Collaborative Regulation ofEscherichia coliGlutamate-Dependent Acid Resistance by Two AraC-Like Regulators, GadX and GadW (YhiW)

Abstract

An important feature of Escherichia coli pathogenesis is an ability to withstand extremely acidic environments of pH 2 or lower. This acid resistance property contributes to the low infectious dose of pathogenic E. coli species. One very efficient E. coli acid resistance system encompasses two isoforms of glutamate decarboxylase (gadA and gadB) and a putative glutamate:␥-amino butyric acid (GABA) antiporter (gadC). The system is subject to complex controls that vary with growth media, growth phase, and growth pH. Previous work has revealed that the system is controlled by two sigma factors, two negative regulators (cyclic AMP receptor protein [CRP] and H-NS), and an AraC-like regulator called GadX. Earlier evidence suggested that the GadX protein acts both as a positive and negative regulator of the gadA and gadBC genes depending on environmental conditions. New data clarify this finding, revealing a collaborative regulation between GadX and another AraC-like regulator called GadW (previously YhiW). GadX and GadW are DNA binding proteins that form homodimers in vivo and are 42% homologous to each other. GadX activates expression of gadA and gadBC at any pH, while GadW inhibits GadX-dependent activation. Regulation of gadA and gadBC by either regulator requires an upstream, 20-bp GAD box sequence. Northern blot analysis further indicates that GadW represses expression of gadX. The results suggest a control circuit whereby GadW interacts with both the gadA and gadX promoters. GadW clearly represses gadX and, in situations where GadX is missing, activates gadA and gadBC. GadX, however, activates only gadA and gadBC expression. CRP also represses gadX expression. It does this primarily by repressing production of sigma S, the sigma factor responsible for gadX expression. In fact, the acid induction of gadA and gadBC observed when rich-medium cultures enter stationary phase corresponds to the acid induction of sigma S production. These complex control circuits impose tight rein over expression of the gadA and gadBC system yet provide flexibility for inducing acid resistance under many conditions that presage acid stress.

GadE (YhiE): a novel activator involved in the response to acid environment in Escherichia coli

“…hdeD, which encodes an integral membrane protein of unknown function, is located immediately downstream of hdeAB and was also induced by overexpression of both gadX and gadE (Table 3). Gel shift experiments demonstrated that both GadX (Ma et al, 2002) and GadE bind to the promoter region of these genes, suggesting that they play a direct role in their activation. Moreover, several genes encoding putative chaperones and proteases were also induced by overexpression of gadX or gadE, e.g.…”

“…Using genome-wide technologies such as transcriptome analysis (Velculescu et al, 1997), GadX was identified as a regulator of the glutamate decarboxylase genes from the AraC family in E. coli Ma et al, 2002). As for ToxR from Vibrio cholerae, which is an acidinduced factor and a virulence regulator, GadX has been shown to be involved in the appropriate expression of genes required for virulence of enteropathogenic E. coli (Shin et al, 2001).…”

The glutamate-dependent acid resistance system in Escherichia coli: essential and dual role of the His–Asp phosphorelay RcsCDB/AF

“…A 25 % reduction of gadWlacZ fusion expression was observed in the rcsB strain, leaving the possibility that RcsB might activate gadAB expression through gadW. This is however unlikely, as GadW was reported to be a negative regulator of gadAB expression (Ma et al, 2002Tucker et al, 2003). In addition, a gadW mutant was reported to resist acid as well as a wild-type strain .…”

“…3(a). The gadA, gadB, gadX and gadE fusions were described by and Ma et al (2002Ma et al ( , 2003; the limits for gadA and gadB are shown in Fig. 3(a).…”

“…Highlighting the importance of the AR2 system is the plethora of regulators involved in its regulation, the relative importance of each regulator depending on environmental pH and growth phase. These regulators include: two phosphorelays, EvgAS (Masuda & Church, 2002) and TorRS (Bordi et al, 2003), the stationary phase s s -factor (De Biase et al, 1999), the global regulators Crp ) and H-NS (Hommais et al, 2001), three AraC-like regulators, GadW (Ma et al, 2002), GadX (Shin et al, 2001) and YdeO (Masuda & Church, 2003), and the Era-like GTPase TrmE (Gong et al, 2004). Central to the AR2 system is the activity of the LuxRlike transcription regulator GadE, as most of the regulators listed target gadE expression.…”