Institute of Medical Science and Technology, National Sun Yat-sen University, No. 70 Lien-hai Road, Kaohsiung 80424, Kaohsiung 804, Taiwan.
Department of Chemistry, National Sun Yat-sen University, No. 70 Lien-hai Road, Kaohsiung 80424, Kaohsiung 804, Taiwan.
ACS Chem Neurosci. 2022 Apr 20;13(8):1143-1164. doi: 10.1021/acschemneuro.1c00646. Epub 2022 Apr 8.
The accumulation of β-sheet-rich α-synuclein (α-Syn) protein in human brain cells is a pathological hallmark of Parkinson's disease (PD). Moreover, it has been reported that familial PD mutations (A30P, E46K, H50Q, G51D, and A53T) accumulate at an accelerated rate both in vivo and in vitro. In addition, accumulations of various C-terminal α-Syn truncations, such as C-terminal-truncated N103 α-synuclein (N103), were found in an aggregated form in the brain tissue of PD patients. Fluorescent protein-tagged wild-type α-Syn, A30P, E46K, H50Q, G51D, A53T, and N103 were transfected into HEK293T and SHSY5Y cells, and their diffusion behaviors were investigated with a custom-built fluorescence microscope system. Based on our experimental results, the oligomerization of α-Syn is a time-dependent process in both HEK293T and SHSY5Y cells, and the oligomer state approaches a plateau after 48 h of transfection. The change in the oligomeric state of E46K, H50Q, and G51D exhibited a similar trend to the wild type at a lower concentration but became intense at a higher concentration. A53T and N103 possess smaller diffusion coefficients than wild-type α-synuclein and other family PD mutations, indicating that these two mutants could form higher oligomeric states or stronger interactions in HEK293T and SHSY5Y cells. In contrast, the smallest oligomer and the lowest intracellular interaction among all investigated α-Syn variants were found for A30P. These phenomena indicated the presence of different pathogeneses among familial PD mutants and C-terminal α-Syn truncations.
β-折叠丰富的α-突触核蛋白（α-Syn）在人脑细胞中的积累是帕金森病（PD）的病理学标志。此外，已经报道家族性 PD 突变（A30P、E46K、H50Q、G51D 和 A53T）在体内和体外都以加速的速度积累。此外，在 PD 患者的脑组织中发现了各种 C 端α-Syn 截断物，如 C 端截断的 N103α-突触核蛋白（N103），以聚集的形式存在。荧光蛋白标记的野生型α-Syn、A30P、E46K、H50Q、G51D、A53T 和 N103 被转染到 HEK293T 和 SHSY5Y 细胞中，并使用定制的荧光显微镜系统研究它们的扩散行为。根据我们的实验结果，α-Syn 的寡聚化是一个在 HEK293T 和 SHSY5Y 细胞中都依赖时间的过程，并且在转染后 48 小时寡聚状态接近平台。E46K、H50Q 和 G51D 的寡聚状态变化在较低浓度下表现出与野生型相似的趋势，但在较高浓度下变得强烈。A53T 和 N103 的扩散系数比野生型α-突触核蛋白和其他家族性 PD 突变体小，这表明这两个突变体在 HEK293T 和 SHSY5Y 细胞中可能形成更高的寡聚状态或更强的相互作用。相比之下，在所有研究的α-Syn 变体中，A30P 的最小寡聚体和细胞内相互作用最低。这些现象表明家族性 PD 突变体和 C 端α-Syn 截断物之间存在不同的发病机制。
