@article{hodgens_flaherty_pullen_khan_english_gillan_rojas-pierce_akpa_2024, title={Model-based inference of a dual role for HOPS in regulating guard cell vacuole fusion}, volume={6}, ISSN={["2517-5025"]}, url={https://doi.org/10.1093/insilicoplants/diae015}, DOI={10.1093/insilicoplants/diae015}, abstractNote={Abstract Guard cell movements depend, in part, on the remodelling of vacuoles from a highly fragmented state to a fused morphology during stomata opening. Indeed, full opening of plant stomata requires vacuole fusion to occur. Fusion of vacuole membranes is a highly conserved process in eukaryotes, with key roles played by two multi-subunit complexes: HOPS (homotypic fusion and vacuolar protein sorting) and SNARE (soluble NSF attachment protein receptor). HOPS is a vacuole tethering factor that is thought to chaperone SNAREs from apposing vacuole membranes into a fusion-competent complex capable of rearranging membranes. In plants, recruitment of HOPS subunits to the tonoplast has been shown to require the presence of the phosphoinositide phosphatidylinositol 3-phosphate. However, chemically depleting this lipid induces vacuole fusion. To resolve this counter-intuitive observation regarding the role of HOPS in regulating plant vacuole morphology, we defined a quantitative model of vacuole fusion dynamics and used it to generate testable predictions about HOPS-SNARE interactions. We derived our model by using simulation-based inference to integrate prior knowledge about molecular interactions with limited, qualitative observations of emergent vacuole phenotypes. By constraining the model parameters to yield the emergent outcomes observed for stoma opening—as induced by two distinct chemical treatments—we predicted a dual role for HOPS and identified a stalled form of the SNARE complex that differs from phenomena reported in yeast. We predict that HOPS has contradictory actions at different points in the fusion signalling pathway, promoting the formation of SNARE complexes, but limiting their activity.}, number={2}, journal={IN SILICO PLANTS}, author={Hodgens, Charles and Flaherty, D. T. and Pullen, Anne-Marie and Khan, Imran and English, Nolan J. and Gillan, Lydia and Rojas-Pierce, Marcela and Akpa, Belinda S.}, editor={Zhu, Xin-GuangEditor}, year={2024}, month={Nov} }
 @article{hodgens_flaherty_pullen_khan_english_gillan_rojas-pierce_akpa_2023, title={Model-based inference of a plant-specific dual role for HOPS in regulating guard cell vacuole fusion}, url={https://doi.org/10.1101/2023.11.07.565947}, DOI={10.1101/2023.11.07.565947}, abstractNote={Stomata are the pores on a leaf surface that regulate gas exchange. Each stoma consists of two guard cells whose movements regulate pore opening and thereby control CO2 fixation and water loss. Guard cell movements depend in part on the remodeling of vacuoles, which have been observed to change from a highly fragmented state to a fused morphology during stomata opening. This change in morphology requires a membrane fusion mechanism that responds rapidly to environmental signals, allowing plants to respond to diurnal and stress cues. With guard cell vacuoles being both large and responsive to external signals, stomata represent a unique system in which to delineate mechanisms of membrane fusion. Fusion of vacuole membranes is a highly conserved process in eukaryotes, with key roles played by two multi-subunit complexes: HOPS (homotypic fusion and vacuolar protein sorting) and SNARE (soluble NSF attachment protein receptor). HOPS is a vacuole tethering factor that is thought to chaperone SNAREs from apposing vacuole membranes into a fusion-competent complex capable of rearranging membranes. To resolve a counter-intuitive observation regarding the role of HOPS in regulating plant vacuole morphology, we derived a quantitative model of vacuole fusion dynamics and used it to generate testable predictions about HOPS-SNARE interactions. We derived our model by applying simulation-based inference to integrate prior knowledge about molecular interactions with limited, qualitative observations of emergent vacuole phenotypes. By constraining the model parameters to yield the emergent outcomes observed for stoma opening - as induced by two distinct chemical treatments - we predicted a dual role for HOPS and identified a stalled form of the SNARE complex that differs from phenomena reported in yeast. We predict that HOPS has contradictory actions at different points in the fusion signaling pathway, promoting the formation of SNARE complexes, but limiting their activity.}, author={Hodgens, Charles and Flaherty, DT and Pullen, Anne-Marie and Khan, Imran and English, Nolan J and Gillan, Lydia and Rojas-Pierce, Marcela and Akpa, Belinda S}, year={2023}, month={Nov} }
