Construction of a full-length cDNA encoding human pro-α2(I) collagen and its expression in pro-α2(I)-deficient W8 rat cells

We have constructed a cDNA encoding the entire human pro-α2(I) collagen molecule. Sequence determination for 2196 base pairs at the 5' end of the cDNA clone, and comparison with previously characterized human α2(I) sequences, identified a number of nucleotide and amino acid polymorphisms. Functionality of the cDNA clone, under control of the long terminal repeat of Rous sarcoma virus, was demonstrated by its introduction into the W8 cell line. The W8 line, a chemically transformed variant of K16 rat liver epithelial cells, has been previously shown to lack detectable levels of α2(I) RNA, but to secrete α1(I) homotrimers. Introduction of the human cDNA into W8 cells, resulted in secretion of chimeric type I collagen comprised of rat α1(I) and human α2(I) chains. Availability of a functional full-length clone of human α2(I) cDNA, combined with the W8 cell line as expression system, will allow detailed analysis, through site-directed mutagenesis, of domains on the pro-α2(I) molecule involved in assembly, transport, secretion, and fibrillogenesis.