coli acts as a negative regulator of the cadBA operon in the absence of exogenous lysine (Neely et al., 1994; Neely & Olson, 1996). A recent study has also shown that E. coli http://www.selleckchem.com/products/erastin.html CadC is inactivated through an interaction with the lysine permease LysP in the absence of exogenous lysine (Tetsch et al., 2008). However, whether LysP functions similarly in Salmonella has not been determined. Prediction of the transmembrane segments using the DAS program (Stockholm University, Sweden) suggests that S. Typhimurium LysP is a multiple membrane-spanning protein (data not shown). To determine whether LysP inhibits the induction of cadBA transcription in S. Typhimurium,

we compared the expression of a chromosomal cadA–lacZ fusion in the JF3068 (wild-type) and YK5006 (ΔlysP mutant) strains using β-galactosidase assays. Figure 4(a) shows that the YK5006 strain expresses a cadA–lacZ transcriptional fusion, even in the absence of exogenous lysine, indicating that a mutation in the lysP gene confers lysine-independent cadBA transcription. Although the lysine signal is not directly involved in the proteolytic processing

of CadC, it is essential for expression of the S. Typhimurium cadBA operon (Fig. 3). To test the effect of the lysine signal on the transcriptional activity of lysP, RT-PCR analysis was conducted on total RNA isolated from UK1 wild-type cells collected at different intervals following the addition of 10 mM lysine. As shown in Fig. 4(b), expression of selleck chemical lysP mRNA was significantly reduced after lysine addition. To further confirm this observation, immunoblot analysis was conducted on the total protein extracts prepared from the ΔlysP strain harboring pACYC184-LysP-HA. C-terminally HA-tagged LysP (LysP-HA) was expressed under the control of its own promoter. Figure 4(b) shows that the cellular level of LysP-HA decreases

rapidly after lysine addition. These results suggest that the lysine signal represses lysP expression, click here thereby eliminating the negative regulation of CadC activation by LysP. In the present study, a genome-wide search revealed a PTS permease STM4538 as a novel component of CadC signaling in S. Typhimurium (Fig. 1). In particular, we demonstrated that inactivation of STM4538 impaired the proteolytic processing of CadC (Fig. 2). Although it is now clear that STM4538 acts as a positive modulator of CadC activity, questions still remain regarding how this PTS permease affects the proteolytic processing of CadC. One likely explanation is that the PTS permease STM4538 might exert its effects either directly or indirectly by controlling the expression of a gene that encodes a CadC-specific protease. It has been recently demonstrated that bacterial enzymes can also act as regulatory proteins.