裂殖酵母monopolin复合体(即Pcs1/Mde4蛋白复合体)在有丝分裂细胞中定位在多处，暗示其多重功能。已有研究发现，定位在动粒处的Pcs1/Mde4负责招募集缩素(condensin)蛋白，并参与抑制姐妹染色单体merotelic连接(一个着丝粒同时受到来自两极的纺锤体微管牵引)，但这种错误连接是否激活纺锤体组装检验点(SAC)尚存争议。Pcs1/Mde4还定位于核仁内，其功能尚不清楚，其是否与核仁内集缩素蛋白功能相关也未见报道。我们的前期结果显示，Pcs1/Mde4缺失导致SAC沉默缺陷，还会引起rDNA重组增加，但仍能维持rDNA区转录沉默，而集缩素突变体则解除rDNA区转录沉默。本项目拟利用GBP-GFP高亲和系统，通过GBP与烟草蚀纹病毒蛋白酶的融合蛋白(GBP-TEV)分别特异降解动粒处或核仁内的Pcs1/Mde4，解析其参与SAC以及维持rDNA拷贝数稳定的分子机制。

The fission yeast monopolin complex (i.e. Pcs1/Mde4 protein complex) localizes at multiple loci during mitotic cell cycle, indicating its multiple functions. Previous studies showed Pcs1/Mde4 at kinetochores can recruit condensin and inhibit merotelic attachment between sister chromatids and spindle microtubules. It is still controversial whether merotelic attachments activate spindle assembly checkpoint (SAC). Pcs1/Mde4 also localize inside nucleolus, but its nucleolar function remains unclear. The potential functional connection between nucleolar Pcs1/Mde4 and condensin has not been studied. Our preliminary results showed that the absence of Pcs1/Mde4 leads to SAC silencing defects and more frequent recombination events between rDNA repeats. In contrast to condensin mutants, which show rDNA silencing defects, our results showed that Pcs1 and Mde4 are not required for rDNA silencing. We have constructed a GBP-TEV fusion carrying GFP-binding protein (GBP) and tobacco etch virus (TEV) protease, which can bind GFP/CFP/YFP-tagged proteins and cleave proteins with TEV recognizing sites. In this project, we intend to dissect the functions of fission yeast monopolin complex in SAC activation or silencing and rDNA repeat maintenance, using the newly developed GBP-GFP high affinity system.