We previously established that DNA methyltransferase 3b (Dnmt3b) is the sole Dnmt responsive to fracture repair and that Dnmt3b expression is induced in progenitor cells during fracture repair. In the current study, we confirmed that Dnmt3b ablation in mesenchymal progenitor cells (MPCs) resulted in impaired endochondral ossification, delayed fracture repair, and reduced mechanical strength of the newly formed bone in Prx1-Cre;Dnmt3bf/f (Dnmt3bPrx1) mice. Mechanistically, deletion of Dnmt3b in MPCs led to reduced chondrogenic and osteogenic differentiation in vitro. We further identified Rbpjκ as a downstream target of Dnmt3b in MPCs. In fact, we located 2 Dnmt3b binding sites in the murine proximal Rbpjκ promoter and gene body and confirmed Dnmt3b interaction with the 2 binding sites by ChIP assays. Luciferase assays showed functional utilization of the Dnmt3b binding sites in murine C3H10T1/2 cells. Importantly, we showed that the MPC differentiation defect observed in Dnmt3b deficiency cells was due to the upregulation of Rbpjκ, evident by restored MPC differentiation upon Rbpjκ inhibition. Consistent with in vitro findings, Rbpjκ blockage via dual antiplatelet therapy reversed the differentiation defect and accelerated fracture repair in Dnmt3bPrx1 mice. Collectively, our data suggest that Dnmt3b suppresses Notch signaling during MPC differentiation and is necessary for normal fracture repair.
Jun Ying, Taotao Xu, Cuicui Wang, Hongting Jin, Peijian Tong, Jianjun Guan, Yousef Abu-Amer, Regis O’Keefe, Jie Shen
This file is in Adobe Acrobat (PDF) format. If you have not installed and configured the Adobe Acrobat Reader on your system.
PDFs are designed to be printed out and read, but if you prefer to read them online, you may find it easier if you increase the view size to 125%.
Many versions of the free Acrobat Reader do not allow Save. You must instead save the PDF from the JCI Online page you downloaded it from. PC users: Right-click on the Download link and choose the option that says something like "Save Link As...". Mac users should hold the mouse button down on the link to get these same options.