Master of Science in Environmental Science, Cleveland State University, 2025, College of Arts and Sciences

Isolated trees in urban green spaces face unique environmental conditions that influence their structural and functional traits, shaping canopy architecture differently than trees in natural forests. This study examined how functional traits, including those associated with Corner's rules and the plant economic spectrum, relate to total twig-scale leaf area (TLA) and leaf area index (LAI) in isolated trees. We aimed to determine whether commonly measured traits could serve as predictors of canopy structure, informing urban forest management. Between June and September 2022, we measured leaf and twig traits for 32 tree species growing in open conditions at Secrest Arboretum. Key traits, including specific leaf area, leaf dry matter content, leaf size, twig diameter, and twig density, were assessed to determine their influence on TLA. Whole-canopy traits, including wood area index, branch angle, and branching intensity (i.e., ‘branchiness') were estimated using terrestrial laser scanning between February and April 2022. Additional canopy traits, including LAI, leaf area density, and diffuse non-interceptance, were estimated using a gap fraction analyzer between September 2022 and August 2023. To further contextualize canopy structure, we also considered phyllotaxy, twig density, and shade tolerance. Our findings revealed partial support for Corner's rules in isolated trees in urban green spaces. While TLA was positively correlated with twig diameter, individual leaf area and twig diameter were not strongly linked, suggesting that certain aspects of Corner's rules may not fully apply to isolated trees. Leaf size and twig diameter emerged as key predictors of TLA, indicating that thicker twigs and larger leaves contribute to greater TLA. At the canopy scale, almost 50 to 60% of LAI variation was explained by phyllotaxy, twig density, and TLA downstream of the second-most-distal branch order, standardized by cross-sectional area. This suggests that simple trait m (open full item for complete abstract)

Bachelor of Science (BS), Ohio University, 2025, Environmental and Plant Biology

Production of offspring is essential for every species of organism on Earth and plants are no exception. Plants have evolved countless techniques for efficient reproduction in the habitats they exist in. Seeds are the reproductive structures of two major groups of plants: gymnosperms (the more ancient group, often keeping their seeds on “scales” in cones) and angiosperms (flowering plants that protect their seeds in fruit). Seeds carry a plant's genetic material and represent the one time in a plant's life cycle that it can spread to new locations. This latter process is called dispersal and is vital for the survival and expansion of plant species. Unlike the vegetation of a plant, which will wilt and die without water, seeds are often the only part of a plant that can survive complete desiccation (or drying out) over long periods of time. With this unique ability to withstand extreme conditions, they ensure the continuation of plant species through time and across different habitats. The capacity of plant species to produce persistent, viable seeds enables them to endure temporally unfavorable environmental conditions. Seeds that have dormancy or are persistent in soil make up the seed bank of a community. The dynamics of a seed bank are essential to the ability of a community to maintain itself and respond to changes. While seed research has existed for many years, vegetative traits have always gotten more attention in the world of plant biology. Only in recent years has interest grown in understanding the ecological and evolutionary significance of seed traits. Tropical and subtropical species have been especially underrepresented in seed trait research, and scientists are only just tapping into the understanding of seed dormancy in the tropics. Understanding the seed bank of a community and the dormancy dynamics and germination requirements that exist within it can help with its restoration efforts. The purpose of this study is to contribute to the sparse data o (open full item for complete abstract)

Doctor of Philosophy, The Ohio State University, 2025, Food, Agricultural and Biological Engineering

