Exp Mol Med.  2005 Apr;37(2):91-100.

Calcium sensing receptor forms complex with and is up-regulated by caveolin-1 in cultured human osteosarcoma (Saos-2) cells

Affiliations
  • 1Department of Physiology and Biophysics, College of Medicine Inha University, Incheon, Korea.
  • 2Department of Pharmacology and Toxicology, College of Medicine Inha University, Incheon, Korea. shcha@inha.ac.kr
  • 3Department of Dentistry, College of Medicine Inha University, Incheon, Korea.
  • 4Hormone Research Center, School of Biological Sciences and Technology, Chonnam National University, 300 Yongbong-dong, Buk-gu, Gwangju, Korea.
  • 5Kidney Disease Research Group in Inha, Research Institute for Medical Sciences, Korea.

Abstract

The calcium sensing receptor (CaSR) plays an important role for sensing local changes in the extracellular calcium concentration ([Ca2+]o) in bone remodeling. Although the function of CaSR is known, the regulatory mechanism of CaSR remains controversial. We report here the regulatory effect of caveolin on CaSR function as a process of CaSR regulation by using the human osteosarcoma cell line (Saos-2). The intracellular calcium concentration ([Ca2+]i) was increased by an increment of [Ca2+]o. This [Ca2+]i increment was inhibited by the pretreatment with NPS 2390, an antagonist of CaSR. RT-PCR and Western blot analysis of Saos-2 cells revealed the presence of CaSR, caveolin (Cav)-1 and -2 in both mRNA and protein expressions, but there was no expression of Cav-3 mRNA and protein in the cells. In the isolated caveolae-rich membrane fraction from Saos-2 cells, the CaSR, Cav-1 and Cav-2 proteins were localized in same fractions (fraction number 4 and 5). The immuno-precipitation experiment using the respective antibodies showed complex formation between the CaSR and Cav-1, but no complex formation of CaSR and Cav-2. Confocal microscopy also supported the co-localization of CaSR and Cav-1 at the plasma membrane. Functionally, the [Ca2+]o- induced [Ca2+]i increment was attenuated by the introduction of Cav-1 antisense oligodeoxynucleotide (ODN). From these results, in Saos-2 cells, the function of CaSR might be regulated by binding with Cav-1. Considering the decrement of CaSR activity by antisense ODN, Cav-1 up-regulates the function of CaSR under normal physiological conditions, and it may play an important role in the diverse pathophysiological processes of bone remodeling or in the CaSR- related disorders in the body.

Keyword

antisense oligodeoxynucleotides; calcium sensing receptor; caveolae; caveolin-1; confocal microscopy; osteosarcoma cell line
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