Recurrent Deep Intronic Mutations in the SLC12A3 Gene Responsible for Gitelman's Syndrome

Summary Background and objectives Gitelman’s syndrome (GS) is an autosomal recessive renal tubular disorder caused by mutations in the SLC12A3 gene encoding the thiazide-sensitive Na-Cl cotransporter (NCC). Despite meticulous sequencing of genomic DNA, approximately one-third of GS patients are negative or heterozygotes for the known mutations. Design, Setting, Participants, & Measurements Because blood leukocytes express NCC mRNA, we evaluate whether deep intronic mutations contribute to GS patients with uniallelic or undetectable SLC12A3 mutations. Twenty-nine patients with GS (men/women 16/13), including eight negative and 21 uniallelic SLC12A3 mutations from 19 unrelated families, and normal controls were enrolled in an academic medical center. Analysis of cDNA from blood leukocytes, sequencing of the corresponding introns of genomic DNA for abnormal transcript, and analysis of NCC protein expression from renal biopsy were performed. Results We identified nine Taiwan aboriginal patients carrying c.1670–191C3T mutations in intron 13 and 10 nonaboriginal patients carrying c.2548253C3T mutations in intron 21 from 14 families (14/ 19). These two mutations undetected in 100 healthy subjects created pseudoexons containing new premature termination codons. Haplotype analysis with markers flanking SLC12A3 revealed that both mutations did not have founder effects. Apical NCC expression in the DCT of renal tissue was markedly diminished in two patients carrying deep intronic mutations. Conclusions Deep intronic mutations in SLC12A3 causing defective NCC expression can be identified with the RNA-based approach in patients with GS. c.1670–191C3T and c.2548253C3T are hot spot mutations that can be screened in GS patients with uniallelic or negative SLC12A3 mutations. Clin J Am Soc Nephrol 6: 630–639, 2011. doi: 10.2215/CJN.06730810