Dissection of progesterone receptor-mediated chromatin remodeling and transcriptional activation in vivo.
We have investigated whether constitutive binding by the progesterone receptor (PR) to a promoter is required for the maintenance of an open chromatin structure in vivo. For these experiments, we used human T47D breast cancer cells in which the mouse mammary tumor virus (MMTV) promoter, stably assembled as chromatin, is constitutively hypersensitive to endonucleolytic cleavage. In vivo footprinting revealed that transcription factors nuclear factor 1 and the PR were constitutively bound to the MMTV promoter in these cells. Treatment of these cells for 1 hr with the steroid antagonist ZK98299 prevented PR binding to chromatin in vivo and reversed hypersensitivity, leading to the loss of transcription factor binding. The reduction in hypersensitivity induced by ZK98299 was readily reversed by treatment with the progestin R5020. The chromatin organization of the promoter could be cycled between the open and closed states by consecutive treatments with agonist or antagonist. The antagonist RU486 also blocked activation of transcription and the assembly of a transcription preinitiation complex, but in contrast to ZK98299, maintained the hypersensitive chromatin state. Taken together, these results suggest that PR binding to chromatin is sufficient to induce hypersensitivity to endonucleolytic cleavage. Furthermore, they indicate that the PR binding to DNA and the resulting chromatin hypersensitivity is functionally separate from transcriptional activation in vivo.
Mymryk
J S
JS
Department of Obstetrics and Gynaecology, University of Western Ontario, London, Canada.
Archer
T K
TK
D000818:Animals; D001943:Breast Neoplasms / Q000473:pathology; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral; D002843:Chromatin / Q000187:drug effects / Q000378:metabolism*; D005260:Female; D006067:Gonanes / Q000494:pharmacology; D006801:Humans; D008324:Mammary Tumor Virus, Mouse / Q000235:genetics; D051379:Mice; D015735:Mifepristone / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D011980:Receptors, Progesterone / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D015533:Transcriptional Activation / Q000187:drug effects
D002843:Chromatin; D006067:Gonanes; D011980:Receptors, Progesterone; D015735:Mifepristone; C053238:onapristone