Browsing by Department "Plant Biology"
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Item Accelerated substitution rates, plastid genome evolution and cytonuclear coevolution in Geraniaceae(2015-08) Weng, Mao-Lun; Jansen, Robert K., 1954-; Chen, Zengjian Jeffery; Linder, Craig Randal; Simpson, Beryl B.; Hillis, David M.Plastids are cytoplasmic organelles that are descendants of free-living prokaryotes. Since endosymbiosis, massive amounts of genetic material have been transferred from plastids to the nucleus. As a result plastids require the import of proteins encoded by nuclear genes that were formerly located in their ancestral bacterial genomes and the coordination between nucleus and plastid becomes vital for the cells. Among seed plants, the plastid genome is relatively conserved in gene order and content and has lower nucleotide substitution rates than the nucleus, but a few unrelated lineages display rearranged genomes and accelerated substitution rates. These unusual lineages present an opportunity to understand the relative roles of natural selection and neutral processes in shaping genome organization and evolutionary rates. They also provide a system to investigate how the accelerated substitution rate in plastids affects cytoplasmic-nuclear (cytonuclear) coevolution. My research uses Geraniaceae as a study system to investigate (1) the evolutionary processes governing accelerated rates of change in nucleotide substitutions and rearrangements in plastid genomes, and (2) coordinated evolution between plastid and nuclear genomes. In chapter two, the monophyly of three major clades within Pelargonium is confirmed. This phylogeny was then used to examine karyotype evolution and nucleotide substitution rates in the genus. A correlation between genome size and chromosome size but not number is observed. Also, both plastid and mitochondrial genes have accelerated substitution rates but with markedly disparate patterns. Chapter three focuses on reconstructing the genomic rearrangement events in the plastid genomes of Geraniaceae. The reconstructed ancestral Geraniaceae plastid genome is relatively unrearranged, but there are a large number of subsequent independent rearrangements in each genus. My analyses show that the distribution and content of repetitive sequences are significantly correlated with the degree of genomic rearrangements, and there is a positive correlation between nonsynonymous substitution rates and genomic rearrangements in Geraniaceae plastid genomes. Chapter four investigates the cytonuclear coevolution in plastid ribosomes of Geraniaceae. My data shows that both plastid-encoded and nuclear-encoded subunits of the plastid ribosome have accelerated nonsynonymous substitutions rates, and the analysis indicates that nuclear substitutions are driving increased substitution in plastid genes.Item Bacterial cellulose in cyanobacteria : enhancement of cellulose production in Synechococcus elongatus with Gluconacetobacter xylinus transgenes(2014-12) Sessler, Tate Hopkins; Brown, R. (Richard Malcolm), 1939-Bacterial cellulose produced by Gluconacetobacter xylinus possesses an abundance of desirable properties which allow for commercial applications in manufacturing, construction, medicine, as a biofuel feedstock and in electronics as an electrical insulator. Large scale production methods are currently limited by expensive media and requirements for sterile conditions. This thesis investigates the potential for using cellulose synthesized by cyanobacteria as an alternative. Cyanobacteria require fewer media components, fix atmospheric carbon dioxide, and some species fix atmospheric nitrogen, reducing the demand for fertilizer and lowering contamination potential. In this study, a suite of genes involved with cellulose synthesis in G. xylinus were successfully transferred to into the unicellular cyanobacterium, Synechococcus elongatus. Cellulose synthesis was increased in multiple transgenic strains by 300-400% and cellulose microfibrils were observed with enhanced crystallization upon addition of the acsCD transgene from G. xylinus. The evolutionary history of the proteins involved in cellulose synthesis was also investigated, with particular interest paid toward the catalytic subunit. It was determined that all current organisms with the capability of cellulose assembly possess synthases from a single ancestral origin at least 2.5 billion years ago. This work provides additional support to the theory that vascular plants acquired cellulose synthases from cyanobacteria.Item Carbon isotope fractionation by ribulose 1,5-bisphosphate carboxylase from various organisms(1977) Estep, Marilyn Louise Fogel; Not availableItem Characterization of underlying transcription factors that regulate betalain pigment formation in beets(2015-08) Akhavan, Neda Attar; Lloyd, Alan M.; Fischer, Janice A.; Mehdy, Mona; Roux, Stanley J.; Simpson, Beryl B.The plant kingdom is a colorful place with most vascular plants producing phenylalanine-based red/violet anthocyanin pigment. Only a single order of flowering plants, the Caryophyllales, is known to produce an unusual pigment known as Betalain. Betalains encompasses an entire range of colors between yellow to