Investigating the roles and functions of plasma membrane E3 ligases and MKP1 in plant immunity
Authors:   , Ellingsen Jared Kenneth 
Published: December 2022

When a plant encounters a pathogen, a complex set of responses occurs on both sides of the interaction. In a process known as pattern-triggered immunity (PTI), the plant uses plasma membrane (PM)-localized proteins, or pattern recognition receptors (PRRs), to perceive the presence of a threat by recognition of pathogen- or microbe-associated molecular patterns (PAMPs/MAMPs), which are non-self molecules endogenous to the pathogen but not the plant. After perception, these PRRs initiate signaling events leading to canonical PTI responses such as activation of the MAP kinase cascade, production of reactive oxygen species (ROS) and regulation of defense-related transcripts. At the same time, the pathogen recognizes a potential host through plant-derived metabolites, which induces its type-III secretion system (T3SS) to inject effector proteins into the plant cell to make nutrient acquisition easier. Due to the effector proteins' key role in virulence, a goal of the plant during these early stages of the interaction is to restrict the delivery of effectors into the plant cell. Decreasing the abundance of T3SS-inducing metabolites from the extracellular space, essentially masking its presence from the pathogen, is sufficient to drastically reduce the number of effectors translocated into the cell, conferring resistance to the plant. Previous work in the lab established a knockout of MAP Kinase Phosphatase 1 (mkp1) as a model for studying this layer of PTI. mkp1 plants restrict effector delivery into the plant, resulting in greater resistance than in wild type plants. This resistance is due to a basal decrease in extracellular T3SS-inducing metabolites compared to wild type. A key question remains as to how the plant is able to modulate the accumulation of T3SS-inducing metabolites. The work in this dissertation establishes the PM-localized E3 ligase, ATL6, as a key positive regulator of PTI, playing an important role in PTI-associated resistance and effector delivery restriction, while not impacting other canonical PTI phenotypes. In addition, using an atl6 mkp1 double mutant, I establish that ATL6 is genetically linked to MKP1, suggesting a specific role in the regulation of T3SS-inducing metabolites and effector delivery restriction. I also identify another E3 ligase, ATL31, as a positive regulator of PTI, with atl31 knockouts showing phenotypes similar to atl6 plants. Together, these mutants define novel regulators of plant immune responses that are required for the resistance observed in mkp1 mutants. When effectors are translocated into the plant cell, they inhibit defense responses by targeting a spectrum of proteins involved in plant immunity. However, plants have evolved another layer of defense called effector-triggered immunity (ETI), which is activated when a class of intracellular receptors (R genes) recognize the presence or the activity of the effector. Recognition results in a more robust defense response, often leading to cell death. Due to the decrease of effectors being delivered to mkp1 plants, we hypothesized that mkp1 plants may be partially compromised in their ability to mount an ETI response because there are fewer effectors to trigger ETI. Surprisingly, this work shows the opposite result: mkp1 plants are more resistant to ETI-triggering bacterial strains, establishing MKP1 as a negative regulator of ETI. In addition, further work with higher order mutations shows that MKP1 acts in a distinct pathway during ETI compared to PTI. Thus, this work identifies a new role for MKP1 in ETI and suggests the existence of a previously undescribed pathway.