Defective Lipoprotein Sorting Induces lolA Expression through the Rcs Stress Response Phosphorelay System

Abstract

cThe Escherichia coli LolA protein is a lipoprotein-specific chaperone that carries lipoproteins from the inner to the outer membrane. A dominant negative LolA mutant, LolA(I93C/F140C), in which both 93 Ile and 140 Phe are replaced by Cys, binds tightly to the lipoprotein-dedicated ABC transporter LolCDE complex on the inner membrane and therefore inhibits the detachment of outer membrane-specific lipoproteins from the inner membrane. We found that the expression of this mutant strongly induced lolA gene transcription. The depletion of the LolA or LolB protein also triggered lolA gene transcription, indicating that the inhibition of outer membrane lipoprotein transport triggers lolA transcription. To elucidate the mechanism, we isolated mutants that are unable to induce lolA transcription using the lacZ gene fused to the lolA promoter as a reporter and found that the Rcs phosphorelay system directly regulates lolA transcription. An outer membrane lipoprotein, RcsF, was essential for this activation, while the coactivator RcsA was dispensable. Taking the observation that an RcsF mutant localized in the inner membrane constitutively activated the Rcs phosphorelay system into consideration, the results shown here strongly suggest that correct lipoprotein sorting to the outer membrane is monitored by RcsF, which plays a key role in the Rcs stress response system.

Multiple Transcriptional Factors Regulate Transcription of the rpoE Gene in Escherichia coli under Different Growth Conditions and When the Lipopolysaccharide Biosynthesis Is Defective

“…Mislocalization of lipoproteins has been shown to cause induction of the Rcs pathway (53). Consistent with the positive regulation of the rpoEP3 promoter by the Rcs system, its activation by overexpression of the lipoproteins is also Rcsdependent.…”

“…We reasoned that the increased synthesis of lipoprotein could cause limitation of the Lol system required for sorting of lipoproteins. Defective lipoprotein is known to induce the Rcs pathway (53). Thus, we tested if the mechanism of activation of the rpoEP3 promoter was also Rcs-dependent.…”

The role for TolA in enterohemorrhagic Escherichia coli pathogenesis and virulence gene transcription

“…While the mechanistic basis of RcsF signal transduction to the sensor kinase RcsC has not yet been determined, it has been shown that both perturbations to the outer-membrane and sensing of mislocalization of outer-membrane lipoproteins stimulate RcsF signaling to RcsC [38,39]. Thus, activation of the LEE in TW14359DtolA may be due to RcsF sensing of pleiotropic cellular effects following loss of TolA.…”

“…Signal transduction to RcsC and Rcs phosphorelay activation typically requires the action of an additional factor, RcsF, which is a~14-kDa outer-membrane bound lipoprotein that orients towards the periplasm [37]. RcsF signaling has been demonstrated for mutations that affect the proper folding of periplasmic proteins, as well as those which disrupt the proper synthesis and localization of lipopolysaccharides and lipoproteins [38,39]. Activity is dependent on both the formation of nonconsecutive disulfides and an unstructured proline rich region to properly convey signals to the sensor kinase RcsC for Rcs phosphorelay activation [40,41].…”

A penicillin-binding protein inhibits selection of colistin-resistant, lipooligosaccharide-deficient Acinetobacter baumannii

“…Our analysis demonstrated that increased transcription of lipoprotein transporters and lipoproteins in LOSdeficient A. baumannii resulted in outer-membrane remodeling that could promote A. baumannii survival without lipid A/LOS. Regulation of lolA in E. coli is controlled by the Rcs phosphorelay system, which detects cell-envelope stress (33). However, A. baumannii does not encode an orthologous system, and regulatory mechanisms controlling lolA gene expression in A. baumannii are not known.…”