red, are nitrogen-containing water-soluble compounds derived from tyrosine, and are acidic in nature due to the presence of several carboxyl groups. The betalain and the anthocyanin pigments are mutually exclusive. The pathway and enzymes for betalain biosynthesis, from tyrosine to the end products, red/violet betacyanins and yellow betaxanthins, has largely been determined. Little however is known about the regulation of the biosnynthetic genes. The strong biological correlation between the anthocyanin and the betalains prompted the suggestion that the molecular regulation of betalains and anthocyanins uses the same MYB and bHLH and WD-repeat regulators, the MBW complex. The work described here strives to understand the regulatory mechanisms controlling betalain pigmentation in the Caryophyllales and how they can be controlled and influenced. To understand the pathway, there was a pressing need for analysis at the biochemical, molecular, and genetic levels. Before the work reported here, two pigment biosynthetic genes were identified. The gene/ enzyme responsible for step two was identified as DOPA 4, 5-dioxygenase (DODA) functioning to produce betalamic acid (BA) from LDOPA (Christinet et al., 2004), and later we showed that a novel cytochrome P450, CYP76AD1, is absolutely required for the red pigment in beets by catalyzing the step producing cyclo-DOPA from the LDOPA substrate (Hatlestad et al., 2012). Through this project I have: (1) discovered and characterized Beet MYB1 (BvMYB1), a MYB evolved from the anthocyanin regulating MYBs, and analyzed how BvMYB1 regulates betalain production by its interaction with DNA and other proteins; (2) determined overlapping functional redundancies of BvMYB1 with two other R2R3 BvMYBs, BvMYB2 and BvMYB3; (3) identified a beet bHLH protein, BvbHLH1, and a beet WD-repeat protein, BvTTG1, that function similar to the Arabidopsis thaliana proteins; and finally (4) I worked towards characterizing a novel BvMYB1 Response Element (MRE) that BvMYB1 directly binds to activate betalain biosynthetic genes. By identifying members of the regulatory complex for the betalain pathway I hope to contribute toward understanding the evolutionary replacement of anthocyanins by betalains within a single flowering order, and fill a lack of knowledge about producing, controlling, and influencing this valuable natural pigment.Item Circadian clock gene expression and growth vigor in arabidopsis hybrids and mRNA stability in arabidopsis allotetraploids(2010-12) Kim, Eun Deok; Chen, Z. JeffreyHybrids and polyploids are very common in plants and some animals. Although hybrid vigor or heterosis has been widely adopted in agricultural practices, the underlying mechanisms are poorly understood partly because of their multigenic nature and the lack of a good model system for the study. Allotetraploidy is an emerging model system for investigating molecular mechanisms of hybrid vigor. An allotetraploid is formed by interspecific hybridization followed by chromosome doubling or hybridization between two autotetraploid parents and is genetically stable. A previous study showed nonadditive expression (different from the mid-parent value) of over 5% of genes in the allotetraploids, suggesting altered transcriptional and post-transcriptional regulation. Here oligo-gene microarray analysis of mRNA stability in allotetraploids was carried out to investigate how nonadditive gene regulation upon allopolyploidization is achieved at the posttranscriptional level. Approximately 1% of annotated genes were identified as unstable transcripts, and their estimated half-life is less than 60 minutes. The unstable transcripts in Arabidopsis allotetraploids are associated with nonadditive gene expression and with stress and environmental responses. The nonadditively expressed genes identified in the previous study include those encoding proteins involved in energy and metabolic pathways, which are putative targets of circadian clock regulators. To test how circadian clock genes affect downstream genes and pathways, expression of CIRCADIAN CLOCK ASSOCIATED1 (CCA1) was up- or down-regulated by overexpressing CCA1 or cca1(RNAi) driven by the promoter of TIMING OF CAB EXPRESSION 1 (TOC1). Upregulation of CCA1 was associated with repression of downstream genes in chlorophyll biosynthesis and starch metabolism, whereas down-regulation of CCA1 correlated with upregulation of these downstream genes. As a result, chlorophyll and starch content was ~10% higher in the TOC1::cca1(RNAi) transgenic plants than the controls, while the growth vigor is lower in the TOC1::CCA1 transgenic plants. To further test the effects of clock genes in growth vigor, CCA1 expression was examined in reciprocal hybrids of A. thaliana ecotypes. The maternal effect on starch content was observed in several combinations of hybrids, which was correlated with preferential expression of maternal CCA1 during early stages of seed development. Although the cause of parent-of-origin effects is still unclear, the data have clearly documented parent-of-origin effects on circadian clock gene expression and starch metabolism in hybrids.Item Clock-regulatory networks contribute to growth vigor in maize hybrids(2016-08) Ko, Dae