A novel mutation of the primary protein kinase C phosphorylation site in the calcium-sensing receptor causes autosomal dominant hypocalcemia.
OBJECTIVE
The calcium-sensing receptor (CASR) is a key controller of calcium homeostasis by regulating parathyroid hormone (PTH) secretion and renal calcium reabsorption. CASR(T888) is a protein kinase C (PKC) phosphorylation site in the receptor's intracellular domain that has previously been identified as a critical negative regulator of CASR downstream signaling in vitro, but whose importance in vivo is unknown.


METHODS
The proband presented with mild symptomatic hypocalcemia following treatment for nephrotic syndrome due to minimal change glomerulonephropathy. Laboratory tests revealed inappropriately normal PTH concentrations and relative hypercalciuria typical of autosomal dominant hypocalcemia. His asymptomatic father had similar laboratory test results.


METHODS
The CASR gene was sequenced. To investigate the molecular consequences of CASR(T888M) mutation, site-directed mutagenesis was used to modify the wild-type (wt)-CASR gene, with the resulting mutant being transfected transiently into HEK-293 cells.


RESULTS
A novel CASR missense mutation, T888M, was identified in both cases. The CASR(T888M) mutant exhibited enhanced sensitivity to extracellular calcium concentration, both for intracellular calcium (Ca(2+)(i)) mobilization and for ERK phosphorylation, despite having unaltered levels of cell surface expression. Furthermore, CASR(T888M) elicited sustained Ca(2+)(i) mobilization rather than high frequency Ca(2+)(i) oscillations, and, unlike the wt-CASR, the response was resistant to acute inhibition by the PKC activator, phorbol 12-myristate 13-acetate.


CONCLUSIONS
The clinical and functional data provide the first genotype-phenotype correlation for a mutation at T888, indicating its critical physiological importance in CASR signaling. Thus, CASR(T888) represents a functionally important, inhibitory phosphorylation site that contributes to the control of PTH secretion.
Department of Endocrinology, Royal Brisbane and Women's Hospital, Herston, Queensland 4029, Australia.
Syndia
S
Lazarus
Carel J
CJ
Pretorius
Frederick
F
Khafagi
Katherine L
KL
Campion
Sarah C
SC
Brennan
Arthur D
AD
Conigrave
Edward M
EM
Brown
Donald T
DT
Ward
D000328:Adult; D006801:Humans; D006996:Hypocalcemia / Q000235:genetics*; D008297:Male; D009154:Mutation; D010766:Phosphorylation / Q000235:genetics; D011493:Protein Kinase C / Q000378:metabolism*; D044169:Receptors, Calcium-Sensing / Q000235:genetics*; D055815:Young Adult
D044169:Receptors, Calcium-Sensing; D011493:Protein Kinase C