Molecular analysis of CYP21 gene mutations carried on HLA-B14 positive haplotypes

HLA-B14 positive haplotypes have increased frequencies in a group of patients with puberty disorders, IgA deficiency and cancer of the ovary. Clinical investigations demonstrated that all these patients have high values of 170H progesteron after the ACTH test which suggests an alterated function of 21 hydroxylase enzyme. In order to investigate whether these B14 positive haplotypes carry the same CYP21 mutation in the various diseases and controls, we have amplified by polymerase chain reaction (PCR) the sections of CYP21B gene which include amino acid positions 172 and 281 where typical mutations are known to occur in 21 hydroxylase deficiency. The presence or absence of the defined mutations was tested by oligonucleotide hybridization using oligonucleotides, labelled with DIG-ddUTP, designed to hybridize with the mutated or with the normal sequence. It was found that regardless of whether the subject tested was a patient or a healthy control the mutation at position 281 was found in all cases carrying HLA-B14, DR1 haplotype. Interestingly, this mutation does not seem to be in association with HLA-B14, DR7 haplotype. These findings suggest that CYP21 gene plays a role in all these differing diseases although it must be stressed that there may be alternative explanations for the observed data.