Kwan; Chen, Z. Jeffrey; Jansen, Robert K.; Atkinson, Nigel; Huq, Enamul; Sung, SibumHeterosis, or hybrid vigor, has been widely used in agriculture for more than a century. Despite extensive investigation and various models proposed, the molecular basis for heterosis remains largely elusive. In Arabidopsis interspecific and intraspecific hybrids, increased photosynthetic and metabolic activities are linked to altered expression of central circadian clock regulators, including CIRCADIAN CLOCK ASSOCIATED 1 (CCA1). It is unknown whether a similar mechanism mediates maize heterosis. In this dissertation, I investigated whether and how the circadian clock regulation contributes to growth heterosis in maize. I reported that higher levels of carbon fixation and starch accumulation in maize hybrids are associated with altered temporal gene expression. Two maize CCA1 homologs, ZmCCA1a and ZmCCA1b, are diurnally up-regulated in the hybrids. In Arabidopsis ZmCCA1 complements the cca1 mutant phenotype, and overexpressing ZmCCA1b disrupts circadian rhythms and heterosis. Furthermore, overexpressing ZmCCA1b in maize reduced chlorophyll content and plant height. Reduced height stems from reduced node elongation but not total node number in both greenhouse and field conditions. Chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) analysis revealed a temporal shift of ZmCCA1-binding targets to the early morning in the hybrids, suggesting that activation of morning-phased genes in the hybrids promotes photosynthesis and growth vigor. This temporal shift of ZmCCA1-binding targets correlated with nonadditive and additive gene expression in early and late stages of seedling development. These results could guide breeding better hybrid crops to meet the growing demand in food and bioenergy.Item Coevolution between nuclear and plastid genomes in Geraniaceae(2015-08) Zhang, Jin, doctor of plant biology; Jansen, Robert K., 1954-; Herrin, David L.; Linder, C. Randy; Wilke, Claus O.Plastid genomes of angiosperms are highly conserved in both genome organization and nucleotide substitution rates. Geraniaceae have highly rearranged genomes and elevated nucleotide substitution rates, which provides an attractive system to study nuclear-plastid genome coevolution. My dissertation research has focused on two areas of nuclear-plastid genome coevolution in Geraniaceae. First, I have investigated the correlation of nucleotide substitution rates between nuclear and plastid genes that encode interacting subunits that form the multi-subunit complex of Plastid Encoded RNA Polymerase (PEP). Second, the hypothesis that the unusual changes of plastid genome organization and elevated nucleotide substitution rates of plastid encoded genes is the result of alterations in nuclear encoded DNA replication, recombination and repair (DNA RRR) genes is tested. The second chapter investigates the optimal methods for transcriptome sequencing/assembly. My findings supported the use of transcriptome assemblers optimized for Illumina sequencing platform (Trinity and SOAPdenovo-trans). The third chapter investigated coevolution of nucleotide substitution rates between plastid encoded RNAP (rpoA, rpoB, rpoC1, rpoC2) and nuclear encoded SIG (sig1-6) genes that are part of the multi-subunit complex PEP. Using the transcriptomes of 27 Geraniales species I extracted the PEP genes and performed a systematic correlation test. I detected strong correlations of dN (nonsynonymous substitutions) but not dS (synonymous substitutions) between RNAP and SIG but no correlations were detected for the control genes, which provides a plausible explanation for the cause of plastome-genome incompatibility in Geraniaceae. The fourth chapter investigated the effect of DNA RRR system on the aberrant evolutionary phenomena in Geraniaceae plastid genomes. I extracted DNA RRR and nuclear control genes with different subcellular locations from 27 Geraniales transcriptomes and estimated genome complexity with various measures from plastid genomes of the same species. I detected significant correlations for dN but not dS for three DNA RRR genes, 10 nuclear encoded plastid targeted (NUCP) and three nuclear encoded mitochondrial targeted (NUMT) genes. The findings of a correlation between dN of DNA RRR genes and genome complexity support the hypothesis that changes of plastid genome complexity in Geraniaceae may be caused by dysfunction of DNA RRR systems.Item Comparative single-cell diatom population genomics across the world's largest lakes and across the world(2017-07-25) Brady, Mariska Catherine; Theriot, Edward C. (Edward Claiborne), 1953-; Cannatella, David; Jansen, Robert K; Juenger, Thomas E; La Claire, John W; Linder, Craig RandalA single-cell method was developed for the comparative population genomics study of five Aulacoseira species from the Laurentian Great Lakes, North America and Lake Baikal, Siberia. This new methodology was necessary to investigate difficult-to-culture diatoms and overcome challenges associated with culture bias and geographically remote sampling locations. Chapter one presents this method, and documents the method's ability to detect strong genetic differentiation in instances where spatial genetic