Manipulation of noncanonical Ire1-dependent MAPK signaling by a Vibrio agonist-antagonist effector pair

Abstract: Many bacterial pathogens employ effector delivery systems to disrupt vital cellular processes in the host. The type III secretion system 1 of the marine pathogen, Vibrio parahaemolyticus, utilizes the sequential action of four effectors to induce a rapid, pro-inflammatory cell death uniquely characterized by a pro-survival host transcriptional response. Herein we show that this pro-survival response is due to the action of the channel-forming effector VopQ that disrupts autophagic flux through two activities: lysosomal deacidification and inhibition of lysosome-autophagosome fusion. Using a point mutation to separate these two functions, we demonstrate that VopQ acts as an agonist of host ERK1/2 MAPK signaling as a result of its fusion-blocking activity. The pulse of ERK1/2 phosphorylation is restricted to early infection by the timed inhibition of Rho GTPase signaling by the antagonist effector VopS. Inhibition of autophagosome-lysosome fusion and activation of MAPK kinase signaling can be linked to the UPR. VopQ not only activates the IRE1 branch of the UPR, but VopQ’s activation of ERK1/2 MAPK signaling is dependent on IRE1. Since VopS can dampen VopQ-induced IRE1-dependent ERK1/2 activation, we suggest that IRE1 must activate ERK1/2 signaling at or above the level of Rho GTPases. This work elucidates new connections between host autophagy, MAPK signaling and the UPR that bacterial pathogens have evolved to manipulate the host. Methods: We employed genome-wide transcriptional profiling methods (RNA-seq) to measure changes in gene expression in response to infection with V. para T3SS1+, T3SS1+ ΔvopQ, or T3SS1+ ΔvopS. Time points were collected 90 minutes post-infection. In total, libraries from 12 samples (triplicates of uninfected, T3SS1+, T3SS1+ ΔvopQ, and T3SS1+ ΔvopS) were sequenced and mapped to the human genome. EdgeR was used for differential expression analysis using statistical cutoffs of of false discovery rate (FDR) ≤0.01, log2counts-per-million (log2CPM) ≥0 and fold change (FC) cutoffs of -1.5≥ FC ≥1.5 Overall 12 samples were analyzed. Each of the 4 conditions was performed in triplicate. There are two sets of reference groups. Group one is uninfected cells and Group 2 is V. para strain T3SS1+-infected cells, because we were trying to understand the individual effects of the effectors vopQ and vopS on host gene expression.