Gladstone Institute of Neurological Disease, University of California, San Francisco, California 94158, USA.
J Biol Chem. 2010 May 7;285(19):14777-90. doi: 10.1074/jbc.M109.093708. Epub 2010 Mar 10.
Huntington disease (HD) is caused by an expansion of more than 35-40 polyglutamine (polyQ) repeats in the huntingtin (htt) protein, resulting in accumulation of inclusion bodies containing fibrillar deposits of mutant htt fragments. Intriguingly, polyQ length is directly proportional to the propensity for htt to form fibrils and the severity of HD and is inversely correlated with age of onset. Although the structural basis for htt toxicity is unclear, the formation, abundance, and/or persistence of toxic conformers mediating neuronal dysfunction and degeneration in HD must also depend on polyQ length. Here we used atomic force microscopy to demonstrate mutant htt fragments and synthetic polyQ peptides form oligomers in a polyQ length-dependent manner. By time-lapse atomic force microscopy, oligomers form before fibrils, are transient in nature, and are occasionally direct precursors to fibrils. However, the vast majority of fibrils appear to form by monomer addition coinciding with the disappearance of oligomers. Thus, oligomers must undergo a major structural transition preceding fibril formation. In an immortalized striatal cell line and in brain homogenates from a mouse model of HD, a mutant htt fragment formed oligomers in a polyQ length-dependent manner that were similar in size to those formed in vitro, although these structures accumulated over time in vivo. Finally, using immunoelectron microscopy, we detected oligomeric-like structures in human HD brains. These results demonstrate that oligomer formation by a mutant htt fragment is strongly polyQ length-dependent in vitro and in vivo, consistent with a causative role for these structures, or subsets of these structures, in HD pathogenesis.
亨廷顿病（HD）是由亨廷顿蛋白（htt）中超过 35-40 个谷氨酰胺（polyQ）重复扩展引起的，导致包含突变 htt 片段纤维状沉积物的包含体的积累。有趣的是，polyQ 长度与 htt 形成纤维的倾向以及 HD 的严重程度成正比，与发病年龄成反比。尽管 htt 毒性的结构基础尚不清楚，但介导 HD 中神经元功能障碍和退化的毒性构象体的形成、丰度和/或持久性也必须依赖于 polyQ 长度。在这里，我们使用原子力显微镜证明突变 htt 片段和合成 polyQ 肽以 polyQ 长度依赖性的方式形成寡聚物。通过延时原子力显微镜，寡聚体在纤维形成之前形成，本质上是瞬态的，并且偶尔是纤维的直接前体。然而，绝大多数纤维似乎通过单体添加形成，与寡聚体的消失同时发生。因此，寡聚体必须在形成纤维之前经历主要的结构转变。在永生化纹状体细胞系和 HD 小鼠模型的脑匀浆中，突变 htt 片段以 polyQ 长度依赖性的方式形成寡聚体，其大小与体外形成的寡聚体相似，尽管这些结构在体内随时间积累。最后，使用免疫电子显微镜，我们在人类 HD 大脑中检测到类似寡聚体的结构。这些结果表明，突变 htt 片段的寡聚体形成在体外和体内强烈依赖于 polyQ 长度，这与这些结构或这些结构的子集在 HD 发病机制中的因果作用一致。