structure is expected (here, between two populations of the same species from across the globe). Additionally, the sensitivity of this method to resolving population genetic signal consistent with known demographics of the organism (here, clonality) is demonstrated. In chapter two, this method is used to explore the population genomics of three Aulacoseira species from the Great Lakes. Findings indicate that genetic differentiation and diversity vary across species, and -within species- across localities. In Chapter three, findings from previous chapters provide a benchmark to contextualize population genomic findings from A. baicalensis, an endemic diatom, which is foundational to Lake Baikal's simple but endemic pelagic food web. Specific temporal and geographic hypotheses were tested under the expectation that geographic population structure may be detected, given heterogeneous warming and nutrient conditions between basins within Lake Baikal. The null hypothesis of panmixia could not be rejected for A. baicalensis and effective population sizes were estimated to be finite, favoring drift rather than natural selection as the force driving evolution in this species. Observed and expected heterozygosity estimates for A. baicalensis were three times lower than for any other Aulacoseira species investigated, including a congener from Lake Baikal. Reduced genetic diversity in A. biacalensis could have grave consequences for the adaptability of this important endemic phytoplankter when faced with the rapid warming predicted for the Lake. This study contributes needed population genetic data on freshwater planktonic diatoms, and is the first in the diatom field to take a comparative population genomics approach. These findings, alongside those from other recent studies, challenge long-standing paradigms about natural microbial eukaryotic populations, including unlimited gene flow and infinite population sizes, emphasizing the need for more population-level, quantitative data in microbial molecular ecology.Item Consequences, repair, and utilization of an induced double-strand break in the chloroplast DNA of Arabidopsis and tobacco(2011-05) Kwon, Taegun; Herrin, David L.; Herrin, David L.; Roux, Stanley J.; Jansen, Robert K.; Huq, Enamul; De Lozanne, Arturo; Somanchi, AravindIn mature chloroplasts, the DNA (cpDNA) is surrounded by a potentially genotoxic environment that would make the mitochondrial DNA milieu look like a “nadree” (picnic). And yet, the slower evolution of cpDNA compared to other cellular genomes suggests that this organelle must have efficient mechanisms for repairing DNA. Unfortunately, those mechanisms have been barely noted, much less studied. This dissertation describes a novel approach that was developed to study how chloroplasts of Arabidopsis repair the most severe form of DNA damage, a double-strand break (described in Chapter 2). The success with this approach also prompted the development of a new method for site-specific modification of tobacco cpDNA that is described in Chapter 3. To study the consequences and repair of a break in the circular plastid genome, we developed an inducible system based on a psbA-intron endonuclease from Chlamydomonas (I-CreII) that specifically cleaves the psbA gene of Arabidopsis. The protein was targeted to the chloroplast using the rbcS1 transit peptide, and activation of the nuclear gene was made dependent on an exogenous inducer (β-estradiol). In Chlamydomonas, I-CreII cleavage at psbA was repaired, in the absence of the intron, by homologous recombination between repeated sequences (20-60 bp) that are abundant in that genome. By comparison, Arabidopsis cpDNA is very repeat-poor. Nonetheless, phenotypically strong and weak transgenic lines were obtained, and shown to correlate with I-CreII expression levels. Southern blot hybridizations indicated a substantial loss of psbA, but not cpDNA as a whole, in the strongly-expressing line. PCR analysis identified deletions nested around the I-CreII cleavage site that were indicative of repair using microhomology (6-12 bp perfect repeats, or 10-16 bp with mismatches) or no homology. The results provide evidence of alternative repair pathways in the Arabidopsis chloroplast that resemble the nuclear microhomology-mediated and nonhomologous end-joining pathways, in terms of the homology requirement. Moreover, when taken together with the results from Chlamydomonas, plus other considerations, the data suggests that an evolutionary relationship may exist between the repeat structure of cpDNA and the organelle’s ability to repair broken chromosomes. Taking advantage of the inducible I-CreII system, I developed a method to delete defined regions of cpDNA in tobacco, which was named DREEM (for direct repeat and endonuclease mediated). Chloroplast transformation was used to introduce an I-CreII cleavage site adjacent to an aadA:gfp marker and flanked by a direct repeat of 84 bp. When chloroplast-targeted I-CreII was induced with β-estradiol during germination, complete loss of the aadA:gfp marker occurred by SSA-type repair involving the 84-bp direct repeat. I obtained additional evidence for DREEM effectiveness by deleting 3.5 kb of native cpDNA that included part of