Direct interaction between yeast spindle pole body components: Kar1p is required for Cdc31p localization to the spindle pole body.
Direct interaction between yeast spindle pole body components: Kar1p is required for Cdc31p localization to the spindle pole body.
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DOI：
10.1083/jcb.125.4.843
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发表时间：
1994-05
期刊：
影响因子：
--
通讯作者：
Rose MD
中科院分区：
文献类型：
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作者：
Biggins S;Rose MD
酿酒酵母（Saccharomycescerevisiae）基因KAR 1和CDC 31是酵母中纺锤体极体（SPB）复制的初始阶段所必需的。Cdc 31蛋白与caltractin/centrin最相关，caltractin/centrin是一种存在于许多生物体微管组织中心的钙结合蛋白。由于CDC 31和KAR 1之间的多种遗传相互作用（Vallen，E.一、W.何M. Winey和M. D.玫瑰1994.遗传学我们想确定Cdc 31 p和Kar 1 p是否发生物理相互作用。Cdc 31 p从大肠杆菌中表达并纯化，并具有结合钙的活性。使用蛋白质印迹技术，Cdc 31 p通过SPB复制所需的Kar 1 p中的必需结构域在体外与Kar 1 p结合（Vallen，E.一、M. A. Hiller，T. Y. Scherson和M. D.玫瑰1992年a。117：1277-1287）。通过免疫荧光显微镜，我们确定这种相互作用也发生在体内。Cdc 31 p定位于野生型细胞中的SPB，但在kar 1突变株中被错误定位。在一个kar 1突变体含有一个显性的CDC 31抑制，Cdc 31 p再次定位于SPB。此外，Cdc 31 p的本地化的SPB的Kar 1 p-β-半乳糖苷酶杂合物的过表达的影响。基于这些数据，我们建议，Kar 1 p的基本功能是本地化Cdc 31 p的SPB，这种相互作用通常是需要SPB复制。
The Saccharomyces cerevisiae genes KAR1 and CDC31 are required for the initial stages of spindle pole body (SPB) duplication in yeast. The Cdc31 protein is most related to caltractin/centrin, a calcium-binding protein present in microtubule organizing centers in many organisms. Because of a variety of genetic interactions between CDC31 and KAR1 (Vallen, E. A., W. Ho. M. Winey, and M. D. Rose. 1994. Genetics. In press), we wanted to determine whether Cdc31p and Kar1p physically interact. Cdc31p was expressed and purified from Escherichia coli and active for binding calcium. Using a protein blotting technique, Cdc31p bound to Kar1p in vitro via an essential domain in Kar1p required for SPB duplication (Vallen, E. A., M. A. Hiller, T. Y. Scherson, and M. D. Rose. 1992a. J. Cell Biol. 117:1277-1287). By immunofluorescence microscopy, we determined that the interaction also occurs in vivo. Cdc31p was localized to the SPB in wild-type cells but was mislocalized in a kar1 mutant strain. In a kar1 mutant containing a dominant CDC31 suppressor, Cdc31p was again localized to the SPB. Furthermore, the localization of Cdc31p to the SPB was affected by the overexpression of Kar1p-beta-galactosidase hybrids. Based on these data, we propose that the essential function of Kar1p is to localize Cdc31p to the SPB, and that this interaction is normally required for SPB duplication.