Ikui Amy E, Furuya Kanji, Yanagida Mitsuhiro, Matsumoto Tomohiro
Department of Radiation Oncology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA.
J Cell Sci. 2002 Apr 15;115(Pt 8):1603-10. doi: 10.1242/jcs.115.8.1603.
To ensure accurate chromosome segregation, the spindle checkpoint delays the onset of sister chromatid separation when the spindle is not attached to a kinetochore. Mad2, a component of the checkpoint, targets fission yeast Slp1/budding yeast Cdc20/human p55CDC and prevents it from promoting proteolysis, which is a prerequisite to sister chromatid separation. The protein is localized to unattached kinetochores in higher eukaryotes, and it is thought to be required for activation of the checkpoint as well. In this study, Mad2 and its target Slp1 were visualized in a tractable organism, fission yeast Schizosaccharomyces pombe. When cells were arrested at a prometaphase-like stage, the Mad2-Slp1 complex was stable and the two proteins were colocalized to unattached kinetochores. When the spindle attachment was completed, the complex was no longer detectable and only Mad2 was found associated to the spindle. These results would suggest that unattached kinetochores provide sites for assembly of the Mad2-Slp1 complex. During interphase, Mad2 was localized to the nuclear periphery as well as to the chromatin domain. This localization was abolished in a yeast strain lacking Mad1, a protein that physically interacts with Mad2. Mad1 may anchor Mad2 to the nuclear membrane and regulate its entry into the nucleus.
为确保染色体准确分离，当纺锤体未附着于动粒时，纺锤体检查点会延迟姐妹染色单体分离的起始。检查点的一个组成部分Mad2作用于裂殖酵母Slp1/芽殖酵母Cdc20/人类p55CDC，并阻止其促进蛋白水解，而蛋白水解是姐妹染色单体分离的一个先决条件。在高等真核生物中，该蛋白定位于未附着的动粒，并且人们认为它也是激活检查点所必需的。在本研究中，在一种易于处理的生物体——裂殖酵母粟酒裂殖酵母中观察到了Mad2及其靶标Slp1。当细胞停滞在类似前中期的阶段时，Mad2-Slp1复合物是稳定的，并且这两种蛋白共定位于未附着的动粒。当纺锤体附着完成后，该复合物不再能被检测到，并且仅发现Mad2与纺锤体相关联。这些结果表明，未附着的动粒为Mad2-Slp1复合物的组装提供了位点。在间期，Mad2定位于核周以及染色质区域。在缺乏与Mad2发生物理相互作用的蛋白Mad1的酵母菌株中，这种定位被消除。Mad1可能将Mad2锚定在核膜上并调节其进入细胞核。
