Kinesin-14 Family Proteins HSET/XCTK2 Control Spindle Length by Cross-Linking and Sliding Microtubules
Kinesin-14 Family Proteins HSET/XCTK2 Control Spindle Length by Cross-Linking and Sliding Microtubules
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DOI：
10.1091/mbc.e08-09-0971
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发表时间：
2009-03-01
影响因子：
3.3
通讯作者：
Walczak, Claire E.
中科院分区：
文献类型：
--
作者：
Cai, Shang;Weaver, Lesley N.;Walczak, Claire E.
Kinesin-14家族蛋白是一种负末端定向马达，在纺锤体组装过程中起着关键作用。我们先前表明，非洲爪蟾驱动蛋白-14 XCTK 2通过XCTK 2尾部的二分NLS与importin α/β的结合受到Ran的调节，该结合调节XCTK 2在纺锤体形成期间交联微管的能力。在这里，我们表明，突变的核定位信号（NLS）的人驱动蛋白-14 HSET引起的积累在细胞质中，这导致了强烈的微管捆绑。HeLa细胞中的HSET过表达导致更长的纺锤体，类似于提取物中XCTK 2的NLS突变体所观察到的，表明驱动蛋白-14蛋白在提取物和体细胞中发挥类似的作用。相反，通过RNAi敲低HSET导致较短的纺锤体，但不影响极的形成。纺锤体长度的变化不依赖于K-纤维，因为Nuf 2 RNAi消除K-纤维导致纺锤体长度增加，这部分地被HSET的co-RNAi挽救。然而，这些纺锤体长度的变化确实需要微管滑动，因为HSET突变体的过表达使其滑动活性与其ATP酶活性解偶联，导致细胞的纺锤体长度短于过表达野生型HSET的细胞。我们的研究结果与Ran调节驱动蛋白-14与输入蛋白α/β的关联以通过在间期期间在细胞核中隔离驱动蛋白-14来防止微管的异常交联和捆绑的模型一致。驱动蛋白-14在有丝分裂过程中起作用，在平行微管之间交联和滑动，以调节纺锤体长度。
Kinesin-14 family proteins are minus-end directed motors that cross-link microtubules and play key roles during spindle assembly. We showed previously that the Xenopus Kinesin-14 XCTK2 is regulated by Ran via the association of a bipartite NLS in the tail of XCTK2 with importin alpha/beta, which regulates its ability to cross-link microtubules during spindle formation. Here we show that mutation of the nuclear localization signal (NLS) of human Kinesin-14 HSET caused an accumulation of HSET in the cytoplasm, which resulted in strong microtubule bundling. HSET overexpression in HeLa cells resulted in longer spindles, similar to what was seen with NLS mutants of XCTK2 in extracts, suggesting that Kinesin-14 proteins play similar roles in extracts and in somatic cells. Conversely, HSET knockdown by RNAi resulted in shorter spindles but did not affect pole formation. The change in spindle length was not dependent on K-fibers, as elimination of the K-fiber by Nuf2 RNAi resulted in an increase in spindle length that was partially rescued by co-RNAi of HSET. However, these changes in spindle length did require microtubule sliding, as overexpression of an HSET mutant that had its sliding activity uncoupled from its ATPase activity resulted in cells with spindle lengths shorter than cells overexpressing wild-type HSET. Our results are consistent with a model in which Ran regulates the association of Kinesin-14s with importin alpha/beta to prevent aberrant cross-linking and bundling of microtubules by sequestering Kinesin-14s in the nucleus during interphase. Kinesin-14s act during mitosis to cross-link and slide between parallel microtubules to regulate spindle length.