the large ycf1 gene. DREEM can be used for other modifications besides gene deletions, partly because it is seamless and leaves no trace of introduced DNA. Since expression of the endonuclease is controlled by steroid application (and concentration), and the deleted cpDNA is probably destroyed during the SSA process, this inducible gene-ablation technique could enable the study of essential chloroplast genes in vivo.Item Conserved modulation of the constitutive photomorphogenic 1 E3 ubiquitin ligase activity by the bHLH transcription factors, phytochrome interacting factors(2016-05) Xu, Xiaosa; Huq, Enamul; Roux, Stanley J; Vokes, Steven A; Lloyd, Alan M; Linder, Craig RAs sessile organism, plants are informed of the time of the day and their place of growth by a collection of photoreceptors that detect changing intensity, quality, and direction of light in the environment. Among these photoreceptors, phytochromes (A, B, C, D, E) are the major ones to drive a developmental switch for initial emergence of seedlings from subterranean darkness into sunlight, called plant photomorphogenesis. Previous studies have identified many regulators in the phytochorme-mediated photomorphogenesis pathway. Among them, CONSTITUTIVELY PHOTOMORPHOGENIC 1/ SUPPRESSOR OF PHYTOCHROME A (COP1/SPA) complex and PHYTOCHROME INTERACTING FACTORs (PIF1, 3, 4, 5, 7, 8) are key negative regulators that can suppress photomorphogenesis individually. However, the functional relationships between the COP1-SPA and the PIFs are still unknown. Here in my dissertation project, I showed that PIFs have nontranscriptional roles by acting as cofactors of the COP1 E3 Ubiquitin ligase to enhance the trans-ubiquitination and subsequent degradation of the substrates of COP1, including LONG HYPOCOTYL 5 (HY5), LONG HYPOCOTYL IN FAR-RED 1 (HFR1) and a newly identified substrate HECATE 2 (HEC2), to suppress photomorphogenesis. HFR1 also promotes the degradation of PIF1 in the dark via direct heterodimerization to trigger rapid seed germination upon light exposure. The reciprocal co-degradation between PIF1 and HFR1 is dependent on the ubi/26S-proteasome pathway in vivo. In addition, the cop1 and pif1, 3, 4, 5 mutant combinations showed overproliferation of stigmatic tissues phenotype similar to HEC overexpression plants. Biochemical and genetic evidence showed that HECs are highly abundant in the cop1 pifs mutant flowers. Moreover, HECs negatively regulate the PIFs’ binding activity to the G-box regions of promoters of flower pattern genes, SEP1 and SEP3. Taken together, these data revealed the conserved modulation of the COP1 Ubiquitin E3 ligase activity by PIFs, uncovered a suicidal co-degradation mechanism between the HFR1 and PIF1 to fine tune seed germination and seedling development, and demonstrated a novel function of COP1 and PIFs in regulating flower pattern development.Item Coordinated response and regulation of carotenogenesis in Thermosynechococcus elongatus (BP-1) : implications for commercial application(2014-12) Knight, Rebecca Anne; Brand, Jerry J. (Jerry Jay), 1941-; Alper, Hal S.If small isoprenoids, the starting component of carotenoids, can be efficiently excreted from thermophilic cyanobacteria, they could help satisfy the demand for sustainably produced hydrocarbons. This is the driving force behind wanting to understand the response and regulation of isoprenoid pathways to environmental stimuli in the thermophilic cyanobacterium, Thermosynechococcus elongatus, BP-1. The portion of the isoprenoid pathway studied here is the carotenoid pathway since these products are critical to adaptation and they encompass the largest pool of isoprenoid compounds in cyanobacteria. Although synthetic biology in cyanboacteria has improved in recent years, there are many undiscovered metabolic complexities that make large-scale commercial production challenging. To address this need, I quantify and report for the first time metabolic shifts within the carotenoid pathway of BP-1 due to combined effects of temperature, pH and blue light. I show that metabolism shifts from the dicyclic into the monocyclic carotenoid pathway in response to pH, and that decreasing temperature drives flux into the end products of both pathways. Also, I report that the productivity of an uncommon carotenoid, 2-hydroxymyxol 2’-fucoside (HMF), approached 500 μg/L-day in cultures grown at 45 °C, high light intensity, and pH 8. In order to further elucidate these responses, I analyzed 42 RNAseq samples taken over time of BP-1 induced by cold and heat stress and compared these results to metabolomics data. I showed that crtR and crtG, two central carotenogenesis genes, are transcriptionally controlled and used weighted gene co-expression network analysis (WGCNA) to determine eight separate co-expressed modules of biological significance. Among the co-regulated heat response and cold response genes there were three and five non-coding RNA, respectively, providing targets for future investigation. Using subtractive genomics and transcriptional data I narrowed the potential missing steps of the myxol pathway in cyanobacteria to seven unknown BP-1 genes, two of which were confirmed not to be involved in the missing step(s). Finally, by