Identification and functional characterization of loss-of-function mutations of the calcium-sensing receptor in four Italian kindreds with familial hypocalciuric hypercalcemia.

[familial hypocalciuric hypercalcemia]

Identification and characterization of calcium-sensing receptor (CASR ) mutations in four unrelated Italian kindreds with familial hypocalciuric hypercalcemia .Clinical evaluation and genetic analysis of CASR gene . Functional characterization of mutated CASRs .Direct sequencing of CASR gene in genomic DNA . Studies of CASR -mediated increases in cytosolic calcium concentration [Ca (2 )(+)](i ) in CASR -transfected COS -7 cells in vitro .Four unreported heterozygous CASR mutations were identified , including three missense (H 595 Y , P 748 H , and C 7 65 W ) and one splice site (IVS 2 +1 G >C ) mutation . The H 595 Y , P 748 H , and C 7 65 W mutant receptors , although expressed at normal levels on the cell surface , showed a reduced response in [Ca (2 )(+)](i ) relative to the wildtype (WT ) CASR to increasing extracellular calcium concentrations . Cotransfection experiments showed that the H 595 Y and P 748 H mutants did not affect the apparent affinity of the WT CASR for calcium , suggesting that they do not exert a dominant-negative effect . On the other hand , the co -transfected C 7 65 W mutant decreased the maximum response of the WT CASR to calcium , suggesting that it may reduce the effective concentration of the normal CASR on the cell surface or impair its maximal signaling capacity .Four CASR mutations were identified . The reduced functional responses to extracellular calcium and normal expression of the mutant receptors suggest that conformational changes account for altered CASR activity . Moreover , a reduced complement of normal CASRs in these heterozygous patients , perhaps combined with a mutant receptor-induced decrease in maximal activity of the WT receptor , may contribute to defective calcium -sensing in vivo .

Diseases
Validation

Diseases presenting "normal expression of the mutant receptors suggest that conformational changes account for altered casr activity" symptom

familial hypocalciuric hypercalcemia

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