Submission
| Title: |
The Emergence of Location Bias in G protein-coupled receptor Pharmacology and Therapeutics |
| Co-Authors: |
Abhyankar, Surabhi, Department of Biochemistry and Molecular Biology, Indiana University School of Medicine; Hongxia Ren, Center for Diabetes and Metabolic Diseases, Indiana University School of Medicine |
Abstract
Background/Significance/Rationale: G protein-coupled receptors are highly druggable targets for diabetes and obesity treatment. Our lab showed that intestinal G protein-coupled receptor 17 (Gpr17) knock-out in mice improved glucose tolerance and increased glucagon-like peptide 1 (GLP-1) secretion and insulin release, suggesting intestinal GPR17 is a potential target for diabetes and obesity intervention. Therefore, we wanted to elucidate 1) tissue distribution and 2) subcellular localization of GPR17.
Methods: 1) Duodenum, jejunum, ileum, and colon paraffin fixed tissue sections were obtained from four human subjects. Immunohistochemical staining was performed to determine tissue distribution of GPR17 with hormones- Cholecystokinin (CCK), and GLP-1. 2)To investigate subcellular localization of GPR17 to physiological stimuli, HeLa cells were transfected with HA-Gpr17-GFP and cultured either in growth medium (GM) or in serum free medium (SFM) and later stained with antibodies specific for different subcellular organelles.
Results/Findings: 1) >90% of the GPR17+ cells are CCK positive in duodenum and jejunum, and ~59% of the GPR17+ cells are GLP-1 positive in ileum. 2)Confocal microscopy revealed that GPR17 is not only localized to the plasma membrane but also to the intracellular membranes. Pearson’s correlation coefficient showed majority of Gpr17 is localized to the Endoplasmic-Reticulum (ER) (p<0.0001) when cells were grown in GM.
Conclusions/Discussion: 1) GPR17 overlaps with CCK in the proximal intestine and with GLP-1 in the distal intestine. 2)The majority of Gpr17 is localized to the ER and, serum in the growth medium increased Gpr17 localization to the ER, which suggests presence of the endogenous ligand in the serum and/or activation by kinases that stimulated internalization.
Translational/Human Health Impact: This study is expected to contribute to our understanding of how GPR17 signaling affects metabolism from various subcellular locations. This will help us to find potential drugs to inhibit GPR17 to improve metabolism and leveraging this for the treatment of diabetes and obesity.
