Differential expression of Listeria monocytogenes virulence genes in mammalian host cells

We have used RT-PCR and GFP-mediated fluorescence to analyse the regulation of PrfA-dependent virulence genes of Listeria monocytogenes during prolife ration in mammalian host cells. Our data show that most of the PrfA-regulat ed virulence genes are more efficiently expressed, as measured by transcrip t levels. when L. monocytogenes is grown in macrophages and macrophage-like cells rather than in epithelial cells, hepatocytes or endothelial cells. T he promoters for hly and plcA are predominantly activated within the phagos omal compartment, while those for uctA and inlC are predominantly activated in the host cell cytosol. Expression of actA and plcB precedes that of inl C after infection of epithelial cells and macrophages. Little transcription of inlA or inlB is observed in epithelial cells and there is only slightly more in macrophages. Tn both cell types the level of transcription of the inlAB operon is lower than is seen under extracellular growth conditions in rich media, which is compatible with the assumption that InlA and InlB are not required during intracellular growth of the bacteria. Activation of th e PrfA-independent lap promoter is also low during intracellular growth, al though the gene product (p60) is required for cell viability. The levels of the PrfA-dependent virulence gene transcripts do not correlate with the am ount of prfA transcript present, which is low under all intracellular condi tions analysed, suggesting that the prfA transcript is either highly unstab le in bacteria that are growing intracellularly, or that the small amount o f PrfA produced is highly activated by additional component(s).