The dynamic hypoosmotic response ofVibrio choleraerelies on the mechanosensitive channel MscS

Abstract

Intestinal bacteria, including the facultative pathogen Vibrio cholerae, can adapt to a wide range of osmotic environments. In high-osmolarity media, bacteria accumulate small compatible metabolites to maintain turgor pressure, but under drastic osmotic down-shifts bacteria are able to avoid mechanical rupture by rapidly releasing these metabolites through mechanosensitive (MS) channels. Previous experiments on V. cholerae have identified two major types of MS channels - MscS and MscL. We functionally examine these channels specific roles in Vibrio osmotic rescuing via genetic modification, bacterial patch-clamp electrophysiology, and stopped-flow light scattering. The light scattering kinetics revealed that WT Vibrio cells were capable of releasing up to 10% of their total non-aqueous content within ~30 ms. To investigate each channels individual contribution to V. cholerae osmotic permeability response, we generated and characterized the first individual ∆mscS, ∆mscL, and double ∆mscL ∆mscS mutants in V. cholerae O395. Both mutants lacking MscS exhibited delayed osmolyte release kinetics and decreased osmotic survival rates compared to WT. Surprisingly, the ∆mscL mutant had comparable kinetics to WT, but a much higher osmotic survival, whereas ∆mscS had low survival, comparable to the double ∆mscL ∆mscS mutant. The data indicate that MscS is much more efficient in osmotic adjustment and is up-regulated in the absence of MscL, whereas in the absence of the low-threshold MscS, MscL even becomes toxic. Kinetic modeling of the cell swelling process and channel activation reveals the advantage of low-threshold MscS in curbing tension surges in Vibrio and its role in proper termination of the osmotic permeability response.