Misregulation effect of a novel allelic variant in the Z promoter region found in cis with the CYP21A2 p.P482S mutation: implications for 21-hydroxylase deficiency

Abstract

Purpose: To search for the presence of genetic variants in a distal regulatory region (Z promoter) of the CYP21A2 gene in patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency. Methods: Screening of the 10 most frequent pseudogene-derived mutations followed by direct sequencing of the entire coding sequence, the proximal promoter and a distal regulatory region in DNAsamples from patients with at least one non-determined allele. The putative pathogenic implication of a novel variant found in a distal regulatory region was assessed by in silico analyses and in vitrotranscriptional activity measurement of the mutant. Results: Three non-classical patients presented a novel genetic variant -g.15626A>G- within the Z promoter regulatory region. In all the patients, the novel variant was found in cis with the mild, less frequent, p.P482S mutation located in the exon 10 of the CYP21A2 gene. Topological analyses showed differences in the curvature and bendability of the DNA region bearing the novel variant. By performing functional studies, a significantly decreased activity of a reporter gene placed downstream from the regulatory region was found by the G transition. Conclusions: Our results may suggest that the activity of an allele bearing the p.P482S mutation may be influenced by the misregulated CYP21A2 transcriptional activity exerted by the Z promoter A>G variation.