cAMP signaling in Mycobacterium tuberculosis

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DESCRIPTION (provided by applicant): The long term objective of this project is to better understand how Mycobacterium tuberculosis (Mtb) establishes infection so that effective strategies can be developed to prevent it. Gene regulation is a critical aspect of Mtb's successful response to the host environment during infection. The recent finding that Mtb encodes as many as 15 adenylate cyclases (AC) suggests that cAMP signaling constitutes an important, but previously unrecognized, regulatory mechanism in Mtb. A multidisciplinary approach will address the role of cAMP signaling in Mtb, at the molecular, genetic and cellular levels, with a focus on conditions associated with latency and Mtb's interaction with macrophages. Specific aims include: Aim 1: identify, using DMA microarrays and isogenic knockout mutants, Mtb genes that are regulated by the putative cAMP-dependent transcription factor, Rv1675c, at various levels of cAMP during hypoxia; Aim 2: define and characterize a second likely cAMP-dependent regulon controlled by putative transcription factor Rv3676 in Mtb. Candidate members of this regulon will be functionally assessed by defining the role of Rv3676 binding to a DMA motif identified in their promoter regions; Aim 3: evaluate the roles of cAMP and a mammalian-type AC, Rv1625c, in Mtb's interaction with macrophages, particularly with respect to bacterial replication and trafficking; Aim 4: assess the regulation and function of a specific macrophage-induced, cAMP regulated gene within macrophages and during latency-associated conditions. Identification and characterization of cAMP-mediated gene regulation in Mtb will contribute to our understanding of the factors needed for the establishment of tuberculosis infection and disease. This work will explore a new gene regulatory network in Mtb and provide an important foundation for future work on the role of cAMP signaling in Mtb virulence gene regulation, leading to identification of potential targets for TB vaccines, therapeutics, and/ or diagnostic purposes.