Gassmann Reto, Kline Susan L, Carvalho Ana, Desai Arshad
Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, UCSD School of Medicine, CMM-East, Rm 3080, 9500 Gilman Dr., La Jolla, CA 92093, USA.
Methods. 2007 Feb;41(2):177-89. doi: 10.1016/j.ymeth.2006.07.027.
All eukaryotes rely on multi-protein assemblies, called kinetochores, to direct the segregation of their chromosomes in mitosis. The list of known kinetochore components has been growing rapidly in the post-genomic era: in animal cells, there are presently more than 80 proteins that show either exclusive or partial localization at kinetochores during mitosis. The future challenge is to elucidate how these proteins contribute to kinetochore structure, spindle microtubule attachment, regulation of microtubule dynamics, and the detection, signaling, and correction of microtubule attachment errors. Cultured human tumor cells, especially HeLa cells, are widely used for the study of kinetochores. Recently, the experimental advantages offered by the nematode Caenorhabditis elegans have been exploited for functional analysis of kinetochore components in the first embryonic division. Here, we discuss basic methods, largely based on fluorescence imaging, to study kinetochore structure and function in these two metazoan model systems.
所有真核生物都依赖于一种称为动粒的多蛋白复合体来在有丝分裂过程中指导其染色体的分离。在后基因组时代,已知的动粒成分列表一直在迅速增长:在动物细胞中,目前有超过80种蛋白质在有丝分裂期间在动粒处表现出排他性或部分定位。未来的挑战是阐明这些蛋白质如何有助于动粒结构、纺锤体微管附着、微管动力学调节以及微管附着错误的检测、信号传导和校正。培养的人类肿瘤细胞,尤其是HeLa细胞,被广泛用于动粒研究。最近,线虫秀丽隐杆线虫所具有的实验优势已被用于首次胚胎分裂中动粒成分的功能分析。在这里,我们讨论主要基于荧光成像的基本方法,以研究这两种后生动物模型系统中的动粒结构和功能。