Coupling of Calcium Receptors to Inositol Phosphate and Cyclic AMP Generation in Mammalian Cells and <i>Xenopus laevis</i> Oocytes and Immunodetection of Receptor Protein by Region-Specific Antipeptide Antisera

Chang, Wenhan; Pratt, Stacy; Chen, Tsui‐Hua; Nemeth, Edward F.; Huang, Zhengmin; Shoback, Dolores

doi:10.1359/jbmr.1998.13.4.570

Cited by 89 publications

(89 citation statements)

References 39 publications

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“…Immunoblotting revealed the expected pattern for bCaR expression in HEK-293 cells (20). The strong and faint bands at 140 and 160 kDa, respectively, likely represent glycosylated variants of the receptor.…”

Section: Resultsmentioning

confidence: 80%

“…Signal transduction by tCaR was evaluated by measuring high [Ca 2ϩ ] o -induced activation of phospholipase C (PLC) in transiently transfected human embryonic kidney (HEK)-293 cells (20). HEK-293 cells were grown and transfected with tCaR or bCaR cDNA as previously described except for the use of transfection reagent, which was 40 l/ml Lipofectamine TM 2000 (Invitrogen) in these experiments (20).…”

Section: Methodsmentioning

confidence: 99%

“…HEK-293 cells were grown and transfected with tCaR or bCaR cDNA as previously described except for the use of transfection reagent, which was 40 l/ml Lipofectamine TM 2000 (Invitrogen) in these experiments (20). At 24 h after transfection, cells were washed, plated into 6-well culture plates, and allowed to attach and grow for 24 -48 Subsequently, the ability of tCaR to activate the mitogen-activated protein (MAP) kinase signaling cascade was assessed by examining phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK1/2).…”

Section: Methodsmentioning

confidence: 99%

“…Cells in the low (50 mM) and high (200 mM) NaCl treatment groups were exposed to varying [Ca 2ϩ ] o (0. (20). Briefly, proteins (5 g for bCaR; 85 g for tCaR) were electrophoresed on 12.5% SDS-PAGE gels and transferred to Immobilon-P membranes (Millipore, Billerica, MA).…”

Section: Methodsmentioning

confidence: 99%

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cDNA Cloning and Functional Expression of a Ca2+-sensing Receptor with Truncated C-terminal Tail from the Mozambique Tilapia (Oreochromis mossambicus)

Loretz¹,

Pollina

Hyodo

et al. 2004

Journal of Biological Chemistry

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The complete cDNA sequence of the tilapia extracellular Ca 2؉ -sensing receptor (CaR) was determined. The transcript length of tilapia CaR (tCaR) is 3.4 kbp and encodes a 940-amino acid, 7-transmembrane domain protein that is consistent in its structural features with known mammalian and piscine CaRs. The tCaR extracellular domain includes a characteristic hydrophobic segment, conserved cysteine residues that are implicated in receptor dimerization (Cys 129 and Cys 131 ) and in coupling to the transmembrane domain (nine conserved cysteine residues), and conserved serine residues (Ser 147 and The roles for ionic calcium (Ca 2ϩ ) in physiological functions are many and varied and essential for survival. Ca 2ϩ -dependent cellular processes include hormone secretion (stimulussecretion coupling), excitability and cell motility (excitationcontraction coupling), and neurotransmission (Ca 2ϩ -based action potentials). This critical involvement of Ca 2ϩ in these and other cellular functions is unique among ions. Ca 2ϩ is simultaneously a controlled variable in physiological systems, an extracellular messenger in multicellular organisms, and an intracellular mediator in a variety of effector cells.Unicellular organisms acquire Ca 2ϩ from the external environment; they have little control over the composition of the surrounding medium and thus had to evolve plasma membrane-bound and other cellular mechanisms to realize cellular Ca 2ϩ homeostasis eventually permitting some degree of habitat survival and selection. In multicellular organisms, active regulation by integumentary tissues of the composition of an "extracellular fluid," which bathes all cells, assures a constant internal milieu and contributes to normal Ca 2ϩ -dependent cell functions. In terrestrial vertebrates, Ca 2ϩ homeostasis depends upon appropriate dietary intake, renal output, and osseous storage and mobilization of Ca 2ϩ . These processes are well known and are regulated by parathyroid hormone, calcitonin, and 1,25-dihydroxyvitamin D acting on kidney, intestine, and bone. Activation of the Ca 2ϩ -sensing receptor (CaR) 1 by its natural ligand, extracellular Ca 2ϩ , alters parathyroid hormone and calcitonin secretion and inhibits renal 1-hydroxylase to retard the synthesis of 1,25-dihydroxyvitamin D (1, 2). Ca 2ϩ balance is achieved through regulated calciotropic hormone secretion in response to the extracellular Ca 2ϩ concentration ([Ca 2ϩ ] o ); the secreted hormones, in turn, control the activity of Ca 2ϩ -transporting tissues to bring about alterations in [Ca 2ϩ ] o . Interestingly, in fishes, the calcium homeostatic process involves ion-transporting tissues not represented in other vertebrate classes (viz., the mitochondria-rich "chloride cells" of the gills and skin) and a different panel of hormones (prolactin and somatolactin from the pituitary, and stanniocalcin from the piscine corpuscles of Stannius (3)). Opportunities for cutaneous

