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Scholarly Works (57 results)

  • Thesis
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UC Berkeley Electronic Theses and Dissertations (2022)

Polyploidy, the condition of having more than two sets of chromosomes, is very common in the plant kingdom. However, the role of polyploidy, or whole-genome duplication (WGD), in plant evolution is far from clear. In some cases, polyploidy appears to be an engine of saltational evolution, while in other cases it seems to have little appreciable effect. To understand the role of polyploidy in plant evolution, we must look to the genetics and genomics of the WGD event itself. The progenitors’ genomes play a crucial role in establishing the polyploid’s genetic stability and ultimate evolutionary trajectory. We have investigated genome structure and evolution in Brachypodium hybridum (2n=4x=30). The purple false brome B. distachyon (2n=2x=10) is a well-established model organism, and it is also one of the progenitors of B. hybridum, the other being B. stacei (2n=2x=20). Despite the availability of many genetic resources for B. distachyon, little is known about the allotetraploid B. hybridum. Here, we describe several genomic features of two independent natural lineages of B. hybridum. In chapter 1, I describe the transcriptional landscape of the model B. hybridum accession ABR113. We found that polyploidy had no appreciable effect on the transcriptome, and differences between progenitor and subgenome were within the level of variation we would expect between accessions of the same diploid species. We used a novel and straightforward analytical approach to cross-species RNA-seq that may be valuable to other researchers. We also demonstrated the importance of including the progenitors in studies of polyploid gene expression, as ‘parental legacy’ appeared to be the main driver of gene expression. This was part of a larger effort to survey the genome of ABR113, which was published in Gordon et al. (2020). Chapter 2 constitutes the main project of my doctoral research. This project describes the genome of B. hybridum accession Bhyb26. Bhyb26 is an older lineage of B. hybridum: the Bhyb26 WGD event is estimated to have occurred 1.4 million years ago, while the ABR113 WGD occurred approximately 140,000 years ago (Gordon et al. 2020). Both lineages possess the expected karyotype, that is, a D subgenome of 10 chromosomes and an S subgenome of 20 chromosomes, with no major rearrangements or sequence loss. However, we found that the Bhyb26 genome shows subtle genomic changes consistent with relaxed natural selection where the younger ABR113 genome did not. When we searched for biased genome evolution favoring one subgenome in Bhyb26, the results were mixed. We conclude that Bhyb26 is evolving toward the diploid state in a very gradual and largely unbiased manner. This study constitutes a rare glimpse of diploidization “caught in the act” on two evolutionary timescales. Chapter 3 is a survey of the 3D genome topology of B. hybridum ABR113 and its progenitors using Hi-C technology. During the course of our survey, we stumbled onto a mysterious 3D chromatin structure in the B. hybridum genome. We find evidence that this 3D chromatin structure, composed of mutually contacting loci akin to the KEEs/IHIs described in Arabidopsis (Feng et al. 2014; Grob et al. 2014), was passed down from progenitors to polyploid. The two separate chromatin interaction networks merge to form a single network in the polyploid, which we refer to as the KNOT, in keeping with Grob et al. (2014). Transposable elements (TEs) in the Brachypodium KNOT bear certain characteristics that are consistent with proposed functions of the Arabidopsis KNOT in TE defense, though the participating loci are not as TE-rich as those hypotheses predict. This study approaches the question of ‘parental legacy’ from an unusual angle: the inheritance of 3D chromatin structural features. In summary, we find that B. hybridum belongs in a class of polyploids that show a subtle and unbiased, rather than a disruptive, genomic response to WGD. We provide theoretical arguments explaining this observation. B. hybridum supports the hypothesis that even an allopolyploid from a very wide cross can undergo gradual, unbiased evolution provided that the progenitor genomes are similar in their TE load.

