EmbR, a regulatory protein with ATPase activity, is a substrate of multiple serine/threonine kinases and phosphatase in Mycobacterium tuberculosis.
Phosphorylation of the mycobacterial transcriptional activator, EmbR, is essential for transcriptional regulation of the embCAB operon encoding cell wall arabinosyltransferases. This signaling pathway eventually affects the resistance to ethambutol (a frontline antimycobacterial drug) and the cell wall Lipoarabinomannan/Lipomannan ratio (an important determinant for averting the host immune response). In this study, further biochemical characterization revealed that EmbR, as a transcriptional regulator, interacts with RNA polymerase and possesses a phosphorylation-dependent ATPase activity that might play a role in forming an open complex between EmbR and RNA polymerase. EmbR was recently shown to be phosphorylated by the cognate mycobacterial serine/threonine (Ser/Thr) kinase, PknH. Using bioinformatic analysis and in vitro assays, we identified additional novel regulators of the signaling pathway leading to EmbR phosphorylation, namely the Ser/Thr protein kinases PknA and PknB. A previously unresolved question raised by this signaling scheme is the fate of phosphorylated kinases and EmbR at the end of the signaling cycle. Here we show that Mstp, a mycobacterial Ser/Thr phosphatase, antagonizes Ser/Thr protein kinase-EmbR signaling by dephosphorylating Ser/Thr protein kinases, as well as EmbR, in vitro. Additionally, dephosphorylation of EmbR reduced its ATPase activity, interaction with Ser/Thr protein kinases and DNA-binding activity, emphasizing the antagonistic role of Mstp in the EmbR-Ser/Thr protein kinase signaling system.
Sharma
Kirti
K
Institute of Genomics and Integrative Biology, Delhi, India.
Gupta
Meetu
M
Krupa
Ananth
A
Srinivasan
Narayanaswamy
N
Singh
Yogendra
Y
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D015964:Gene Expression Regulation, Bacterial; D008954:Models, Biological; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D013379:Substrate Specificity; D013997:Time Factors; D015534:Trans-Activators / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D015534:Trans-Activators; C484411:PknH protein, Mycobacterium tuberculosis; D017346:Protein Serine-Threonine Kinases; D012321:DNA-Directed RNA Polymerases; D010749:Phosphoprotein Phosphatases; D000251:Adenosine Triphosphatases