Because of their cytosolic localization, stimuli corresponding to variations in central metabolites are thought to affect the expression of CpxR targets in a CpxA-independent way (Strozen et al., 2005; Wolfe et al., 2008; Kinnersley et al., 2009; Lima et al., Belnacasan 2011). Decreased cAMP levels (Strozen et al., 2005), glucose (Kinnersley

et al., 2009) and intermediates of the acetyl-CoA pathway (Wolfe et al., 2008; Lima et al., 2011) induce the expression of degP and cpxP, respectively. For intermediates of the acetyl-CoA pathway, two mechanisms exist: acetyl phosphate is known to act as a direct phosphor donor for CpxR in vitro (Raivio & Silhavy, 1997) and in vivo (Klein et al., 2007; Groban et al., 2009), and acetyl-CoA promotes the acetylation of RNA polymerase, which is critical for the glucose-dependent induction of cpxP transcription (Lima et al., 2011). In contrast to cytosolic stimuli, Ku0059436 phosphatidylethanolamine depletion, indole, alcohols, acetone and phenethyl alcohol are likely sensed by the TMD of CpxA (Mileykovskaya & Dowhan, 1997; Garbe et al., 2000; Rutherford et al., 2010;

Clarke & Voigt, 2011). All these stimuli are proposed to modulate the physical properties of the inner membrane (Dombek & Ingram, 1984) and result in conformational changes within the membrane helices of CpxA (Anbazhagan et al., 2010). For phosphatidylethanolamine depletion, two specific mechanisms that result in the activation of CpxA are also conceivable: (1) direct influence IKBKE by lipids and (2) indirect effects through alteration of a cell envelope component that is modified in a phosphatidylethanolamine-dependent manner such as LPS (Mileykovskaya & Dowhan, 1997). Alternatively, all these stimuli

might influence CpxA in an indirect way by inducing misfolding of inner membrane proteins (Shimohata et al., 2002, 2007; Akiyama, 2009). Another Cpx-inducing signal that modulates the physical properties of the outer membrane is the attachment to hydrophobic surfaces (Otto & Silhavy, 2002). Surface attachment–induced Cpx activation depends on the outer membrane lipoprotein new lipoprotein E (NlpE; Otto & Silhavy, 2002), suggesting that NlpE might serve as a second accessory protein to deliver signalling information to CpxA. The metals zinc (Lee et al., 2005) and copper (Yamamoto & Ishihama, 2005) are excellent inducers of the Cpx system. Based on the presence of zinc in the CpxP crystal structure (Thede et al., 2011) and the observation that CpxP shares high homology with the metal sensor CnrX (Grass et al., 2000, 2005), it was suggested that CpxP might act as a zinc sensor (Thede et al., 2011). In contrast, it has been suggested that sensing of copper by the Cpx-TCS occurs via NlpE (also known as copper homeostasis protein CutF), because mutation of nlpE results in a decrease in copper tolerance (Gupta et al.