Global Regulator MorA Affects Virulence-Associated Protease Secretion in Pseudomonas aeruginosa PAO1

Abstract

Bacterial invasion plays a critical role in the establishment of Pseudomonas aeruginosa infection and is aided by two major virulence factors – surface appendages and secreted proteases. The second messenger cyclic diguanylate (c-di-GMP) is known to affect bacterial attachment to surfaces, biofilm formation and related virulence phenomena. Here we report that MorA, a global regulator with GGDEF and EAL domains that was previously reported to affect virulence factors, negatively regulates protease secretion via the type II secretion system (T2SS) in P. aeruginosa PAO1. Infection assays with mutant strains carrying gene deletion and domain mutants show that host cell invasion is dependent on the active domain function of MorA. Further investigations suggest that the MorA-mediated c-di-GMP signaling affects protease secretion largely at a post-translational level. We thus report c-di-GMP second messenger system as a novel regulator of T2SS function in P. aeruginosa. Given that T2SS is a central and constitutive pump, and the secreted proteases are involved in interactions with the microbial surroundings, our data broadens the significance of c-di-GMP signaling in P. aeruginosa pathogenesis and ecological fitness.

A Genome-Wide Screen Identifies Genes in Rhizosphere-Associated Pseudomonas Required to Evade Plant Defenses

“…The WCS365_05664 gene product is similar to the previously characterized P. aeruginosa PAO1 protein MorA, a known diguanylate cyclase/phosphodiesterase (68% identity, 99% coverage) ( 51 , 52 ).…”

Flagellin FliC Phosphorylation Affects Type 2 Protease Secretion and Biofilm Dispersal in Pseudomonas aeruginosa PAO1

“…We investigated the effect of loss of FliC phosphorylation on the type 2 secretome profile of PAO1 WT, Δ fliC , Δ fliC -FL T27A, Δ fliC -FL and Δ fliC -FL S28A strains. We have previously profiled the type 2 secretome of PAO1 [ 34 ]. The secretome profile showed the presence of proteases such as elastases LasA & LasB, alkaline metalloproteinase AprA, chitin binding protein CbpD, endoprotease PrpL and PA0572, a zinc metallopeptidase-like protein in all five strains, consistent with previous observation.…”

“…The precipitated proteins were pelleted and washed twice with ice-cold acetone each time to remove salts and then air-dried. The solubilisation buffer was optimised to a denaturing buffer containing 40mM Tris, 40mM Dithiothreitol (DTT) and 2% SDS [ 34 ]. Densitometry analysis of protein bands from the various lanes were performed using the image analysis tool ImageJ 1.43 ( http://rsbweb.nih.gov/ij/ ) developed by Wayne Rasband, National Institutes of Health, Bethesda, MD.…”

CryoEM map ofPseudomonas aeruginosaPilQ enables structural characterization of TsaP

“…Reduced extracellular protease secretion, decreased twitching motility zones, and increased biofilm formation are all associated with cyclic di-GMP induction 4,[38][39][40] . Measurement of luciferase expression from a cyclic di-GMP sensitive promotor verified that TsaP overexpression increases cellular cyclic di-GMP concentrations ( Figure 3C).…”