title: Evidence for Osteocalcin Binding and Activation of GPRC6A in β-Cells.
abstract: The possibility that G protein-coupled receptor family C member A (GPRC6A) is the osteocalcin (Ocn)-sensing G protein-coupled receptor that directly regulates pancreatic β-cell functions is controversial. In the current study, we found that Ocn and an Ocn-derived C-terminal hexapeptide directly activate GPRC6A-dependent ERK signaling in vitro. Computational models probe the structural basis of Ocn binding to GPRC6A and predict that the C-terminal hexapeptide docks to the extracellular side of the transmembrane domain of GPRC6A. Consistent with the modeling, mutations in the computationally identified binding pocket of GPRC6A reduced Ocn and C-terminal hexapeptide activation of this receptor. In addition, selective deletion of Gprc6a in β-cells (Gprc6a(β)(-cell-cko)) by crossing Gprc6a(flox/flox) mice with Ins2-Cre mice resulted in reduced pancreatic weight, islet number, insulin protein content, and insulin message expression. Both islet size and β-cell proliferation were reduced in Gprc6a(β)(-cell-cko) compared with control mice. Gprc6a(β)(-cell-cko) exhibited abnormal glucose tolerance, but normal insulin sensitivity. Islets isolated from Gprc6a(β)(-cell-cko) mice showed reduced insulin simulation index in response to Ocn. These data establish the structural basis for Ocn direct activation of GPRC6A and confirm a role for GPRC6A in regulating β-cell proliferation and insulin secretion.
authors.affiliation: Departments of Medicine (M.P., R.Y., L.D.Q.) and Microbiology, Immunology and Biochemistry (S.K.N.), University of Tennessee Health Science Center, Memphis, Tennessee 38163; University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics (K.K., J.C.S., J.B.), Oak Ridge, Tennessee 37830; and Department of Biochemistry and Cellular and Molecular Biology (J.C.S.), University of Tennessee, Knoxville, Tennessee 37996.
authors.forename: Min
authors.initials: M
authors.lastname: Pi
authors.affiliation: Departments of Medicine (M.P., R.Y., L.D.Q.) and Microbiology, Immunology and Biochemistry (S.K.N.), University of Tennessee Health Science Center, Memphis, Tennessee 38163; University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics (K.K., J.C.S., J.B.), Oak Ridge, Tennessee 37830; and Department of Biochemistry and Cellular and Molecular Biology (J.C.S.), University of Tennessee, Knoxville, Tennessee 37996.
authors.forename: Karan
authors.initials: K
authors.lastname: Kapoor
authors.affiliation: Departments of Medicine (M.P., R.Y., L.D.Q.) and Microbiology, Immunology and Biochemistry (S.K.N.), University of Tennessee Health Science Center, Memphis, Tennessee 38163; University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics (K.K., J.C.S., J.B.), Oak Ridge, Tennessee 37830; and Department of Biochemistry and Cellular and Molecular Biology (J.C.S.), University of Tennessee, Knoxville, Tennessee 37996.
authors.forename: Ruisong
authors.initials: R
authors.lastname: Ye
authors.affiliation: Departments of Medicine (M.P., R.Y., L.D.Q.) and Microbiology, Immunology and Biochemistry (S.K.N.), University of Tennessee Health Science Center, Memphis, Tennessee 38163; University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics (K.K., J.C.S., J.B.), Oak Ridge, Tennessee 37830; and Department of Biochemistry and Cellular and Molecular Biology (J.C.S.), University of Tennessee, Knoxville, Tennessee 37996.
authors.forename: Satoru Kenneth
authors.initials: SK
authors.lastname: Nishimoto
authors.affiliation: Departments of Medicine (M.P., R.Y., L.D.Q.) and Microbiology, Immunology and Biochemistry (S.K.N.), University of Tennessee Health Science Center, Memphis, Tennessee 38163; University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics (K.K., J.C.S., J.B.), Oak Ridge, Tennessee 37830; and Department of Biochemistry and Cellular and Molecular Biology (J.C.S.), University of Tennessee, Knoxville, Tennessee 37996.
authors.forename: Jeremy C
authors.initials: JC
authors.lastname: Smith
authors.affiliation: Departments of Medicine (M.P., R.Y., L.D.Q.) and Microbiology, Immunology and Biochemistry (S.K.N.), University of Tennessee Health Science Center, Memphis, Tennessee 38163; University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics (K.K., J.C.S., J.B.), Oak Ridge, Tennessee 37830; and Department of Biochemistry and Cellular and Molecular Biology (J.C.S.), University of Tennessee, Knoxville, Tennessee 37996.
authors.forename: Jerome
authors.initials: J
authors.lastname: Baudry
authors.affiliation: Departments of Medicine (M.P., R.Y., L.D.Q.) and Microbiology, Immunology and Biochemistry (S.K.N.), University of Tennessee Health Science Center, Memphis, Tennessee 38163; University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics (K.K., J.C.S., J.B.), Oak Ridge, Tennessee 37830; and Department of Biochemistry and Cellular and Molecular Biology (J.C.S.), University of Tennessee, Knoxville, Tennessee 37996.
authors.forename: Leigh Darryl
authors.initials: LD
authors.lastname: Quarles
mesh_terms: D000818:Animals; D000242:Cyclic AMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D007328:Insulin / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D015675:Osteocalcin / Q000378:metabolism* / Q000494:pharmacology; D010179:Pancreas / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*
chemical_list: C515750:GPRC6A protein, mouse; D007328:Insulin; D012333:RNA, Messenger; D043562:Receptors, G-Protein-Coupled; D015675:Osteocalcin; D000242:Cyclic AMP