Premature termination codons are present on both alleles of the bullous pemphigoid antigen 2 type XVII collagen gene in five Austrian families with generalized atrophic benign epidermolysis bullosa

Patients with generalized atrophic benign epidermolysis bullosa (GABEB), an inherited subepidermal blistering disease, often have no immunologically detectable bullous pemphigoid antigen 2 (BPAG2) in their epidermal basement membrane. Recently, we analyzed the BPAG2 gene (GenBank no. M91669) in an Austrian family with GABEB and identified a homozygous deletion mutation, 4003de1TC, that results in a downstream premature termination codon (PTC). This mutation has now been identified in additional descendants, suggesting transmission of this mutant allele through at least six generations. Screening of four other Austrian GABEB families revealed that affected members were homozygous for 4003de1TC in tow cases and heterozygous in two others. In the latter, mutational analysis identified two novel nonsense mutations, Q1403X and G803X, that were confirmed by restriction endonuclease digestions. Thus, PTCs on both alleles of PBAG2 are present in all of these GABEB families. Immunoprecipitation and northern blot studies of cultured keratinocytes from homozygous GABEB patients show that 4003de1TC results in undetectable levels of BPAG2 protein and mRNA-findings consistent with the process of nonsense-mediated mRNA decay. Incubating keratinocytes with cycloheximide increased BPAG2 mRNA to a level detectable by northern analysis. When the latter was used in reverse transcription-PCR studies, the mutation was demonstrated, suggesting that cycloheximide may allow mutational analysis in cases where low transcript levels have previously thwarted RT-PCR studies. These findings account for the absence of BPAG2 in GABEB patients in adhesion of epidermis to epidermal basement membrane.