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 ?-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 ?-cells (Gprc6a<
sup>
?-cell-cko<
/sup>
) by crossing Gprc6a<
sup>
flox/flox<
/sup>
mice with Ins2-Cre mice resulted in reduced pancreatic weight, islet number, insulin protein content, and insulin message expression. Both islet size and ?-cell proliferation were reduced in Gprc6a<
sup>
?-cell-cko<
/sup>
compared with control mice. Gprc6a<
sup>
?-cell-cko<
/sup>
exhibited abnormal glucose tolerance, but normal insulin sensitivity. Islets isolated from Gprc6a<
sup>
?-cell-cko<
/sup>
mice showed reduced insulin simulation index in response to Ocn. Here, these data establish the structural basis for Ocn direct activation of GPRC6A and confirm a role for GPRC6A in regulating ?-cell proliferation and insulin secretion.