Three novel type I collagen mutations in osteogenesis imperfecta type IV probands are associated with discrepancies between electrophoretic migration of osteoblast and fibroblast collagen

In three cases of type IV osteogenesis imperfecta (OI), we identified unique point mutations in type I collagen α1 (I) cDNA. In two cases, the appearance of dimers indicated the presence of cysteine substitutions in the α1 (I) protein chain. Cyanogen bromide digestion localized these cross- links to CB8 and 3, respectively. In the third case, the overmodification pattern of the CNBr peptides was compatible with a substitution in the aa 123-402 region of either type I collagen chain. We identified a unique point mutation in each proband, which resulted in substitutions for glycine residues in a 300-aa region of the αl(I) helix, specifically, Gly to Ala at codon 220 (GGT→CT), Gly to Cys at codon 349 (GGT→TGT) and Gly to Cys at codon 523 (GGT→TGT). We compared each proband\u27s fibroblast and osteoblast collagen directly, as well as with fibroblast and osteoblast controls. For all cases, the OI osteoblast collagen was more electrophoretically delayed than OI fibroblast collagen. In the patient with G349C, OI fibroblast and osteoblast collagen synthesized in the presence of α,α\u27-dipyridyl co- migrated on gels, demonstrating that the electrophoretic discrepancy resulted from differences in post-translational modification. Melting temperature curves for stability of the collagen helix yielded an identical T(m) for control fibroblast and osteoblast collagen (41.2°C). By contrast, for collagen with the gly349→cys substitution, the T(m) of the fibroblast collagen was 1°C lower than the T(m) of the osteoblast collagen. These data indicate that the metabolism of mutant collagen might be cell-specific and has significant implications for understanding the phenotype/genotype correlations and the pathophysiology of OI