Expression of calcium sensing receptor and receptor activity modifying proteins in cells involved in calcium homeostasis and fluorescence resonance energy transfer based stoichiometric analysis of their interactions

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Event Abstract Back to Event Expression of calcium sensing receptor and receptor activity modifying proteins in cells involved in calcium homeostasis and fluorescence resonance energy transfer based stoichiometric analysis of their interactions AJ Desai1*, DJ Roberts1, GO Richards1 and TM Skerry1 1 University of Sheffield, Department of Human Metabolism, United Kingdom {BR}The G-protein coupled receptor known as the Calcium Sensing Receptor (CaSR) is known to play an important role in calcium homeostasis by sensing changes in extracellular calcium and modulating secretion of calcitropic hormones. A recent study revealed that cell surface expression of the CaSR requires its association with single transmembrane accessory protein called Receptor Activity Modifying Protein (RAMP), specifically either RAMP1 or 3.{BR}We hypothesize that efficiency of interaction and co-localization of RAMPs with CaSR could be determined using stoichiometric Fluorescence Resonance Energy Transfer (FRET) analysis. {BR}In transfected COS7 cells, CaSR+RAMP1/3 complexes are highly co-localized around the peri-nuclear region indicative of co-localization in the endoplasmic reticulum and the Golgi bodies and also in regions of the cell membrane. The CaSR+RAMP3 FRET complex is ~25% more efficient than CaSR+RAMP1 FRET complex. Furthermore, the fraction of RAMP3 involved in FRET is ~18.8% more than RAMP1 whereas the fraction of CaSR involved in FRET with RAMP3 is ~33% more than with RAMP1. {BR}To determine whether RAMP interactions with CaSR in cells involved in calcium homeostasis had roles beyond trafficking, we explored their expression in thyroid and bone cell lines.{BR}We detected the expression of mRNA and protein for CaSR, RAMP1 and 2 in thyroid medullary carcinoma cell line (TT cells) using quantitative PCR, immunocytochemistry and western blotting. However, we could not detect expression of CaSR and RAMP3 mRNA transcripts in pre-osteoblastic MG63, SAOS-2 and TE85 cell lines using quantitative PCR. A high dose of extracellular calcium and differentiation of these cells into mature osteoblasts did not induce expression of CaSR or RAMP3. This suggests that osteosarcoma cell lines are not ideal models of osteoblasts to study CaSR expression and function.{BR}The novel finding of the current study is that it provides a stoichiometric insight into CaSR-RAMP interaction and also indicates that CaSR interacts more efficiently with RAMP3 efficiently than RAMP1. This data is a step forward from the existing knowledge of CaSR-RAMP interaction. The increased understanding of their interaction will clarify their role in physiology and patho-physiology. Keywords: Bones, Bone Research Conference: 2011 joint meeting of the Bone Research Society & the British Orthopaedic Research Society, Cambridge, United Kingdom, 27 Jun - 29 Jun, 2011. Presentation Type: Poster Topic: Abstracts Citation: Desai A, Roberts D, Richards G and Skerry T (2011). Expression of calcium sensing receptor and receptor activity modifying proteins in cells involved in calcium homeostasis and fluorescence resonance energy transfer based stoichiometric analysis of their interactions. Front. Endocrinol. Conference Abstract: 2011 joint meeting of the Bone Research Society & the British Orthopaedic Research Society. doi: 10.3389/conf.fendo.2011.02.00013 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 30 Sep 2011; Published Online: 30 Sep 2011. * Correspondence: Mr. AJ Desai, University of Sheffield, Department of Human Metabolism, United Kingdom, a.desai@sheffield.ac.uk Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers AJ Desai DJ Roberts GO Richards TM Skerry Google AJ Desai DJ Roberts GO Richards TM Skerry Google Scholar AJ Desai DJ Roberts GO Richards TM Skerry PubMed AJ Desai DJ Roberts GO Richards TM Skerry Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.