Compelling evidence of a cell surface receptor sensitive to extracellular calcium was observed as early as the 1980s and was finally realized in 1993 when the calcium-sensing receptor (CaR) was cloned from bovine parathyroid tissue. Initial studies relating to the CaR focused on its key role in extracellular calcium homeostasis, but as the amount of information about the receptor grew it became evident that it was involved in many biological processes unrelated to calcium homeostasis. The CaR responds to a diverse array of stimuli extending well beyond that merely of calcium, and these stimuli can lead to the initiation of a wide variety of intracellular signaling pathways that in turn are able to regulate a diverse range of biological processes. It has been through the examination of the molecular characteristics of the CaR that we now have an understanding of how this single receptor is able to convert extracellular messages into specific cellular responses. Recent CaR-related reviews have focused on specific aspects of the receptor, generally in the context of the CaR's role in physiology and pathophysiology. This review will provide a comprehensive exploration of the different aspects of the receptor, including its structure, stimuli, signalling, interacting protein partners, and tissue expression patterns, and will relate their impact on the functionality of the CaR from a molecular perspective.

The C-terminal domain of Hsp90 displays independent chaperone activity, mediates dimerization, and contains the MEEVD motif essential for interaction with tetratricopeptide repeat-containing immunophilin cochaperones assembled in mature steroid receptor complexes. An ␣-helical region, upstream of the MEEVD peptide, helps form the dimerization interface and includes a hydrophobic microdomain that contributes to the Hsp90 interaction with the immunophilin cochaperones and corresponds to the binding site for novobiocin, a coumarin-related Hsp90 inhibitor. Mutation of selected residues within the hydrophobic microdomain significantly impacted the chaperone function of a recombinant C-terminal Hsp90 fragment and novobiocin inhibited wild-type chaperone activity. Prior incubation of the Hsp90 fragment with novobiocin led to a direct blockade of immunophilin cochaperone binding. However, the drug had little influence on the pre-formed Hsp90-immunophilin complex, suggesting that bound cochaperones mask the novobiocin-binding site. We observed a differential effect of the drug on Hsp90-immunophilin interaction, suggesting that the immunophilins make distinct contacts within the C-terminal domain to specifically modulate Hsp90 function. Novobiocin also precluded the interaction of full-length Hsp90 with the p50 There is now increasing evidence that receptor function is critically dependent on the selection of immunophilin within steroid receptor complexes (2-4). This may be governed, in part, by the selective preference of receptors for specific immunophilins. PP5 has been reported to have a modulating role in glucocorticoid signaling (5), and there are strong indications that FKBP51 inhibits glucocorticoid receptor function. Elevated expression of FKBP51, resulting in greatly increased incorporation of FKBP51 into glucocorticoid receptor complexes, reduces hormone-binding affinity and promotes glucocorticoid resistance in primates (6, 7). FKBP51 also sequesters glucocorticoid receptor within the cytoplasm (8, 9), but hormone binding induces a functional exchange of FKBP52 for FKBP51 in receptor complexes allowing translocation of the complex to the nucleus (8). In a yeast model, FKBP52 was shown to dramatically potentiate glucocorticoid-dependent reporter gene activity through a mechanism that results in increased receptor hormone-binding affinity (10). Consistent with previous findings, coexpression of FKBP51 blocked the potentiating effects of FKBP52. These potentiating properties require FKBP52 catalytic activity as well as a functional interaction of the immunophilin with Hsp90. Receptor function, then, can be directly influenced by the prolyl isomerase activity of a TPR immunophilin. The Smith laboratory has now extended the study of FKBP52 function to a FKBP52 knock-out mouse model (11-13). Male mice exhibit many features in common with partial androgen insensitivity, reflecting loss of FKBP52-mediated potentiation of androgen receptor function (11). Female mice display a maternal defect linked to progesterone ins...