The global economy relies on natural rubber (NR) for transportation of goods and people via tires and automobile components. Rubber is also critical to a plethora of industrial and medical products including gloves, condoms, and catheters. New and emerging uses of NR include space exploration, innovative medical procedures and specialized electronics. Despite the fact that thousands of plants produce NR, the tropical rubber tree, Hevea brasiliensis, remains the sole commercial source of NR. H. brasiliensis is unable to keep up with projected increases in NR demand due to many factors but especially small growing regions that are unable to expand without illegal deforestation of rainforests and high susceptibility of rubber trees to diseases. Concerns for the NR supply have sparked interest in alternative NR-producing plants with potential to supplement or replace the Hevea supply chain. There are three species besides Hevea which have been commercialized and had rubber extracted in bulk quantities during World War II: 1) Parthenium argentatum Gray (guayule), 2) Scorzonera tau-saghyz Lipsch. & G.G.Bosse (STS, mountain gum), and 3) Taraxacum kok-saghyz Rodin (TK, rubber dandelion). The work of this dissertation focuses on the potential of TK as an alternative rubber-producing crop. Of the three alternative rubber plants which have been successfully commercialized in the past, it is the only one capable of hydroponic growth, opening the door to unique NR production systems such as indoor and vertical farms. TK is also well suited to outdoor field production in temperate regions such as northern United States, much of Europe and a similar temperate region south of the equator, meaning it has a much larger habitable zone than Hevea. Additionally, at 6 months old TK has rubber of commercial quality, whilst Hevea is not tappable until 7 years old, making it a much more adaptable species for scaling up production in the event of a rubber crisis. The focus of this work wa (open full item for complete abstract)

Doctor of Philosophy, The Ohio State University, 2025, Plant Pathology

Rice (Oryza sativa L.), a staple crop for nearly 50% of the global population, particularly in Asia, is highly susceptible to various pathogens, leading to significant yield losses. To understand the molecular mechanisms underlying rice defense, two projects were carried out in this dissertation. The first study (Chapter 2) utilized rice as a model to examine transcriptional and metabolomic changes associated with the resistance (R) gene, Piz-t. Transcriptomic analyses revealed 2,791 Piz-t-specific differentially expressed genes (DEGs) following inoculation with Magnaporthe oryzae strain RO1-1. These findings highlighted correlations between gene expression and metabolite alterations, including changes in terpenoids, phenylpropanoids, flavonoids, fatty acids, amino acids, glycolysis/TCA intermediates, and phenylalanine pathways, providing critical insights into how R gene-mediated resistance enhances disease resistance in rice. The focus of the second study (Chapter 3) is to understand the role of programmed cell death (PCD) in rice immunity. Rice SPL11 cell-death suppressor 2 (SDS2), a receptor-like protein kinase, regulates PCD by phosphorylating and interacting with the U-box E3 ligase SPL11, as negative regulator of cell death, OsRLCK118 and OsRLCK176. The phosphorylated OsRLCK118 interacts with OsRbohB to mediate reactive oxygen species (ROS) generation. Despite these insights, the downstream pathways of SDS2-mediated resistance remain unclear. RNA-seq analysis of SDS2-ACT indicated higher expression of OsBDG1, suggesting possible interactor of SDS2, localizing to both the non-nuclear and nuclear area. OsBDG1 is a small protein with an LRR N-terminal domain and three LRR domains at its C-terminus, regulating in leaf inclination and grain size but no study was reported in disease resistance. Reduced luciferase and GUS activity were found by the co-expression of 35s::SDS2-HA and 35s::OsBDG1-GFP in rice protoplasts relative to the co-expression of 3 (open full item for complete abstract)

Master of Science (MS), Wright State University, 2024, Biological Sciences

Emerald ash borer (EAB), Agrilus planipennis, is an invasive pest that significantly impacts olive trees (Olea europaea). This study aimed to assess the role of abiotic stress factors such as drought and salinity in influencing plant defenses and resistance to EAB, while also examining cultivar variation in resistance. The study specifically aimed to address: (1) What effects do varying degrees of abiotic stress (drought, salinity) have on olive trees' antioxidant and peroxidase activity in olive tree? (2) Do distinct olive cultivars exhibit varying levels of resistance to EAB under various stressors? We used a controlled experimental design with two stress treatments (drought and salinity) and 20 olive cultivars to conduct the bioassay. Growth metrics (height and stem diameter), peroxidase activity, and antioxidant activity were measured, while EAB survival and performance were assessed by monitoring larval feeding and growth. Results indicated no significant differences in antioxidant activity between treatments, although low salinity slightly enhanced antioxidant responses. Peroxidase activity was highest under high salinity, and growth responses varied by treatment and cultivar, with Leccino showing the highest antioxidant activity. EAB larvae performed best under drought conditions but exhibited poor survival on certain cultivars. These findings suggest that abiotic stress influences both plant defense mechanisms and pest resistance, with cultivar-specific responses to stress and pest pressure.

