Recessive types VII and VIII osteogenesis imperfecta (OI) are severe bone dysplasias caused by null mutations in prolyl 3-hydroxylase 1 (P3H1) or cartilage-associated protein (CRTAP), two mutually supportive components of the prolyl 3-hydroxylase complex expressed in bone and cartilage. Individuals with types VII/VIII OI are missing the complex components as well as 3-hydroxylation of the collagen substrate residue α1(I)P986. This primary modification defect leads to increased post-translational lysine hydroxylation of the type I collagen helix, consistent with delayed folding. Since α1(I)P986 has been proposed to play a role in fine-tuning the alignment of collagen helices within fibrils, we sought to clarify the role of the P986 substrate modification in bone dysplasia and collagen overmodification by generating a knock-in mouse model with an α1(I)P986A substitution that cannot be 3-hydroxylated. We verified exclusive expression of α1(I)A986 collagen in murine tissues and cellular cDNA and protein. Neither heterozygous (986P/A) nor homozygous (986A/A) mice recapitulate critical features of types VII/VIII OI. Mutant mice had normal growth rates, femoral biomechanical properties (stiffness, ultimate load, brittleness) and collagen folding kinetics. However, the P986A substitution affected collagen biochemistry and higher order structures. Skeletal staining and radiographs revealed flared rib cages and delayed calvarial mineralisation in 986A/A pups, with kyphosis by 2 months. Despite normal 3-hydroxylase complex levels, 986P/A and 986A/A osteoblast type I collagen was moderately overmodified on PAGE. Dermal fibrils of 986A/A displayed decreased diameters and heterogeneity. Interestingly, although femoral aBMD and TMD was reduced in 986A/A versus WT, bone matrix mineralisation density distribution assessed by qBEI was normal. Together, these data suggest that 3-hydroxylation of α1(I)P986 is important for regulating type I collagen modification, crosslinking and mineral organisation in bone, but does not cause the severe bone pathology of collagen 3-hydroxylation defects, which likely result from absence of the ER complex and cartilage α1(II)P986 3-hydroxylation.
Disclosure: The authors declared no competing interests. This work was supported by NIH intramural funding.