Homozygous nonsense mutation in helix 2 of K14 causes severe recessive epidermolysis bullosa simplex

We have studied a consanguineous family containing two children with severe, generalized epidermolysis bullosa simplex (EBS). Electron microscopy of skin biopsies from the affected individuals showed that basal keratinocytes were devoid of tonofilament bundles, although some single intermediate filament were visible. Genetic linkage analysis with the microsatellite probe D12S96 excluded the type II keratin gene cluster in this family. However, homozygosity by descent was observed with the polymorphic probes KRT9, KRT 10 Ava II, and D 17S1787 in both affected children, consistent with a recessive defect in a type 1 keratin. Immunoreactivity to keratin K5 and K15 was normal, but monoclonal antibodies LL001 and RCK107 against K14 showed no staining, suggesting a deficiency of K14 in these individuals. mRNA extracted from biopsy material was amplified by RT-PCR to obtain full-length K14 cDNA. Direct automated sequencing identified a homozygous nonsense mutation, W305X. A Hinf I restriction enzyme site is created by this nucleotide transition, which was used to confirm the presence of the mutation in this kindred and exclude it from 100 normal chromosomes. This is the fourth kindred with severe recessive EBS for whom a mutation has been found in the K14 gene. In this instance, the premature termination codon is the farthest downstream of the reported cases, occurring in the helix 2 domain and so giving a much longer translation product. Nevertheless, the heterozygous carriers are unaffected by the disease and display no epidermal fragility. We postulate that translation of the potentially dominant-negative truncated K14 might be down-regulated due to instability of the mutant mRNA, as observed in previous cases with similar mutations.