Aberrant Splicing Is the Pathogenicity Mechanism of the p.Glu314Lys Variant in Gene.

The cholesterol side chain cleavage enzyme (CYP11A1) catalyzes the conversion of cholesterol to pregnenolone, the first rate-limiting step of steroidogenesis. mutations are associated with primary adrenal insufficiency (PAI) as well as disorders of sex development (DSD) in 46,XY patients. To define the pathogenicity mechanism for the p.Glu314Lys variant, previously reported, and found in four additional patients with CYP11A1 deficiency. DNA of four patients presenting with delayed PAI and/or 46,XY DSD were studied by Sanger or Massively Parallel sequencing. Three mutations were characterized and , and one by mRNA analysis on testicular tissue. All patients were compound heterozygous for the previously described p.Glu314Lys variant. studies predicted this mutation as benign with no effect on splicing but mRNA analysis found that it led to incomplete exon 5 skipping. This mechanism was confirmed by minigene experiment. The protein carrying this mutation without exon skipping should conserve almost normal activity, according to studies. Two other mutations found in , the p.Arg120Gln and p.Arg465Trp, had similar activity compared to negative control, consistent with the studies. We provide biological proof that the p. Glu314Lys variant is pathogenic due to its impact on splicing and seems responsible for the moderate phenotype of the four patients reported herein. The present study highlights the importance of considering the potential effect of a missense variant on splicing when it is not predicted to be disease causing.