The goal of this project is to understand the mechanism by which the progesterone receptor (PR) interacts with and remodels chromatin at target genes in vivo using the mouse mammary tumor virus (MMTV) promoter as a model system. In the non-activated state, this promoter has a chromatin structure repressive to transcription when it exists in a stably-replicating form, either integrated into the cellular genome or as an episome. Upon binding by the liganded glucocorticoid receptor (GR), the promoter undergoes a chromatin remodeling event which is mechanistically involved in the activation of transcription. Our previous work has shown that the GR and PR have different requirements for chromatin remodeling at the MMTV promoter even though they bind to the same DNA sequences in the promoter. Our observations may form the basis for a mechanism by which the GR and PR control expression of distinct sets of target genes in vivo. This is particularly relevant in the mammary gland where the GR and PR can coexist in the same cell types. We have established that the PR exists in two distinct states in cultured mammary adenocarcinoma cells. In one state, it can neither remodel chromatin nor activate transcription at the MMTV promoter; thus its action may be restricted to target genes which do not require remodeling. In addition, this form of the PR can be activated by other signal transduction pathways in a progestin-independent fashion. In the second state, the PR is able to remodel and activate the MMTV promoter in chromatin, but is refractory to ligand-independent activation. Thus, this form of the PR responds only to its ligand but would be able to activate target genes even in a repressive chromatin environment. We have also shown that the PR can be converted from the first state to the second by some form of cellular processing. This may represent a mechanism by which cells can restrict or expand the activity of the PR in vivo. To elucidate the biochemical basis for the two distinct functional states, we have been developing an immunoprecipitation method to isolate the PR in its native forms and examine associated proteins and post-translational processing. This has been done in collaboration with Dr. David Smith who has provided us with reagents and expertise invaluble to the project. We have successfully developed this method to the point where we can begin comparative studies of the PR in its two functional states. In collaboration with Dr. Stoney Simons (NIDDK), we have also initiated a project to examine the activity of the two functional forms of the PR in the presence of anti-progestins. Given recent studies linking progestin exposure to increased risk of breast cancer in postmenopausal women, it is probable that antiprogestins may be used in a chemopreventive role in the future. A better understanding of PR behavior in response to these agents specifically in the context of mammary-derived cells may provide important information to clinicians.
