Identification of a novel centrosomal protein CrpF46 involved in cell cycle progression and mitosis.
Identification of a novel centrosomal protein CrpF46 involved in cell cycle progression and mitosis.
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DOI：
10.1016/j.yexcr.2008.02.021
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发表时间：
2008-05
影响因子：
3.7
通讯作者：
Yi Wei;E. Shen;N. Zhao;Qian Liu;Jinling Fan;J. Marc;Yongchao Wang;Le Sun;Qianjin Liang
中科院分区：
文献类型：
--
作者：
Yi Wei;E. Shen;N. Zhao;Qian Liu;Jinling Fan;J. Marc;Yongchao Wang;Le Sun;Qianjin Liang
使用进行性系统性硬化症患者的血清F46检测到一种新的中心体相关蛋白crpf46。我们通过免疫沉淀和Western blotting以及串联质谱测序鉴定了该蛋白。蛋白质crpf46的表观分子质量约为60 kDa，与蛋白质Golgin-245的c端527个氨基酸序列高度同源，似乎是Golgin-245的剪接变体。用抗crpf46单克隆抗体标记的同步HeLa细胞的免疫荧光显微镜显示，尽管crpf46在有丝分裂开始时分散在整个细胞质中，但在间期仅定位于中心体。在转化为HeLa细胞的gfp表达载体中使用crpf46片段进行结构域分析，发现中心体靶向是由c端线圈结构域赋予的。反义crpf46敲低抑制细胞生长和增殖，细胞周期通常停滞在S期。这种敲除还导致多中心体和多核细胞的形成，最终导致细胞凋亡。这些结果表明，crpf46是一种新的中心体相关蛋白，以细胞周期依赖的方式与中心体结合，并参与细胞周期的进展和M期机制。
A novel centrosome-related protein CrpF46was detected using a serum F46 from a patient suffering from progressive systemic sclerosis. We identified the protein by immunoprecipitation and Western blotting followed by tandem mass spectrometry sequencing. The protein CrpF46has an apparent molecular mass of ~60 kDa, is highly homologous to a 527 amino acid sequence of the C-terminal portion of the protein Golgin-245, and appears to be a splice variant of Golgin-245. Immunofluorescence microscopy of synchronized HeLa cells labeled with an anti-CrpF46monoclonal antibody revealed that CrpF46localized exclusively to the centrosome during interphase, although it dispersed throughout the cytoplasm at the onset of mitosis. Domain analysis using CrpF46fragments in GFP-expression vectors transformed into HeLa cells revealed that centrosomal targeting is conferred by a C-terminal coiled-coil domain. Antisense CrpF46knockdown inhibited cell growth and proliferation and the cell cycle typically stalled at S phase. The knockdown also resulted in the formation of poly-centrosomal and multinucleate cells, which finally became apoptotic. These results suggest that CrpF46is a novel centrosome-related protein that associates with the centrosome in a cell cycle-dependent manner and is involved in the progression of the cell cycle and M phase mechanism.