Background Spontaneous read-through of a premature termination codon (PTC) has so far not been observed in patients carrying nonsense mutations. This report describes a patient with junctional epidermolysis bullosa who was expected to die because of compound heterozygous nonsense mutations in the gene LAMA3 (R943X/R1159X), but was rescued by spontaneous readthrough of the R943X allele. Results and conclusion FACS analysis of cells carrying various PTCs surrounded by their natural neighbouring codons revealed significant reporter gene expression despite the PTC only for this patient's genetic context. Gene expression could be abolished by replacing the first or third nucleotide before, or one of the two nucleotides following the PTC. Site-directed mutagenesis was used to identify genotypes allowing PTC read-through. The genetic context of the LAMA3 mutation R943X is close to a hypothetical consensus sequence for maximum PTC read-through. Bioinformatic analysis showed that this consensus sequence is present in four sequences from the NCBI reference database, each of which contains another in-frame termination codon three or four codons apart. This indicates strong selective pressure against leaky termination codons in the human genome. This patient's mutated full length mRNA escaped nonsensemediated decay, leading to LAMA3 mRNA levels similar to those of a healthy control, and full length laminin a3 could be detected in culture supernatant of the patient's keratinocytes. Immunofluorescence analyses of skin biopsies and continuous clinical improvement of the patient's condition suggested accumulation of intact laminin-332 in the epidermal basement membrane. These findings provide important clues for the prediction of PTC read-through in human genetic disease.
ASJC Scopus subject areas