Splicing modulation of integrin beta4 pre-mRNA carrying a branch point mutation underlies epidermolysis bullosa with pyloric atresia undergoing spontaneous amelioration with ageing.

A general improvement with ageing has been reported in a few cases of epidermolysis bullosa with pyloric atresia (PA-JEB), an autosomal recessive skin disease characterized by extensive disadhesion of epithelia. In a patient who improved from severe to mild PA-JEB, a search for mutations in the integrin beta4 gene (IGTB4) detected heterozygosity for a novel base substitution 3986-19T-->A in the putative branchpoint sequence of intron 31, and a point mutation 3802+1G-->A in the donor splice site of intron 30 previously associated with severe PA-JEB. Analysis of mRNA showed that the intronic mutation prevents legitimate splicing of the beta4 pre-mRNA. Functional splicing can be restored in vitro by seeding the proband's keratinocytes on feeders of irradiated fibroblasts. Study of mRNA in wild-type keratinocytes transfected with IGTB4 minigenes containing intron 31 with or without mutation 3986-19T-->A, confirmed the causative role of the intronic mutation in PA-JEB, and highlighted the influence of feeders on the maturation process of the mutated beta4 pre-mRNA. Our results show that in a context of overall reduction of the beta4 mRNA levels, activation of the legitimate splice site in the aberrant beta4 pre-mRNA underlies the transient severity of the condition. The results also point to the relevance which the interaction between epithelial and stromal cells may have in modulating expression of integrin receptors.