Doctor of Philosophy (PhD), Ohio University, 2024, Plant Biology (Arts and Sciences)

Rapid advancements in genetic methods and bioinformatic tools are leading to broader applications across various organisms and research questions, often becoming more cost effective for non-model species. In this dissertation, I utilize these current genetic resources to achieve three overall goals: (i) test the utility of chloroplast genomic data to infer phylogenetic relationships to fully resolve relationships among genera and add to the growing information about the chloroplast genome characteristics in the Batrachospermales, (ii) assess the genetic variation of across fiver major drainage basins in eastern North America and determine the origin of dispersal of Batrachospermum gelatinosum following the Pleistocene glaciation and (iii) document the bacterial community diversity and composition of Batrachospermum gelatinosum across spatial and temporal scales. Adding eight new chloroplast genomes to the Batrachospermales, I established that chloroplast genomic data provided strong support for relationships among a clade of seven genera. My phylogeographic study of Batrachospermum gelatinosum revealed genetic partitioning among five major drainage basins, with no admixture detected within basins; unexpectedly, genetic diversity did not show a geographic pattern and varied widely across sites, likely due to intragametophytic selfing contributing to reduced genetic diversity in some streams. I found that the origin for B. gelatinosum following the Pleistocene glaciation was likely the Mid-Atlantic, though more sampling is needed to clarify these patterns as intragametophytic selfing likely eroded genetic diversity following these dispersal events, complicating demographic history analysis. Finally, I determined the diversity and composition of the bacterial community associated with B. gelatinosum varied across both spatial and temporal scales. However, much of this variation remains could not be explained by physical and chemical stream condi (open full item for complete abstract)

Doctor of Philosophy (PhD), Ohio University, 2024, Biological Sciences (Arts and Sciences)

Plant communities that occur on restrictive soils are characterized by stressful soil conditions and isolated patches of habitat, both of which have important consequences for the ecology and evolution of the species that occur on them. Despite their restrictive nature, edaphic communities contain high biodiversity, comprising a unique assemblage of plants, many of which are rare. Edaphic communities contain numerous, distantly related species that evolved under similar stressful conditions, but we still do not understand how the evolutionary process of edaphic specialization and speciation unfolds or the myriad of ecological and evolutionary consequences of occurring on restrictive soils. I examined species that occur on and off gypsum-edaphic communities to answer four questions, each as its own chapter: (Chapter 2) What fitness consequences do plants that occur on restrictive soils experience, (Chapter 3) how do diversification rates change for clades where gypsum endemism occur, (Chapter 4) how have dispersal syndromes and dispersion changed in edaphic communities because of their restrictive, fragmented substrate, and (Chapter 5) has selection favor limited dispersal in gypsum endemics? To answer those questions, I compared plant communities on gypsum outcrops (which contained both endemics and tolerators [= plants that grow on and off gypsum]) with surrounding, non-gypsum communities, and analysed selection and diversification rates of various gypsum associated clades. To determine the fitness consequences of inhabiting gypsum, I measured fitness for gypsum tolerating species across an edaphic gradient of gypsum to non-gypsum soils. I found negative and neutral fitness effects for species growing on gypsum soils. Various physical and chemical properties control fitness of tolerator species, but no common soil property was identified between the species that explained fitness changes on gypsum soil. To answer my second question, I gathered pre-constructed clado (open full item for complete abstract)

Doctor of Philosophy (PhD), Ohio University, 2024, Plant Biology (Arts and Sciences)