  • 1 supplemental ZIP
Cover page: Multiple origins, one evolutionary trajectory: gradual genome evolution in the allotetraploid grass Brachypodium hybridum
  • Article
  • Peer Reviewed
UC Berkeley Previously Published Works (2019)

Background

Our understanding of polyploid genomes is limited by our inability to definitively assign sequences to a specific subgenome without extensive prior knowledge like high resolution genetic maps or genome sequences of diploid progenitors. In theory, existing methods for assigning sequences to individual species from metagenome samples could be used to separate subgenomes in polyploid organisms, however, these methods rely on differences in coarse genome properties like GC content or sequences from related species. Thus, these approaches do not work for subgenomes where gross features are indistinguishable and related genomes are lacking. Here we describe a method that uses rapidly evolving repetitive DNA to circumvent these limitations.

Results

By using short, repetitive, DNA sequences as species-specific signals we separated closely related genomes from test datasets and subgenomes from two polyploid plants, tobacco and wheat, without any prior knowledge.

Conclusion

This approach is ideal for separating the subgenomes of polyploid species with unsequenced or unknown progenitor genomes.
    • Article
    • Peer Reviewed
    UC Berkeley Previously Published Works (2021)
    In recent years, the root microbiome (i.e., microorganisms growing inside, on, or in close proximity to plant roots) has been shown to play an important role in plant health and productivity. Despite its importance, the root microbiome is challenging to study because of its complexity, heterogeneity, and subterranean location. Fortunately, root microbiome research has seen a tremendous influx of novel technologies (e.g., imaging tools, robotics, and molecular analyses), experimental platforms (e.g., micro- and mesocosms), and data integration, modeling, and prediction tools in the past decade that have greatly increased our ability to dissect the complex network of interactions between above- and belowground environmental parameters, plants, bacteria, and fungi that dictate soil and broader ecosystem health. Herein, we discuss methods that are currently used in root microbiome research and that can be expanded to phytobiome research in general ranging from laboratory studies to mesocosm-scale studies and, finally, to field studies; evaluate their relevance to ecosystem studies; and discuss future root microbiome research directions.
      Cover page: Novel and Emerging Capabilities that Can Provide a Holistic Understanding of the Plant Root MicrobiomeCreative Commons 'BY-NC-ND' version 4.0 license
      • Article
      • Peer Reviewed
      UC Berkeley Previously Published Works (2016)
      The genetic transformation of monocot grasses is a resource intensive process, the quality and efficiency of which is dependent in part upon the method of DNA introduction, as well as the ability to effectively separate transformed from wildtype tissue. Agrobacterium-mediated transformation of Brachypodium has relied mainly on Agrobacterium tumefaciens strain AGL1. Currently the antibiotic hygromycin B has been the selective agent of choice for robust identification of transgenic calli in Brachypodium distachyon and Brachypodium sylvaticum but few other chemicals have been shown to work as well for selection of transgenic Brachypodium cells in tissue culture. This study demonstrates that Agrobacterium rhizogenes strain 18r12v and paromomycin selection can be successfully used for the efficient generation of transgenic B. distachyon and B. sylvaticum. Additionally we observed that the transformation rates were similar to or higher than those obtained with A. tumefaciens strain AGL1 and hygromycin selection. The A. rhizogenes strain 18r12v harboring the pARS1 binary vector and paromomycin selection is an effective means of generating transgenic Brachypodium plants. This novel approach will facilitate the transgenic complementation of T-DNA knockout mutants of B. distachyon which were created using hygromycin selection, as well as aid the implementation of more complex genome manipulation strategies which require multiple rounds of transformation.
        Cover page: Use of Agrobacterium rhizogenes Strain 18r12v and Paromomycin Selection for Transformation of Brachypodium distachyon and Brachypodium sylvaticum
        • Article
        • Peer Reviewed
        UC Berkeley Previously Published Works (2019)
        Predicting phenotypic expression from genomic and environmental information is arguably the greatest challenge in today's biology. Being able to survey genomic content, e.g., as single-nucleotide polymorphism data, within a diverse population and predict the phenotypes of external traits, represents the holy grail across genome-informed disciplines, from personal medicine and nutrition to plant breeding. In the present study, we propose a two-step procedure in bridging the genome to phenome gap where external phenotypes are viewed as emergent properties of internal phenotypes, such as molecular profiles, in interaction with the environment. Using biomass accumulation and shoot-root allometry as external traits in diverse genotypes of the model grass Brachypodium distachyon, we established correlative models between genotypes and metabolite profiles (metabotypes) as internal phenotypes, and between metabotypes and external phenotypes under two contrasting watering regimes. Our results demonstrate the potential for employing metabotypes as an integrator in predicting external phenotypes from genomic information.
          Cover page: Metabotyping as a Stopover in Genome-to-Phenome Mapping
          • Article
          • Peer Reviewed
          UC Berkeley Previously Published Works (2019)
          Maize domestication and breeding have resulted in drastic and well documented changes in aboveground traits, but belowground effects on root system functioning and rhizosphere microbial communities remain poorly understood, despite their critical importance for nutrient and water acquisition. We investigated the rhizosphere microbial community composition and structure of ten Zea mays accessions along an evolutionary transect (two teosinte, three inbred maize lines, and five modern maize hybrids) grown in nutrient depleted soil from a low input agricultural system. Microbial community analysis revealed significant differences in community composition between soil compartments (proximal vs. distal rhizosphere) and between plant genetic groups (teosinte, inbred, and modern hybrid). Only a small portion of the microbial community was differentially selected across plant genetic groups: 3.7% of prokaryotic community members and 4.9% of fungal community members were significantly associated with a specific plant genetic group. Indicator species analysis showed the greatest differentiation between modern hybrids and the other two plant genetic groups. Co-occurrence network analysis revealed that microbial co-occurrence patterns of the inbred maize lines' rhizosphere were significantly more similar to those of the teosintes than to the modern hybrids. Our results suggest that advances in hybrid development significantly impacted rhizosphere microbial communities and network assembly.
            Cover page: Impacts of Maize Domestication and Breeding on Rhizosphere Microbial Community Recruitment from a Nutrient Depleted Agricultural Soil
            • Article
            • Peer Reviewed
            UC Berkeley Previously Published Works (2007)