generating a ΔcrtG mutant and testing it under different environmental parameters, I showed that HMF does not protect against high pH or low temperature (despite up-regulation at these conditions), and that CrtG has a higher affinity for monocyclic than dicyclic carotenoids.Item Critical studies of soil and vegetation relations(1932) Morrow, Marie Betzner; Goldsmith, Glenn W. (Glenn Warren), 1886-Item Determination of the genetic basis of seed oil composition and melting point—adaptive quantitative traits—and their fitness effects in Arabidopsis thaliana(2013-12) Pelc, Sandra Elaine; Linder, C. RandalEvidence indicates seed oil melting point is likely an adaptive quantitative trait in many flowering plant species. An adaptive hypothesis suggests selection has changed the melting point of seed oils to covary with germination temperatures because of a trade-off between total energy stores and the rate of energy acquisition during germination under competition. The predicted differences in relative fitness under different temperatures have not yet been tested and little is known about the genetic basis of differences in oil composition. I used Arabidopsis thaliana to: (1) assess the fitness consequences of high and low melting point seeds germinating at different temperatures, (2) assess what genes underlie natural variation in seed oil composition, and (3) consider how these genes may be used to create oils with particular characteristics. To assess the effects of seed oil melting point on timing of seedling emergence and fitness, I competed high and low melting point lines of A. thaliana under cold and warm temperatures. Emergence timing between these lines was not significantly different at either temperature, which comported with warm temperature predictions but not cold temperature predictions. Under all conditions, plants competing against high melting point lines had lower fitness relative to those against low melting point lines, which matched expectations for undifferentiated emergence times. To assess the genetic basis of naturally occurring variation in seed oil melting point, the seed oil compositions of 391 accessions of A. thaliana were used in a genome-wide association study. Twelve genes were tightly linked with SNPs significantly associated with seed oil melting point variation. Seven encoded lipid synthesis enzymes or regulatory products. The remaining 5 encoded products with no clear relation to seed oil melting point. Results suggest selection can alter quantitative trait variation in response to local conditions through a small set of genes. 268 seed-expressed, candidate genes were linked to 103 SNPs associated with A. thaliana seed oil fatty acids. Eight genes were involved in lipid metabolism, and thirty-four encoded regulatory products. I discuss how knowledge of these genes can be used to breed and engineer desirable oil compositions for industry and nutrition.Item Early responses of the marine diatom Phaeodactylum tricornutum to herbivory-related decadienal : effects on the transcriptome, proteome and regulatory mechanisms of two light harvesting complex genes(2022-10-06) Islam, Shahima; Mehdy, Mona Cynthia, 1955-; Roux, Stanley J; Herrin, David; Sullivan, Christopher; Theriot, EdwardWorldwide food security is based on crop production which largely depends on plants’ stress tolerance capacities at the onset of diverse biotic and abiotic stresses. Photosynthetic organisms have developed many sophisticated defense responses that balance maintenance of central metabolic processes such as photosynthesis with new metabolic demands for acclimation to the stress. However, in both higher plants and algae, how central metabolic pathway-related genes are regulated and the functional significance of the adjustments are poorly understood. Marine diatoms are known to cope with continuous challenges of stress in the ocean and can provide general insights on the roles of rapid photosynthetic and core carbon metabolism gene expression changes in acclimation. During zooplankton grazing, damaged diatom cells release polyunsaturated aldehyde (PUA) from membranes, which impair reproduction in grazers and at the same time affect neighboring undamaged diatoms. In response to PUA, undamaged diatoms cells exhibit remarkable alteration in the photosynthesis-related gene expression. In this study, early response dynamics in the marine diatom Phaeodactylum tricornutum were investigated in response to sublethal concentrations of a model PUA, 2-E, 4-E/Z-decadienal (DD). Genome-wide analysis showed mRNAs encoding photosynthetic Light Harvesting Complex (LHC) antenna proteins were differentially downregulated except for a few upregulated mRNAs at 3 and 6 h of DD treatments. Our findings revealed gene regulatory mechanisms of two differentially regulated LHC transcripts. In cells treated with DD, he downregulation of Lhcf2 mRNA was due to reduced transcription and increased mRNA turnover, and Lhcf15 upregulation depended on increased transcription and mRNA stability. Further, I compared changes in the transcriptome and proteome, especially for photosynthesis-and central carbon metabolism-related pathways to investigate early molecular responses during short-term