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Open Access Publications from the University of California

Scholarly Works (11 results)

  • Article
  • Peer Reviewed
UC Berkeley Previously Published Works (2021)
Unlike animals, plants do not have specialized immune cells and lack an adaptive immune system. Instead, plant cells rely on their unique innate immune system to defend against pathogens and coordinate beneficial interactions with commensal and symbiotic microbes. One of the major convergent points for plant immune signaling is the nucleus, where transcriptome reprogramming is initiated to orchestrate defense responses. Mechanisms that regulate selective transport of nuclear signaling cargo and chromatin activity at the nuclear boundary play a pivotal role in immune activation. This review summarizes the current knowledge of how nuclear membrane-associated core protein and protein complexes, including the nuclear pore complex, nuclear transport receptors, and the nucleoskeleton participate in plant innate immune activation and pathogen resistance. We also discuss the role of their functional counterparts in regulating innate immunity in animals and highlight potential common mechanisms that contribute to nuclear membrane-centered immune regulation in higher eukaryotes.
    Cover page: Regulation of Plant Immunity by Nuclear Membrane-Associated Mechanisms
    • Article
    • Peer Reviewed
    UC Berkeley Previously Published Works (2017)
    Plants employ sophisticated mechanisms to interact with pathogenic as well as beneficial microbes. Of those, membrane trafficking is key in establishing a rapid and precise response. Upon interaction with pathogenic microbes, surface-localized immune receptors undergo endocytosis for signal transduction and activity regulation while cell wall components, antimicrobial compounds, and defense proteins are delivered to pathogen invasion sites through polarized secretion. To sustain mutualistic associations, host cells also reprogram the membrane trafficking system to accommodate invasive structures of symbiotic microbes. Here, we provide an analysis of recent advances in understanding the roles of secretory and endocytic membrane trafficking pathways in plant immune activation. We also discuss strategies deployed by adapted microbes to manipulate these pathways to subvert or inhibit plant defense.
      Cover page: Membrane Trafficking in Plant Immunity
      • Thesis
      • Peer Reviewed
      UC Berkeley Electronic Theses and Dissertations (2024)

      Plant pathogens cause significant crop losses, and current pathogen control methods are often detrimental to human and environmental health. Identifying pathogen resistance in cultivars or wild relatives of crops remains the primary approach for achieving pathogen resistance. However, mining natural resistance is time consuming, and population level resistance may not exist for emerging pathogens. As an alternative to traditional resistance breeding, natural molecular diversity can be used to understand the “rules” governing pathogen recognition, which can enable the design of novel resistance within existing recognition frameworks.The intracellular angiosperm immune receptor, HOPZ-ACTIVATED RESISTANCE 1 (ZAR1), recognizes a remarkable diversity of pathogen secreted proteins, known as type 3 secreted effectors (T3SE). ZAR1 interacts with and monitors RLCKs for T3SE modification, including HOPZ ETI-DEFICIENT 1 (ZED1) and ZED1-RELATED KINASES (ZRKs). By monitoring diverse RLCKs, ZAR1 mediates broad immunodiversity through indirect effector interaction. Moreover, ZAR1 is the most highly conserved plant NLR described to date, suggesting the mechanism ZAR1 uses to recognize effectors is effective and warrants further investigation. Chapter 1 introduces the rationale for my approach using natural protein diversity to understand the molecular mechanisms of plant pathogen resistance. In Chapter 2, I describe a natural diversity screen using Arabidopsis thaliana ecotypes to investigate the molecular mechanism of ZAR1 and ZED1-mediated recognition of the Pseudomonas syringae T3SE HopZ1a. This work demonstrates key regions in ZAR1 that are required for ZED1 interaction and HopZ1a recognition. In Chapter 3, I used natural protein diversity of A. thaliana RLCKs to investigate ZAR1 interaction specificity. I showed that single mutations in ZAR1 guardees can increase interaction strength with ZAR1, which is a crucial first step towards rational design of a ZAR1 guardee. Moreover, I showed that a Nicotiana benthamiana ZAR1 ortholog causes an autoactive HR phenotype in response to expression of certain A. thaliana ZRKs. In Chapter 4, I generated a ZED1 point mutant library, and screened for HopZ1a recognition through A. thaliana ZAR1 and a Nicotiana benthamiana ZAR1 ortholog. While I 1 identified some functional divergence between ZAR1 orthologs, these ZAR1 orthologs are highly similar. I identified multiple mutations that abolish interspecies ZAR1-ZRK autoactivity. I also identified a mutation in ZED1 that abolishes autoactivity through tomato ZAR1, while still maintaining the ability to mediate HopZ1a recognition through tomato ZAR1. This advance is the first example of rational expansion of effector recognition through ZAR1 orthologs from non-model species. In Chapter 5, I explored natural resistance to the highly virulent bacterial pathogen P. syringae pv. tomato race 1 (Pst race 1). Using a mapping population created by crossing a resistant wild tomato relative Solanum pimpinellifolium with a susceptible cultivated tomato variety, I used a novel sequencing approach to identify candidate genes contributing to Pst race 1 resistance. Traditional QTL mapping approaches assisted in narrowing down the list of candidate genes, for future evaluation in breeding programs. In Chapter 6, I used single chain antibodies to attempt to achieve ZAR1 activation through antigen binding. Using two linker design approaches, I generated ZRK-GFP nanobody fusions with tailored conformations. I showed that ZRK-GFP nanobody fusions can successfully interact with ZAR1 and the GFP antigen. Although no ZRK-GFP nanobody fusions activate ZAR1 in the presence of the GFP antigen, future protein design tools may enable the success of this approach. Overall, the work presented here elucidates key molecular mechanisms of ZAR1 mediated recognition of P. syringae effectors by leveraging natural diversity. It identifies candidate genes contributing to P. syringae resistance in a tomato mapping population. Finally, it explores approaches to novel ZAR1-mediated effector recognition.