            Background

            The hypersensitive necrosis response (HR) of resistant plants to avirulent pathogens is a form of programmed cell death in which the plant sacrifices a few cells under attack, restricting pathogen growth into adjacent healthy tissues. In spite of the importance of this defense response, relatively little is known about the plant components that execute the cell death program or about its regulation in response to pathogen attack.

            Results

            We isolated the edr2-6 mutant, an allele of the previously described edr2 mutants. We found that edr2-6 exhibited an exaggerated chlorosis and necrosis response to attack by three pathogens, two powdery mildew and one downy mildew species, but not in response to abiotic stresses or attack by the bacterial leaf speck pathogen. The chlorosis and necrosis did not spread beyond inoculated sites suggesting that EDR2 limits the initiation of cell death rather than its spread. The pathogen-induced chlorosis and necrosis of edr2-6 was correlated with a stimulation of the salicylic acid defense pathway and was suppressed in mutants deficient in salicylic acid signaling. EDR2 encodes a novel protein with a pleckstrin homology and a StAR transfer (START) domain as well as a plant-specific domain of unknown function, DUF1336. The pleckstrin homology domain binds to phosphatidylinositol-4-phosphate in vitro and an EDR2:HA:GFP protein localizes to endoplasmic reticulum, plasma membrane and endosomes.

            Conclusion

            EDR2 acts as a negative regulator of cell death, specifically the cell death elicited by pathogen attack and mediated by the salicylic acid defense pathway. Phosphatidylinositol-4-phosphate may have a role in limiting cell death via its effect on EDR2. This role in cell death may be indirect, by helping to target EDR2 to the appropriate membrane, or it may play a more direct role.
              Cover page: EDR2 negatively regulates salicylic acid-based defenses and cell death during powdery mildew infections of Arabidopsis thaliana.
              • Article
              • Peer Reviewed
              UC Berkeley Previously Published Works (2016)
              Stomata, epidermal valves facilitating plant-atmosphere gas exchange, represent a powerful model for understanding cell fate and pattern in plants. Core basic helix-loop-helix (bHLH) transcription factors regulating stomatal development were identified in Arabidopsis, but this dicot's developmental pattern and stomatal morphology represent only one of many possibilities in nature. Here, using unbiased forward genetic screens, followed by analysis of reporters and engineered mutants, we show that stomatal initiation in the grass Brachypodium distachyon uses orthologs of stomatal regulators known from Arabidopsis but that the function and behavior of individual genes, the relationships among genes, and the regulation of their protein products have diverged. Our results highlight ways in which a kernel of conserved genes may be alternatively wired to produce diversity in patterning and morphology and suggest that the stomatal transcription factor module is a prime target for breeding or genome modification to improve plant productivity.
                Cover page: Grasses use an alternatively wired bHLH transcription factor network to establish stomatal identity
                • Article
                • Peer Reviewed
                UC Berkeley Previously Published Works (2018)
                Transposable element (TE) activity has emerged as a major cause of variation in genome size and structure among species. To what extent TEs contribute to genetic variation and divergence within species, however, is much less clear, mainly because population genomic data have so far only been available for the classical model organisms. In this study, we use the annual Mediterranean grass Brachypodium distachyon to investigate TE dynamics in natural populations. Using whole-genome sequencing data for 53 natural accessions, we identified more than 5,400 TE polymorphisms across the studied genomes. We found, first, that while population bottlenecks and expansions have shaped genetic diversity in B. distachyon, these events did not lead to lineage-specific activations of TE families, as observed in other species. Instead, the same families have been active across the species range and TE activity is homogeneous across populations, indicating the presence of conserved regulatory mechanisms. Second, almost half of the TE insertion polymorphisms are accession-specific, most likely because of recent activity in expanding populations and the action of purifying selection. And finally, although TE insertion polymorphisms are underrepresented in and around genes, more than 1,000 of them occur in genic regions and could thus contribute to functional divergence. Our study shows that while TEs in B. distachyon are "well-behaved" compared with TEs in other species with larger genomes, they are an abundant source of lineage-specific genetic variation and may play an important role in population divergence and adaptation.
                  Cover page: Recent activity in expanding populations and purifying selection have shaped transposable element landscapes across natural accessions of the Mediterranean grass Brachypodium distachyon
                  • Article
                  • Peer Reviewed
                  UC Berkeley Previously Published Works (2010)
                  Abstract Background Glycoside hydrolases cleave the bond between a carbohydrate and another carbohydrate, a protein, lipid or other moiety. Genes encoding glycoside hydrolases are found in a wide range of organisms, from archea to animals, and are relatively abundant in plant genomes. In plants, these enzymes are involved in diverse processes, including starch metabolism, defense, and cell-wall remodeling. Glycoside hydrolase genes have been previously cataloged for Oryza sativa (rice), the model dicotyledonous plant Arabidopsis thaliana, and the fast-growing tree Populus trichocarpa (poplar). To improve our understanding of glycoside hydrolases in plants generally and in grasses specifically, we annotated the glycoside hydrolase genes in the grasses Brachypodium distachyon (an emerging monocotyledonous model) and Sorghum bicolor (sorghum). We then compared the glycoside hydrolases across species, at the levels of the whole genome and individual glycoside hydrolase families. Results We identified 356 glycoside hydrolase genes in Brachypodium and 404 in sorghum. The corresponding proteins fell into the same 34 families that are represented in rice, Arabidopsis, and poplar, helping to define a glycoside hydrolase family profile which may be common to flowering plants. For several glycoside hydrolase familes (GH5, GH13, GH18, GH19, GH28, and GH51), we present a detailed literature review together with an examination of the family structures. This analysis of individual families revealed both similarities and distinctions between monocots and eudicots, as well as between species. Shared evolutionary histories appear to be modified by lineage-specific expansions or deletions. Within GH families, the Brachypodium and sorghum proteins generally cluster with those from other monocots. Conclusions This work provides the foundation for further comparative and functional analyses of plant glycoside hydrolases. Defining the Brachypodium glycoside hydrolases sets the stage for Brachypodium to be a grass model for investigations of these enzymes and their diverse roles in planta. Insights gained from Brachypodium will inform translational research studies, with applications for the improvement of cereal crops and bioenergy grasses.
                  • 6 supplemental files
                  Cover page: Annotation and comparative analysis of the glycoside hydrolase genes in Brachypodium distachyon
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