Orphan Adhesion GPCR GPR64/ADGRG2 Is Overexpressed in Parathyroid Tumors and Attenuates Calcium‐Sensing Receptor‐Mediated Signaling

• Published in: Journal of bone and mineral research Vol. 32; no. 3; pp. 654 - 666
• Main Authors: Balenga, Nariman, Azimzadeh, Pedram, Hogue, Joyce A, Staats, Paul N, Shi, Yuhong, Koh, James, Dressman, Holly, Olson, John A
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01.03.2017
• Subjects: Adenoma - metabolism; Adenoma - pathology; Adenosine kinase; Amino acids; AMP; Calcium (blood); Calcium (intracellular); Calcium signalling; Calcium-sensing receptors; Cell Separation; Cell surface; CELL/TISSUE SIGNALING; Cyclic AMP - metabolism; Cyclic AMP response element-binding protein; Cyclic AMP Response Element-Binding Protein - metabolism; Cyclic AMP-Dependent Protein Kinases - metabolism; Cyclin D1; DISORDERS OF CALCIUM/PHOSPHATE METABOLISM; G protein-coupled receptors; Gene expression; HEK293 Cells; HORMONE REPLACEMENT/RECEPTOR MODULATORS; Humans; Hyperparathyroidism; Hyperparathyroidism, Primary - pathology; Immunohistochemistry; Intracellular signalling; Kinases; Neuroendocrine tumors; Parathyroid hormone; Parathyroid Neoplasms - metabolism; Parathyroid Neoplasms - pathology; Phosphates; Protein Binding; Protein kinase A; Proteins; Proteolysis; PTH/VIT D/FGF23; Receptors, Calcium-Sensing - metabolism; Receptors, G-Protein-Coupled - metabolism; Reporter gene; Retinoblastoma; Signal Transduction; TRANSCRIPTION FACTORS; Tumors; Up-Regulation; Western blotting; CELL/TISSUE SIGNALING; TRANSCRIPTION FACTORS; PTH/VIT D/FGF23; DISORDERS OF CALCIUM/PHOSPHATE METABOLISM; HORMONE REPLACEMENT/RECEPTOR MODULATORS
• ISSN: 0884-0431; 1523-4681
• DOI: 10.1002/jbmr.3023

ABSTRACT Abnormal feedback of serum calcium to parathyroid hormone (PTH) secretion is the hallmark of primary hyperparathyroidism (PHPT). Although the molecular pathogenesis of parathyroid neoplasia in PHPT has been linked to abnormal expression of genes involved in cell growth (e.g., cyclin D1, retinoblastoma, and β‐catenin), the molecular basis of abnormal calcium sensing by calcium‐sensing receptor (CaSR) and PTH hypersecretion in PHPT are incompletely understood. Through gene expression profiling, we discovered that an orphan adhesion G protein‐coupled receptor (GPCR), GPR64/ADGRG2, is expressed in human normal parathyroid glands and is overexpressed in parathyroid tumors from patients with PHPT. Using immunohistochemistry, Western blotting, and coimmunoprecipitation, we found that GPR64 is expressed on the cell surface of parathyroid cells, is overexpressed in parathyroid tumors, and physically interacts with the CaSR. By using reporter gene assay and GPCR second messenger readouts we identified Gαs, 3′,5′‐cyclic adenosine monophosphate (cAMP), protein kinase A, and cAMP response element binding protein (CREB) as the signaling cascade downstream of GPR64. Furthermore, we found that an N‐terminally truncated human GPR64 is constitutively active and a 15–amino acid–long peptide C‐terminal to the GPCR proteolysis site (GPS) of GPR64 activates this receptor. Functional characterization of GPR64 demonstrated its ability to increase PTH release from human parathyroid cells at a range of calcium concentrations. We discovered that the truncated constitutively active, but not the full‐length GPR64 physically interacts with CaSR and attenuates the CaSR‐mediated intracellular Ca2+ signaling and cAMP suppression in HEK293 cells. Our results indicate that GPR64 may be a physiologic regulator of PTH release that is dysregulated in parathyroid tumors, and suggest a role for GPR64 in pathologic calcium sensing in PHPT. © 2016 American Society for Bone and Mineral Research.