Regulation of co-repressive activity of and HDAC recruitment to RIP140 by site-specific phosphorylation.
Receptor interacting protein 140 (RIP140) is a versatile transcriptional co-repressor that contains several autonomous repressive domains (RDs). The N-terminal RD acts by recruiting histone deacetylases (HDACs). In a comprehensive proteomic analysis of RIP140 by MS, 11 phosphorylation sites of RIP140 are identified; among them five sites are located in the N-terminal RD including Ser104, Thr202, Thr207, Ser358, and Ser380. The role of phosphorylation of RIP140 in regulating its biological activity and the underlying mechanism are examined using a site-directed mutagenesis approach. Mutations mimicking constitutive phosphorylation or dephosphorylation are introduced. The N-terminal RD phosphorylation, mediated by the mitogen-activated protein kinase (MAPK), enhances its repressive activity through increased recruitment of HDAC. Mutations mimicking constitutive dephosphorylation at Thr202 or Thr207 significantly impair its repressive activity and HDAC recruitment, whereas mutation at Ser358 only slightly affects its HDAC recruitment and the repressive activity. Consistently, mutations mimicking constitutive phosphorylation at either Thr202 or Thr207 convert RIP140 into a more potent repressor, which is less responsive to a disturbance in the MAPK system. Furthermore, constitutive phosphorylation at both Thr202 and Thr207 residues renders RIP140 fully repressive and strongly interacting with HDAC. The activity of this mutant is resistant to the MAPK inhibitor, indicating an essential role for Thr202 and Thr207 in MAPK-mediated modulation of RIP140 function. The study provides insights into the modulation of RIP140 biological activity through a specific cellular signaling pathway that augments phosphorylation at specific residues of RIP140 molecule and alters its cofactor recruitment.
Gupta
Pawan
P
Department of Pharmacology, University of Minnesota Medical School, Minneapolis, Minnesota 55455, USA.
Huq
M D Mostaqul
MD
Khan
Shaukat Ali
SA
Tsai
Nien-Pei
NP
Wei
Li-Na
LN
D048868:Adaptor Proteins, Signal Transducing; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D015536:Down-Regulation / Q000187:drug effects; D006655:Histone Deacetylases / Q000378:metabolism*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D000078723:Nuclear Receptor Interacting Protein 1; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D021381:Protein Transport; D040901:Proteomics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D015203:Reproducibility of Results
D048868:Adaptor Proteins, Signal Transducing; D009687:Nuclear Proteins; D000078723:Nuclear Receptor Interacting Protein 1; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D012097:Repressor Proteins; D010770:Phosphotransferases; D006655:Histone Deacetylases