Escherichia coli SPFH membrane microdomain proteins HflKC contribute to aminoglycoside and oxidative stress tolerance

Many eukaryotic membrane-dependent functions are often spatially and temporally regulated by membrane microdomains (FMMs) also known as lipid rafts. These domains are enriched in polyisoprenoid lipids and scaffolding proteins belonging to the Stomatin, Prohibitin, Flotillin, and HflK/C (SPFH) protein superfamily that was also identified in Gram-positive bacteria. By contrast, little is still known about FMMs in Gram-negative bacteria. In Escherichia coli K12, 4 SPFH proteins, YqiK, QmcA, HflK, and HflC, were shown to localize in discrete polar or lateral inner-membrane locations, raising the possibility that E. coli SPFH proteins could contribute to the assembly of inner-membrane FMMs and the regulation of cellular processes. Here we studied the determinant of the localization of QmcA and HflC and showed that FMM-associated cardiolipin lipid biosynthesis is required for their native localization pattern. Using Biolog phenotypic arrays, we showed that a mutant lacking all SPFH genes displayed increased sensitivity to aminoglycosides and oxidative stress that is due to the absence of HflKC. Our study therefore provides further insights into the contribution of SPFH proteins to stress tolerance in E. coli. IMPORTANCEEukaryotic cells often segregate physiological processes in cholesterol-rich functional membrane micro-domains. These domains are also called lipid rafts and contain proteins of the Stomatin, Prohibitin, Flotillin, and HflK/C (SPFH) superfamily, which are also present in prokaryotes but were mostly studied in Gram-positive bacteria. Here, we showed that the cell localization of the SPFH proteins QmcA and HflKC in the Gram-negative bacteria E. coli is altered in absence of cardiolipin lipid synthesis. This suggests that cardiolipins contribute to E. coli membrane microdomain assembly. Using a broad phenotypic analysis, we also showed that HflKC contribute to E. coli tolerance to aminoglycosides and oxidative stress. Our study, therefore, provides new insights into the cellular processes associated with SPFH proteins in E. coli.