This dissertation enhances a forest gap model (ForClim) by incorporating seed production and seedling establishment processes, addressing a critical gap in understanding forest regeneration under climate change. The regeneration of forests in the Pacific Northwest (PNW) is a key driver of biodiversity, shaping species composition and ecosystem structure, and climate change is expected to significantly alter these processes, leading to shifts in both biodiversity and timber productivity. Simulations in this study revealed that seedling survival plays a more critical role than seed production in determining future species composition, particularly as climate variability increases. Resilient species like Pseudotsuga menziesii and Pinus ponderosa may sustain or increase their dominance, while species such as Abies grandis and Tsuga mertensiana face declines due to reduced seedling survival. Additionally, current forest management practices may need adjustment, with "no management" maximizing harvest volume for Coastal Douglas fir, while Mountain Douglas fir may experience reduced yields under future extreme climate scenarios. These findings highlight the importance of integrating regeneration processes into forest models to predict forest biodiversity and timber industry outcomes.

Doctor of Philosophy (PhD), Ohio University, 2024, Molecular and Cellular Biology (Arts and Sciences)

Gravity is a fundamental driving force of plant evolution, profoundly influencing numerous developmental and growth processes in plants. Gravity's most evident impact is the provision of directional cues to germinating seeds, guiding the roots downward and shoots upward. Known as gravitropism, this directional response to gravity is crucial to plants' overall health and productivity. Although biochemical and physiological studies have identified key ionic, chemical, and genetic factors involved in gravitropic signaling, the coordination of these actors remains poorly understood. Recent advances in omics technologies and the emergence of ways to isolate the effects of gravity on plants such as spaceflight experiments aboard the International Space Station (ISS), ground-based simulated gravities using clinostat and random positioning machines, and simple reorientation experiments have provided opportunities to investigate the molecular intricacies of this signaling cascade. Hence, this dissertation utilized both transcriptomics and proteomics to investigate gravitropic signaling in Arabidopsis plants during spaceflight in the Biological Research In Canister – Light Emitting Diode (BRIC LED) hardware. The results revealed key adaptive responses to the spaceflight environment, including destabilization and rearrangement of cell wall components, increased metabolic energy demands, and hypersensitivity of the photosystem. These adaptations were accompanied by a lack of direct correlation between transcriptomics and proteomics datasets, prompting further analysis using statistical and machine learning models. It was found that comparisons at the metabolic pathways level provided more comprehensive insights than simple gene-to-protein correlations. In addition, a meta-analysis of four existing plant proteomics datasets from spaceflight experiments aboard the ISS was conducted to assess variability. Factors such as spaceflight hardware, seedling age, li (open full item for complete abstract)

Master of Science, The Ohio State University, 2024, Human Ecology: Human Nutrition

Regular cleaning and sanitation of hydroponic systems is critical to ensure the safety of hydroponic crops. There are currently no validated protocols for sanitation of deep-water culture (DWC) hydroponic production surfaces. The objectives of this study were to evaluate the effectiveness of sanitizers in eliminating Listeria monocytogenes and its biofilms from DWC hydroponic surfaces including floaters made from high density polyethylene (HDPE) and polystyrene, Oasis dry foam, HDPE liner and rockwool. DWC coupon surfaces were inoculated with ~107 CFU/ml L. monocytogenes (ATCC 19111) rendered resistant to 100g/ml nalidixic acid. For biofilm formation, coupons were inoculated with ~108 CFU/ml L. monocytogenes and incubated for seven days. Inoculated coupons were treated with sanitizers for a contact time per manufacturer instructions: sodium hypochlorite (100 and 200ppm), SaniDate 12.0 (100 and 200ppm) composed of hydrogen peroxide and peroxyacetic acid, Virkon (1%) composed of potassium peroxymonosulphate and sodium chloride, KleenGrow (2%) composed of didecyl dimethyl ammonium and hydrogen peroxide (3%). Coupon surfaces were processed, and serial dilutions were enumerated on LB agar after incubation at 35C for 24h. Scanning electron microscopy was used to visualize biofilm structures on untreated and treated surfaces of HDPE liner, floater and polystyrene Styrofoam following biofilm formation. SaniDate 12.0 (100 and 200 ppm), Virkon (1%), KleenGrow (2%) and hydrogen peroxide (3%) eliminated L. monocytogenes from all five surfaces achieving greater than 5 log reductions. In comparison, sodium hypochlorite treatments (100 and 200 ppm) and water treatment were significantly less effective on all surfaces (p < 0.05). Sodium hypochlorite (100 and 200 ppm) did not eliminate L. monocytogenes from all surfaces tested in the study. Less than 3 log reduction was achieved on most surfaces when sodium hypochlorite (100 and 200 ppm) was used: HDPE liner (2.050.29 a (open full item for complete abstract)

