Characterization of a cAMP responsive transcription factor, Cmr (Rv1675c), in TB complex mycobacteria reveals overlap with the DosR (DevR) dormancy regulon

Abstract

Mycobacterium tuberculosis (Mtb) Cmr (Rv1675c) is a CRP/FNR family transcription factor known to be responsive to cAMP levels and during macrophage infections. However, Cmr's DNA binding properties, cellular targets and overall role in tuberculosis (TB) complex bacteria have not been characterized. In this study, we used experimental and computational approaches to characterize Cmr's DNA binding properties and identify a putative regulon. Cmr binds a 16-bp palindromic site that includes four highly conserved nucleotides that are required for DNA binding. A total of 368 binding sites, distributed in clusters among ∼200 binding regions throughout the Mycobacterium bovis BCG genome, were identified using ChIP-seq. One of the most enriched Cmr binding sites was located upstream of the cmr promoter, and we demonstrated that expression of cmr is autoregulated. cAMP affected Cmr binding at a subset of DNA loci in vivo and in vitro, including multiple sites adjacent to members of the DosR (DevR) dormancy regulon. Our findings of cooperative binding of Cmr to these DNA regions and the regulation by Cmr of the DosR-regulated virulence gene Rv2623 demonstrate the complexity of Cmr-mediated gene regulation and suggest a role for Cmr in the biology of persistent TB infection.

Cmr is a redox-responsive regulator of DosR that contributes to M. tuberculosis virulence

“…This suggests that Cmr-mediated gene regulation is complex and its influence over M. tuberculosis stress responses extends beyond de-repression of the DosR-regulon, as indicated by the ChIP-seq data reported by Ranganathan et al. for M. bovis BCG (13). Moreover, Cmr-mediated gene expression could be further complicated by its capacity to respond to both oxidation and nitrosation.…”

AbmR (Rv1265) is a novel transcription factor of Mycobacterium tuberculosis that regulates host cell association and expression of the non‐coding small RNA Mcr11

“…AbmR's putative operator contains a biochemically validated Cmr-binding site (Gazdik et al, 2009) and a predicted CRP Mt -binding site (Arnvig et al, 2011). While neither TF was found within the Rv1264-mcr11-abmR region in recent ChIP-seq studies (Kahramanoglou et al, 2014, Knapp et al, 2015Ranganathan et al, 2016), regulation of abmR by both Cmr and CRP Mt have been reported (Gazdik et al, 2009;Arnvig et al, 2011;Kahramanoglou et al, 2014;Knapp et al, 2015). The possibility that AbmR, Cmr and/or CRP Mt engage in competitive or cooperative DNA binding in this region warrants further investigation, as does the potential for indirect regulation by AbmR of gene targets downstream of Mcr11.…”

“…The universal second messenger molecule 3′, 5′ cyclic adenosine monophosphate (cAMP) is a global regulator of gene expression within Mtb, while secreted cAMP modulates Mtb's interactions with host macrophages (Lowrie et al, 1979;Agarwal et al, 2009;Bai et al, 2009;Nambi et al, 2013;Kahramanoglou et al, 2014;Knapp et al, 2015;Ranganathan et al, 2016;Johnson et al, 2017). cAMP levels affect the gene regulatory activities of two virulence-associated CRP/FNR family transcription factors in Mtb, Cmr and CRP Mt (Bai et al, 2005;Rickman et al, 2005;Kahramanoglou et al, 2014;Ranganathan et al, 2016). We previously identified gene of unknown function Rv1265 as a direct regulatory target of Cmr (Gazdik and McDonough, 2005;Ranganathan et al, 2016).…”

“…cAMP levels affect the gene regulatory activities of two virulence-associated CRP/FNR family transcription factors in Mtb, Cmr and CRP Mt (Bai et al, 2005;Rickman et al, 2005;Kahramanoglou et al, 2014;Ranganathan et al, 2016). We previously identified gene of unknown function Rv1265 as a direct regulatory target of Cmr (Gazdik and McDonough, 2005;Ranganathan et al, 2016). Rv1265 encodes a CHP whose expression is induced early during macrophage infection and in response to starvation or elevated levels of cAMP during hypoxia (Betts et al, 2002;Hobson et al, 2002;Gazdik et al, 2009).…”

Chemical activation of adenylyl cyclase Rv1625c inhibits growth of Mycobacterium tuberculosis on cholesterol and modulates intramacrophage signaling

“…Bioinformatic analyses indicate that most of these ACs contain different combinations of functional protein domains, suggesting a capacity to integrate multiple activities with cAMP signaling in Mtb (McCue et al ., ). cAMP‐responsive transcription factors Crp MT (Rv3676) and Cmr (Rv1675c) in Mtb regulate gene expression (Rickman et al ., ; Gazdik et al ., ; Stapleton et al ., 2010; 2012; Knapp et al ., ; Ranganathan et al ., ) to control metabolism, pathogenicity, dormancy and stress responses (Rickman et al ., ; Gazdik et al ., ; Stapleton et al ., ; Xu et al ., ; Pelly et al ., ; Hayden et al ., ; Nambi et al ., ; Choudhary et al ., ; Knapp et al ., ; Ranganathan et al ., ). In addition, elevated cAMP levels in Mtb can stimulate activity of the lysine acetyl‐transferase MtPat (Rv0998), which modulates the function of key metabolic enzymes via post‐translational modifications (Xu et al ., ; Hayden et al ., ; Nambi et al ., ).…”