Loss of CHFR in human mammary epithelial cells causes genomic instability by disrupting the mitotic spindle assembly checkpoint
Loss of CHFR in human mammary epithelial cells causes genomic instability by disrupting the mitotic spindle assembly checkpoint
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DOI：
10.1593/neo.08176
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发表时间：
2008-07-01
期刊：
影响因子：
4.8
通讯作者：
Petty, Elizabeth M.
中科院分区：
文献类型：
--
作者：
Privette, Lisa M.;Weier, Jingly F.;Petty, Elizabeth M.
CHFR是一种E3泛素连接酶，是一种早期有丝分裂检查点蛋白，与许多癌症有关，并在维持基因组稳定性方面发挥着重要作用。为了分析CHFR在基因组稳定性中的作用，我们通过siRNA降低了CHFR在基因组稳定的MCF10A细胞中的表达。由于许多有丝分裂缺陷，CHFR表达降低很快导致非整倍体增加。首先，我们证实了CHFR与有丝分裂激酶Aurora A相互作用来调节其表达。此外，我们发现CHFR的减少会导致多极有丝分裂纺锤体的无序排列。这一发现得到了CHFR与α-微管蛋白的相互作用并可以调节其泛素化的发现的支持，以响应诺可达唑和乙酰化的α-微管蛋白的量，α-微管蛋白是有丝分裂纺锤体的组成部分。最后，我们发现了一个新的CHFR相互作用蛋白，纺锤体检查点蛋白MAD2。CHFR表达降低导致MAD2和BUBR1在有丝分裂过程中定位错误，并损害MAD2/CDC20复合体的形成。纺锤体检查点受损的进一步证据是CHFR基因敲除细胞中存在中期染色体错位、后期染色体滞后和胞质分裂缺陷。重要的是，我们的结果表明了CHFR调节染色体分离的新作用，其中表达减少，如在癌细胞中所见，通过损害纺锤体组装检查点而导致基因组不稳定。
CHFR is an E3 ubiquitin ligase and an early mitotic checkpoint protein implicated in many cancers and in the maintenance of genomic stability. To analyze the role of CHFR in genomic stability, by siRNA, we decreased its expression in genomically stable MCF10A cells. Lowered CHFR expression quickly led to increased aneuploidy due to many mitotic defects. First, we confirmed that CHFR interacts with the mitotic kinase Aurora A to regulate its expression. Furthermore, we found that decreased CHFR led to disorganized multipolar mitotic spindles. This was supported by the finding that CHFR interacts with alpha-tubulin and can regulate its ubiquitination in response to nocodazole and the amount of acetylated alpha-tubulin, a component of the mitotic spindle. Finally, we found a novel CHFR interacting protein, the spindle checkpoint protein MAD2. Decreased CHFR expression resulted in the mis-localization of both MAD2 and BUBR1 during mitosis and impaired MAD2/CDC20 complex formation. Further evidence of a compromised spindle checkpoint was the presence of misaligned metaphase chromosomes, lagging anaphase chromosomes, and defective cytokinesis in CHFR knockdown cells. Importantly, our results suggest a novel role for CHFR regulating chromosome segregation where decreased expression, as seen in cancer cells, contributes to genomic instability by impairing the spindle assembly checkpoint.