RT Journal Article SR Electronic T1 The Progestin Receptor Interactome in the Female Mouse Hypothalamus: Interactions with Synaptic Proteins Are Isoform-Specific and Ligand-Dependent JF eneuro JO eNeuro FD Society for Neuroscience SP ENEURO.0272-17.2017 DO 10.1523/ENEURO.0272-17.2017 A1 Kalpana D. Acharya A1 Sabin A. Nettles A1 Katherine J. Sellers A1 Dana D. Im A1 Moriah Harling A1 Cassandra Pattanayak A1 Didem Vardar-Ulu A1 Cheryl F Lichti A1 Shixia Huang A1 Dean P. Edwards A1 Deepak P. Srivastava A1 Larry Denner A1 Marc J. Tetel YR 2017 UL http://www.eneuro.org/content/early/2017/09/11/ENEURO.0272-17.2017.abstract AB Progestins bind to the progestin receptor (PR) isoforms, PR-A and PR-B, in brain to influence development, female reproduction, anxiety and stress. Hormone-activated PR associate with multiple proteins to form functional complexes. In the present study, proteins from female mouse hypothalamus that associate with PR were isolated using affinity pull-down assays with GST-tagged mouse PR-A and PR-B. Using complementary proteomics approaches, reverse phase protein array (RPPA) and mass spectrometry, we identified hypothalamic proteins that interact with PR in ligand-dependent and isoform-specific manner, which were confirmed by Western blot. Synaptic proteins, including synapsin-I and synapsin-II, interacted with agonist-bound PR isoforms, suggesting that both isoforms function in synaptic plasticity. In further support, synaptogyrin-III and synapsin-III associated with PR-A and PR-B, respectively. PR also interacted with kinases, including c-Src, mTOR and MAPK1, confirming phosphorylation as an integral process in rapid effects of PR in brain. Consistent with a role in transcriptional regulation, PR associated with transcription factors and coactivators in a ligand-specific and isoform-dependent manner. Interestingly, both PR isoforms associated with a key regulator of energy homeostasis, FoxO1, suggesting a novel role for PR in energy metabolism. Because many identified proteins in this PR interactome are synaptic proteins, we tested the hypothesis that progestins function in synaptic plasticity. Indeed, progesterone enhanced synaptic density, by increasing synapsin-I positive synapses, in rat primary cortical neuronal cultures. This novel combination of RPPA and mass spectrometry allowed identification of PR action in synaptic remodeling and energy homeostasis, and provide unique roles for progestins in brain function and disease.Significance Statement Progesterone binds to the progestin receptor (PR) isoforms, PR-A and PR-B, in brain to profoundly influence female reproduction, brain development, neuroprotection and stress. We combined mass spectrometry and reverse phase protein arrays to identify mouse hypothalamic proteins that function in synaptic plasticity, transcription, translation and energy metabolism that interacted with PR in a ligand-dependent and isoform-specific manner. In addition, progesterone increased synaptic density in primary cortical neurons, suggesting a function for progesterone in synapse formation in cortical neurons. Taken together, these findings provide evidence for novel functions of PR in synaptic modulation and energy homeostasis to affect health and disease.