title: The Mycobacterium tuberculosis transcriptional repressor EthR is negatively regulated by Serine/Threonine phosphorylation.
abstract: Recent efforts have underlined the role of Serine/Threonine Protein Kinases (STPKs) in growth, pathogenesis and cell wall metabolism in mycobacteria. Herein, we demonstrated that the Mycobacterium tuberculosis EthR, a transcriptional repressor that regulates the activation process of the antitubercular drug ethionamide (ETH) is a specific substrate of the mycobacterial kinase PknF. ETH is a prodrug that must undergo bioactivation by the monooxygenease EthA to exert its antimycobacterial activity and previous studies reported that EthR represses transcription of ethA by binding to the ethA-ethR intergenic region. Mass spectrometry analyses and site-directed mutagenesis identified a set of four phosphoacceptors, namely Thr2, Thr3, Ser4 and Ser7. This was further supported by the complete loss of PknF-dependent phosphorylation of a phosphoablative EthR mutant protein. Importantly, a phosphomimetic version of EthR, in which all phosphosites were replaced by Asp residues, exhibited markedly decreased DNA-binding activity compared with the wild-type protein. Together, these findings are the first demonstration of EthR phosphorylation and indicate that phosphorylation negatively affects its DNA-binding activity, which may impact ETH resistance levels in M. tb.
authors.affiliation: CNRS-UMR5235, DIMNP, Université de Montpellier 2, Place Eugène Bataillon, 34095 Montpellier Cedex 5, France.
authors.forename: Jade
authors.initials: J
authors.lastname: Leiba
authors.affiliation: CNRS-UMR5235, DIMNP, Université de Montpellier 2, Place Eugène Bataillon, 34095 Montpellier Cedex 5, France.
authors.forename: Séverine
authors.initials: S
authors.lastname: Carrère-Kremer
authors.affiliation: U1019-UMR8204, Univ Lille Nord de France, Institut Pasteur de Lille, 59019 Lille, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Blondiaux
authors.affiliation: U1019-UMR8204, Univ Lille Nord de France, Institut Pasteur de Lille, 59019 Lille, France.
authors.forename: Martin Moune
authors.initials: MM
authors.lastname: Dimala
authors.affiliation: Structural Biology Brussels and Molecular and Cellular Interactions, VIB, 1050 Brussels, Belgium.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Wohlkönig
authors.affiliation: U1019-UMR8204, Univ Lille Nord de France, Institut Pasteur de Lille, 59019 Lille, France.
authors.forename: Alain
authors.initials: A
authors.lastname: Baulard
authors.affiliation: CNRS-UMR5235, DIMNP, Université de Montpellier 2, Place Eugène Bataillon, 34095 Montpellier Cedex 5, France; Inserm, DIMNP, Place Eugène Bataillon, 34095 Montpellier Cedex 05, France.
authors.forename: Laurent
authors.initials: L
authors.lastname: Kremer
authors.affiliation: CNRS-UMR5235, DIMNP, Université de Montpellier 2, Place Eugène Bataillon, 34095 Montpellier Cedex 5, France. Electronic address: virginie.molle@univ-montp2.fr.
authors.forename: Virginie
authors.initials: V
authors.lastname: Molle
mesh_terms: D000595:Amino Acid Sequence; D000995:Antitubercular Agents / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D005000:Ethionamide / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013912:Threonine / Q000378:metabolism; D014376:Tuberculosis / Q000382:microbiology
chemical_list: D000995:Antitubercular Agents; D001426:Bacterial Proteins; C481661:EthR protein, Mycobacterium tuberculosis; D012097:Repressor Proteins; D013912:Threonine; D012694:Serine; D017346:Protein Serine-Threonine Kinases; D005000:Ethionamide
keywords: EthR; Ethionamide; Mycobacterium; Phosphorylation; Ser/Thr protein kinases