Osteoblast energy flux response in the molecular consequence of osteogenesis imperfecta type V

Abstract

Osteogenesis Imperfecta type V (OIV) is a rare genetic bone disease characterized by bone fragility, deformity, hyperplastic callus formation and interosseous membrane ossification. The disease is caused by an autosomal dominant mutation in the 5’UTR of IFITM5, c.-14C>T, resulting in an addition of five amino acids at the N-terminus of the protein. While the mutation was discovered in 2012, the precise function of IFITM5 and the molecular consequence of the mutation remain unclear. Here, I leverage the use of clinical bone specimens and primary human bone cells from OIV patients to investigate the cellular and molecular changes in OIV bone cells to gain further insights.

In vivo assessment of OIV bone showed abnormal collagen matrix deposition, associated with an increase in osteocyte lacunae density and maturation to osteocytes was impaired. In vitro assessment of osteoblasts in culture showed enhanced biomineralization but reduced type I collagen in OIV cells, implicating an imbalance in inorganic and organic components. Interestingly, cultured primary osteoblasts no longer express IFTM5 in controls or OIV osteoblasts, suggesting a complex regulatory requirement for endogenous gene expression, and that an intrinsic change has occurred in OIV osteoblasts leading to observed outcomes. Multi-omic analyses of gene and protein profiles showed the isolated OIV osteoblasts were “primed” towards a more osteogenic/mineralization state, linked to up-regulations of the unfolded protein response (UPR), de novo serine biosynthesis, mTORC1 signalling pathways and oxidative phosphorylation as intrinsic changes compared to controls.

The elevated UPR was validated in vivo, with activated expressions of key UPR genes such as BiP, EIF2 and ATF4 in OIV osteoblast and osteocytes, that were not detectable in controls. Consistent with all OV samples is activation of BiP at the early stages of osteogenesis when IFITM5 is first expressed. As BiP up-regulation is the first response of the UPR, it is likely linked to mutant IFITM5 expression. UPR activation is a cell survival mechanism that may require metabolic and other cellular changes, but apoptosis can occur if unable to adapt. The increased cell number in OIV bones suggests an adaptation, supported by intrinsic changes consisting of elevated energy metabolism, increased de novo serine synthesis and the mTOC1 signalling pathway in primary OIV osteoblasts.

While the precise mechanism by which the observed changes in osteogenesis and osteoblast/osteocyte function is still unclear, the data suggest a link to an intrinsic cellular response from the mutant IFITM5 expression resulting in a gain-of-function outcome. The intrinsic changes have led to an impairment in osteocyte maturation, but the switch to a higher energy output in the osteoblasts placed these cells at a “primed” state for enhanced mineralization relative to collagen deposition, leading to a more brittle bone. Further, the adapted metabolic state to the UPR may be captured as epigenetic marks on the genome, changing the activity of osteoblasts even in the absence of the mutant IFITM5. This study has provided new insights into the molecular and cellular consequence of OIV that has opened new research areas to gain a full understanding of the disease mechanism.