Doctor of Philosophy (PhD), Ohio University, 2024, Plant Biology (Arts and Sciences)

Plants encounter various biotic and abiotic stresses daily and have developed defense mechanisms to overcome these challenges. One key system involved in these defense mechanisms is the ubiquitin (Ub)-26S proteasome system (UPS), which targets malfunctioning proteins for degradation through Ub-tagged proteasomal pathways. The E3 ligases, specifically S-phase kinase-associated protein 1 (SKP1), Cullin 1 (CUL1), and F-box (SCF) complexes, play crucial roles in this process by recognizing and tagging specific protein substrates. Arabidopsis thaliana, with over 700 F-box proteins, has the largest group of E3 ligases, yet only 5% have been functionally characterized. Phylogenetic relationships among 111 plant species have identified four clusters of F-box genes, including a cluster with more conserved F-boxes, referred to as core Arabidopsis F-box (CAF) genes. Given that CAF genes have more known functions compared to other clusters, this dissertation hypothesizes significant potential for discovering new functions among the uncharacterized F-boxes within this group. Considering the evolutionary conservation of most CAFs, I adopted a genetic approach to investigate the roles of CAFs during seed germination and seed development. To address the challenges posed by functional redundancy of duplicated CAF genes and the lethality associated with constitutive F-box overexpression in transgenic plants, I created a library of inducible overexpression lines for 40 CAF genes, many of which lacked known biological functions. By systematically examining the effects of conditional overexpression of these 40 CAFs, I found that CAF overexpression during seed germination and seed development can positively or negatively regulate radicle rupture growth, thus controlling the germination process. Specifically, I identified 24 CAFs that enhance radicle rupture and two that inhibited it by interfering with abscisic acid (ABA)-mediated germination suppression. Induction of CAFs during seed (open full item for complete abstract)

Master of Science, Miami University, 2024, Biology

Recent studies show root trait plasticity within species, suggesting differences in root response to ecosystem heterogeneity. The multi-axis root economic spectrum framework (RES) proposes that most root trait variation is explained by phylogeny and ecosystem conditions. We evaluated architecture and morphology in tree species in a long term, nitrogen (N) x phosphorus (P) fertilization study. Root traits differed between taxa and responded to nutrient addition. Red maple roots were less branched, produced fewer tips, and were thicker than roots of the species beech and birch. In both taxa, branching density, total root tips, average diameter, and the root length fraction in the 0-0.25 mm diameter class were lower in response to P. Most of these effects were reversed in response to N+P. Our findings show these species shift root growth to acquisitive traits when P limitations is alleviated but become conservative in response to N+P. This highlights the need to consider whether root growth is single or co-limited in ecosystems due to different effects of N x P interaction on root traits. We must continue refining the multi-axis RES framework to better account for within species plasticity in response to ecosystem conditions.

Bachelor of Science (BS), Ohio University, 2024, Environmental and Plant Biology

Caryophyllaceae flowers have a wide range of morphological appearances, particularly in regards to their stamens and petaloids. In order to determine how these structures evolved, I reviewed the phylogenetic relationships, mature morphology, and developmental pathways of flowers in the family. I concluded that in addition to an antesepalous whorl of stamens, there was likely a whorl of some ancestral structure in the alternisepalous position. This structure may have been a petal, a stamen, or a transitionary staminode, but it diversified in various lineages into the numerous forms we see today.

