CLONING OF THE HUMAN TYPE-XVII COLLAGEN GENE (COL17A1), AND DETECTIONOF NOVEL MUTATIONS IN GENERALIZED ATROPHIC BENIGN EPIDERMOLYSIS-BULLOSA

Generalized atrophic benign epidermolysis bullosa (GABEB) is a nonleth al variant of junctional epidermolysis bullosa (JEB). Previous finding s have suggested that type XVII collagen is the candidate gene for mut ations in this disease. We now have cloned the entire human type XVII collagen gene (COL17A1) and have elucidated its intron-exon organizati on. The gene comprises 56 distinct exons, which span similar to 52 kb of the genome, on the long arm of chromosome 10. It encodes a polypept ide, the alpha 1(XVII) chain, consisting of an intracellular globular domain, a transmembrane segment, and an extracellular domain that cont ains 15 separate collagenous subdomains, the largest consisting of 242 amino acids. We also have developed a strategy to identify mutations in COL17A1 by use of PCR amplification of genomic DNA, using primers p laced on the flanking introns. The PCR products are scanned for sequen ce variants by heteroduplex analysis using conformation-sensitive gel electrophoresis and then are subjected to direct automated sequencing. We have identified several intragenic polymorphisms in COL17A1, as we ll as mutations, in both alleles, in two Finnish families with GABEB. The probands in both families showed negative immunofluorescence stain ing with an anti-type XVII collagen antibody. In one family, the proba nd was homozygous for a 5-bp deletion, 2944del5, which resulted in fra meshift and a premature termination codon of translation. The proband in the other family was a compound heterozygote, with one allele conta ining the 2944del5 mutation and the other containing a nonsense mutati on, Q1023X. These results expand the mutation database in different va riants of JEB, and they attest to the functional importance of type XV II collagen as a transmembrane component of the hemidesmosomes at the dermal/epidermal junction.