title: Progesterone receptor deficient in chromatin binding has an altered cellular state.
abstract: Our previous work has shown that the progesterone receptor (PR) can exist in two distinct functional states in mammary adenocarcinoma cells. The differences in function included the ability to activate a promoter in organized chromatin, sensitivity to ligand, and ligand-independent activation. To determine whether these functional differences were because of altered cellular processing, we carried out biochemical analyses of the functionally distinct PRs. Although the majority of PR is localized to the nucleus, biochemical partitioning resulted in a loosely bound (cytosolic) fraction, and a tightly bound (nuclear) fraction. In the absence of progestins, the functionally distinct PRs differed significantly in partitioning between the two fractions. To characterize these fractions further, we analyzed interactions of unliganded PR with chaperones by coimmunoprecipitation. We determined that PR in the cytosolic fraction associated with hsp90 and p23. In contrast, PR in the nuclear fraction consisted of complexes containing hsp90, p23, and FKBP51 as well as PR that was dimerized and highly phosphorylated. Hormone treatment significantly reduced the formation of all PR-chaperone complexes. The hsp90 inhibitor, geldanamycin, similarly blocked transcriptional activity of both functionally distinct receptors. However, the two forms of the PR differed in their ability to associate with the mouse mammary tumor virus promoter in organized chromatin. These findings provide new information about the composition and distribution of mature progesterone receptor complexes in mammary adenocarcinoma cells, and suggest that differences in receptor subcellular distribution have a significant impact on their function. These findings also reveal that transiently expressed steroid receptors may not always be processed like their endogenous counterparts.
authors.affiliation: Laboratory of Receptor Biology and Gene Expression, NCI, National Institutes of Health, Bethesda, Maryland 20892-5055, USA. smithcat@exchange.nih.gov
authors.forename: Jeannine
authors.initials: J
authors.lastname: Botos
authors.forename: Wenjuan
authors.initials: W
authors.lastname: Xian
authors.forename: David F
authors.initials: DF
authors.lastname: Smith
authors.forename: Catharine L
authors.initials: CL
authors.lastname: Smith
mesh_terms: D000818:Animals; D016227:Benzoquinones; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019084:Fluorescent Antibody Technique, Indirect; D005814:Genes, Viral; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D006728:Hormones / Q000378:metabolism; D017404:In Situ Hybridization, Fluorescence; D019746:Intramolecular Oxidoreductases; D047029:Lactams, Macrocyclic; D008024:Ligands; D008156:Luciferases / Q000378:metabolism; D008324:Mammary Tumor Virus, Mouse / Q000235:genetics; D051379:Mice; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D018832:Molecular Chaperones / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D000071876:Prostaglandin-E Synthases; D011485:Protein Binding; D011487:Protein Conformation; D011809:Quinones / Q000494:pharmacology; D011980:Receptors, Progesterone / Q000737:chemistry* / Q000378:metabolism; D022021:Tacrolimus Binding Proteins / Q000737:chemistry / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D001616:beta-Galactosidase / Q000378:metabolism; D000099159:Tacrolimus Binding Protein 5
chemical_list: D016227:Benzoquinones; D002843:Chromatin; D004247:DNA; D004791:Enzyme Inhibitors; D018841:HSP90 Heat-Shock Proteins; D006728:Hormones; D019746:Intramolecular Oxidoreductases; D047029:Lactams, Macrocyclic; D008024:Ligands; D008156:Luciferases; D018832:Molecular Chaperones; D010750:Phosphoproteins; D000071876:Prostaglandin-E Synthases; D011809:Quinones; D011980:Receptors, Progesterone; D022021:Tacrolimus Binding Proteins; D001616:beta-Galactosidase; D000099159:Tacrolimus Binding Protein 5; C001277:geldanamycin