LncRNA ODIR1通过组蛋白修饰调控Osterix参与干细胞成骨分化的 机制研究

诱导干细胞定向成骨分化是骨再生医学领域的研究前沿。申请人前期研究发现间充质干细胞诱导成骨分化时LncRNA表达谱发生改变，尤其是Lnc ODIR1显著下调，敲低其表达可通过上调成骨特异性转录因子Osterix促进成骨，而过表达则抑制成骨分化；并能结合组蛋白H2A，H2B和H4及泛素连接酶FBXO25、CUL3、BARD1和甲基转移酶ZNF518A，因此提出了Lnc ODIR1可能通过组蛋白修饰影响Osterix调控干细胞成骨分化的科学假说。本项目拟利用CRISPR/Cas9敲低、CHIRP和CLIP等新技术研究Lnc ODIR1如何招募FBXO25、CUL3和BARD1调控组蛋白H2A/H2B的泛素化和招募ZNF518A调控组蛋白H4的甲基化进而影响Osterix表达，提出通过敲低Lnc ODIR1的表达水平促进成骨分化的策略，将为再生医学领域骨缺损修复和成骨障碍性疾病的治疗提供新的思路。

Osteogenic lineage commitment of mesenchymal stem cells is the research focus and the frontier of regenerative medicine of bone repair. Our previous studies have found that the expression profile of long non-coding RNAs was altered when the umbilical cord mesenchymal stem cells (hUC-MSCs) commit osteogenic differentiation. Among those, a long non-coding RNA, ODIR1 (osteogenic differentiation inhibitory RNA 1,briefly named as Lnc ODIR1) was significantly down-regulated. Knockdown of Lnc ODIR1 promotes the osteogenesis of MSCs. Over-expression of Lnc ODIR1 decreases the expression of osterix, a specific activated transcription factor in osteoblasts and inhibits osteogenesis of MSCs. RNA-pull down assay combined with mass spectrometry analysis and Western Blot analysis confirmed that Lnc ODIR1 was bound to histone H2A, H2B, H4, and ubiquitination-associated protein FBXO25, BARD1, CUL3 and methylation-associated protein ZNF518A. Therefore, it is proposed that Lnc ODIR1 regulates the osterix-mediated osteogenic differentiation of hUC-MSCs through controlling histone modification. This project aims to further explore how Lnc ODIR1 affects histone H2A/H2B ubiquitination through FBXO25, BARD1 and CUL3 and methylation of histone H4 through ZNF518A, thereby inhibiting the transcriptional osterix during osteogenic differentiation of MSCs with a series of new advanced technologies(CRISPR, CHIRP and CLIP,etc). The implementation of this project supports that knockdown Lnc ODIR1 in mesenchymal stem cells promotes the osteogenic differentiation, which provides novel ideas and strategies for the treatment of bone defects and osteogenic disorders.