dai, Shaojun and Cheng, Yuxiang}, year={2015}, month={Aug}, pages={94–108} } @article{loziuk_parker_li_lin_wang_li_sederoff_chiang_muddiman_2015, title={Elucidation of Xylem-Specific Transcription Factors and Absolute Quantification of Enzymes Regulating Cellulose Biosynthesis in Populus trichocarpa}, volume={14}, ISSN={["1535-3907"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84942916917&partnerID=MN8TOARS}, DOI={10.1021/acs.jproteome.5b00233}, abstractNote={Cellulose, the main chemical polymer of wood, is the most abundant polysaccharide in nature.1 The ability to perturb the abundance and structure of cellulose microfibrils is of critical importance to the pulp and paper industry as well as for the textile, wood products, and liquid biofuels industries. Although much has been learned at the transcript level about the biosynthesis of cellulose, a quantitative understanding at the proteome level has yet to be established. The study described herein sought to identify the proteins directly involved in cellulose biosynthesis during wood formation in Populus trichocarpa along with known xylem-specific transcription factors involved in regulating these key proteins. Development of an effective discovery proteomic strategy through a combination of subcellular fractionation of stem differentiating xylem tissue (SDX) with recently optimized FASP digestion protocols, StageTip fractionation, as well as optimized instrument parameters for global proteomic analysis using the quadrupole-orbitrap mass spectrometer resulted in the deepest proteomic coverage of SDX protein from P. trichocarpa with 9,146 protein groups being identified (1% FDR). Of these, 20 cellulosic/hemicellulosic enzymes and 43 xylem-specific transcription factor groups were identified. Finally, selection of surrogate peptides led to an assay for absolute quantification of 14 cellulosic proteins in SDX of P. trichocarpa.}, number={10}, journal={JOURNAL OF PROTEOME RESEARCH}, author={Loziuk, Philip L. and Parker, Jennifer and Li, Wei and Lin, Chien-Yuan and Wang, Jack P. and Li, Quanzi and Sederoff, Ronald R. and Chiang, Vincent L. and Muddiman, David C.}, year={2015}, month={Oct}, pages={4158–4168} } @article{wang_chuang_loziuk_chen_lin_shi_qu_muddiman_sederoff_chiang_2015, title={Phosphorylation is an on/off switch for 5-hydroxyconiferaldehyde O-methyltransferase activity in poplar monolignol biosynthesis}, volume={112}, ISSN={0027-8424 1091-6490}, url={http://dx.doi.org/10.1073/PNAS.1510473112}, DOI={10.1073/pnas.1510473112}, abstractNote={Significance}, number={27}, journal={Proceedings of the National Academy of Sciences}, publisher={Proceedings of the National Academy of Sciences}, author={Wang, Jack P. and Chuang, Ling and Loziuk, Philip L. and Chen, Hao and Lin, Ying-Chung and Shi, Rui and Qu, Guan-Zheng and Muddiman, David C. and Sederoff, Ronald R. and Chiang, Vincent L.}, year={2015}, month={Jun}, pages={8481–8486} } @article{lin_li_chen_li_sun_shi_lin_wang_chen_chuang_et al._2014, title={A simple improved-throughput xylem protoplast system for studying wood formation}, volume={9}, ISSN={["1750-2799"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84907026654&partnerID=MN8TOARS}, DOI={10.1038/nprot.2014.147}, abstractNote={Isolated protoplasts serve as a transient expression system that is highly representative of stable transgenics in terms of transcriptome responses. They can also be used as a cellular system to study gene transactivation and nucleocytoplasmic protein trafficking. They are particularly useful for systems studies in which stable transgenics and mutants are unavailable. We present a protocol for the isolation and transfection of protoplasts from wood-forming tissue, the stem-differentiating xylem (SDX), in the model woody plant Populus trichocarpa. The method involves tissue preparation, digestion of SDX cell walls, protoplast isolation and DNA transfection. Our approach is markedly faster and provides better yields than previous protocols; small (milligrams)- to large (20 g)-scale SDX preparations can be achieved in ~60 s, with isolation of protoplasts and their subsequent transfection taking ~50 min. Up to ten different samples can be processed simultaneously in this time scale. Our protocol gives a high yield (~2.5 × 10(7) protoplasts per g of SDX) of protoplasts sharing 96% transcriptome identity with intact SDX.}, number={9}, journal={NATURE PROTOCOLS}, author={Lin, Ying-Chung and Li, Wei and Chen, Hao and Li, Quanzi and Sun, Ying-Hsuan and Shi, Rui and Lin, Chien-Yuan and Wang, Jack P. and Chen, Hsi-Chuan and Chuang, Ling and et al.