The immunophilin cochaperones, cyclophilin 40 (CyP40), FKBP51 and FKBP52 and PP5, a serine/threonine protein phosphatase, have been implicated as modulators of steroid receptor function through their association with Hsp90, a molecular chaperone with a key role in steroid hormone signalling. Although progress towards a satisfying definition for the role of these components in steroid receptor complexes has been slow, recent developments arising from novel approaches in both yeast and mammalian systems, together with available crystal structures for Hsp90 and some of these cochaperones, are beginning to provide important clues about their function. Hsp90, recently identified as a member of the GHKL superfamily of ATPases, is the central player in receptor assembly, an energy-driven process that allows receptor and the immunophilins to be proximally located, or to interact directly, on a Hsp90 scaffold. Immunophilin structure, relative abundance, their binding affinity for Hsp90 and their ability to interact with specific receptors may all contribute to a selective preference of the immunophilins for individual receptors. Association of receptors with different immunophilins leads to differential functional consequences for receptor activity. Observations of glucocorticoid resistance in New World primates, attributed to FKBP51 overexpression and incorporation into glucocorticoid receptor complexes, have provided the first evidence that these cochaperones can control hormone-binding affinity. Application of a yeast model to FKBP52 function in the glucocorticoid receptor system has now provided crucial evidence that this immunophilin enhances receptor transcriptional activity by increasing receptor avidity for hormone through PPIase-mediated conformational changes in the ligand-binding domain. A recent novel finding suggests that hormone binding may induce a functional exchange of immunophilins in receptor complexes and that the modified complex directs receptor to the nucleus.

Sequestosome 1/p62 (p62) mutations are associated with PDB; however, there are limited data regarding functional consequences. We report a novel mutation in exon 7 (K378X) in a patient with polyostotic Paget's disease of bone. p62 mutants increased NF-B activation and significantly potentiated osteoclast formation and bone resorption in human primary cell cultures.Introduction: Sequestosome 1/p62 (p62) mutations are associated with Paget's disease of bone (PDB); however, there are limited data regarding functional consequences. One report has linked the common P392L mutation in the p62 ubiquitin binding associated (UBA) domain with increases in NF-B activity, a transcription factor essential for osteoclastogenesis. To further clarify the functional impact of p62 mutations associated with PDB, we assessed the effect of p62 mutation (a novel mutation: K378X, and previously reported mutations: P392L and E396X) on RANK-induced NF-B activation and compared this with the effect of wildtype p62. In addition, we studied the effect of p62 mutation on osteoclast formation and bone resorption. Materials and Methods:We performed co-transfection experiments with expression plasmids for p62 (wildtype or mutated) and RANK and an NF-B luciferase reporter gene. Luciferase activities were recorded after addition of luciferin to cellular lysates. RAW 264.7 cells stably expressing enhanced green fluorescent protein (EGFP)-tagged p62 (wildtype, K378X, or P392L) or EGFP alone were assessed for changes in cell proliferation. Additionally, these cells were stimulated with RANKL to produce osteoclast-like cells (OLCs). Primary human monocytes collected from the K378X-affected patient and a control subject were stimulated to form OLCs and bone resorption data were obtained. Results:The novel mutation introduces a premature stop codon in place of Lys-378 and thereby eliminates the entire p62 UBA domain; this and two additional natural mutations (P392L, E396X) increased NF-B activation compared with wildtype p62. Wildtype p62 consistently inhibited NF-B activation compared with empty vector. UBA mutations (K378X and P392L) significantly increased the number of OLCs formed in response to RANKL and also the number of nuclei of the OLCs. K378X-affected human monocytes formed more OLCs with more nuclei and increased bone resorption compared with control monocytes. Conclusions: Our data show that mutation of the p62 UBA domain results in increased activation of NF-B and osteoclast formation and function compared with wildtype p62. These results may partially explain the mechanism by which p62 mutation contributes to the pathogenesis of PDB.