 @article{function of the pseudo phosphotransfer proteins has diverged between rice and arabidopsis_2021, volume={106}, url={http://dx.doi.org/10.1111/tpj.15156}, DOI={10.1111/tpj.15156}, abstractNote={SUMMARY The phytohormone cytokinin plays a significant role in nearly all aspects of plant growth and development. Cytokinin signaling has primarily been studied in the dicot model Arabidopsis, with relatively little work done in monocots, which include rice ( Oryza sativa ) and other cereals of agronomic importance. The cytokinin signaling pathway is a phosphorelay comprised of the histidine kinase receptors, the authentic histidine phosphotransfer proteins (AHPs) and type‐B response regulators (RRs). Two negative regulators of cytokinin signaling have been identified: the type‐A RRs, which are cytokinin primary response genes, and the pseudo histidine phosphotransfer proteins (PHPs), which lack the His residue required for phosphorelay. Here, we describe the role of the rice PHP genes. Phylogenic analysis indicates that the PHPs are generally first found in the genomes of gymnosperms and that they arose independently in monocots and dicots. Consistent with this, the three rice PHPs fail to complement an Arabidopsis php mutant ( aphp1/ahp6 ). Disruption of the three rice PHPs results in a molecular phenotype consistent with these elements acting as negative regulators of cytokinin signaling, including the induction of a number of type‐A RR and cytokinin oxidase genes. The triple php mutant affects multiple aspects of rice growth and development, including shoot morphology, panicle architecture, and seed fill. In contrast to Arabidopsis, disruption of the rice PHPs does not affect root vascular patterning, suggesting that while many aspects of key signaling networks are conserved between monocots and dicots, the roles of at least some cytokinin signaling elements are distinct.}, number={1}, journal={The Plant Journal}, publisher={Wiley}, year={2021}, month={Apr}, pages={159–173} }
 @misc{hodgens_akpa_long_2021, title={Solving the puzzle of Fe homeostasis by integrating molecular, mathematical, and societal models}, volume={64}, ISSN={["1879-0356"]}, url={https://doi.org/10.1016/j.pbi.2021.102149}, DOI={10.1016/j.pbi.2021.102149}, abstractNote={To ensure optimal utilization and bioavailability, iron uptake, transport, subcellular localization, and assimilation are tightly regulated in plants. Herein, we examine recent advances in our understanding of cellular responses to Fe deficiency. We then use intracellular mechanisms of Fe homeostasis to discuss how formalizing cell biology knowledge via a mathematical model can advance discovery even when quantitative data is limited. Using simulation-based inference to identify plausible systems mechanisms that conform to known emergent phenotypes can yield novel, testable hypotheses to guide targeted experiments. However, this approach relies on the accurate encoding of domain-expert knowledge in exploratory mathematical models. We argue that this would be facilitated by fostering more "systems thinking" life scientists and that diversifying your research team may be a practical path to achieve that goal.}, journal={CURRENT OPINION IN PLANT BIOLOGY}, publisher={Elsevier BV}, author={Hodgens, Charles and Akpa, Belinda S. and Long, Terri A.}, year={2021}, month={Dec} }
 @phdthesis{hodgens_2020, title={Innovations in cloning genes identified in genetic screens and their application to cytokinin signaling in Arabidopsis thaliana}, url={https://cdr.lib.unc.edu/concern/dissertations/cr56n655b}, DOI={10.17615/W0MT-T167}, school={The University of North Carolina at Chapel Hill University Libraries}, author={Hodgens, Charles}, year={2020} }
 @article{hodgens_chang_schaller_kieber_2020, title={Mutagenomics: A Rapid, High-Throughput Method to Identify Causative Mutations from a Genetic Screen}, volume={9}, url={https://doi.org/10.1104/pp.20.00609}, DOI={10.1104/pp.20.00609}, abstractNote={Genetic screens are powerful tools to dissect complex biological processes, but a rate-limiting step is often the cloning of targeted genes. Here, we present a strategy, "mutagenomics," to identify causal mutations from a screen in a high throughput fashion in the absence of backcrossing. Mutagenomics is initiated by sequencing the genomes of the mutants identified, which are then subjected to a three-stage pipeline. The first stage identifies sequence changes in genes previously linked to the targeted pathway. The second stage uses heuristics derived from a simulation strategy to identify genes that are represented by multiple independent alleles more often than expected by chance. The third stage identifies candidate genes for the remaining lines by sequencing multiple lines of common descent. Our simulations indicate that sequencing as few as three to four sibling lines generally results in fewer than five candidate genes. We applied mutagenomics to a screen for Arabidopsis (Arabidopsis thaliana) mutants involved in the response to the phytohormone cytokinin. Mutagenomics identified likely causative genes for many of the mutant lines analyzed from this screen, including 13 alleles of the gene encoding the ARABIDOPSIS HIS KINASE4 cytokinin receptor. The screen also identified 1-AMINOCYCLOPROPANE-1-CARBOXYLATE (ACC) SYNTHASE7, an ACC synthase homolog involved in ethylene biosynthesis, and ELONGATED HYPOCOTYL5 (HY5), a master transcriptional regulator of photomorphogenesis. HY5 was found to mediate a subset of the transcriptional response to cytokinin. Mutagenomics has the potential to accelerate the pace and utility of genetic screens in Arabidopsis.}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Hodgens, Charles and Chang, Nicole and Schaller, G. Eric and Kieber, Joseph J.}, year={2020}, month={Dec}, pages={pp.00609.2020} }
