CcpC-dependent regulation of citB and lmo0847 in Listeria monocytogenes

In Bacillus subtilis, the catabolite control protein C (CcpC) plays a critical role in regulating the genes encoding the enzymes of the tricarboxylic acid branch of the Krebs citric acid cycle. A gene encoding a potential CcpC homolog and two potential target genes were identified in the Listeria monocytogenes genome. In vitro gel mobility shift assays and DNase I footprinting experiments showed that L. monocytogenes CcpC (CcpC(Lm)) interacts with the promoter regions of citB(Lm) (the gene that is likely to encode aconitase) and lmo0847 (encoding a possible glutamine transporter) and that citrate is a specific inhibitor of this interaction. To study in vivo promoter activity, a new lacZ reporter system was developed. This system allows stable integration into the chromosome of a promoter region transcriptionally fused to a promoterless lacZ gene at a nonessential, ectopic locus. Analysis of strains carrying a citB(Lm)-lacZ or lmo0847-lacZ fusion revealed that CcpC(Lm) represses citB(Lm) and lmo0847 in media containing an excess of glucose and glutamine. In addition, regulation of citB(Lm) expression in rich medium was growth phase dependent; during exponential growth phase, expression was very low even in the absence of CcpC(Lm), but a higher level of citB(Lm) expression was induced in stationary phase, suggesting the involvement of another, as yet unidentified regulatory factor.