Bachelor of Science (BS), Ohio University, 2024, Environmental and Plant Biology

A plant's survival is largely impacted by its ability to sense and respond to gravitational changes, such as the changes that occur in spaceflight. However, the gravitropic signaling pathway is not fully understood. In the BRIC-20 experiment aboard the International Space Station, the Arabidopsis proton pump AHA2 was found to be differentially phosphorylated in microgravity compared to ground controls. AHA2 is hypothesized to be involved between plant hormone movement and differential growth during gravity signaling. When subject to reorientation, aha2 mutant seedlings had increased root curvature compared to wild type. Furthermore, wild type seedlings treated with fusicoccin, a phytotoxin that increases AHA2 phosphorylation and activity, had increased root curvature compared to untreated plants. To determine the role of AHA2 phosphorylation in gravitropic signaling, several lines of transgenic Arabidopsis with modifications in phosphorylation sites are being developed. The transformants will then be phenotyped for altered gravity response, showing if the phosphorylation of the AHA2 protein is involved in a plant's gravity response. These findings will further knowledge on the molecular mechanisms of gravitropism to develop space-tolerant plants for life support in spaceflight and for habitation of the moon, Mars, and beyond.

Doctor of Philosophy, The Ohio State University, 2024, Translational Plant Sciences

Grapevine (Vitis) is the world's most valuable fruit crop, therefore, reducing yield losses to stressors is paramount. Vitis labrusca, a wild North American grapevine, is well adapted to its local environment, exhibiting stout pathogen resistance. Meanwhile, Vitis vinifera grapevine, grown worldwide for winemaking, is native to Europe and is highly susceptible to biotic stressors, particularly fungal and insect pests. V. labrusca has been long utilized in Vitis breeding programs to imbue resistance. Therefore, in this dissertation, we determined if V. labrusca acc. ‘GREM4' was more insect herbivory resistant than V. vinifera cv. ‘PN40024' and investigated the morphological, genetic, and metabolomic factors which may contribute to resistance. In an herbivory choice assay, Japanese beetles (Popillia japonica), a major pest of grapevine, preferred to feed upon ‘PN40024' compared to ‘GREM4'. Further, increased leaf area was consumed on ‘PN40024' compared to ‘GREM4' in a time course (30min, 1h, and 4h) feeding assay. These results reported ‘GREM4' is resistant to Japanese beetle herbivory compared to ‘PN40024'. To determine morphological adaptations that may impact defense, trichomes were next investigated. Trichome densities were greater on ‘GREM4' compared to ‘PN40024' leaves. In trichome-focused herbivory studies, beetles exhibited a preference for lower trichome density sides of leaves and, when provided tissues with equal trichome densities for both ‘GREM4' and ‘PN40024', more leaf tissue was still lost from ‘PN40024' compared to ‘GREM4'. These results report that trichomes play a role in resistance but are not the sole factor. Therefore, we conducted a comparative transcriptomic analysis to identify differences in gene expression upon insect herbivory between the two species. When comparing constitutive expression differences prior to insect herbivory, genes with greater expression in ‘GREM4' were enriched in secondary metabolite biosynthesis while enr (open full item for complete abstract)

Doctor of Philosophy, The Ohio State University, 2024, Translational Plant Sciences

Field pennycress (Thlaspi arvense L.) is a winter annual with extreme cold hardiness and seed oil properties desirable for sustainable aviation fuel production. Integration of pennycress as an off-season biofuel cash crop into Midwest corn and soybean rotations could lead to the production of 1 billion liters of seed oil annually, therefore boosting farmer revenue and offsetting carbon emissions. Pennycress fields are vulnerable to heavy spring precipitation events, which can lead to waterlogged soils where the root system is submerged under water. However, it is unknown if growth, development, or yield of pennycress is affected by waterlogging at the reproductive developmental stage which occurs during April. This work aimed to characterize the morphological and transcriptomic responses of pennycress under one week of waterlogging during the reproductive stage. This was done with two core pennycress lines: one is the reference genome (MN106), and one is commonly used for gene editing (SP32-10). Additionally, natural populations of pennycress with predicted variation in soil water availability were included. One week of waterlogging at the reproductive stage under controlled conditions significantly reduced total seed weight in seven accessions, including SP32-10, whereas three accessions did not have reduced seed weight, such as MN106. Therefore, natural phenotypic variation in waterlogging responses existed between the two core lines, so they were further investigated to determine the transcriptomic responses contributing to waterlogging tolerance. Twice as many genes were differentially expressed between waterlogged and control roots in MN106 (3,424 genes) compared to SP32-10 (1,767 genes) after one week of waterlogging at the reproductive stage. Functional enrichment analysis of upregulated differentially expressed genes in both lines revealed Gene Ontology (GO) terms associated with hypoxia and decreased oxygen, including genes involved in alcoholic fermentatio (open full item for complete abstract)

