Novel NCC mutants and functional analysis in a new cohort of patients with Gitelman syndrome

Abstract

Gitelman syndrome (GS) is an autosomal recessive disorder characterized by hypokalemic metabolic alkalosis in conjunction with significant hypomagnesemia and hypocalciuria. The GS phenotype is caused by mutations in the solute carrier family 12, member 3 (SLC12A3) gene that encodes the thiazide-sensitive NaCl cotransporter (NCC). We analyzed DNA samples of 163 patients with a clinical suspicion of GS by direct sequencing of all 26 exons of the SLC12A3 gene. In total, 114 different mutations were identified, 31 of which have not been reported before. These novel variants include 3 deletions, 18 missense, 6 splice site and 4 nonsense mutations. We selected seven missense mutations to investigate their effect on NCC activity and plasma membrane localization by using the Xenopus laevis oocyte expression system. The Thr392Ile mutant did not display transport activity (probably class 2 mutation), while the Asn442Ser and Gln1030Arg NCC mutants showed decreased plasma membrane localization and consequently function, likely due to impaired trafficking (class 3 mutation). Even though the NaCl uptake was hampered for NCC mutants Glu121Asp, Pro751Leu, Ser475Cys and Tyr489His, the transporters reached the plasma membrane (class 4 mutation), suggesting an effect on NCC regulation or ion affinity. The present study shows the identification of 38 novel mutations in the SLC12A3 gene and provides insight into the mechanisms that regulate NCC.

Gitelman’s Syndrome Associated with Chondrocalcinosis: A Case Study from the Azores Islands (Portugal)

“…The assumption that GS is caused by a defect in the NCCT cotransporter in the renal distal tubule has been proven by the identification of numerous variations in the SLC12A3 gene in patients with GS [1,4,19,21]. In the present study the specific involvement of this cotransporter in the etiology of this disorder is further substantiated by the finding that the proband is homozygous for the S615L variation.…”

“…Onset is usually in adult life, but cases with onset in childhood are also known [1]. GS is characterized by hypomagnesemia, hypokalemia, metabolic alkalosis, hypocalciuria and hyperreninemic hyperaldosteronism.…”

“…The clinical spectrum is wide and includes: cramps, myalgies, muscle weakness, tetania, and paralysis [2]. GS is mostly caused by loss of function mutations in the solute carrier family 12, member 3 (SLC12A3 gene) which consists of 26 exons and is located on the long arm of 16 th chromosome [1]; this gene encodes the thiazide-sensitive sodium-chloride cotransporter (NCCT), expressed in the distal convoluted tubule of the kidney [3].…”

The sodium chloride cotransporter SLC12A3: new roles in sodium, potassium, and blood pressure regulation

“…Gender and the type of mutation contribute to phenotypic variability, with males and patients with homozygous deep intronic mutations exhibiting a more severe phenotype [64,65]. Novel mutations are still being identified in Gitelman syndrome and these genetic defects lead to impaired production, processing, insertion or regulation of the NCC protein (type I, II, III or IV mutations) [32,113,123]. Although Gitelman syndrome is usually a relatively benign disorder that often remains subclinical for many years, a recent report suggests that the electrolyte disorders associated with Gitelman syndrome may result in chronic kidney disease and glucose intolerance [113].…”

Exonic Mutations in the SLC12A3 Gene Cause Exon Skipping and Premature Termination in Gitelman Syndrome

“…1 To date, .160 mutations, including missense, nonsense, frameshift, and splice-site mutations, as well as gene rearrangements, have been documented in GS. 2 Among them, only a small number of missense mutations have been reported as exhibiting reduced function or as being nonfunctional. [3][4][5][6] Because the precise molecular basis of the residual missense mutations in GS remains unknown, novel approaches to clarify these issues are urgently needed.…”