        • Article
        • Peer Reviewed
        UC Berkeley Previously Published Works (2022)
        The nuclear basket (NB) is an essential structure of the nuclear pore complex (NPC) and serves as a dynamic and multifunctional platform that participates in various critical nuclear processes, including cargo transport, molecular docking, and gene expression regulation. However, the underlying molecular mechanisms are not completely understood, particularly in plants. Here, we identified a guanylate-binding protein (GBP)-like GTPase (GBPL3) as a novel NPC basket component in Arabidopsis. Using fluorescence and immunoelectron microscopy, we found that GBPL3 localizes to the nuclear rim and is enriched in the nuclear pore. Proximity labeling proteomics and protein-protein interaction assays revealed that GBPL3 is predominantly distributed at the NPC basket, where it physically associates with NB nucleoporins and recruits chromatin remodelers, transcription apparatus and regulators, and the RNA splicing and processing machinery, suggesting a conserved function of the NB in transcription regulation as reported in yeasts and animals. Moreover, we found that GBPL3 physically interacts with the nucleoskeleton via disordered coiled-coil regions. Simultaneous loss of GBPL3 and one of the 4 Arabidopsis nucleoskeleton genes CRWNs led to distinct development- and stress-related phenotypes, ranging from seedling lethality to lesion development, and aberrant transcription of stress-related genes. Our results indicate that GBPL3 is a bona fide component of the plant NPC and physically and functionally connects the NB with the nucleoskeleton, which is required for the coordination of gene expression during plant development and stress responses.
          Cover page: GBPL3 localizes to the nuclear pore complex and functionally connects the nuclear basket with the nucleoskeleton in plants
          • Article
          • Peer Reviewed
          UC Berkeley Previously Published Works (2016)
          Nuclear transport of immune receptors, signal transducers, and transcription factors is an essential regulatory mechanism for immune activation. Whether and how this process is regulated at the level of the nuclear pore complex (NPC) remains unclear. Here, we report that CPR5, which plays a key inhibitory role in effector-triggered immunity (ETI) and programmed cell death (PCD) in plants, is a novel transmembrane nucleoporin. CPR5 associates with anchors of the NPC selective barrier to constrain nuclear access of signaling cargos and sequesters cyclin-dependent kinase inhibitors (CKIs) involved in ETI signal transduction. Upon activation by immunoreceptors, CPR5 undergoes an oligomer to monomer conformational switch, which coordinates CKI release for ETI signaling and reconfigures the selective barrier to allow significant influx of nuclear signaling cargos through the NPC. Consequently, these coordinated NPC actions result in simultaneous activation of diverse stress-related signaling pathways and constitute an essential regulatory mechanism specific for ETI/PCD induction.
            Cover page: Nuclear Pore Permeabilization Is a Convergent Signaling Event in Effector-Triggered Immunity
            • Article
            • Peer Reviewed
            UC Berkeley Previously Published Works (2014)
            Effector-triggered immunity (ETI), the major host defense mechanism in plants, is often associated with programmed cell death (PCD). Plants lack close homologs of caspases, the key mediators of PCD in animals. So although the NB-LRR receptors involved in ETI are well studied, how they activate PCD and confer disease resistance remains elusive. We show that the Arabidopsis nuclear envelope protein, CPR5, negatively regulates ETI and the associated PCD through a physical interaction with cyclin-dependent kinase inhibitors (CKIs). Upon ETI induction, CKIs are released from CPR5 to cause overactivation of another core cell-cycle regulator, E2F. In cki and e2f mutants, ETI responses induced by both TIR-NB-LRR and CC-NB-LRR classes of immune receptors are compromised. We further show that E2F is deregulated during ETI, probably through CKI-mediated hyperphosphorylation of retinoblastoma-related 1 (RBR1). This study demonstrates that canonical cell-cycle regulators also play important noncanonical roles in plant immunity.