}, year={2014}, month={Sep}, pages={2194–2205} } @article{wang_naik_chen_shi_lin_liu_shuford_li_sun_tunlaya-anukit_et al._2014, title={Complete Proteomic-Based Enzyme Reaction and Inhibition Kinetics Reveal How Monolignol Biosynthetic Enzyme Families Affect Metabolic Flux and Lignin in Populus trichocarpa}, volume={26}, ISSN={1040-4651 1532-298X}, url={http://dx.doi.org/10.1105/tpc.113.120881}, DOI={10.1105/tpc.113.120881}, abstractNote={A proteomic-based predictive kinetic metabolic-flux model was developed for monolignol biosynthesis in Populus trichocarpa. Absolute quantities of all monolignol pathway proteins and 189 kinetic parameters were generated to construct the model, which was experimentally validated in transgenic P. trichocarpa and provides a comprehensive description of the monolignol biosynthetic pathway. We established a predictive kinetic metabolic-flux model for the 21 enzymes and 24 metabolites of the monolignol biosynthetic pathway using Populus trichocarpa secondary differentiating xylem. To establish this model, a comprehensive study was performed to obtain the reaction and inhibition kinetic parameters of all 21 enzymes based on functional recombinant proteins. A total of 104 Michaelis-Menten kinetic parameters and 85 inhibition kinetic parameters were derived from these enzymes. Through mass spectrometry, we obtained the absolute quantities of all 21 pathway enzymes in the secondary differentiating xylem. This extensive experimental data set, generated from a single tissue specialized in wood formation, was used to construct the predictive kinetic metabolic-flux model to provide a comprehensive mathematical description of the monolignol biosynthetic pathway. The model was validated using experimental data from transgenic P. trichocarpa plants. The model predicts how pathway enzymes affect lignin content and composition, explains a long-standing paradox regarding the regulation of monolignol subunit ratios in lignin, and reveals novel mechanisms involved in the regulation of lignin biosynthesis. This model provides an explanation of the effects of genetic and transgenic perturbations of the monolignol biosynthetic pathway in flowering plants.}, number={3}, journal={The Plant Cell}, publisher={American Society of Plant Biologists (ASPB)}, author={Wang, Jack P. and Naik, Punith P. and Chen, Hsi-Chuan and Shi, Rui and Lin, Chien-Yuan and Liu, Jie and Shuford, Christopher M. and Li, Quanzi and Sun, Ying-Hsuan and Tunlaya-Anukit, Sermsawat and et al.}, year={2014}, month={Mar}, pages={894–914} } @misc{li_song_peng_wang_qu_sederoff_chiang_2014, title={Plant biotechnology for lignocellulosic biofuel production}, volume={12}, ISSN={["1467-7652"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84911961843&partnerID=MN8TOARS}, DOI={10.1111/pbi.12273}, abstractNote={Summary}, number={9}, journal={PLANT BIOTECHNOLOGY JOURNAL}, author={Li, Quanzi and Song, Jian and Peng, Shaobing and Wang, Jack P. and Qu, Guan-Zheng and Sederoff, Ronald R. and Chiang, Vincent L.}, year={2014}, month={Dec}, pages={1174–1192} } @article{chen_song_wang_lin_ducoste_shuford_liu_li_shi_nepomuceno_et al._2014, title={Systems Biology of Lignin Biosynthesis in Populus trichocarpa: Heteromeric 4-Coumaric Acid:Coenzyme A Ligase Protein Complex Formation, Regulation, and Numerical Modeling}, volume={26}, ISSN={1040-4651 1532-298X}, url={http://dx.doi.org/10.1105/tpc.113.119685}, DOI={10.1105/tpc.113.119685}, abstractNote={This work shows that 4CL, an enzyme in monolignol biosynthesis, is found as a heterotetrameric complex of two isoforms in Populus trichocarpa. The activity of the heterotetramer can be described by a mathematical model that explains the effects of each isoform with mixtures of substrates and three types of inhibition, providing insights into the regulation of metabolic flux for this pathway. As a step toward predictive modeling of flux through the pathway of monolignol biosynthesis in stem differentiating xylem of Populus trichocarpa, we discovered that the two 4-coumaric acid:CoA ligase (4CL) isoforms, 4CL3 and 4CL5, interact in vivo and in vitro to form a heterotetrameric protein complex. This conclusion is based on laser microdissection, coimmunoprecipitation, chemical cross-linking, bimolecular fluorescence complementation, and mass spectrometry. The tetramer is composed of three subunits of 4CL3 and one of 4CL5. 