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J Biol Chem, Vol. 275, Issue 5, 3256-3263, February 4, 2000

Sensing of Extracellular Cations in CasR-deficient Osteoblasts
EVIDENCE FOR A NOVEL CATION-SENSING MECHANISM^*

Min Pi, Sanford C. Garner, Patrick Flannery, Robert F. Spurney, and L. Darryl Quarles

From the Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710

We isolated osteoblastic cell lines from wild-type (CasR^+/+) and receptor null (CasR^/) mice to investigate whether CasR is present in osteoblasts and accounts for their responses to extracellular cations. Osteoblasts from both CasR^+/+ and CasR^/ mice displayed an initial period of cell replication followed by a culture duration-dependent increase in alkaline phosphatase activity, expression of osteocalcin, and mineralization of extracellular matrix. In addition, a panel of extracellular cations, including aluminum and the CasR agonists gadolinium and calcium, stimulated DNA synthesis, activated a transfected serum response element-luciferase reporter construct, and inhibited agonist-induced cAMP in CasR^/ osteoblasts. The functional responses to these cations were identical in CasR^+/+ and CasR^/ osteoblasts. Thus, the absence of CasR alters neither the maturational profile of isolated osteoblast cultures nor their in vitro responses to extracellular cations. In addition, CasR transcripts could not be detected by reverse transcription-polymerase chain reaction with mouse specific primers in either CasR^+/+ or CasR^/ osteoblasts, and immunoblot analysis with a CasR-specific antibody was negative for CasR protein expression in osteoblasts. The presence of a cation-sensing response in osteoblasts from CasR^/ mice indicates the existence of a novel osteoblastic extracellular cation-sensing mechanism.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank^TM/EMBL Data Bank with accession number(s) AF128842.

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