Novel mutations in the SCNN1A gene causing Pseudohypoaldosteronism type 1

Abstract

Pseudohypoaldosteronism type 1 (PHA1) is a rare inherited disease characterized by resistance to the actions of aldosterone. Mutations in the subunit genes (SCNN1A, SCNN1B, SCNN1G) of the epithelial sodium channel (ENaC) and the NR3C2 gene encoding the mineralocorticoid receptor, result in systemic PHA1 and renal PHA1 respectively. Common clinical manifestations of PHA1 include salt wasting, hyperkalaemia, metabolic acidosis and elevated plasma aldosterone levels in the neonatal period. In this study, we describe the clinical and biochemical manifestations in two Chinese patients with systemic PHA1. Sequence analysis of the SCNN1A gene revealed a compound heterozygous mutation (c.1311delG and c.1439+1G>C) in one patient and a homozygous mutation (c.814_815insG) in another patient, all three variants are novel. Further analysis of the splicing pattern in a minigene construct showed that the c.1439+1G>C mutation can lead to the retainment of intron 9 as the 5'-donor splice site disappears during post-transcriptional processing of mRNA. In conclusion, our study identified three novel SCNN1A gene mutations in two Chinese patients with systemic PHA1.

Figure 1. Three novel mutations were identified in the SCNN1A gene.

(A) Sequences showing a compound heterozygous mutations (c.1311delG in exon 8 c.1439+1G>C in intron 9) in PHA1 patient of case 1. (B) Sequences showing a homozygous mutation (c.814_815insG in exon 4) in PHA1 patient of case 2.

Figure 2. In vitro splicing assay of the c.1439+1G>C mutation.

(A) The splice region (c.1439+1, intron 9) of SCNN1A gene was amplified and products were ligated into the pcDNA3.1/Myc-His B vector. (B) RT-PCR of HEK293 cells transfected with either wild-type or mutant SCNN1A. Minigenes showed that the mutation c.1439+1G>C was sufficient to produce a longer band. (C) Lane 1: Empty pcDNA3.1 vector; Lane 2: wild-type SCNN1A (256 bp); Lane 3: c.1439+1G>C mutant (361 bp); Lanes 4, 5, and 6: GAPDH used as control (245 bp).