Calcium-sensing Receptor In Cells Research Articles

The mTORC2 Regulator Homer1 Modulates Protein Levels and Sub-Cellular Localization of the CaSR in Osteoblast-Lineage Cells.

We recently found that, in human osteoblasts, Homer1 complexes to Calcium-sensing receptor (CaSR) and mediates AKT initiation via mechanistic target of rapamycin complex (mTOR) complex 2 (mTORC2) leading to beneficial effects in osteoblasts including β-catenin stabilization and mTOR complex 1 (mTORC1) activation. Herein we further investigated the relationship between Homer1 and CaSR and demonstrate a link between the protein levels of CaSR and Homer1 in human osteoblasts in primary culture. Thus, when siRNA was used to suppress the CaSR, we observed upregulated Homer1 levels, and when siRNA was used to suppress Homer1 we observed downregulated CaSR protein levels using immunofluorescence staining of cultured osteoblasts as well as Western blot analyses of cell protein extracts. This finding was confirmed in vivo as the bone cells from osteoblast specific CaSR−/− mice showed increased Homer1 expression compared to wild-type (wt). CaSR and Homer1 protein were both expressed in osteocytes embedded in the long bones of wt mice, and immunofluorescent studies of these cells revealed that Homer1 protein sub-cellular localization was markedly altered in the osteocytes of CaSR−/− mice compared to wt. The study identifies additional roles for Homer1 in the control of the protein level and subcellular localization of CaSR in cells of the osteoblast lineage, in addition to its established role of mTORC2 activation downstream of the receptor.

The Calcium-Sensing Receptor and the Reproductive System.

Active placental transport of maternal serum calcium (Ca2+) to the offspring is pivotal for proper development of the fetal skeleton as well as various organ systems. Moreover, extracellular Ca2+ levels impact on distinct processes in mammalian reproduction. The calcium-sensing receptor (CaSR) translates changes in extracellular Ca2+-concentrations into cellular reactions. This review summarizes current knowledge on the expression of CaSR and its putative functions in reproductive organs. CaSR was detected in placental cells mediating materno-fetal Ca2+-transport such as the murine intraplacental yolk sac (IPYS) and the human syncytiotrophoblast. As shown in casr knock-out mice, ablation of CaSR downregulates transplacental Ca2+-transport. Receptor expression was reported in human and rat ovarian surface epithelial (ROSE) cells, where CaSR activation stimulates cell proliferation. In follicles of various species a role of CaSR activation in oocyte maturation was suggested. Based on studies in avian follicles, the activation of CaSR expressed in granulosa cells may support the survival of follicles after their selection. CaSR in rat and equine sperms was functionally linked to sperm motility and sperm capacitation. Implantation involves complex interactions between the blastocyst and the uterine epithelium. During early pregnancy, CaSR expression at the implantation site as well as in decidual cells indicates that CaSR is important for blastocyst implantation and decidualization in the rat uterus. Localization of CaSR in human extravillous cytotrophoblasts suggests a role of CaSR in placentation. Overall, evidence for functional involvement of CaSR in physiologic mammalian reproductive processes exists. Moreover, several studies reported altered expression of CaSR in cells of reproductive tissues under pathologic conditions. However, in many tissues we still lack knowledge on physiological ligands activating CaSR, CaSR-linked G-proteins, activated intracellular signaling pathway, and functional relevance of CaSR activation. Clearly, more work is required in the future to decode the complex physiologic and pathophysiologic relationship of CaSR and the mammalian reproductive system.

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

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.