Congenital erythropoietic porphyria: a novel uroporphyrinogen III synthase branchpoint mutation reveals underlying wild-type alternatively spliced transcripts

Splicing mutations account for approximately 10% of lesions causing genetic diseases, but few branchpoint sequence BPS) lesions have been reported. In 3 families with autosomal recessive congenital erythropoietic porphyria CEP) resulting from uroporphyrinogen III synthase URO-synthase) deficiency, sequencing the promoter, all 10 exons and the intron/exon boundaries did not detect a mutation. Northern analyses of lymphoblast mRNAs from 2 patients and reversetranscribed polymerase chain reaction (RT-PCR) of lymphoblast mRNAs from all 3 patients revealed multiple longer transcripts involving intron 9 and low levels of wild-type message. Sequencing intron 9 RT-PCR products and genomic DNA in each case revealed homozygosity for a novel BPS mutation (c.661-31T -> G) and alternatively spliced transcripts containing 81, 246, 358, and 523 nucleotides from intron 9. RT-PCR revealed aberrant transcripts in both wild-type and CEP lymphoblasts, whereas BPS mutation reduced the wild-type transcript and enzyme activity in CEP lymphoblasts to approximately 10% and 15% of normal, respectively. Although the +81-nucleotide alternative transcript was in-frame, it only contributed approximately 0.2% of the lymphoblast URO-synthase activity. Thus, the BPS mutation markedly reduced the wildtype transcript and enzyme activity, thereby causing the disease. This is the first BPS mutation in the last intron, presumably accounting for the observed 100% intron retention without exon skipping. (Blood. 2010;115:1062-1069)