4CL5 appears to have a regulatory role. This protein–protein interaction affects the direction and rate of metabolic flux for monolignol biosynthesis in P. trichocarpa. A mathematical model was developed for the behavior of 4CL3 and 4CL5 individually and in mixtures that form the enzyme complex. The model incorporates effects of mixtures of multiple hydroxycinnamic acid substrates, competitive inhibition, uncompetitive inhibition, and self-inhibition, along with characteristic of the substrates, the enzyme isoforms, and the tetrameric complex. Kinetic analysis of different ratios of the enzyme isoforms shows both inhibition and activation components, which are explained by the mathematical model and provide insight into the regulation of metabolic flux for monolignol biosynthesis by protein complex formation.}, number={3}, journal={The Plant Cell}, publisher={Oxford University Press (OUP)}, author={Chen, Hsi-Chuan and Song, Jina and Wang, Jack P. and Lin, Ying-Chung and Ducoste, Joel and Shuford, Christopher M. and Liu, Jie and Li, Quanzi and Shi, Rui and Nepomuceno, Angelito and et al.}, year={2014}, month={Mar}, pages={876–893} } @article{chen_song_williams_shuford_liu_wang_li_shi_gokce_ducoste_et al._2013, title={Monolignol Pathway 4-Coumaric Acid: Coenzyme A Ligases in Populus trichocarpa: Novel Specificity, Metabolic Regulation, and Simulation of Coenzyme A Ligation Fluxes}, volume={161}, ISSN={["0032-0889"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84874626790&partnerID=MN8TOARS}, DOI={10.1104/pp.112.210971}, abstractNote={Abstract}, number={3}, journal={PLANT PHYSIOLOGY}, author={Chen, Hsi-Chuan and Song, Jina and Williams, Cranos M. and Shuford, Christopher M. and Liu, Jie and Wang, Jack P. and Li, Quanzi and Shi, Rui and Gokce, Emine and Ducoste, Joel and et al.}, year={2013}, month={Mar}, pages={1501–1516} } @article{shi_shuford_wang_sun_yang_chen_tunlaya-anukit_li_liu_muddiman_et al._2013, title={Regulation of phenylalanine ammonia-lyase (PAL) gene family in wood forming tissue of Populus trichocarpa}, volume={238}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84882877816&partnerID=MN8TOARS}, DOI={10.1007/s00425-013-1905-1}, abstractNote={Phenylalanine ammonia-lyase (PAL) catalyzes the initial step of phenylpropanoid biosynthesis in plants. Five PAL genes (PtrPAL1 to 5) have been identified in Populus trichocarpa. These genes are classified into two subgroups according to their transcript sequence similarity and tissue specificity. However, the regulation of these genes and their protein functions are not well understood. In this study, enzymatic properties of each PtrPALs were characterized based on their recombinant proteins expressed in E.coli. Subcellular localizations of each PtrPALs in stem wood forming tissue were investigated and individual PtrPAL protein abundances in cytosol and membrane protein fractions were measured using protein cleavage-isotope dilution mass spectrometry (PC-IDMS). Protein/mRNA ratios of PtrPALs were further verified using RNA-Seq and gel-enhanced liquid chromatography mass spectrometry (GeLC-MS). All PtrPALs have similar catalytic properties for the deamination of L-phenylalanine, their major substrate. All PtrPALs have similar subcellular locations in stem wood forming tissue, with major amount in the cytosol (93-96 %) and less in the membrane (4-7 %). However, the protein/mRNA ratios of subgroup A (PtrPAL2, 4 and 5) are about five times that of subgroup B (PtrPAL1 and 3) in stem wood forming tissue, while all PtrPALs have similar transcript abundances. These results indicate a greater functional significance of subgroup A PtrPALs for stem wood formation, and highlight the role of gene post-transcriptional regulation.}, number={3}, journal={Planta}, author={Shi, R. and Shuford, C. M. and Wang, Jack P. and Sun, Y. H. and Yang, Z. C. and Chen, H. C. and Tunlaya-Anukit, S. and Li, Q. Z. and Liu, J. and Muddiman, David and et al.}, year={2013}, pages={487–497} } @article{loziuk_wang_li_sederoff_chiang_muddiman_2013, title={Understanding the Role of Proteolytic Digestion on Discovery and Targeted Proteomic Measurements Using Liquid Chromatography Tandem Mass Spectrometry and Design of Experiments}, volume={12}, ISSN={["1535-3907"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84890109136&partnerID=MN8TOARS}, DOI={10.1021/pr4008442}, abstractNote={Workflows in bottom-up proteomics have traditionally implemented the use of proteolysis during sample preparation; enzymatic digestion is most commonly performed using trypsin. This results in the hydrolysis of peptide bonds forming tryptic peptides, which can then be subjected to LC-MS/MS analysis. While the structure, specificity, and kinetics of trypsin are well characterized, a