Lim Liangzhong, Wei Yuanyuan, Lu Yimei, Song Jianxing
Department of Biological Sciences, Faculty of Science, National University of Singapore, Singapore.
NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, Singapore.
PLoS Biol. 2016 Jan 6;14(1):e1002338. doi: 10.1371/journal.pbio.1002338. eCollection 2016 Jan.
TAR-DNA-binding protein-43 (TDP-43) C-terminus encodes a prion-like domain widely presented in RNA-binding proteins, which functions to form dynamic oligomers and also, amazingly, hosts most amyotrophic lateral sclerosis (ALS)-causing mutations. Here, as facilitated by our previous discovery, by circular dichroism (CD), fluorescence and nuclear magnetic resonance (NMR) spectroscopy, we have successfully determined conformations, dynamics, and self-associations of the full-length prion-like domains of the wild type and three ALS-causing mutants (A315E, Q331K, and M337V) in both aqueous solutions and membrane environments. The study decodes the following: (1) The TDP-43 prion-like domain is intrinsically disordered only with some nascent secondary structures in aqueous solutions, but owns the capacity to assemble into dynamic oligomers rich in β-sheet structures. By contrast, despite having highly similar conformations, three mutants gained the ability to form amyloid oligomers. The wild type and three mutants all formed amyloid fibrils after incubation as imaged by electron microscopy. (2) The interaction with nucleic acid enhances the self-assembly for the wild type but triggers quick aggregation for three mutants. (3) A membrane-interacting subdomain has been identified over residues Met311-Gln343 indispensable for TDP-43 neurotoxicity, which transforms into a well-folded Ω-loop-helix structure in membrane environments. Furthermore, despite having very similar membrane-embedded conformations, three mutants will undergo further self-association in the membrane environment. Our study implies that the TDP-43 prion-like domain appears to have an energy landscape, which allows the assembly of the wild-type sequence into dynamic oligomers only under very limited condition sets, and ALS-causing point mutations are sufficient to remodel it to more favor the amyloid formation or irreversible aggregation, thus supporting the emerging view that the pathologic aggregation may occur via the exaggeration of functionally important assemblies. Furthermore, the coupled capacity of TDP-43 in aggregation and membrane interaction may critically account for its high neurotoxicity, and therefore its decoupling may represent a promising therapeutic strategy to treat TDP-43 causing neurodegenerative diseases.
TAR-DNA结合蛋白43（TDP-43）的C末端编码一个朊病毒样结构域，该结构域广泛存在于RNA结合蛋白中，其功能是形成动态寡聚体，而且令人惊讶的是，大多数导致肌萎缩侧索硬化症（ALS）的突变也集中在此区域。在此，基于我们之前的发现，通过圆二色性（CD）、荧光和核磁共振（NMR）光谱技术，我们成功测定了野生型和三种导致ALS的突变体（A315E、Q331K和M337V）全长朊病毒样结构域在水溶液和膜环境中的构象、动力学及自聚集情况。该研究揭示了以下几点：（1）TDP-43朊病毒样结构域在水溶液中本质上是无序的，仅具有一些新生二级结构，但具有组装成富含β-折叠结构的动态寡聚体的能力。相比之下，尽管三种突变体的构象高度相似，但它们获得了形成淀粉样寡聚体的能力。野生型和三种突变体在孵育后通过电子显微镜成像均形成了淀粉样纤维。（2）与核酸的相互作用增强了野生型的自组装，但引发了三种突变体的快速聚集。（3）已鉴定出一个位于Met311-Gln343残基上的与膜相互作用的亚结构域，它对TDP-43神经毒性不可或缺，在膜环境中会转变为折叠良好的Ω-环-螺旋结构。此外，尽管三种突变体在膜中的嵌入构象非常相似，但它们在膜环境中会进一步自聚集。我们的研究表明，TDP-43朊病毒样结构域似乎具有一种能量态势，这使得野生型序列仅在非常有限的条件下组装成动态寡聚体，而导致ALS的点突变足以将其重塑为更有利于淀粉样形成或不可逆聚集的状态，从而支持了一种新出现的观点，即病理性聚集可能是通过夸大功能重要的组装体而发生的。此外，TDP-43在聚集和膜相互作用方面的耦合能力可能是其高神经毒性的关键原因，因此将其解耦可能是治疗TDP-43所致神经退行性疾病的一种有前景的治疗策略。
