端锚聚合酶Tankyrase1介导的PAR修饰在细胞有丝分裂纺锤体组装的功能与机理研究

有丝分裂纺锤体的正常组装保证了细胞内遗传物质的均等分配和染色体的稳定性，由端锚聚合酶Tankyrase1(TNKS1)介导的聚ADP-核糖（PAR）修饰发挥重要调控作用，但其调控机理尚不清楚。最近我们发现TNKS1与纺锤体组装因子BRCC36(B36)、接头蛋白MERIT40 相互作用，破坏它们的相互作用将导致纺锤体组装异常。我们推测TNKS1可能通过调节B36的PAR水平，从而在纺锤体组装中发挥重要作用。本研究拟利用活细胞实时摄影、免疫共沉淀、免疫荧光、RNA干扰和体外PAR反应等技术探索在纺锤体组装过程中，1）B36是否存在PAR修饰及其生物学功能影响；2）B36的PAR水平是否受TNKS1酶学调控；3）MERIT40是否及如何介导TNKS1对B36的调控。此研究对于阐明细胞有丝分裂纺锤体组装调控的分子基础和针对BRCA1/2突变肿瘤的PAR抑制剂的靶向药物研发都有重要意义。

The proper mitotic spindle assembly is essential for the chromosome segregation and genome stability during cell division in eukaryotes. Tankyrase1, a poly(ADP-ribose) polymerase (PARP) that plays an important role in spindle assembly, but its mechanism is largely unknown. However, We found that tankyrase1 interacts with BRCC36/ MERIT40 and disruption of their interaction causes abnormal mitotic spindle assembly recently. it was suggested that tankyrase1-mediated PARsylation of BRCC36 may have a critical role in the regulation of mitosis. In this study, we will use live-cell imaging, Co-IP,immunofluorescence,RNA interference and PAR assays in vitro to demonstrate that whether the PARsylation accumulated on BRCC36; Whether the PARsylation of BRCC36 is mediated by tankyrase1 and whether MERIT40 regulates tankyrase1-mediated PARsylation of BRCC36. Thus, in this study we propose to further elucidate the mechanism underlying of mitotic spindle assembly and have important implications for the development of PARP inhibitors to treat BRCA1/2 mutant cancers.