 @article{burr_sun_yamburenko_willoughby_hodgens_boeshore_elmore_atkinson_nimchuk_bishopp_et al._2020, title={The HK5 and HK6 cytokinin receptors mediate diverse developmental pathways in rice}, volume={1}, url={http://dx.doi.org/10.1242/dev.191734}, DOI={10.1242/dev.191734}, abstractNote={The phytohormone cytokinin regulates diverse aspects of plant growth and development. Our understanding of the metabolism and perception of cytokinin has made great strides in recent years, mostly from studies of the model dicot}, journal={Development}, publisher={The Company of Biologists}, author={Burr, Christian A. and Sun, Jinjing and Yamburenko, Maria V. and Willoughby, Andrew and Hodgens, Charles and Boeshore, Samantha Louise and Elmore, Agustus and Atkinson, Jonathan and Nimchuk, Zachary L. and Bishopp, Anthony and et al.}, year={2020}, month={Jan} }
 @article{reed_wu_reeves_hodgens_yadav_hayes_pierik_2018, title={Three Auxin Response Factors Promote Hypocotyl Elongation}, volume={178}, url={https://doi.org/10.1104/pp.18.00718}, DOI={10.1104/pp.18.00718}, abstractNote={The hormone auxin regulates growth largely by affecting gene expression. By studying Arabidopsis (Arabidopsis thaliana) mutants deficient in AUXIN RESPONSE FACTORS (ARFs), we have identified three ARF proteins that are required for auxin-responsive hypocotyl elongation. Plants deficient in these factors have reduced responses to environmental conditions that increase auxin levels, including far-red-enriched light and high temperature. Despite having decreased auxin responses, the ARF-deficient plants responded to brassinosteroid and gibberellin, indicating that different hormones can act partially independently. Aux/IAA proteins, encoded by IAA genes, interact with ARF proteins to repress auxin response. Silencing expression of multiple IAA genes increased hypocotyl elongation, suggesting that Aux/IAA proteins modulate ARF activity in hypocotyls in a potential negative feedback loop.}, number={2}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Reed, Jason W. and Wu, Miin-Feng and Reeves, Paul H. and Hodgens, Charles and Yadav, Vandana and Hayes, Scott and Pierik, Ronald}, year={2018}, month={Oct}, pages={864–875} }
 @article{hodgens_nimchuk_kieber_2017, title={indCAPS: A tool for designing screening primers for CRISPR/Cas9 mutagenesis events}, url={https://doi.org/10.1101/196121}, DOI={10.1101/196121}, abstractNote={Abstract Genetic manipulation of organisms using CRISPR/Cas9 technology generally produces small insertions/deletions (indels) that can be difficult to detect. Here, we describe a technique to easily and rapidly identify such indels. Sequence-identified mutations that alter a restriction enzyme recognition site can be easily distinguished from wild-type alleles using a cleaved amplified polymorphic sequence (CAPS) technique. If a restriction site is created or altered by the mutation such that only one allele contains the restriction site, a polymerase chain reaction (PCR) followed by a restriction digest can be used to distinguish the two alleles. However, in the case of most CRISPR-induced alleles, no such restriction sites are present in the target sequences. In this case, a derived CAPS (dCAPS) approach can be used in which mismatches are purposefully introduced in the oligonucleotide primers to create a restriction site in one, but not both, of the amplified templates. Web-based tools exist to aid dCAPS primer design, but when supplied sequences that include indels, the current tools often fail to suggest appropriate primers. Here, we report the development of a Python-based, species-agnostic web tool, called indCAPS, suitable for the design of PCR primers used in dCAPS assays that is compatible with indels. This tool should have wide utility for screening editing events following CRISPR/Cas9 mutagenesis as well as for identifying specific editing events in a pool of CRISPR-mediated mutagenesis events. This tool was field-tested in a CRISPR mutagenesis experiment targeting a cytokinin receptor ( AHK3 ) in Arabidopsis thaliana . The tool suggested primers that successfully distinguished between wild-type and edited alleles of a target locus and facilitated the isolation of two novel ahk3 null alleles. Users can access indCAPS and design PCR primers to employ dCAPS to identify CRISPR/Cas9 alleles at http://indcaps.kieber.cloudapps.unc.edu/ .}, author={Hodgens, Charles and Nimchuk, Zachary L. and Kieber, Joseph J.}, year={2017}, month={Sep} }