sublethal DD treatment. The majority of the differentially expressed transcripts involved in photosynthetic light harvesting, chlorophyll biosynthesis were downregulated and central carbon metabolism exhibited a mixed expression. In contrast, few of the corresponding proteins exhibited differential expression. Thus, during the early exposure to DD, proteins involved in photosynthesis-and energyrelated pathway remained predominantly stable despite extensive differential expression of their encoding transcripts. I have postulated that transient suppression of transcripts is an early defense response to save energy and resources to support synthesizing important components needed for acclimation to DD induced stress. Hence, our study provides novel insight into the photosynthetic gene regulatory mechanism and metabolic network reprogramming during early stress response of herbivory-related DD stress condition.Item Ecology and evolution of cold tolerance in Panicum hallii(2018-12-07) Palacio Mejía, Juan Diego; Juenger, Thomas; Jha, Shalene; Lowry, David B; Linder, Randy; Roux, StanGiven that plants are sessile organism, they are continuously challenged by both biotic and abiotic stresses in natural and agricultural conditions which they are forced to acclimate or adapt. One of the major abiotic stressors affecting plant growth and development, and thereby limiting distribution of plants and crop production, is low temperature. It is estimated that crop yield is affected by cold stress in 57% of the global land area. To study the ecology and evolution of cold tolerance in plants, Hall's panicgrass (Panicum hallii), a perennial C₄ grass native to North America, was used to investigate cold tolerance due to its adaptation to diverse environments across its natural distribution. Three approaches were followed in this study: 1. The genetic diversity and population structure as well the response to cold stress were evaluated in samples collected across the natural distributions of the species. 2. Then, genomic regions associated to response to cold temperatures were found by QTL analysis of a RIL population. 3. Finally, a global transcriptome analysis was conducted in a set of selected phenotypes to identify candidate genes involved in cold responses. For the population genetic approach, we evaluated the genomic and morphological diversity as well as the genetic structure of the C₄ grass Panicum hallii using ddRAD molecular approaches. We found strong genetic and morphological divergence between varieties and strong genetic structure between the seven populations that was strongly correlated with geological and ecological conditions. Following the genetic diversity analyses, we utilized QTL and global gene expression approaches to determine the early chilling stress transcriptome response of P. hallii. This analysis revealed 16 genes occurring within a major QTL interval and exhibiting expression responses. Six genes were previously associated with responses to cold stress in studies with other plant species, and five genes were identified with unknown functions. Accordingly, the results here not only aid in the discovery of the genetic mechanism that underline local adaptation but also provide a foundation to improve switchgrass yield under cold conditionsItem The ecology and vegetation of Chihuahua, Mexico, north of parallel twenty-eight(1938) LeSueur, Hardeman David, 1909-1942; Not availableItem Ecophysiology and ecosystem-level impacts of an invasive C4 perennial grass, Bothriochloa ischaemum(2013-12) Basham, Tamara Sue; Fowler, Norma L.; Litvak, Marcy E.The anthropogenic introduction of species into new ecosystems is a global phenomenon, and identifying the mechanisms by which some introduced species become dominant in their introduced ranges (i.e., invasive) is crucial to predicting, preventing, and mitigating the impacts of biological invasions. Introduced perennial C₄ grasses are invading semi-arid grassland and savanna ecosystems throughout the south-central U.S. We hypothesized that in these semi-arid ecosystems, where variable precipitation patterns strongly influence vegetation dynamics, the success of an invasive plant species may be due in part to ecophysiological traits that enable high performance in response to unpredictable water availability. We also hypothesized that increased primary productivity and decreased plant input quality associated with these grass invasions have the potential to alter ecosystem carbon and nitrogen cycling and storage by altering the ratio of inputs (productivity) to outputs (decomposition/respiration). We tested the first hypothesis by quantifying ecophysiological performance differences between an invasive C₄ grass, Bothriochloa ischaemum, and co-occurring C₃ and C₄ native grasses under wet and dry conditions in the field and under two levels of simulated precipitation frequencies in a greenhouse experiment. We tested the second hypothesis by examining whether increased primary productivity and decreased C₃:C₄ grass ratios in savanna grass-matrices associated with B. ischaemum invasion altered (1) plant input quality and thus nutrient cycling and/or (2) net ecosystem carbon uptake in invaded areas. B. ischaemum's success