Polyphosphate kinase 1, a central node in the stress response network of Mycobacterium tuberculosis, connects the two-component systems MprAB and SenX3-RegX3 and the extracytoplasmic function sigma factor, sigma E.
Polyphosphate (poly P) metabolism regulates the stress response in mycobacteria. Here we describe the regulatory architecture of a signal transduction system involving the two-component system (TCS) SenX3-RegX3, the extracytoplasmic function sigma factor sigma E (SigE) and the poly P-synthesizing enzyme polyphosphate kinase 1 (PPK1). The ppk1 promoter of Mycobacterium tuberculosis is activated under phosphate starvation. This is attenuated upon deletion of an imperfect palindrome likely representing a binding site for the response regulator RegX3, a component of the two-component system SenX3-RegX3 that responds to phosphate starvation. Binding of phosphorylated RegX3 to this site was confirmed by electrophoretic mobility shift assay. The activity of the ppk1 promoter was abrogated upon deletion of a putative SigE binding site. Pull-down of SigE from M. tuberculosis lysates of phosphate-starved cells with a biotinylated DNA harbouring the SigE binding site confirmed the likely binding of SigE to the ppk1 promoter. In vitro transcription corroborated the involvement of SigE in ppk1 transcription. Finally, the overexpression of RseA (anti-SigE) attenuated ppk1 expression under phosphate starvation, supporting the role of SigE in ppk1 transcription. The regulatory elements identified in ppk1 transcription in this study, combined with our earlier observation that PPK1 is itself capable of regulating sigE expression via the MprAB TCS, suggest the presence of multiple positive-feedback loops in this signalling circuit. In combination with the sequestering effect of RseA, we hypothesize that this architecture could be linked to bistability in the system that, in turn, could be a key element of persistence in M. tuberculosis.
Department of Chemistry, Bose Institute, 93/1 Acharya Prafulla Chandra Road, Kolkata 700009, India.
Sourav
S
Sanyal
Department of Chemistry, Bose Institute, 93/1 Acharya Prafulla Chandra Road, Kolkata 700009, India.
Srijon Kaushik
SK
Banerjee
Department of Chemistry, Bose Institute, 93/1 Acharya Prafulla Chandra Road, Kolkata 700009, India.
Rajdeep
R
Banerjee
Department of Chemistry, Bose Institute, 93/1 Acharya Prafulla Chandra Road, Kolkata 700009, India.
Jayanta
J
Mukhopadhyay
Department of Chemistry, Bose Institute, 93/1 Acharya Prafulla Chandra Road, Kolkata 700009, India.
Manikuntala
M
Kundu
D001426:Bacterial Proteins / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D015964:Gene Expression Regulation, Bacterial; D053263:Gene Regulatory Networks; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000502:physiology*; D010710:Phosphates / Q000378:metabolism; D010770:Phosphotransferases / Q000378:metabolism*; D017856:Phosphotransferases (Phosphate Group Acceptor) / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism; D012808:Sigma Factor / Q000378:metabolism*; D015398:Signal Transduction; D013312:Stress, Physiological; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004269:DNA, Bacterial; C480224:MprA protein, Mycobacterium tuberculosis; D010710:Phosphates; C110608:RegX3 protein, Mycobacterium; D012808:Sigma Factor; C111259:sigE protein, Bacteria; D010770:Phosphotransferases; D011494:Protein Kinases; C480223:MprB protein, Mycobacterium tuberculosis; C110607:SenX3 protein, Mycobacterium; D017856:Phosphotransferases (Phosphate Group Acceptor); C022424:polyphosphate kinase