Phosphorylation of human progesterone receptors at serine-294 by mitogen-activated protein kinase signals their degradation by the 26S proteasome.
Ligand-dependent down-regulation that leads to rapid and extensive loss of protein is characteristic of several nuclear steroid receptors, including human progesterone receptors (PRs). In breast cancer cells, >95% of PRs are degraded 6 h after the start of progestin treatment. The mechanism for down-regulation is unknown. We examined the role of PR phosphorylation by mitogen-activated protein kinases (MAPKs) in this process. Lactacystin and calpain inhibitor I, specific inhibitors of the 26S proteasome, blocked progestin-induced down-regulation, and ubiquitinated conjugates of PR accumulated in cells. Ligand-dependent PR degradation was also blocked by specific inhibition of p42 and p44 MAPKs. To define the targets of phosphorylation by this kinase, two serine/proline MAPK consensus sites on PR were mutated. We demonstrate that mutation of PR serine-294 to alanine (S294A) specifically and completely prevents ligand-dependent receptor down-regulation. We also find that rapid, ligand-independent degradation of immature PR intermediates occurs by a proteasome-mediated pathway. These results demonstrate that PR destruction, by either of two alternate routes, is mediated by the 26S proteasome. Specifically, down-regulation of mature PRs occurs by a mechanism in which ligand binding activates PR phosphorylation by MAPKs at a unique serine residue, which then targets the receptors for degradation.
Lange
C A
CA
Department of Medicine, The Molecular Biology Program, and The Colorado Cancer Center, University of Colorado Health Sciences Center, Denver, CO 80262, USA.
Shen
T
T
Horwitz
K B
KB
D000111:Acetylcysteine / Q000031:analogs & derivatives / Q000494:pharmacology; D001943:Breast Neoplasms / Q000737:chemistry; D016384:Consensus Sequence; D003546:Cysteine Endopeptidases / Q000187:drug effects / Q000378:metabolism*; D015536:Down-Regulation / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006023:Glycoproteins / Q000494:pharmacology; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D020935:MAP Kinase Signaling System; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009097:Multienzyme Complexes / Q000187:drug effects / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000378:metabolism*; D010766:Phosphorylation; D010768:Phosphoserine / Q000737:chemistry*; D011397:Promegestone / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex; D020033:Protein Isoforms / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011725:Pyridines / Q000494:pharmacology; D011980:Receptors, Progesterone / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014452:Ubiquitins / Q000378:metabolism
D004791:Enzyme Inhibitors; D006023:Glycoproteins; D007093:Imidazoles; D009097:Multienzyme Complexes; D009363:Neoplasm Proteins; D020033:Protein Isoforms; D011725:Pyridines; D011980:Receptors, Progesterone; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; C039304:calpain inhibitors; C067713:lactacystin; D010768:Phosphoserine; D011397:Promegestone; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C093642:SB 203580; C090942:4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole; D000111:Acetylcysteine