UC Berkeley

One of the largest and longest standing societal challenges is acquiring enough food to support the human population. Our current era has the added complexities of the population nearing 9 billion, a globalized industrial agricultural production system, and a clearer understanding of how environmentally harmful and unsustainable that system is. While these issues are multifaceted, agriculture is fundamentally coupled with plant and microbial biology and there has been a diversity of avenues pursued by researchers seeking potential solutions and disruptive innovations to address our need to provide healthy food for the world without destroying it in the process. Modern agricultural practices currently require high energetic costs through extensive irrigation, application of synthetic fertilizers and pesticides, and the use of large, mechanized equipment. The lack of biodiversity in large-scale monoculture systems also presents issues in crop nutritional quality, disease pressure, and soil health. Recent advancements in our understanding of plant root systems and the relationships they form with soil microbiota have demonstrated a wealth of potential applications regarding these issues as well as in further illuminating the complex processes at play in plant health and development in an ecological context. It has become increasingly clear that plant roots are essential components to local and global nutrient cycling and drive much of the microbial activity in soils. In order to ensure a future with sustainable, climate-resilient agriculture which enables nutritious diets, we must continue to explore the wealth of knowledge underground. This dissertation seeks to clarify some of the socioeconomic reasons why we grow food the way we do and the limitations of material available for study, as well as underlying biochemical and genetic mechanisms which influence plant root biology and their relationships with microorganisms. The results of this research contribute to the growing body of knowledge in root biology and a paradigm shift in how we understand plants and agriculture to be connected to the wider ecosystem.Chapter 1 seeks to understand the historical and contemporary contexts which influence the scale, scope, and direction of research in agricultural and plant sciences. Analyzing the socioeconomic institution of modern Science, from the conditions leading to the Scientific Revolution in the 16th century to modern day, reveals the conserved influence of western European colonial-imperialism on global agricultural production. From its inception, the institution of Science is shown to be integral to the expansion and maintenance of Western imperial powers materially and socially, by driving critical revenue generation via the enabling and adoption of cash crop agriculture and through controlling the value and direction of intellectual pursuits. From the 19th century we see Science increasingly entangled with emerging capitalist corporate and state interests, which further entrench practices that began with cash crop agriculture and prove to be detrimental to environmental health while distancing crop production from nutritional needs. I describe how these relationships ultimately limit the resources we have available for research in the modern day, hindering our abilities to address the myriad socio-scientific challenges of this century. This clarity is important when making the critical and strategic assessments necessary to direct on-going and future research and teaching efforts. Chapter 2 is an effort to detail that plant roots are essential to plant health and adaptation as well as important contributors to numerous ecological and biogeochemical processes. Despite this, they have been comparatively understudied aspects of plant biology. Recent developments have indicated that a better understanding of root systems can reveal breeding and engineering applications towards plants more well-suited for low-input agricultural systems and frequent stresses. I highlight four co-authored publications concerning the study of root systems: Detailing the limitations and considerations of root system study as well as demonstrations of progress to address these challenges, metabarcoding combined with genomics and data analytics programs to inform the basis of rhizosphere microbiome heritability in Sorghum bicolor, constitutive promoters to fine tune gene expression in synthetic biology with demonstrated function in root systems, and utilizing root system imaging methods and software to inform the effect of a bacterial metabolite on plant-bacterial associations in Arabidopsis thaliana. These studies hope to provide examples of and inspire further efforts to understand root systems, thus enabling improvements in plant performance under new and changing agricultural practices better suited for the world we live in today. Chapter 3 seeks to further illuminate the underlying mechanisms of pectin biosynthesis. Pectin is an abundant component of plant cell walls demonstrated to be important in cell to cell adhesion, plant development, and a variety of signaling pathways. Understanding has so far remained limited due to the wide diversity and redundancy of enzymes involved in the process. The pectin component rhamnogalacturonan I (RG-I) is biosynthesized in part by rhamnosyltransferases (RRTs) in the GT106 family. While it is known that RRTs are expressed in a range of tissues, few studies have demonstrated their function outside of RG-I biosynthesis in seed coat mucilage. In this study we show Lotus japonicus RRT1 contributes to the addition of rhamnose monosaccharide residues to RG-I in root tissues. Mutants contained ~19% less rhamnose in their roots and pectin from aboveground tissues had less galactose and more xylose. RG-I in root tissue from Ljrrt1 also had a larger molecular weight and altered structure compared to wild type (WT) Gifu plants, but this was not due to transcriptional differences in other GTs responsible for RG-I biosynthesis. Mutants exhibited altered root morphology, impacted stem and root growth, and impairment of nodule formation when inoculated with Mesorhizobium loti. These findings constitute the first demonstration of RRT function in vascular plants outside of seed coat mucilage and contribute to the increasingly nuanced understanding of RG-I in cell wall biosynthesis and intersecting processes. Chapter 4 is an effort to characterize an unknown subclass of plant GTPase-related signaling proteins which appear to influence symbiotic relationships formed in root systems. Plants possess a unique class of heterotrimeric G? subunits called extra-large GTPases (XLGs) which contribute to numerous developmental and stress responses. XLGs have an uncharacterized N-terminal domain, a G?-like C-terminal domain, and overlapping and distinct functions compared to conventional G? subunits. In this study, we identified homologs of XLG3 in Lotus japonicus responsive to rhizobial and mycorrhizal symbiosis. However, these proteins were approximately one-third the size of conventional XLGs and only aligned to the N-terminal domain, containing a putative NLS and the cysteine-rich domain of unknown function. Multiple sequence alignment and phylogenetic analysis determined SXLGs did not share domains with other mono- or heterotrimeric G-protein classes and exhibited a pattern of duplication and neofunctionalization typical of genes involved in symbiotic signaling pathways. Transient expression of LjSXLGs in tobacco demonstrated their potential for localization to the plasma membrane, nucleus, and nucleolus. Analysis of L. japonicus sxlg2 mutants revealed transient impairment of immature nodule formation in a destructive experimental setup and inhibition of infection events in a nutrient-limited non-destructive experimental setup, with no observed difference in nodule maturation rate. Additionally, sxlg2 mutants showed a potential impairment of the root growth response in N-limited conditions. SXLGs present an ideal opportunity to better understand the evolution, function, and structure of XLGs and are another example of G-protein involvement in symbiotic relationships. Ultimately, this research supports growing efforts to develop more resilient and sustainable agricultural practices through a focus on root systems biology by providing assessments of the technical and methodological resources in the field, demonstrating dynamics of pectin biosynthesis in root tissue, and uncovering new elements of plant symbiotic and G-protein related signaling pathways. These findings will promote further mechanistic and evolutionary discoveries in aspects of root biology that remain filled with questions and unknowns, but also notable potential in the development of roots more amenable to regenerative agricultural approaches, maximizing benefits from microbial associations, and utilization in the biosynthesis of valuable products and biofuels. The situating of historical and contemporary socioeconomic contexts that have heavily influenced the progression of agriculture and plant sciences over the past half a millennium is a critical addition to our ability to wholly assess the materials, practices, and technologies available for further research. This clarity is essential if we truly wish to address the systemic challenges of our era. Technical solutions have limited ability to resolve complex socio-scientific issues without understanding the broader social contexts they were born from and operate in. In the same way that systems biology has begun to permeate many scientific disciplines, our perspectives must shift to accommodate the nuance and complexity of how interconnected this world is