              Cover page: A Noncanonical Role for the CKI-RB-E2F Cell-Cycle Signaling Pathway in Plant Effector-Triggered Immunity
              • Article
              • Peer Reviewed
              UC Berkeley Previously Published Works (2020)
              The inner nuclear membrane (INM) selectively accumulates proteins that are essential for nuclear functions; however, overaccumulation of INM proteins results in a range of rare genetic disorders. So far, little is known about how defective, mislocalized, or abnormally accumulated membrane proteins are actively removed from the INM, especially in plants and animals. Here, via analysis of a proximity-labeling proteomic profile of INM-associated proteins in Arabidopsis, we identify critical components for an INM protein degradation pathway. We show that this pathway relies on the CDC48 complex for INM protein extraction and 26S proteasome for subsequent protein degradation. Moreover, we show that CDC48 at the INM may be regulated by a subgroup of PUX proteins, which determine the substrate specificity or affect the ATPase activity of CDC48. These PUX proteins specifically associate with the nucleoskeleton underneath the INM and physically interact with CDC48 proteins to negatively regulate INM protein degradation in plants.
                Cover page: Proximity labeling proteomics reveals critical regulators for inner nuclear membrane protein degradation in plants
                • Article
                • Peer Reviewed
                UC Berkeley Previously Published Works (2015)
                NPR1, a master regulator of basal and systemic acquired resistance in plants, confers immunity through a transcriptional cascade, which includes transcription activators (e.g., TGA3) and repressors (e.g., WRKY70), leading to the massive induction of antimicrobial genes. How this single protein orchestrates genome-wide transcriptional reprogramming in response to immune stimulus remains a major question. Paradoxically, while NPR1 is essential for defense gene induction, its turnover appears to be required for this function, suggesting that NPR1 activity and degradation are dynamically regulated. Here we show that sumoylation of NPR1 by SUMO3 activates defense gene expression by switching NPR1's association with the WRKY transcription repressors to TGA transcription activators. Sumoylation also triggers NPR1 degradation, rendering the immune induction transient. SUMO modification of NPR1 is inhibited by phosphorylation at Ser55/Ser59, which keeps NPR1 stable and quiescent. Thus, posttranslational modifications enable dynamic but tight and precise control of plant immune responses.
                  Cover page: Posttranslational Modifications of the Master Transcriptional Regulator NPR1 Enable Dynamic but Tight Control of Plant Immune Responses
                  • Article
                  • Peer Reviewed
                  UC Berkeley Previously Published Works (2016)
                  It is an apparent conundrum how plants evolved effector-triggered immunity (ETI), involving programmed cell death (PCD), as a major defence mechanism against biotrophic pathogens, because ETI-associated PCD could leave them vulnerable to necrotrophic pathogens that thrive on dead host cells. Interestingly, during ETI, the normally antagonistic defence hormones, salicylic acid (SA) and jasmonic acid (JA) associated with defence against biotrophs and necrotrophs respectively, both accumulate to high levels. In this study, we made the surprising finding that JA is a positive regulator of RPS2-mediated ETI. Early induction of JA-responsive genes and de novo JA synthesis following SA accumulation is activated through the SA receptors NPR3 and NPR4, instead of the JA receptor COI1. We provide evidence that NPR3 and NPR4 may mediate this effect by promoting degradation of the JA transcriptional repressor JAZs. This unique interplay between SA and JA offers a possible explanation of how plants can mount defence against a biotrophic pathogen without becoming vulnerable to necrotrophic pathogens.
                    Cover page: Salicylic acid receptors activate jasmonic acid signalling through a non-canonical pathway to promote effector-triggered immunity
                    • Article
                    • Peer Reviewed
                    UC Berkeley Previously Published Works (2021)
                    Transcription initiation has long been considered a primary regulatory step in gene expression. Recent work, however, shows that downstream events, such as transcription elongation, can also play important roles.1-3 A well-characterized example from animals is promoter-proximal pausing, where transcriptionally engaged Pol II accumulates 30-50 bp downstream of the transcription start site (TSS) and is thought to enable rapid gene activation.2 Plants do not make widespread use of promoter-proximal pausing; however, in a phenomenon known as 3' pausing, a significant increase in Pol II is observed near the transcript end site (TES) of many genes.4-6 Previous work has shown that 3' pausing is promoted by the BORDER (BDR) family of negative transcription elongation factors. Here we show that BDR proteins play key roles in gene repression. Consistent with BDR proteins acting to slow or pause elongating Pol II, BDR-repressed genes are characterized by high levels of Pol II occupancy, yet low levels of mRNA. The BDR proteins physically interact with FPA,7 one of approximately two dozen genes collectively referred to as the autonomous floral-promotion pathway,8 which are necessary for the repression of the flowering time gene FLOWERING LOCUS C (FLC).9-11 In early-flowering strains, FLC expression is repressed by repressive histone modifications, such as histone H3 lysine 27 trimethylation (H3K27me3), thereby allowing the plants to flower early. These results suggest that the repression of transcription elongation by BDR proteins may allow for the temporary pausing of transcription or facilitate the long-term repression of genes by repressive histone modifications.
                      Cover page: The BORDER family of negative transcription elongation factors regulates flowering time in Arabidopsis
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