Gardner Melissa K, Haase Julian, Mythreye Karthikeyan, Molk Jeffrey N, Anderson Marybeth, Joglekar Ajit P, O'Toole Eileen T, Winey Mark, Salmon E D, Odde David J, Bloom Kerry
Department of Biomedical Engineering, University of Minnesota, Minneapolis, MN 55455, USA.
J Cell Biol. 2008 Jan 14;180(1):91-100. doi: 10.1083/jcb.200710164. Epub 2008 Jan 7.
In budding yeast, the mitotic spindle is comprised of 32 kinetochore microtubules (kMTs) and approximately 8 interpolar MTs (ipMTs). Upon anaphase onset, kMTs shorten to the pole, whereas ipMTs increase in length. Overlapping MTs are responsible for the maintenance of spindle integrity during anaphase. To dissect the requirements for anaphase spindle stability, we introduced a conditionally functional dicentric chromosome into yeast. When centromeres from the same sister chromatid attach to opposite poles, anaphase spindle elongation is delayed and a DNA breakage-fusion-bridge cycle ensues that is dependent on DNA repair proteins. We find that cell survival after dicentric chromosome activation requires the MT-binding proteins Kar3p, Bim1p, and Ase1p. In their absence, anaphase spindles are prone to collapse and buckle in the presence of a dicentric chromosome. Our analysis reveals the importance of Bim1p in maintaining a stable ipMT overlap zone by promoting polymerization of ipMTs during anaphase, whereas Kar3p contributes to spindle stability by cross-linking spindle MTs.
在出芽酵母中,有丝分裂纺锤体由32条动粒微管(kMTs)和约8条极间微管(ipMTs)组成。在后期开始时,kMTs向两极缩短,而ipMTs长度增加。重叠的微管负责在后期维持纺锤体的完整性。为了剖析后期纺锤体稳定性的要求,我们将一条条件性功能双着丝粒染色体引入酵母中。当来自同一条姐妹染色单体的着丝粒附着于相反两极时,后期纺锤体伸长会延迟,随后会出现一个依赖于DNA修复蛋白的DNA断裂-融合-桥循环。我们发现,双着丝粒染色体激活后的细胞存活需要微管结合蛋白Kar3p、Bim1p和Ase1p。在缺乏这些蛋白的情况下,后期纺锤体在双着丝粒染色体存在时容易坍塌和弯曲。我们的分析揭示了Bim1p在后期通过促进ipMTs聚合来维持稳定的ipMT重叠区的重要性,而Kar3p则通过交联纺锤体微管来促进纺锤体稳定性。