Doctor of Philosophy (PhD), Ohio University, 2024, Plant Biology (Arts and Sciences)

Transcription factors involved in regulating plant cell wall biosynthesis are diverse and come from various families, including but not limited to AP2/ERF, MYB, NAC, bZIP, and WRKY. These TFs play crucial roles in modulating the expression of genes responsible for cell wall synthesis, composition, and structure, thereby influencing plant development and responses to environmental stimuli. Using gene association networks (GAN) analysis, four transcription factors co-expressed with cellulose synthases and glycosyltransferases for xylan biosynthesis were identified, namely, Os06g0194000 (OsERF071), Os02g0782700 (OsERF070), Os09g0434500 (OsERF072) from the AP2-ERF family, and Os08g0549000 (OsMYB102) from the MYB family. Loss-of-function allelic mutants were created using CRISPR/Cas9 technology. Allelic mutants that are homozygous and Cas9-free were evaluated morphologically, physiologically, and biochemically in the T2 generation. Mutants in Os06g0194000 (OsERF071) exhibited the most significant alterations compared to the CRISPR control plants, prompting the generation of overexpression lines for this gene to enable a more comprehensive functional analysis of this gene. Mutants lacking OSERF071 were taller plants with larger seed sizes, panicle lengths, thousand grain weights, biological yields, and grain yields compared to the CRISPR control plants, while the opposite phenotypes were observed in overexpression lines. Additionally, the allelic mutants exhibited higher content of lignin in their tissues compared to the control and showed higher expression levels of the OsCesA genes related to secondary cell wall biosynthesis compared to those linked to primary cell wall biosynthesis, suggesting OsERF071 may function as a repressor of lignin biosynthesis during primary cell wall deposition. Taken together, my results suggest that OsERF071 may function as a repressor of secondary cell wall formation in tissues that are dividing and growing. Another gene of interest wa (open full item for complete abstract)

Doctor of Philosophy (PhD), Ohio University, 2024, Plant Biology (Arts and Sciences)

Heteroxylans (HXs) are major hemicellulosic polymers, that provide structural integrity of plant cell walls which directly impact the growth and development of the plant. The backbone of HXs is comprised of β-linked xylose (Xyl) residues with different substitution patterns (acetyl, glucuronic acid/methylglucuronic acid [GlcA/[Me]GlcA] and/or arabinofuranosyl [Ara] side chains). In Arabidopsis, glucuronidation of HXs is mediated by GlucUronic acid substitution of Xylan (GUX) enzymes which are members of the GT8 family (CAZy). Reduced GlcA/[Me]GlcA side chains in HXs have been found to improve saccharification by reducing biomass recalcitrance and improving biofuel production. Therefore, identifying GUXs is of importance, offering insights into enhancing biofuel production and ensuring agricultural sustainability. Recently, GUXs have been identified in other species like conifers, Brachypodium and sugarcane. However, several outstanding questions about HX glucuronidation in grasses remain unanswered. Thus, this work focuses on the understanding of HX glucuronidation in rice, a significant cereal crop renowned for its nutritional value and substantial biomass, holding great potential for biofuel conversion and food security challenges. Through comprehensive phylogenetic and bioinformatics analyses, I identified three OsGUXs in rice, designated as OsGUX-A, OsGUX-B, and OsGUX-C. These OsGUX proteins exhibited structural conservation with counterparts from various species and harbored specific motifs and domains characteristic of GUX enzymes. Subcellular localization studies confirmed their presence within the Golgi apparatus, consistent with the site of HX biosynthesis. Using in vitro assays, I demonstrated that these OsGUXs have glucuronosyltransferase activity and exclusively utilize UDP-GlcA as a substrate. Further investigation through genetic complementation of Arabidopsis gux1/2/3 mutants, characterized by HX lacking GlcA/[Me]GlcA side chains, indicated that OsG (open full item for complete abstract)

