We have screened the elastin gene for mutations responsible for supravalvular aortic stenosis (SVAS) in two large, independently collected families with isolated (nonsyndromic) SVAS. By single-strand conformation polymorphism and heteroduplex analysis, we have identified a C to G transversion within the acceptor splice site of exon 16 in SVAS patients from both families. This mutation segregates in both families with high penetrance of SVAS, and all affected individuals carry the mutation. Haplotype analysis by using closely linked polymorphisms, including a previously unreported BfaI restriction fragment length polymorphism within the 3'-UTR of the elastin gene, indicates that the mutations found in the two apparently nonoverlapping kindreds are identical by descent. To study the effect of the mutation on the expression of the mutant allele, we have established a primary skin fibroblast culture from one of the affected individuals. Reverse transcription/polymerase chain reaction analysis of elastin mRNA species indicates that the mutation results in two abnormal elastin mRNA species. One mutant elastin mRNA is generated by the activation of a cryptic splice site that lies within intron 15 and that adds 44 bp of intronic sequence to the sequence encoded by exon 16. This insertion creates a frame shift that results in a 59-amino-acid-long abnormal protein sequence and leads to a termination codon in the mRNA sequence encoded by exon 17. The smaller abnormal mRNA species arises as a consequence of the skipping of exon 16. This study demonstrates, for the first time, the expression of mutant alleles of the elastin gene in patients with isolated SVAS.
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