as an invader was not directly related to its ability to cope with precipitation variability and availability, but its ability to rapidly produce large amounts of biomass may allow it to directly out-compete native species. B. ischaemum invasion decreased plant input quality and soil nitrogen availability. B. ischaemum invasion shifted ecosystem C-uptake from being nearly year-round to occurring predominantly in the summer. Greater C-uptake during the summer and under drier conditions compensated for a shorter growing seasons in B. ischaemum-invaded areas and cumulative annual NEE was similar between invaded and native-dominated areas. We conclude that B. ischaemum's impacts on soil nitrogen availability and plant-canopy microhabitat may allow it to exclude native species from invaded areas, but that its impacts on ecosystem C sequestration may be small.Item The effects of extracellular ATP on growth in Arabidopsis thaliana(2007-08) Butterfield, Timothy Scott; Roux, Stanley J.Exciting contributions to the field of plant biology have challenged traditional views of and modes of signaling molecule action and activities. In addition to startling and widely touted discoveries such as F-box protein auxin receptors and endomembrane ethylene receptors, evidence has accumulated indicating that ATP may possess a non-canonical function external to the plant cell as a signaling agent. Recent publications have detailed the presence of extracellular ATP at the growing end of root tips, extracellular ATP-induce fluxes in cytosolic calcium, and the effects of extracellular ATP upon root growth and pollen tube elongation. Within these pages, I present evidence for concentration-dependent effects of extracellular ATP upon etiolated hypocotyl elongation and gene expression changes. These findings prompted us to measure the ATP concentration in wound serum of WT and mutant plants that produce an NTDPase, or apyrase (atAPYOE) protein in greater abundance than WT, and we found that the ATP concentration in wound serum derived from the atAPYOE mutants is lower than WT. I also report that apyrase protein levels decrease following exposure to red light, that it is present on the plasma membrane, and the active site likely faces the extracellular matrix. Finally, I describe my attempts to directly label a putative ATP-receptor protein predicted to reside on the external face of the plasma membrane. I have labeled a small number of plasma membrane proteins using a photoactive, radioactive ATP-derivative. As described more fully in the text, the predicted sequence of the labeled proteins was generated using mass-spec fingerprinting. In sum, I present novel data that contributes to our understanding of how plants and the plant cell perceive and respond to the external environment.Item Elucidating molecular mechanisms of seed development in Arabidopsis and fiber development in cotton(2020-09-03) Ando, Atsumi; Chen, Z. Jeffrey; Jansen, Robert K; Qiao, Hong; Roux, Stanley J; Sung, Sibum; Vokes, Steven ASeeds are important energy resource and source of fabric materials for humans. We intake more than 40% of calories as well as nutrients from various angiosperm seeds, and use cotton fibers derived from seed coats for the majority natural fabric materials. In this dissertation, I investigated two seed development mechanisms; imprinting effects on seed size in Arabidopsis and fiber cell differentiation from seed coats in cotton. Seed size is affected by genetic mutations and imprinting. Imprinting is a widespread epigenetic phenomenon in mammals and flowering plants, in which selected genes are differentially expressed between alleles in a parent-of-origin manner. ETHYLENE INSENSITIVE2 (EIN2) encodes an essential signal molecule that links the ethylene perception on endoplasmic reticulum (ER) to transcriptional regulation in the nucleus. Interestingly, EIN2 is an imprinted gene in both Arabidopsis and maize, and is maternally expressed in the endosperm. The function of most imprinted genes including EIN2 is largely unknown in plants. In chapter two, I show how the imprinted expression of EIN2 regulates seed size in Arabidopsis. Cotton (Gossypium spp.) is the largest renewable source of textile fiber in the world and it is also an important oil crop. Cotton fibers are made up of individual cells that are derived from ~30% of epidermal cells on surface of the cotton seed, which each elongate up to several centimeters. Efforts to understand the causes of this shift has been hampered by the difficulty of isolating fiber cells from epidermal cells at the earliest stages of development. In chapter three and four, I show the gene expression and RNA modification of various tissues as well as different stages of fibers including early stage fibers collected by Laser Capture Microdissection technology, to identify the underlying mechanism of fiber development in cotton. These results provide mechanistic insights into the imprinting effect of EIN2 on seed development and shed light on the early changes in gene activity controlling fiber development in cotton, which may help manipulate seed and fiber yields.Item The endangered plants of Texas(1979) Conner-Ogorzaly, Molly; Turner, B. L. (Billie Lee), 1925-