RECURRENT NONSENSE MUTATIONS WITHIN THE TYPE-VII COLLAGEN GENE IN PATIENTS WITH SEVERE RECESSIVE DYSTROPHIC EPIDERMOLYSIS-BULLOSA

The generalized mutilating form of recessive dystrophic epidermolysis bullosa (i.e., the Hallopeau-Siemens type; HS-RDEB) is a life-threaten ing disease characterized by extreme mucocutaneous fragility associate d with absent or markedly altered anchoring fibrils (AF). Recently, we reported linkage between HS-RDEB and the type VII collagen gene (COL7 A1), which encodes the major component of AF. In this study, we invest igated 52 unrelated HS-RDEB patients and 2 patients with RDEB inversa for the presence, at CpG dinucleotides, of mutations changing CGA argi nine codons to premature stop codons TGA within the COL7A1 gene. Eight exons containing 10 CGA codons located in the amino-terminal domain o f the COL7A1 gene were studied. Mutation analysis was performed using denaturing gradient gel electrophoresis of PCR-amplified genomic fragm ents. Direct sequencing of PCR-amplified products with altered electro phoretic mobility led to the characterization of three premature stop codons, each in a single COL7A1 allele, in four patients. Two patients (one affected with HS-RDEB and the other with RDEB inversa) have the same C-to-T transition at arginine codon 109. Two other HS-RDEB patien ts have a C-to-T transition at arginine 1213 and 1216, respectively. T hese nonsense mutations predict the truncation of similar to 56%-32% o f the polypeptide, including the collagenous and the noncollagenous NC -2 domains. On the basis of linkage analysis, which showed no evidence for locus heterogeneity in RDEB, it is expected that these patients a re compound heterozygotes and have additional mutations on the other C OL7A1 allele, leading to impaired AF formation. These results indicate that stop mutations within the COL7A1 gene can underlie both HS-RDEB and RDEB inversa, thus providing further evidence for the implication of this gene in RDEB.