title: Progesterone receptor membrane component 1 is a functional part of the glucagon-like peptide-1 (GLP-1) receptor complex in pancreatic β cells.
abstract: Glucagon-like peptide-1 (GLP-1) is an incretin hormone that regulates glucose homeostasis. Because of their direct stimulation of insulin secretion from pancreatic β cells, GLP-1 receptor (GLP-1R) agonists are now important therapeutic options for the treatment of type 2 diabetes. To better understand the mechanisms that control the insulinotropic actions of GLP-1, affinity purification and mass spectrometry (AP-MS) were employed to uncover potential proteins that functionally interact with the GLP-1R. AP-MS performed on Chinese hamster ovary cells or MIN6 β cells, both expressing the human GLP-1R, revealed 99 proteins potentially associated with the GLP-1R. Three novel GLP-1R interactors (PGRMC1, Rab5b, and Rab5c) were further validated through co-immunoprecipitation/immunoblotting, fluorescence resonance energy transfer, and immunofluorescence. Functional studies revealed that overexpression of PGRMC1, a novel cell surface receptor that associated with liganded GLP-1R, enhanced GLP-1-induced insulin secretion (GIIS) with the most robust effect. Knockdown of PGRMC1 in β cells decreased GIIS, indicative of positive interaction with GLP-1R. To gain insight mechanistically, we demonstrated that the cell surface PGRMC1 ligand P4-BSA increased GIIS, whereas its antagonist AG-205 decreased GIIS. It was then found that PGRMC1 increased GLP-1-induced cAMP accumulation. PGRMC1 activation and GIIS induced by P4-BSA could be blocked by inhibition of adenylyl cyclase/EPAC signaling or the EGF receptor-PI3K signal transduction pathway. These data reveal a dual mechanism for PGRMC1-increased GIIS mediated through cAMP and EGF receptor signaling. In conclusion, we identified several novel GLP-1R interacting proteins. PGRMC1 expressed on the cell surface of β cells was shown to interact with the activated GLP-1R to enhance the insulinotropic actions of GLP-1.
authors.affiliation: From the ‡Department of Physiology, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 1A8; §Division of Advanced Diagnosis, Toronto General Research Institute, Toronto, Canada, M5G 1C7;
authors.forename: Ming
authors.initials: M
authors.lastname: Zhang
authors.affiliation: ¶Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, and Department of Biochemistry, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 3M2;
authors.forename: Mélanie
authors.initials: M
authors.lastname: Robitaille
authors.affiliation: ‖Endocrine Discovery, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana, 46285;
authors.forename: Aaron D
authors.initials: AD
authors.lastname: Showalter
authors.affiliation: From the ‡Department of Physiology, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 1A8; §Division of Advanced Diagnosis, Toronto General Research Institute, Toronto, Canada, M5G 1C7;
authors.forename: Xinyi
authors.initials: X
authors.lastname: Huang
authors.affiliation: From the ‡Department of Physiology, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 1A8;
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.affiliation: From the ‡Department of Physiology, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 1A8;
authors.forename: Alpana
authors.initials: A
authors.lastname: Bhattacharjee
authors.affiliation: ‖‖Quantitative Biology, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana, 46285;
authors.forename: Francis S
authors.initials: FS
authors.lastname: Willard
authors.affiliation: **Department of Endocrinology and Metabolism, Shanghai Jiaotong University Affiliated Sixth People's Hospital, Shanghai, China 200233;
authors.forename: Junfeng
authors.initials: J
authors.lastname: Han
authors.affiliation: From the ‡Department of Physiology, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 1A8; §Division of Advanced Diagnosis, Toronto General Research Institute, Toronto, Canada, M5G 1C7;
authors.forename: Sean
authors.initials: S
authors.lastname: Froese
authors.affiliation: **Department of Endocrinology and Metabolism, Shanghai Jiaotong University Affiliated Sixth People's Hospital, Shanghai, China 200233;
authors.forename: Li
authors.initials: L
authors.lastname: Wei
authors.affiliation: ‡‡Department of Medicine, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 1A8.
authors.forename: Herbert Y
authors.initials: HY
authors.lastname: Gaisano
authors.affiliation: ¶Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, and Department of Biochemistry, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 3M2;
authors.forename: Stéphane
authors.initials: S
authors.lastname: Angers
authors.affiliation: ‖Endocrine Discovery, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana, 46285;
authors.forename: Kyle W
authors.initials: KW
authors.lastname: Sloop
authors.affiliation: From the ‡Department of Physiology, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 1A8; michael.wheeler@utoronto.ca f.dai@utoronto.ca.
authors.forename: Feihan F
authors.initials: FF
authors.lastname: Dai
authors.affiliation: From the ‡Department of Physiology, Faculty of Medicine, University of Toronto, Toronto, Canada, M5S 1A8; §Division of Advanced Diagnosis, Toronto General Research Institute, Toronto, Canada, M5G 1C7; michael.wheeler@utoronto.ca f.dai@utoronto.ca.
authors.forename: Michael B
authors.initials: MB
authors.lastname: Wheeler
mesh_terms: D000067956:Adenylyl Cyclase Inhibitors; D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000096:biosynthesis / Q000378:metabolism; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism; D052216:Glucagon-Like Peptide 1 / Q000378:metabolism*; D000067757:Glucagon-Like Peptide-1 Receptor; D020662:Guanine Nucleotide Exchange Factors / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051381:Rats; D018027:Receptors, Glucagon / Q000378:metabolism*; D011980:Receptors, Progesterone / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism
chemical_list: D000067956:Adenylyl Cyclase Inhibitors; D000242:Cyclic AMP; D066246:ErbB Receptors; D052216:Glucagon-Like Peptide 1; D000067757:Glucagon-Like Peptide-1 Receptor; D020662:Guanine Nucleotide Exchange Factors; D007328:Insulin; D008565:Membrane Proteins; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D018027:Receptors, Glucagon; D011980:Receptors, Progesterone; D020696:rab5 GTP-Binding Proteins; C000593833:GLP1R protein, human; C000593834:Glp1r protein, mouse; C000593835:Glp1r protein, rat; C508868:PGRMC1 protein, mouse