Naturally-occurring mutation in the calcium-sensing receptor reveals the significance of extracellular domain loop III region for class C G-protein-coupled receptor function.

[familial hypocalciuric hypercalcemia]

Inactivating mutations of the calcium-sensing receptor (CaSR ) cause familial hypocalciuric hypercalcemia and neonatal severe hyperparathyroidism . Most mutations are clustered in the N-terminal and Cys-rich regions of the extracellular domain (ECD ) and seven -transmembrane domain . Disease-causing mutations are uncommon in the C terminus of ECD .T he aim of the study was to characterize the CaSR mutations causing neonatal severe hyperparathyroidism in a consanguineous family .Parathyroid glands from the index patient were stained for CaSR protein . The CaSR gene was sequenced , mutations were recreated in CaSR cDNA , and HEK 293 cells were transfected with the CaSR mutant expression vector . Cellular CaSR targeting was detected by immunoblotting and immunocytochemistry ; CaSR activity was assayed by inositol phosphate accumulation , MAPK activation , and single -cell microfluorimetry .Immunocytochemistry showed reduced intracellular CaSR in patient parathyroids . An in -frame homozygous deletion /insertion mutation , c .1031 > 1034 (delACAAinsT ), replaced His 344 -Asn 345 with a single Leu in CaSR loop III . The mutant reduced cell surface expression of CaSR in transfected HEK 293 cells . Inositol phosphate accumulation , MAPK activation , and single -cell microfluorimetry revealed blunted signaling responses of the mutant receptor to changes in extracellular Ca (2 +) concentration .Deletion of His 344 -Asn 345 in the ECD loop III region affects cell surface targeting of CaSR in transfected cells and in affected parathyroid glands . Absence of conserved Asn 345 may interfere with CaSR folding or glycosylation , leading to poor protein targeting to the cell membrane . This loss -of-function mutant indicates that the ECD loop III is required for CaSR activity .

Diseases
Validation

Diseases presenting "c" symptom

adrenomyeloneuropathy

alexander disease

cadasil

coats disease

cohen syndrome

dedifferentiated liposarcoma

epidermolysis bullosa simplex

erythropoietic protoporphyria

familial hypocalciuric hypercalcemia

gm1 gangliosidosis

homocystinuria without methylmalonic aciduria

junctional epidermolysis bullosa

kallmann syndrome

oligodontia

papillon-lefÃ¨vre syndrome

phenylketonuria

pyruvate dehydrogenase deficiency

von hippel-lindau disease

x-linked adrenoleukodystrophy

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