title: Heat shock alters the composition of heteromeric steroid receptor complexes and enhances receptor activity in vivo.
abstract: Under normal cellular conditions, human progesterone receptors (PR), immune-isolated from cytosols of T47D breast cancer cells, associate with two heat shock proteins (hsps), hsp 90 and hsp 70. Receptors activated by hormone binding in vivo and extracted from nuclei with 0.5 M NaCl no longer associate with hsp 90 but retain association with hsp 70. We have examined the effect of heat shock treatment of cells on hsp-receptor interactions and on receptor function. Heat shock resulted in a partial reduction in cellular levels of PR, but receptors that remained were functional for both steroid and DNA binding activities. By steady-state [35S]methionine labeling prior to heat shock treatment, it was determined that heat shock did not affect the composition or maintenance of preexisting cytosolic PR.hsp 90.hsp 70 complexes. By contrast, immune isolation of PR complexes from cells pulse-labeled with [35S]methionine showed that heat shock altered the composition of newly synthesized hsps associated with PR. After heat shock, both the highly inducible form of hsp 70 (72K hsp) and a 100K hsp were bound to cytosol PR, and inducible 72K hsp remained bound with the nuclear-activated PR. Neither of these hsps were associated in detectable amounts with PR under normal cellular conditions. With respect to receptor function, heat shock treatment substantially enhanced the activity of PR in vivo as determined by measuring hormone-dependent PR-mediated transcription of a target reporter gene (MMTV-CAT) that was stably transfected into T47D cells. Heat shock treatment alone, in the absence of hormone, did not stimulate MMTV-CAT expression nor did it affect transcription from a control reporter gene, pSV2-CAT, suggesting that enhanced receptor activity was due to an effect on PR-mediated processes and not to a general effect on transcription. Induction of the heat shock response by a related chemical stress (sodium arsenite) also enhanced PR activity in vivo. Interestingly, sodium arsenite produced both a greater induction of hsp 90 and hsp 70 synthesis and a greater fold enhancement of PR-mediated gene transcription than did heat shock. This suggests that enhancement of PR activity is related not only to induction of hsp synthesis but also to the severity of the stress response. The present results provide an indication that in certain cells there may exist an interrelationship between the activation pathways by which cells respond to stress and to steroid hormones. Possible mechanisms responsible for heat shock effects on PR activity are discussed.
authors.affiliation: Department of Pathology, University of Colorado Health Sciences Center, Denver 80262.
authors.forename: D P
authors.initials: DP
authors.lastname: Edwards
authors.forename: P A
authors.initials: PA
authors.lastname: Estes
authors.forename: V A
authors.initials: VA
authors.lastname: Fadok
authors.forename: B J
authors.initials: BJ
authors.lastname: Bona
authors.forename: S
authors.initials: S
authors.lastname: Oñate
authors.forename: S K
authors.initials: SK
authors.lastname: Nordeen
authors.forename: W J
authors.initials: WJ
authors.lastname: Welch
mesh_terms: D001151:Arsenic / Q000494:pharmacology; D018053:Arsenites; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D006360:Heat-Shock Proteins / Q000096:biosynthesis / Q000378:metabolism*; D006358:Hot Temperature; D006801:Humans; D008324:Mammary Tumor Virus, Mouse / Q000235:genetics; D010766:Phosphorylation; D011397:Promegestone / Q000494:pharmacology; D011980:Receptors, Progesterone / Q000302:isolation & purification / Q000502:physiology*; D017670:Sodium Compounds; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
chemical_list: D018053:Arsenites; D006360:Heat-Shock Proteins; D011980:Receptors, Progesterone; D017670:Sodium Compounds; C017947:sodium arsenite; D004247:DNA; D011397:Promegestone; D015500:Chloramphenicol O-Acetyltransferase; D001151:Arsenic