Increased transcription of a potential sigma factor regulatory gene Rv1364c in Mycobacterium bovis BCG while residing in macrophages indicates use of alternative promoters.

ABSTRACT

Alternative sigma factors are key global regulators that coordinate bacterial responses to environmental changes necessary for adaptation and survival. In turn these sigma factors are controlled by regulators such as anti-sigma and anti-anti-sigma factors. In this report, using a cDNA-total RNA subtractive hybridisation strategy that we have developed previously, we identified increased transcription of a potential sigma factor regulatory gene, Rv1364c, in Mycobacterium bovis BCG upon phagocytosis by macrophages and this was confirmed by Northern blot analysis. Primer extension analysis revealed the use of alternative promotors, P1 and P2, and that the increased expression inside macrophages coincided with promoter switching from P2 to P1. Rv1364c (653 amino acids), originally annotated as RsbU, contains structural domains homologous to the PAS redox sensor, the protein phosphatases anti-anti-sigma factor RsbU/SpoIIE, the protein kinase anti-sigma factor RsbW/SpoIIAB and the anti-anti-sigma factor RsbV/SpoIIAA found in other bacteria. These findings have important implications for understanding coordination of the expression of sigma factors under intra-macrophage conditions. Other potentially differentially expressed genes, including genes for fatty acid metabolism, membrane transportors, heat shock proteins, potential sigma factors and energy metabolic pathways are also listed and their biological significance discussed.