A highly conserved transcriptional repressor controls a large regulon involved in lipid degradation in Mycobacterium smegmatis and Mycobacterium tuberculosis

Abstract

SummaryThe Mycobacterium tuberculosis TetR-type regulator Rv3574 has been implicated in pathogenesis as it is induced in vivo, and genome-wide essentiality studies show it is required for infection. As the gene is highly conserved in the mycobacteria, we deleted the Rv3574 orthologue in Mycobacterium smegmatis (MSMEG_6042) and used real-time quantitative polymerase chain reaction and microarray analyses to show that it represses the transcription both of itself and of a large number of genes involved in lipid metabolism. We identified a conserved motif within its own promoter (TnnAACnnGTTnnA) and showed that it binds as a dimer to 29 bp probes containing the motif. We found 16 and 31 other instances of the motif in intergenic regions of M. tuberculosis and M. smegmatis respectively. Combining the results of the microarray studies with the motif analyses, we predict that Rv3574 directly controls the expression of 83 genes in M. smegmatis, and 74 in M. tuberculosis. Many of these genes are known to be induced by growth on cholesterol in rhodococci, and palmitate in M. tuberculosis. We conclude that this regulator, designated elsewhere as kstR, controls the expression of genes used for utilizing diverse lipids as energy sources, possibly imported through the mce4 system.

Mycobacterium smegmatis is a suitable cell factory for the production of steroidic synthons

“…Ksh1 (KshA1B1), Ksh2 (KshA2B2), Ksh3 (KshA1B2), Ksh4 (KshA2B1). However, a detailed analysis of the upstream sequences of these genes showed that only the MSMEG_5925 ( kshA1 ) and MSMEG_6039 ( kshB1 ) genes are preceded by promoters inducible by cholesterol, having the consensus operator sequence for the binding of the KstR repressor, one of the regulators of cholesterol catabolism (named KstR regulon) (Kendall et al ., 2007). Moreover, microarray expression experiments carried out on M. smegmatis indicate that kshA1 and kshB1 are induced 1.9‐fold and 7.0‐fold, respectively, in cholesterol with respect to glycerol (Table 1) (Uhía et al ., 2012).…”

Characterization of the KstR-dependent promoter of the gene for the first step of the cholesterol degradative pathway in Mycobacterium smegmatis

“…In spite of the relevant role that the kstR regulon seems to play, the mechanisms involved in this regulation are still poorly understood. Gel retardation assays have shown that KstR binds as a dimer to DNA probes containing an in silico-identified motif (TNNAACNNGTTNNA) (Kendall et al, 2007). However, experimental evidence that this motif represents the authentic KstR operator remains to be presented.…”

“…Although recent biochemical and structural studies have assigned a role for some of the mycobacterial genes in cholesterol catabolism, the complete degradative pathway and its specific regulation remain to be fully established (Capyk et al, 2009;Knol et al, 2008;Lack et al, 2010;Yam et al, 2009). Recently, it has been shown that cholesterol utilization in mycobacteria is controlled by two TetR-type transcriptional repressors (Kendall et al, 2007(Kendall et al, , 2010. One of these repressors, named KstR, is encoded by the Rv3574 gene in M. tuberculosis and by the MSMEG_6042 gene in M. smegmatis.…”

“…One of these repressors, named KstR, is encoded by the Rv3574 gene in M. tuberculosis and by the MSMEG_6042 gene in M. smegmatis. KstR controls the expression of 83 cholesterol catabolic genes (kstR regulon) (Kendall et al, 2007); many of these genes are essential for virulence, highlighting the importance of cholesterol catabolism in the pathogenicity of M. tuberculosis (Chang et al, 2007;Hu et al, 2010;Nesbitt et al, 2010;Rengarajan et al, 2005;Sassetti & Rubin, 2003;Schnappinger et al, 2003;Yam et al, 2009). In spite of the relevant role that the kstR regulon seems to play, the mechanisms involved in this regulation are still poorly understood.…”

Transcription factors Rv0081 and Rv3334 connect the early and the enduring hypoxic response of Mycobacterium tuberculosis

“…Rv3334 is a member of the MerR family of transcriptional regulators, and a recent study has found that Rv3334 is an auto repressor but positively regulates the expression of kstR , which encodes a transcription factor [35]. Cholesterol catabolism is essential for M. tb persistence [36,37] and KstR controls a large regulon mediating cholesterol degradation and lipid metabolism [36]. As such, Rv3334 may play an active role in the response of M. tb to hypoxia.…”