Incorporation of Structurally Defective Type II Collagen into Cartilage Matrix in Kniest Chondrodysplasia

Kniest dysplasia, a human chondrodysplasia that severely affects skeletal growth, is caused by mutations in the type II collagen gene, COL2A1. We report here on abnormal type II collagen in the cartilage from a lethal Kniest dysplasia case and identify a novel exon-skipping mutation. Screening of cyanogen bromide (CB) peptides from the cartilage samples by SDS–PAGE indicated an abnormality in peptide α1(II)CB11. Further peptide mapping and N-terminal sequence analysis showed a 15-amino-acid deletion encoded by exon 15 in about 25% of the α1(II) chains in the cartilage. The mutation responsible for exon skipping was found by sequencing amplified genomic DNA. The baby was heterozygous for a G to A transition at the first position of the splice donor of intron 15. Pepsin-solubilized type II collagen from the cartilage matrix contained both normal α1(II) and shortened chains expressed from the mutant allele. Trypsin cleaved the native molecules below 37°C selectively at a site within the exon 15-encoded domain of the normal α1(II) chains. This is best explained by the coassembly of normal and truncated α1(II) chains into heterotrimers in which the triple helix is normally folded in both directions from the deletion site but the latter presents a region of local disruption. The findings support an emerging pattern of COL2A1 mutations that can cause Kniest dysplasia. Short deletions (single or partial exon) clustered in one region of the α1(II) chain are favored, resulting in abnormal heterotrimeric molecules that become a significant component of the cartilage extracellular matrix.