lack of consensus and understanding has remained regarding fundamental parameters critical to obtaining optimal data from a proteomics experiment. These include the type of trypsin used, pH during digestion, incubation temperature as well as enzyme-to-substrate ratio. Through the use of design of experiments (DOE), we optimized these parameters, resulting in deeper proteome coverage and a greater dynamic range of measurement. The knowledge gained from optimization of a discovery-based proteomics experiment was applied to targeted LC-MS/MS experiments using protein cleavage-isotope dilution mass spectrometry for absolute quantification. We demonstrated the importance of these digest parameters with respect to our limit of detection as well as our ability to acquire more accurate quantitative measurements. Additionally, we were able to quantitatively account for peptide decay observed in previous studies, caused by nonspecific activity of trypsin. The tryptic digest optimization described here has eliminated this previously observed peptide decay as well as provided a greater understanding and standardization for a common but critical sample treatment used across the field of proteomics.}, number={12}, journal={JOURNAL OF PROTEOME RESEARCH}, author={Loziuk, Philip L. and Wang, Jack and Li, Quanzi and Sederoff, Ronald R. and Chiang, Vincent L. and Muddiman, David C.}, year={2013}, month={Dec}, pages={5820–5829} } @article{shuford_li_sun_chen_wang_shi_sederoff_chiang_muddiman_2012, title={Comprehensive Quantification of Monolignol-Pathway Enzymes in Populus trichocarpa by Protein Cleavage Isotope Dilution Mass Spectrometry}, volume={11}, ISSN={1535-3893 1535-3907}, url={http://dx.doi.org/10.1021/pr300205a}, DOI={10.1021/pr300205a}, abstractNote={The economic value of wood/pulp from many tree species is largely dictated by the quantity and chemical properties of lignin, which is directly related to the composition and linkages of monolignols comprising the polymer. Although much is known regarding the monolignol biosynthetic pathway, our understanding is still deficient due to the lack of quantitative information at the proteomic level. We developed an assay based on protein cleavage isotope dilution mass spectrometry (PC-IDMS) for the determination of all potential, primary enzymes involved in the biosynthesis of monolignols and the peroxidases responsible for their polymerization to form lignin in the model tree species, Populus trichocarpa. Described is the identification of quantitative surrogate peptides through shotgun analysis of native and recombinant proteins, optimization of trypsin proteolysis using fractional factorial design of experiments, and development of a liquid chromatography-selected reaction monitoring method for specific detection of all targeted peptides. Of the 25 targeted enzymes, three were undetected in the normal xylem tissues, and all but two of the detectable species showed good day-to-day precision (CV < 10%). This represents the most comprehensive assay for quantification of proteins regulating monolignol biosynthesis and will lead to a better understanding of lignin formation at a systems level.}, number={6}, journal={Journal of Proteome Research}, publisher={American Chemical Society (ACS)}, author={Shuford, Christopher M. and Li, Quanzi and Sun, Ying-Hsuan and Chen, Hsi-Chuan and Wang, Jack and Shi, Rui and Sederoff, Ronald. R. and Chiang, Vincent L. and Muddiman, David C.}, year={2012}, month={May}, pages={3390–3404} } @article{wang_shuford_li_song_lin_sun_chen_williams_muddiman_sederoff_et al._2012, title={Functional redundancy of the two 5-hydroxylases in monolignol biosynthesis of Populus trichocarpa: LC-MS/MS based protein quantification and metabolic flux analysis}, volume={236}, ISSN={["1432-2048"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84865584315&partnerID=MN8TOARS}, DOI={10.1007/s00425-012-1663-5}, abstractNote={Flowering plants have syringyl and guaiacyl subunits in lignin in contrast to the guaiacyl lignin in gymnosperms. The biosynthesis of syringyl subunits is initiated by coniferaldehyde 5-hydroxylase (CAld5H). In Populus trichocarpa there are two closely related CAld5H enzymes (PtrCAld5H1 and PtrCAld5H2) associated with lignin biosynthesis during wood formation. We used yeast recombinant PtrCAld5H1 and PtrCAld5H2 proteins to carry out Michaelis-Menten and inhibition kinetics with LC-MS/MS based absolute protein quantification. CAld5H, a monooxygenase, requires a cytochrome P450 reductase (CPR) as an electron donor. We cloned and expressed three