title: 8-Bromo-cyclic AMP induces phosphorylation of two sites in SRC-1 that facilitate ligand-independent activation of the chicken progesterone receptor and are critical for functional cooperation between SRC-1 and CREB binding protein.
abstract: Elevation of intracellular 8-bromo-cyclic AMP (cAMP) can activate certain steroid receptors and enhance the ligand-dependent activation of most receptors. During ligand-independent activation of the chicken progesterone receptor (cPR(A)) with the protein kinase A (PKA) activator, 8-bromo-cAMP, we found no alteration in cPR(A) phosphorylation (W. Bai, B. G. Rowan, V. E. Allgood, B. W. O'Malley, and N. L. Weigel, J. Biol. Chem. 272:10457-10463, 1997). To determine if other receptor-associated cofactors were targets of cAMP-dependent signaling pathways, we examined the phosphorylation of steroid receptor coactivator 1 (SRC-1). We detected a 1.8-fold increase in SRC-1 phosphorylation in transfected COS-1 cells incubated with 8-bromo-cAMP. Phosphorylation was increased on two mitogen-activated protein kinase (MAPK) sites, threonine 1179 and serine 1185. PKA did not phosphorylate these sites in vitro. However, blockage of PKA activity in COS-1 cells with the PKA inhibitor (PKI) prevented the 8-bromo-cAMP-mediated phosphorylation of these sites. Incubation of COS-1 cells with 8-bromo-cAMP resulted in activation of the MAPK pathway, as determined by Western blotting with antibodies to the phosphorylated (active) form of Erk-1/2, suggesting an indirect pathway to SRC-1 phosphorylation. Mutation of threonine 1179 and serine 1185 to alanine in COS-1 cells coexpressing cPR(A) and the GRE(2)E1bCAT reporter resulted in up to a 50% decrease in coactivation during both ligand-independent activation and ligand-dependent activation. This was due, in part, to loss of functional cooperation between SRC-1 and CREB binding protein for coactivation of cPR(A). This is the first demonstration of cross talk between a signaling pathway and specific phosphorylation sites in a nuclear receptor coactivator that can regulate steroid receptor activation.
authors.affiliation: Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.
authors.forename: B G
authors.initials: BG
authors.lastname: Rowan
authors.forename: N
authors.initials: N
authors.lastname: Garrison
authors.forename: N L
authors.initials: NL
authors.lastname: Weigel
authors.forename: B W
authors.initials: BW
authors.lastname: O'Malley
mesh_terms: D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology*; D000818:Animals; D050882:CREB-Binding Protein; D002645:Chickens; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D051548:Histone Acetyltransferases; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008957:Models, Genetic; D009687:Nuclear Proteins / Q000378:metabolism*; D056920:Nuclear Receptor Coactivator 1; D010449:Peptide Mapping; D010748:Phosphopeptides; D010766:Phosphorylation; D011374:Progesterone / Q000494:pharmacology; D011485:Protein Binding; D011980:Receptors, Progesterone / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction; D013912:Threonine / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
chemical_list: D009687:Nuclear Proteins; D010748:Phosphopeptides; D011980:Receptors, Progesterone; D011994:Recombinant Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D015124:8-Bromo Cyclic Adenosine Monophosphate; D013912:Threonine; D012694:Serine; D011374:Progesterone; D050882:CREB-Binding Protein; D051548:Histone Acetyltransferases; D056920:Nuclear Receptor Coactivator 1; D017868:Cyclic AMP-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases