σB contributes to Listeria monocytogenes invasion by controlling expression of inlA and inlB

The ability of Listeria monocytogenes to invade non-phagocytic cells is important for development of a systemic listeriosis infection. The authors previously reported that a L. monocytogenes Delta sigB strain is defective in invasion into human intestinal epithelial cells, in part, due to decreased expression of a major invasion gene, inIA. To characterize additional invasion mechanisms under the control of sigma(B), mutants were generated carrying combinations of in-frame deletions in inIA, inlB and sigB. Quantitative assessment of bacterial invasion into the human enterocyte Caco-2 and hepatocyte HepG-2 cell lines demonstrated that sigma(B) contributes to both InIA and InIB-mediated invasion of L. monocytogenes. Previous identification of the sigma(B)-dependent P2(prfA) promoter upstream of the major virulence gene regulator, positive regulatory factor A (PrfA), suggested that the contributions of sigma B to expression of various virulence genes, including inIA, could be at least partially mediated through PrfA. To test this hypothesis, relative invasion capabilities of Delta sigB and Delta prfA strains were compared. Exponential-phase cells of the Delta sigB and Delta prfA strains were similarly defective at invasion; however, stationary-phase Delta sigB cells were significantly less invasive than stationary-phase Delta prfA cells, suggesting that the contributions of a B to invasion extend beyond those mediated through PrfA in stationary-phase L. monocytogenes. TaqMan quantitative reverse-transcriptase PCRs further demonstrated that expression of in/A and inlB was greatly increased in a sigma(B)-dependent manner in stationary-phase L. monocytogenes. Together, results from this study provide strong biological evidence of a critical role for sigma(B) in L. monocytogenes invasion into non-phagocytic cells, primarily mediated through control of inIA and inlB expression.