Draviam V M, Shapiro I, Aldridge B, Sorger P K
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
EMBO J. 2006 Jun 21;25(12):2814-27. doi: 10.1038/sj.emboj.7601168. Epub 2006 Jun 8.
The correct formation of stable but dynamic links between chromosomes and spindle microtubules (MTs) is essential for accurate chromosome segregation. However, the molecular mechanisms by which kinetochores bind MTs and checkpoints monitor this binding remain poorly understood. In this paper, we analyze the functions of six kinetochore-bound MT-associated proteins (kMAPs) using RNAi, live-cell microscopy and quantitative image analysis. We find that RNAi-mediated depletion of two kMAPs, the adenomatous polyposis coli protein (APC) and its binding partner, EB1, are unusual in affecting the movement and orientation of paired sister chromatids at the metaphase plate without perturbing kinetochore-MT attachment per se. Quantitative analysis shows that misorientation phenotypes in metaphase are uniform across chromatid pairs even though chromosomal loss (CIN) during anaphase is sporadic. However, errors in kinetochore function generated by APC or EB1 depletion are detected poorly if at all by the spindle checkpoint, even though they cause chromosome missegregation. We propose that impaired EB1 or APC function generates lesions invisible to the spindle checkpoint and thereby promotes low levels of CIN expected to fuel aneuploidy and possibly tumorigenesis.
染色体与纺锤体微管（MTs）之间形成稳定而动态的连接对于准确的染色体分离至关重要。然而，动粒结合MTs以及检查点监测这种结合的分子机制仍知之甚少。在本文中，我们使用RNA干扰、活细胞显微镜检查和定量图像分析来分析六种与动粒结合的MT相关蛋白（kMAPs）的功能。我们发现，RNA干扰介导的两种kMAPs，即腺瘤性息肉病大肠杆菌蛋白（APC）及其结合伴侣EB1的缺失，在影响中期板上配对姐妹染色单体的运动和方向方面表现异常，而本身并不干扰动粒与MT的附着。定量分析表明，尽管后期的染色体丢失（CIN）是零星的，但中期的方向错误表型在染色单体对中是一致的。然而，由APC或EB1缺失产生的动粒功能错误即使导致染色体错分离，也很难被纺锤体检查点检测到。我们提出，EB1或APC功能受损会产生纺锤体检查点不可见的损伤，从而促进低水平的CIN，预计这会助长非整倍体并可能导致肿瘤发生。
