ALS-Causing Mutations Significantly Perturb the Self-Assembly and Interaction with Nucleic Acid of the Intrinsically Disordered Prion-Like Domain of TDP-43.
ALS-Causing Mutations Significantly Perturb the Self-Assembly and Interaction with Nucleic Acid of the Intrinsically Disordered Prion-Like Domain of TDP-43.
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DOI：
10.1371/journal.pbio.1002338
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发表时间：
2016-01
期刊：
影响因子：
9.8
通讯作者：
Song J
中科院分区：
文献类型：
--
作者：
Lim L;Wei Y;Lu Y;Song J
TAR-DNA 结合蛋白 43 (TDP-43) C 末端编码广泛存在于 RNA 结合蛋白中的朊病毒样结构域，其功能是形成动态寡聚体，而且令人惊奇的是，它还包含大多数引起肌萎缩侧索硬化症 (ALS) 的突变。在这里，在我们之前的发现的推动下，通过圆二色性（CD）、荧光和核磁共振（NMR）光谱，我们成功地确定了野生型和三种引起ALS的突变体（A315E、Q331K和M337V）的全长朊病毒样结构域在水溶液和膜环境中的构象、动力学和自缔合。该研究解码了以下内容：（1）TDP-43朊病毒样结构域本质上是无序的，仅在水溶液中具有一些新生的二级结构，但具有组装成富含β-折叠结构的动态低聚物的能力。相比之下，尽管具有高度相似的构象，但三个突变体获得了形成淀粉样蛋白寡聚体的能力。通过电子显微镜观察，野生型和三种突变体在孵育后均形成淀粉样原纤维。 (2) 与核酸的相互作用增强了野生型的自组装，但引发了三个突变体的快速聚集。 (3) 在 TDP-43 神经毒性所必需的残基 Met311-Gln343 上发现了一个膜相互作用亚结构域，该亚结构域在膜环境中转化为良好折叠的 Ω-环-螺旋结构。此外，尽管具有非常相似的膜嵌入构象，但三个突变体将在膜环境中经历进一步的自缔合。我们的研究表明，TDP-43 朊病毒样结构域似乎具有能量景观，仅在非常有限的条件下才允许野生型序列组装成动态寡聚体，并且引起 ALS 的点突变足以对其进行重塑，以更有利于淀粉样蛋白形成或不可逆聚集，从而支持新出现的观点，即病理性聚集可能通过功能上重要的组装的夸大而发生。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，因此其解偶联可能代表了治疗TDP-43引起的神经退行性疾病的一种有前景的治疗策略。 TDP-43 的朊病毒结构域似乎具有仅在非常有限的条件下才允许寡聚化的能量景观；然而，导致肌萎缩侧索硬化症的 TDP-43 突变足以重塑该蛋白质，有利于淀粉样蛋白的形成。肌萎缩侧索硬化症（ALS）是最常见的致命运动神经元疾病。它在约 140 年前就被发现了，但该疾病的确切机制仍不清楚。 TAR-DNA 结合蛋白-43 (TDP-43) 被确定为约 97% ALS 和约 45% 额颞叶痴呆 (FTD) 患者中蛋白质内含物的主要成分，并且还在越来越多的其他神经退行性疾病（包括阿尔茨海默病）中观察到。 TDP-43 C 末端是一个关键结构域，它编码类似朊病毒的结构域，并且至关重要的是，它包含几乎所有导致 ALS 的突变。在这里，我们成功地确定了野生型和三种引起 ALS 的突变体的朊病毒样结构域在水溶液和膜环境中的构象、动力学和自关联。该研究表明，TDP-43 朊病毒样结构域似乎具有独特的能量景观，仅在非常有限的条件下才允许野生型序列组装成特定的寡聚体。有趣的是，引起 ALS 的点突变重塑了能量景观，有利于淀粉样蛋白的形成或不可逆的聚集，从而支持了新的观点，即病理聚集可能是通过功能上重要的组装的夸大而发生的。此外，TDP-43 的聚集和膜相互作用的耦合能力可能部分解释了其高神经毒性。因此，将这些因素解耦可能是治疗 TDP-43 介导的神经退行性疾病的一种有前途的治疗策略。
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. The prion-like domain of TDP-43 appears to have an energy landscape that allows oligomerisation only under very limited conditions; however, TDP-43 mutations that cause amyotrophic lateral sclerosis are sufficient to remodel the protein in favor of amyloid formation. Amyotrophic lateral sclerosis (ALS) is the most prevalent fatal motor neuron disease. It was identified ~140 years ago, but the exact mechanism underlying the disease has still not been well defined. TAR-DNA-binding protein-43 (TDP-43) was identified as the major component of the proteinaceous inclusions present in ~97% ALS and ~45% frontotemporal dementia (FTD) patients, and has also been observed in an increasing spectrum of other neurodegenerative disorders, including Alzheimer disease. The TDP-43 C-terminus is a key domain—it encodes a prion-like domain and, crucially, hosts almost all ALS-causing mutations. Here we have successfully determined the conformations, dynamics, and self-associations of the prion-like domains of both wild type and three ALS-causing mutants in both aqueous solutions and membrane environments. The study suggests that the TDP-43 prion-like domain appears to have a unique energy landscape, which allows the assembly of the wild-type sequence into specific oligomers only under very limited conditions. Intriguingly, ALS-causing point mutations remodel the energy landscape to favor amyloid formation or irreversible aggregation, thus supporting the emerging view that pathologic aggregation may occur via the exaggeration of functionally important assemblies. Furthermore, the coupled capacity of TDP-43 in aggregation and membrane interaction may partly account for its high neurotoxicity; decoupling these may therefore represent a promising therapeutic strategy to treat TDP-43-mediated neurodegenerative diseases.