Doctor of Philosophy (PhD), Ohio University, 2024, Plant Biology (Arts and Sciences)

Protein degradation through the Ubiquitin (Ub)-26S Proteasome System (UPS) is a major gene expression regulatory pathway in plants. In this pathway, the 76-amino acid Ub proteins are covalently linked onto a large array of UPS substrates with the help of three enzymes (E1 activating, E2 conjugating and E3 ligating enzymes) and direct them for turnover in the 26S proteasome complex or through autophagy-mediated degradation. The S-Phase Kinase-Associated Protein 1 (Skp1), cullin (CUL)1, and F-box (FBX) protein (SCF) complexes have been identified as the largest E3 ligase group in plants. Since the FBX proteins recognize and determine the specificity of SCF substrates, much effort has made to characterize their genomic, physiological, and biochemical roles in the past two decades of functional genomic studies. The co-immunoprecipitation failed to capture transient interaction while the yeast two-hybrid only test protein interaction in yeast cell instead of plant tissue. Therefore, the sheer number and high sequence diversity of the FBX gene family demands new approaches to uncover unknown functions. In this research, first, from the proteomic level, I developed and tested a proximity labeling and co-immunoprecipitation approach for determining the proteome of active FBX proteins and their associated complexes in Arabidopsis. We demonstrate that ASK1-TurboID is not fully functioning, which led us to discover a novel antagonism between biotinylation and ubiquitylation in regulating protein stability in vivo. This discovery lowers the effectiveness of proximity labeling in ubiquitylation studies. Second, from the FBX level, I characterized a new FBX protein that is involved in reproductive development. The mutant was created using CRISPR/Cas9. And it was confirmed that mutation of this FBX gene impacted seed development. The potential substrates were identified and confirmed using yeast two-hybrid and split-luciferase assay. Third, from the substrate (open full item for complete abstract)

Doctor of Philosophy (PhD), Ohio University, 2024, Molecular and Cellular Biology (Arts and Sciences)

Functional studies of the ubiquitin (Ub)-26S proteasome system (UPS) have demonstrated that virtually all aspects of the plant's life involve UPS-mediated turnover of abnormal or short-lived proteins. However, developmental characterization of the UPS, including in seeds and fruits, remains scarce. Unfortunately, early termination of embryogenesis limits the scope for characterizing the UPS activities in reproductive organs. In this dissertation, I utilized a biochemical approach to tackle the developmental role of UPS in plants, using Arabidopsis thaliana (Arabidopsis hereafter) as a model. The overarching goal of my research is to unravel the molecular mechanisms of UPS underpinning seed development so that new molecular breeding technologies could be developed to promote seed production. First, I systematically compared expression changes of multiple 26S proteasome subunits along with the dynamics of proteasome activity and total protein ubiquitylation in seedlings and developing siliques of Arabidopsis. Because autophagy plays the second largest role in maintaining proteome stability, I parallelly studied three late-limiting enzymes that are involved in autophagy influx. My experiments unexpectedly discovered that, in opposite to the activities in seedlings, both protein and transcript levels of six selected 26S proteasome subunits gradually decline in immature siliques toward maturation while the autophagy influx rises, albeit in a nutrient-rich condition. I also discovered a reciprocal turnover pathway between the proteasome and autophagy. While the autophagy influx is suppressed in seedlings by UPS-mediated degradation of its three key enzymes, transcriptional reprogramming dampens this process in siliques that in turn stimulates a bulk autophagy degradation of proteasomes. Collectively, my discovery about the developmental changes of the UPS and autophagy activities suggests that they relay the proteome homeostasis regulation in early seed development, w (open full item for complete abstract)