Recurrent COL3A1 mutation results in EDS IV or familial aneurysms

Abstract

EIV is a lethal genetic disease of collagen in which mutations in COL3A1 give rise to fragile skin, easy bruisability, and hollow organ ruptures (Kuivaniemi et al., 1991). Familial aneurysms is a vascular disease characterized by late-onset saccular aneurysms, probably multifactorial in origin, but rare cases may be due to COL3A1 mutations. We investigated the cause of EDS IV in a 10-year-old white male with soft skin easy bruisability, and a family history of vascular ruptures. S! nuclease analysis of RNA from patients fibroblasts indicated a defect around exon 20. Sequence analysis of cDNA clones identified 3 out of 5 clones that retained a portion of intron 20 and revealed a G+1 to A point mutation in the splice donor site. The same mutation has been observed in a patient with familial aneurysms (Kontussari et al., 1990). In both patients, RT-PCR analysis revealed the presence of several RNA species: mRNA lacking exon 20, mRNA with 24 bases of intron 20 sequnce in frame, mRNA containing all 132 bases of intron 20, and normally spliced mRNA. The four mRNA species can give rise to three translation products: protein lacking exon 20 sequence, protein with 8 noncollagenous amino acids in the middle of the triple helical domain, and normal protein chains. The mRNA with all 132bp of intron 20 results in a truncated protein incapable of assembly into trimers. Consistent with these findings, cultured fibroblasts from the patient secreted less type III procollagen and retained abnormal timers in the cell layer. Pepsin digestion of the cell layer proteins revealed multiple di sulfide bonded trimers of type III procollagen, presumably the product of protein assembly from a mixture of normal and multiple abnormal type III procollagen monomers.