MUTATIONAL SPECTRUM OF THE STEROID 21-HYDROXYLASE GENE

Lesions in the gene encoding steroid 21-hydroxylase (CYP21) result in defective adrenal steroid synthesis; the severe forms are known as congenital adrenal hyperplasia. To facilitate.complete characterization of mutations in this region of landemly repeated genes, we have developed selective PCR amplification and direct sequencing of full-length steroid 21-hydroxylase genes. This technique identifies known mutations, characterizes or excludes unknown mutations, and gives an estimate of gene copy number. Genetic defects in the 21-hydroxylase genes in a patient material representing 182 unrelated chromosomes have been studied. For 138 of these, HLA class II typing was performed, and for 86 chromosomes the gross structure of the C4 / 21-hydroxylase locus was determined. Thus, the location of the different mutations on different haplotypes are described. Functional consequences of individual alleles and combinations of alleles could be determined in vivo by studying individuals with known gene copy number, including hemizygous individuals. Genotypes showed good correlation to the clinical course of the disease. Six additional defective alleles were found, and several polymorphisms were shown to be neutral. The six mutations found are not present in the pseudogenes hitherto reported. Sequencing of pseudogenes (CYP21P) showed that this gene displays an equal degree of polymorphism to that of CYP21, and that two of the six above-mentioned mutations were present at low frequency. This implies that also the rare mutations can spread via CYP21P and can be expected to arise independently in unrelated individuals.