Pathogen suppression of host defense : weapons and mechanisms of XopX
Abstract/Contents
- Abstract
- One of the mechanisms phytopathogenic bacteria use to ensure successful infection is to inject proteins called effectors into plant cells using a type III secretion system. The study of effector biology over the past 30 years has revealed that most of these proteins function as virulence factors by perturbing the function of key components of the plant immune system. Xanthomonas euvesicatoria (Xe), the causal agent of Bacterial Spot Disease on tomato and pepper, utilizes approximately 35 effectors to facilitate its colonization of leaf tissue and transmission. Several of these effectors are required for maximal Xe growth and symptom development in susceptible leaves; however, little is known about their unique biochemical activities in planta. One Xe effector whose function has remained elusive is XopX. Homologs of XopX can be found in all sequenced Xanthomonas strains. Such conservation suggests that XopX likely plays an essential role in Xanthomonas pathogenesis. XopX is known to suppress specific aspects of plant immunity, including the production of reactive oxygen species (ROS) generation in response to bacterial flagellin, and the activation of the hypersensitive response (HR) triggered by disease resistance receptors. XopX perturbation within the plant cell also leads to the activation of plant defense responses including defense gene transcription, ethylene production and eventually the induction of plant cell death. How XopX suppresses plant immunity while activating cell death was unclear. In my thesis work, I have used an interdisciplinary approach to elucidate the biochemical activity XopX during Xe infection. Using mutational analyses and physiological assays, I identified a key domain in XopX (amino acids 202-211) that is necessary for XopX to suppress flagellin induced ROS production and activate plant cell death. I then determined that XopX localizes to two cellular compartments in plant cells -- the cytoplasm and chloroplasts using microscopy and subcellular fractionation assays. I purified XopX protein complexes from plant cells using immunoprecipitation and determined their identities by protein mass spectrometry. I then used a genetic approach to determine if a subset of the interacting proteins identified are required for XopX-dependent phenotypes. I discovered that the plant protein NAD kinase 1 (NADK1) was necessary for full XopX ROS suppression and cell death activation, establishing a genetic link between NADK1 and XopX. Additionally, I found that co-overexpression of XopX with NADK1 strengthened the cell death phenotype in a way that was dependent on NADK1 being an active enzyme. Using an in vitro enzyme assay, I was able to demonstrate that NADK1 is a functional kinase that phosphorylates NAD to produce NADP. XopX however does not alter NADK1's activity in vitro, suggesting that the mechanism behind XopX virulence phenotypes is not the direct inhibition of NADK1 enzyme activity. Notably, I found that XopX alone has weak NAD kinase activity, and this activity is dependent on the 202-211 amino acid domain. Overall, my thesis work uncovers several new characteristics of XopX to help explain its biological function in Xe pathogenesis. It also highlights the importance of NADK1 in plant immune responses and suggests that NADK1 is a target of XopX. My findings further support emerging research that the nucleotides NAD and NADP are key metabolites that regulate plant immune responses and cellular homeostasis.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2021; ©2021 |
| Publication date | 2021; 2021 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Duncan, Anne Elizabeth |
|---|---|
| Degree supervisor | Mudgett, Mary Beth, 1967- |
| Thesis advisor | Mudgett, Mary Beth, 1967- |
| Thesis advisor | Dixon, Scott |
| Thesis advisor | Long, Sharon R |
| Degree committee member | Dixon, Scott |
| Degree committee member | Long, Sharon R |
| Associated with | Stanford University, Department of Biology |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Anne E. Duncan. |
|---|---|
| Note | Submitted to the Department of Biology. |
| Thesis | Thesis Ph.D. Stanford University 2021. |
| Location | https://purl.stanford.edu/qd054zw0429 |
Access conditions
- Copyright
- © 2021 by Anne Elizabeth Duncan
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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