Identification of a novel dynein binding domain in nudel essential for spindle pole organization in Xenopus egg extract.
Identification of a novel dynein binding domain in nudel essential for spindle pole organization in Xenopus egg extract.
复制标题
DOI：
10.1074/jbc.m110.181578
复制
发表时间：
2011-01-07
期刊：
影响因子：
--
通讯作者：
Zheng Y
中科院分区：
文献类型：
--
作者：
Wang S;Zheng Y
核分布蛋白E(Nude)和核分布蛋白E样蛋白(Nudel或Ndel1)与LissBray1(LIS1)和dynein相互作用。这些相互作用被认为是动力蛋白功能所必需的。以往的研究表明，nude/Nudel的高度保守的N端直接与Lis1结合，这种结合对dynein的活性至关重要。相比之下，尽管nude/Nudel的C末端被报道与动力蛋白结合，但这种结合的功能意义仍不清楚。利用精子在非洲爪哇卵子提取物中的纺锤体组装试验和广泛的突变研究，我们已经在Nudel的前80个氨基酸中鉴定出一个高度保守的动力蛋白结合域。我们进一步证明，动力蛋白复合体中的动力蛋白中间链直接参与了这种相互作用。重要的是，我们证明了Nudel的dynein和Lis1结合域都是纺锤体极点组织所必需的。最后，我们报告了由Nudel免疫耗竭引起的纺锤体缺陷可以通过在非洲爪蛙卵提取液中增加1倍的Lis1浓度来挽救。这表明Nudel的N端的一个重要功能是在纺锤体组装过程中促进Lis1和dynein之间的相互作用。总之，我们的发现为进一步破译Nudel和Lis1调控动力蛋白的机制开辟了新的途径。
The nuclear distribution protein E (NudE) and nuclear distribution protein E-like (Nudel or Ndel1) interact with both lissencephaly 1 (Lis1) and dynein. These interactions are thought to be essential for dynein function. Previous studies have shown that the highly conserved N terminus of NudE/Nudel directly binds to Lis1, and such binding is critical for dynein activity. By contrast, although the C terminus of NudE/Nudel was reported to bind to dynein, the functional significance of this binding has remained unclear. Using the sperm-mediated spindle assembly assay in Xenopus egg extracts and extensive mutagenesis studies, we have identified a highly conserved dynein binding domain within the first 80 amino acids of Nudel. We further demonstrate that the dynein intermediate chain in the dynein complex is directly involved in this interaction. Importantly, we show that both the dynein and Lis1 binding domains of Nudel are required for spindle pole organization. Finally, we report that spindle defects caused by immuno-depletion of Nudel could be rescued by a 1-fold increase of Lis1 concentration in Xenopus egg extracts. This suggests that an important function of the N terminus of Nudel is to facilitate the interaction between Lis1 and dynein during spindle assembly. Together, our findings open up new avenues to further decipher the mechanism of dynein regulation by Nudel and Lis1.