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Scholarly Works (5 results)

  • Thesis
  • Peer Reviewed
UC Riverside Electronic Theses and Dissertations (2015)

Gram negative pathogens rely on the Type III secretion system to inject virulence proteins (aka secrete effectors) into eukaryotic hosts to manipulate defense. YopJ is one of the largest effector family prevalently produced by both animal and plant pathogens. HopZ1a from the plant pathogen Pseudomonas syringae pv. syringae A2 possesses an acetyltransferase activity and acetylates target proteins in plant hosts for virulence.

In chapter I, I used biochemical approaches to characterize the residues required for the enzymatic activity of HopZ1a. I identified a threonine residue T346 as the autoacetylation site of HopZ1a. I also found two serine residues S349 and S351 required for the avirulence and virulence function of HopZ1a, through their contributions to the acetyltransferase activity. The acetyltransferase activity of HopZ1a can be activated by the eukaryotic cofactor inositol hexakisphosphate (IP6). My experiments showed that S349 and S351, together with a previously identified lysine residue K289, were essential to mediate IP6-induced conformational change of HopZ1a. The conservation of S349 and S351 and the requirement of them in the acetyltransferase activity of other YopJ family effectors suggest that additional residues in addition to the indispensable catalytic triad are required for the biological function of HopZ1a.

In chapter II, I used traditional genetics and molecular biology approaches to study the function of an interacting partner of HopZ1a in soybean called GmZINP2. By using transgenic soybeans over-expressing GmZINP2, I showed that GmZINIP2 is a negative regulator of plant immunity and suppresses early and late defense responses. GmZINP2 contains domains that are likely associated with chromatin remodeling functions. I showed that GmZINP2 could bind DNA and also interacts with a transcription factor GmMYB56 in soybean. Interestingly, HopZ1a could acetylate GmZINP2 and affect the interaction between GmZINP2 and GmMYB56. As such, HopZ1a may manipulate soybean defense through its interaction with GmZINP2.

This thesis represents a comprehensive study on the enzymatic activity and virulence functions of an important class of type III effectors. The study on GmZINP2 revealed possible manipulation of host defense by bacterial pathogens through chromatin remodeling process, which is a novel strategy utilized by bacterial pathogens to enhance virulence.

    Cover page: Biochemical and Functional Characterization of the Pseudomonas syringae Effector HopZ1a
    • Article
    • Peer Reviewed
    UC Riverside Previously Published Works (2016)
    Plants are constantly threatened by potential pathogens. In order to optimize the output of defense against pathogens with distinct lifestyles, plants depend on hormonal networks to fine-tune specific responses and regulate growth-defense tradeoffs. To counteract, pathogens have evolved various strategies to disturb hormonal homeostasis and facilitate infection. Many pathogens synthesize plant hormones; more importantly, toxins and effectors are produced to manipulate hormonal crosstalk. Accumulating evidence has shown that pathogens exert extensive effects on plant hormone pathways not only to defeat immunity, but also modify habitat structure, optimize nutrient acquisition, and facilitate pathogen dissemination. In this review, we summarize mechanisms by which a wide array of pathogens gain benefits from manipulating plant hormone pathways.
      Cover page: Phytohormone pathways as targets of pathogens to facilitate infection
      • Article
      • Peer Reviewed
      UC Riverside Previously Published Works (2017)
      The Yersinia outer protein J (YopJ) family of bacterial effectors depends on a novel acetyltransferase domain to acetylate signalling proteins from plant and animal hosts. However, the underlying mechanism is unclear. Here, we report the crystal structures of PopP2, a YopJ effector produced by the plant pathogen Ralstonia solanacearum, in complex with inositol hexaphosphate (InsP6), acetyl-coenzyme A (AcCoA) and/or substrate Resistance to Ralstonia solanacearum 1 (RRS1-R)WRKY. PopP2 recognizes the WRKYGQK motif of RRS1-RWRKY to position a targeted lysine in the active site for acetylation. Importantly, the PopP2-RRS1-RWRKY association is allosterically regulated by InsP6 binding, suggesting a previously unidentified role of the eukaryote-specific cofactor in substrate interaction. Furthermore, we provide evidence for the reaction intermediate of PopP2-mediated acetylation, an acetyl-cysteine covalent adduct, lending direct support to the 'ping-pong'-like catalytic mechanism proposed for YopJ effectors. Our study provides critical mechanistic insights into the virulence activity of YopJ class of acetyltransferases.
        Cover page: Mechanism of host substrate acetylation by a YopJ family effector
        • Article
        • Peer Reviewed
        UC Riverside Previously Published Works (2015)
        Gram-negative bacteria inject type III secreted effectors (T3SEs) into host cells to manipulate the immune response. The YopJ family effector HopZ1a produced by the plant pathogen Pseudomonas syringae possesses acetyltransferase activity and acetylates plant proteins to facilitate infection. Using mass spectrometry, we identified a threonine residue, T346, as the main autoacetylation site of HopZ1a. Two neighboring serine residues, S349 and S351, are required for the acetyltransferase activity of HopZ1a in vitro and are indispensable for the virulence function of HopZ1a in Arabidopsis thaliana. Using proton nuclear magnetic resonance (NMR), we observed a conformational change of HopZ1a in the presence of inositol hexakisphosphate (IP6), which acts as a eukaryotic co-factor and significantly enhances the acetyltransferase activity of several YopJ family effectors. S349 and S351 are required for IP6-binding-mediated conformational change of HopZ1a. S349 and S351 are located in a conserved region in the C-terminal domain of YopJ family effectors. Mutations of the corresponding serine(s) in two other effectors, HopZ3 of P. syringae and PopP2 of Ralstonia solanacerum, also abolished their acetyltransferase activity. These results suggest that, in addition to the highly conserved catalytic residues, YopJ family effectors also require conserved serine(s) in the C-terminal domain for their enzymatic activity.
          Cover page: Two serine residues in Pseudomonas syringae effector HopZ1a are required for acetyltransferase activity and association with the host co-factor.
          • Article
          • Peer Reviewed
          UC Riverside Previously Published Works (2013)
          Gram-negative bacterial pathogens deliver a variety of virulence proteins through the type III secretion system (T3SS) directly into the host cytoplasm. These type III secreted effectors (T3SEs) play an essential role in bacterial infection, mainly by targeting host immunity. However, the molecular basis of their functionalities remains largely enigmatic. Here, we show that the Pseudomonas syringae T3SE HopZ1a, a member of the widely distributed YopJ effector family, directly interacts with jasmonate ZIM-domain (JAZ) proteins through the conserved Jas domain in plant hosts. JAZs are transcription repressors of jasmonate (JA)-responsive genes and major components of the jasmonate receptor complex. Upon interaction, JAZs can be acetylated by HopZ1a through a putative acetyltransferase activity. Importantly, P. syringae producing the wild-type, but not a catalytic mutant of HopZ1a, promotes the degradation of HopZ1-interacting JAZs and activates JA signaling during bacterial infection. Furthermore, HopZ1a could partially rescue the virulence defect of a P. syringae mutant that lacks the production of coronatine, a JA-mimicking phytotoxin produced by a few P. syringae strains. These results highlight a novel example by which a bacterial effector directly manipulates the core regulators of phytohormone signaling to facilitate infection. The targeting of JAZ repressors by both coronatine toxin and HopZ1 effector suggests that the JA receptor complex is potentially a major hub of host targets for bacterial pathogens.
            Cover page: Bacterial Effector Activates Jasmonate Signaling by Directly Targeting JAZ Transcriptional Repressors
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