Characterization of mutations of the type VII collagen gene (COL7A1) in recessive dystrophic epidermolysis bullosa mitis (M-RDEB) from three Korean patients.

In recent years, the molecular basis for the main subtypes of epidermolysis bullosa (EB) has been elucidated with pathogenetic mutations delineated in ten different genes encoding structural components of the dermal-epidermal junction. Both the autosomal dominant and recessive forms of dystrophic EB (DEB) is caused by mutations in the COL7A1 gene. Type VII collagen is a major component of anchoring fibrils, structural elements that stabilize the attachment of the basement membrane to underlying dermis. Recent delineation of the exon-intron organization of the COL7A1 gene provided the basis for the comprehensive design of PCR primer pairs that amplified exons in genomic DNA by placing the primers on the flanking introns. A number of COL7A1 mutations have been reported and some genotype-phenotype correlations are starting to emerge. In this study, we examined mutational analyses from three Korean patients with recessive dystrophic EB (RDEB) mitis. We designed and optimized primers according to the previously reported sequences. Such PCR amplification products can be examined by electrophoretic scanning technique, CSGE heteroduplex analyses. Utilizing heteroduplex analyses, we have identified a number of sequence variants in COL7A1 both in unaffected individuals and in patients with M-RDEB. Mutation detection of the COL7A1 gene revealed six allelic mutations (V6677E, P6685S, Y3749S, P6084S, P6695R and G6697C). We suggest that the full length of type VII collagen polypeptide are synthesized, but those missense mutations, that may affect a critical amino acid, can alter the conformation of the protein and interferes with the assembly and packing of type VII collagen molecules into anchoring fibrils. Immunohistochemical study of skin biopsies by use of anti-type VII collagen antibody showed markedly reduced staining and presence of a dermo/epidermal cleavage. This is the first report of a COL7A1 mutation study in DEB from Korean patients. We hope that these data contribute to the expanding database on COL7A1 mutations in dystrophic epidermolysis bullosa, and further illustrate the extensive diversity of mutational events that led to the RDEB phenotype.