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Section: Resultsmentioning

confidence: 80%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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cDNA Cloning and Functional Expression of a Ca2+-sensing Receptor with Truncated C-terminal Tail from the Mozambique Tilapia (Oreochromis mossambicus)

Loretz¹,

Pollina

Hyodo

et al. 2004

Journal of Biological Chemistry

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“…(GPCR) superfamily and couple to PLC activation, inhibition of cyclic AMP formation, and opening of nonselective cation channels (1)(2)(3)(4). CaRs share modest sequence homology with the metabotropic glutamate receptors (mGluRs) (5,6), the type B ␥-aminobutyric acid receptor (GABA B ) (7), and a large group of pheromone receptors (8,9) and, thus, constitute the family 3 of GPCRs (10).…”

mentioning

confidence: 99%

Amino Acids in the Second and Third Intracellular Loops of the Parathyroid Ca2+-sensing Receptor Mediate Efficient Coupling to Phospholipase C

Chang

Chen

Pratt

et al. 2000

Journal of Biological Chemistry

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To determine the role of amino acids in the second and third intracellular (IC) loops of the Ca 2؉ -sensing receptor (CaR) in phospholipase C (PLC) activation, we mutated residues in these loops either singly or in tandem to Ala and assessed PLC activity by measuring high extracellular [ (7), and a large group of pheromone receptors (8, 9) and, thus, constitute the family 3 of GPCRs (10). Receptors in the CaR/mGluR subfamily share several structural features. These include a large extracellular amino-terminal domain, seven membrane-spanning regions, three IC loops, and a large cytoplasmic tail (see Fig. 1a). The extracellular domains of CaRs and mGluRs are known to be critical for ligand recognition (11-13). Naturally occurring mutants of the CaR, implicated in the pathogenesis of abnormal Ca 2ϩ -sensing in vivo, occur predominantly in the large amino-terminal domain of this receptor (14,15). Point mutations in this domain, responsible for either gain-of-function or loss-of-function, indicate its key role in the Ca 2ϩ -sensing function of the receptor. IC domains of receptors in the CaR/mGluR subfamily are likely, by analogy to other GPCRs, to be responsible for coupling to G-protein-mediated signal transduction (10,16,17). A comparison of CaRs with the mGluR 1-8 indicates limited sequence conservation in their second IC loops (Ͻ10%) but striking conservation (67 to 85%) in their third IC loops (see Fig. 1b). This observation suggests these latter regions likely share similar function.Mutagenesis of mGluR1 and R5 previously demonstrated that specific residues in IC loops 2 and 3 contribute to PLC activation, whereas other residues were involved in the regulation of cyclic AMP formation (5). Domains of comparable functional significance in the CaR have, to date, not been identified. Studies of kindred with familial benign hypercalcemia and neonatal severe hyperparathyroidism indicated that a CaR with a mutation at residue 795 (R795W) in the aminoterminal portion of the third IC loop had a reduced ability to mobilize intracellular Ca 2ϩ (18). The remaining residues within the second and third IC loops have not been carefully examined. In these studies, we mutated amino acids in IC loops 2 and 3 of the bovine CaR to identify the positions of key signaling residues and structural requirements at those sites. Phe-707 in the second IC loop and 2 residues in the third IC loop, Leu-798 and Phe-802, proved critical to the activation of PLC. Glu-804 proved essential for efficient cell surface expression of CaRs. This work supports the presence of several functional determinants in IC loops 2 and 3 in the stimulation of PLC by and expression of CaRs in mammalian cells.

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Magnesium Homeostasis

Khan¹,

Sbayi²,

Schlingmann³

2018

Primer on the Metabolic Bone Diseases and Disorders of Mineral Metabolism

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Coupling of Calcium Receptors to Inositol Phosphate and Cyclic AMP Generation in Mammalian Cells and Xenopus laevis Oocytes and Immunodetection of Receptor Protein by Region-Specific Antipeptide Antisera

Cited by 89 publications

References 39 publications

cDNA Cloning and Functional Expression of a Ca2+-sensing Receptor with Truncated C-terminal Tail from the Mozambique Tilapia (Oreochromis mossambicus)

cDNA Cloning and Functional Expression of a Ca2+-sensing Receptor with Truncated C-terminal Tail from the Mozambique Tilapia (Oreochromis mossambicus)

Amino Acids in the Second and Third Intracellular Loops of the Parathyroid Ca2+-sensing Receptor Mediate Efficient Coupling to Phospholipase C

Magnesium Homeostasis

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