P. trichocarpa CPRs in yeast and show that all are active with both CAld5Hs. The kinetic analysis shows both CAld5Hs have essentially the same biochemical functions. When both CAld5Hs are coexpressed in the same yeast membranes, the resulting enzyme activities are additive, suggesting functional redundancy and independence of these two enzymes. Simulated reaction flux based on Michaelis-Menten kinetics and inhibition kinetics confirmed the redundancy and independence. Subcellular localization of both CAld5Hs as sGFP fusion proteins expressed in P. trichocarpa differentiating xylem protoplasts indicate that they are endoplasmic reticulum resident proteins. These results imply that during wood formation, 5-hydroxylation in monolignol biosynthesis of P. trichocarpa requires the combined metabolic flux of these two CAld5Hs to maintain adequate biosynthesis of syringyl lignin. The combination of genetic analysis, absolute protein quantitation-based enzyme kinetics, homologous CPR specificity, SNP characterization, and ER localization provides a more rigorous basis for a comprehensive systems understanding of 5-hydroxylation in lignin biosynthesis.}, number={3}, journal={PLANTA}, publisher={Springer Science + Business Media}, author={Wang, Jack P. and Shuford, Christopher M. and Li, Quanzi and Song, Jina and Lin, Ying-Chung and Sun, Ying-Hsuan and Chen, Hsi-Chuan and Williams, Cranos M. and Muddiman, David C. and Sederoff, Ronald R. and et al.}, year={2012}, month={Sep}, pages={795–808} } @article{li_min_wang_peszlen_horvath_horvath_nishimura_jameel_chang_chiang_2011, title={Down-regulation of glycosyltransferase 8D genes in Populus trichocarpa caused reduced mechanical strength and xylan content in wood}, volume={31}, ISSN={["1758-4469"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-79953831391&partnerID=MN8TOARS}, DOI={10.1093/treephys/tpr008}, abstractNote={Members of glycosyltransferase protein families GT8, GT43 and GT47 are implicated in the biosynthesis of xylan in the secondary cell walls of Arabidopsis. The Arabidopsis mutant irx8 has a 60% reduction in xylan. However, over-expression of an ortholog of Arabidopsis IRX8, poplar PoGT8D, in Arabidopsis irx8 mutant could not restore xylan synthesis. The functions of tree GT8D genes remain unclear. We identified two GT8 gene homologs, PtrGT8D1 and PtrGT8D2, in Populus trichocarpa. They are the only two GT8D members and are abundantly and specifically expressed in the differentiating xylem of P. trichocarpa. PtrGT8D1 transcript abundance was >7 times that of PtrGT8D2. To elucidate the genetic function of GT8D in P. trichocarpa, the expression of PtrGT8D1 and PtrGT8D2 was simultaneously knocked down through RNAi. Four transgenic lines had 85-94% reduction in transcripts of PtrGT8D1 and PtrGT8D2, resulting in 29-36% reduction in stem wood xylan content. Xylan reduction had essentially no effect on cellulose quantity but caused an 11-25% increase in lignin. These transgenics exhibit a brittle wood phenotype, accompanied by increased vessel diameter and thinner fiber cell walls in stem xylem. Stem modulus of elasticity and modulus of rupture were reduced by 17-29% and 16-23%, respectively, and were positively correlated with xylan content but negatively correlated with lignin quantity. These results suggest that PtrGT8Ds play key roles in xylan biosynthesis in wood. Xylan may be a more important factor than lignin affecting the stiffness and fracture strength of wood.}, number={2}, journal={TREE PHYSIOLOGY}, publisher={Oxford University Press (OUP)}, author={Li, Quanzi and Min, Douyong and Wang, Jack Peng-Yu and Peszlen, Ilona and Horvath, Laszlo and Horvath, Balazs and Nishimura, Yufuko and Jameel, Hasan and Chang, Hou-Min and Chiang, Vincent L.}, year={2011}, month={Feb}, pages={226–236} } @article{yeh_wang_shi_sun_chiang_2007, title={A novel O-methyl transferase-like gene with a drastic ectopic expression in response to tension wood formation in Populus trichocarpa}, volume={41}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-60749092120&partnerID=MN8TOARS}, number={9-10}, journal={Cellulose Chemistry and Technology}, author={Yeh, T.-F. and Wang, J. and Shi, R. and Sun, Y.-H. and Chiang, V.L.}, year={2007}, pages={521–528} } @article{yeh t.f._shi r._v.l._2007, title={A novel O-methyl transferase-like gene with drastic ectopic expression in response to tension wood formation in Populus trichocarpa}, volume={41}, journal={Cellulose Chemistry and Technology}, author={Yeh T.F., Wang J.P. and Shi R., Sun Y.H. and V.L., Chiang}, year={2007}, pages={521–528} }