Identification of host components targeted by the type III effector PopP2 and involved in an R/Avr complex – PERCEPtome

Effector-triggered immunity (ETI) is elicited by host plant resistance (R) proteins upon recognition of a cognate microbial effector (avirulence (Avr) factor) or its action. A growing body of evidence indicates that pathogen perception involves dynamic associations of multiple host proteins, especially in the plant nucleus, a battlefield of plant immunity. Despite intensive research efforts, the components of such multiprotein complexes or “Resistasomes” remain obscure. Resistasome involved in Arabidopsis thaliana immunity to the root bacterial pathogen R. solanacearum requires at least three partners: (i) the RRS1-R resistance protein, (ii) PopP2, its cognate avirulence partner, and (iii) RD19, a vacuolar cysteine protease playing a key role in the RRS1-R-triggered immunity. RRS1-R is an atypical resistance protein that combines classical resistance protein domains (TIR-NBS-LRR) with a WRKY DNA binding domain thereby representing a «ready-to-act» transcription machinery-associated receptor. Together with RRS1-R, RD19 is the second PopP2-interacting partner (PIP) whose nuclear relocalization is triggered by PopP2. Activation of a pre-immune RRS1-R/PopP2 perception complex might require other host components. Several Arabidopsis candidate proteins (PIPs), not previously described as participating in resistance signalling, were identified in a yeast two-hybrid screen. They might correspond to (i) PopP2-targets guarded by the RRS1-R protein or (ii) host components involved in the elaboration of an active RRS1-R/PopP2 perception complex.
By taking some interdisciplinary approaches, one of the main goals of this proposal is to make significant progress in the validation and the functional characterization of these PIP proteins. We will investigate their respective involvement in the formation of a Resistasome perception complex together with PopP2, RRS1-R and RD19, and evaluate their role in the RRS1-R-mediated immunity.
The second exciting aspect of this proposal concerns the characterization of the acetyltransferase activity of PopP2. Among the PIP candidates, we identified PIP3 as being acetylated by PopP2. With the exception of histone modification and DNA repair, in which acetylation has a well established role, impact of acetylation in other nuclear processes that are involved in plant immunity remains uncovered. To date, PIP3 is the first example of a host component acetylated by a plant bacterial effector. Indeed, PopP2 constitutes a powerful tool for the study of the role of this post-translational modification in the regulation of plant innate immunity signalling. PopP2 acetyltransferase activity on its interacting partners will be investigated in vitro and in vivo thanks to the use of different molecular approaches. This study will be extended to the mapping of the acetylated residue(s) of PopP2 substrate(s) and the role of acetylation in the RRS1-R-mediated resistance. In addition, an Acetyllysine subproteome survey aimed at the identification of Arabidopsis proteins acetylated by PopP2 will be performed.
This ambitious proposal should yield important insights into the elucidation of yet-unknown strategies developed either by pathogens to manipulate plant immunity signalling or by host plants to countermeasure their actions.