Interaction of WDR60 intermediate chain with TCTEX1D2 light chain of the dynein-2 complex is crucial for ciliary protein trafficking.
Interaction of WDR60 intermediate chain with TCTEX1D2 light chain of the dynein-2 complex is crucial for ciliary protein trafficking.
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DOI：
10.1091/mbc.e18-03-0173
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发表时间：
2018-07-01
影响因子：
3.3
通讯作者：
Nakayama K
中科院分区：
文献类型：
--
作者：
Hamada Y;Tsurumi Y;Nozaki S;Katoh Y;Nakayama K
动力蛋白 -2复合物通过为包含IFT - A和IFT - B复合物的纤毛内运输（IFT）机制提供动力来介导纤毛蛋白的运输。尽管已知有11个亚基组成动力蛋白 -2复合物，其中一些轻链亚基与动力蛋白 -1复合物共享，但动力蛋白 -2复合物的整体结构尚未完全阐明。利用可见免疫沉淀试验，我们展示了动力蛋白 -2亚基之间的相互作用模式，包括以前未明确的相互作用，例如WDR60与TCTEX1D2 - DYNLT1 / DYNLT3二聚体之间的相互作用。动力蛋白 -2复合物可分为三个亚复合物，即DYNC2H1 - DYNC2LI1、WDR34 - DYNLL1 / DYNLL2 - DYNLRB1 / DYNLRB2和WDR60 - TCTEX1D2 - DYNLT1 / DYNLT3。我们建立了缺乏WDR60或TCTEX1D2的细胞系，这两种都是由引起纤毛病的基因编码的动力蛋白 -2特异性亚基，并且发现WDR60敲除（KO）和TCTEX1D2 - KO细胞在纤毛蛋白逆行运输中都表现出缺陷，WDR60 - KO细胞表现出更严重的缺陷，可能是由于动力蛋白 -2复合物组装失败。缺乏TCTEX1D2结合的WDR60突变体的外源表达部分恢复了逆行运输，使其达到与TCTEX1D2 - KO细胞相当的水平。因此，我们的结果表明，在动力蛋白 -2复合物中，WDR60起主要作用，TCTEX1D2起辅助作用，以介导纤毛蛋白的逆行运输。
The dynein-2 complex mediates trafficking of ciliary proteins by powering the intraflagellar transport (IFT) machinery containing IFT-A and IFT-B complexes. Although 11 subunits are known to constitute the dynein-2 complex, with several light-chain subunits shared by the dynein-1 complex, the overall architecture of the dynein-2 complex has not been fully clarified. Utilizing the visible immunoprecipitation assay, we demonstrated the interaction modes among the dynein-2 subunits, including previously undefined interactions, such as that between WDR60 and the TCTEX1D2–DYNLT1/DYNLT3 dimer. The dynein-2 complex can be divided into three subcomplexes, namely DYNC2H1–DYNC2LI1, WDR34–DYNLL1/DYNLL2–DYNLRB1/DYNLRB2, and WDR60–TCTEX1D2–DYNLT1/DYNLT3. We established cell lines lacking WDR60 or TCTEX1D2, both of which are dynein-2–specific subunits encoded by ciliopathy-causing genes, and found that both WDR60-knockout (KO) and TCTEX1D2-KO cells show defects in retrograde ciliary protein trafficking, with WDR60-KO cells demonstrating more severe defects probably due to failed assembly of the dynein-2 complex. The exogenous expression of a WDR60 mutant lacking TCTEX1D2 binding partially restored retrograde trafficking to a level comparable to that of TCTEX1D2-KO cells. Thus, our results demonstrated that WDR60 plays a major role and TCTEX1D2 plays an auxiliary role in the dynein-2 complex to mediate retrograde ciliary protein trafficking.