O-GlcNAcylation in Gli1(+) Mesenchymal Stem Cells Is Indispensable for Bone Formation and Fracture Healing.

Adult mesenchymal stem cells (MSCs) play a crucial role in maintaining bone health and promoting regeneration. In our previous research, we identified Gli1(+) MSCs as key contributors to the formation of most trabecular bone in adulthood and as essential for healing bicortical fractures. However, the mechanisms behind the maintenance and differentiation of Gli1(+) MSCs are still not fully understood. O-linked N-acetylglucosamine modification (O-GlcNAcylation), mediated by O-GlcNAc glycosyltransferase (OGT), is involved in various biological processes and diseases. Our earlier work also demonstrated that O-GlcNAcylation is necessary for Wnt-stimulated bone formation. Nonetheless, the specific functions of O-GlcNAcylation in MSCs have not been completely elucidated. In this study, we found that the absence of OGT in Gli1(+) MSCs led to a decrease in O-GlcNAcylation, which impaired both the bone formation and regeneration following fractures. Mechanistically, the Hedgehog signaling pathway induced O-GlcNAcylation through the insulin-like growth factor (Igf)-mTORC2 axis. This process stabilized the Gli2 protein at a specific site Ser355 and promoted osteogenesis in MSCs in vitro. Our findings reveal a significant mechanism by which O-GlcNAcylation regulates bone development and repair in mammals.