College of Chemistry and Materials Science, Sichuan Normal University, Chengdu, Sichuan 610068, China; State Key Laboratory of Chemical Biology, Shanghai Institute of Organic Chemistry, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
State Key Laboratory of Chemical Biology, Shanghai Institute of Organic Chemistry, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai 200032, China; School of Chemistry and Materials Science, Hangzhou Institute for Advanced Study, University of Chinese Academy of Sciences, 1 Sub-lane Xiangshan, Hangzhou 310024, China.
J Mol Biol. 2024 Nov 15;436(22):168811. doi: 10.1016/j.jmb.2024.168811. Epub 2024 Oct 5.
Optineurin (OPTN), a multifunctional adaptor protein in mammals, plays critical roles in many cellular processes, such as vesicular trafficking and autophagy. Notably, mutations in optineurin are directly associated with many human diseases, such as amyotrophic lateral sclerosis (ALS). OPTN can specifically recognize Rab8a and the GTPase-activating protein TBC1D17, and facilitate the inactivation of Rab8a mediated by TBC1D17, but with poorly understood mechanism. Here, using biochemical and structural approaches, we systematically characterize the interaction between OPTN and Rab8a, revealing that OPTN selectively recognizes the GTP-bound active Rab8a through its leucine-zipper domain (LZD). The determined crystal structure of OPTN LZD in complex with the active Rab8a not only elucidates the detailed binding mechanism of OPTN with Rab8a but also uncovers a unique binding mode of Rab8a with its effectors. Furthermore, we demonstrate that the central coiled-coil domain of OPTN and the active Rab8a can simultaneously interact with the TBC domain of TBC1D17 to form a ternary complex. Finally, based on the OPTN LZD/Rab8a complex structure and relevant biochemical analyses, we also evaluate several known ALS-associated mutations found in the LZD of OPTN. Collectively, our findings provide mechanistic insights into the interaction of OPTN with Rab8a, expanding our understanding of the binding modes of Rab8a with its effectors and the potential etiology of diseases caused by OPTN mutations.
视神经病变诱导蛋白（Optineurin，OPTN）是哺乳动物中的一种多功能衔接蛋白，在许多细胞过程中发挥着关键作用，如囊泡运输和自噬。值得注意的是，OPTN 中的突变与许多人类疾病直接相关，如肌萎缩侧索硬化症（Amyotrophic Lateral Sclerosis，ALS）。OPTN 可以特异性识别 Rab8a 和 GTP 酶激活蛋白 TBC1D17，并促进 TBC1D17 介导的 Rab8a 失活，但具体机制尚不清楚。在这里，我们使用生化和结构方法系统地研究了 OPTN 和 Rab8a 之间的相互作用，揭示了 OPTN 通过其亮氨酸拉链结构域（Leucine-zipper Domain，LZD）选择性识别 GTP 结合的活性 Rab8a。确定的 OPTN LZD 与活性 Rab8a 复合物的晶体结构不仅阐明了 OPTN 与 Rab8a 结合的详细机制，还揭示了 Rab8a 与其效应物的独特结合模式。此外，我们证明 OPTN 的中心卷曲螺旋结构域和活性 Rab8a 可以同时与 TBC1D17 的 TBC 结构域相互作用，形成三元复合物。最后，基于 OPTN LZD/Rab8a 复合物结构和相关生化分析，我们还评估了 OPTN LZD 中发现的几种与 ALS 相关的突变。总之，我们的研究结果为 OPTN 与 Rab8a 的相互作用提供了机制上的见解，扩展了我们对 Rab8a 与其效应物结合模式的理解以及由 OPTN 突变引起的疾病的潜在病因。
