Electrostatic interaction between polyglutamylated tubulin and the nexin-dynein regulatory complex regulates flagellar motility.
Electrostatic interaction between polyglutamylated tubulin and the nexin-dynein regulatory complex regulates flagellar motility.
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DOI：
10.1091/mbc.e17-05-0285
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发表时间：
2017-08-15
影响因子：
3.3
通讯作者：
Oda T
中科院分区：
文献类型：
--
作者：
Kubo T;Oda T
鉴定了真核生物鞭毛中多聚谷氨酰化微管蛋白的3D定位。多聚谷氨酰化微管蛋白位于微管和其交叉桥结构N-DRC之间的界面上。鞭毛运动受带负电荷的微管蛋白和带正电荷的N-DRC之间的静电相互作用调节。微管蛋白经历各种翻译后修饰。其中，多聚谷氨酰化参与真核生物鞭毛的运动和轴丝微管的稳定性。然而，它仍然不清楚，在多聚谷氨酰化微管蛋白精确定位在轴丝内，以及微管蛋白多聚谷氨酰化如何影响鞭毛运动。在这项研究中，我们确定了三维定位的多聚谷氨酰微管蛋白在衣原体鞭毛抗体标记和冷冻电子断层扫描。多聚谷氨酰化微管蛋白特异性地位于被称为连接蛋白-动力蛋白调节复合物（N-DRC）的微管跨桥结构附近。由于N-DRC带正电荷，我们假设多聚谷氨酰化微管蛋白和N-DRC之间存在静电相互作用，因此我们突变了DRC 4的氨基酸序列以修饰N-DRC的电荷。我们发现，增加和减少DRC 4上的正电荷导致鞭毛运动减少。此外，在一个结构缺陷的N-DRC的突变体减少运动部分恢复增加DRC 4上的正电荷。这些结果清楚地表明，鞭毛的跳动运动是由带负电荷的聚谷氨酰化微管蛋白和带正电荷的N-DRC之间形成的静电交叉桥维持的。
The 3D localization of polyglutamylated tubulin in eukaryotic flagella is identified. Polyglutamylated tubulins are located on the interface between the microtubule and its cross-bridging structure, the N-DRC. Flagellar motility is regulated by electrostatic interaction between negatively charged tubulins and positively charged N-DRC. Tubulins undergo various posttranslational modifications. Among them, polyglutamylation is involved in the motility of eukaryotic flagella and the stability of the axonemal microtubules. However, it remains unclear where polyglutamylated tubulin localizes precisely within the axoneme and how tubulin polyglutamylation affects flagellar motility. In this study, we identified the three-dimensional localization of the polyglutamylated tubulin in Chlamydomonas flagella using antibody labeling and cryo–electron tomography. Polyglutamylated tubulins specifically located in close proximity to a microtubule-cross-bridging structure called the nexin–dynein regulatory complex (N-DRC). Because N-DRC is positively charged, we hypothesized that there is an electrostatic interaction between the polyglutamylated tubulin and the N-DRC, and therefore we mutated the amino acid sequences of DRC4 to modify the charge of the N-DRC. We found that both augmentation and reduction of the positive charge on DRC4 resulted in reduced flagellar motility. Moreover, reduced motility in a mutant with a structurally defective N-DRC was partially restored by increasing the positive charge on DRC4. These results clearly indicate that beating motion of flagella is maintained by the electrostatic cross-bridge formed between the negatively charged polyglutamylated tubulins and the positively charged N-DRC.