Yoo Myungsik, Bunkowski Klaudia, Lie Andrew, Junn Eunsung
RWJMS Institute for Neurological Therapeutics, Department of Neurology, Rutgers -Robert Wood Johnson Medical School, Piscataway, NJ, 08854, USA.
Mol Biol Rep. 2024 Jul 13;51(1):797. doi: 10.1007/s11033-024-09725-w.
Parkinson's disease (PD) is a common neurodegenerative disorder characterized by a multifaceted genetic foundation. Genome-Wide Association Studies (GWAS) have played a crucial role in pinpointing genetic variants linked to PD susceptibility. Current study aims to delve into the mechanistic aspects through which the PD-associated Single Nucleotide Polymorphism (SNP) rs329648, identified in prior GWAS, influences the pathogenesis of PD.
Employing the CRISPR/Cas9-mediated genome editing mechanism, we demonstrated the association of the disease-associated allele of rs329648 with increased expression of miR-4697-3p in differentiated SH-SY5Y cells. We revealed that miR-4697-3p contributes to the formation of high molecular weight complexes of α-Synuclein (α-Syn), indicative of α-Syn aggregate formation, as evidenced by Western blot analysis. Furthermore, our study unveiled that miR-4697-3p elevates SNCA112 mRNA levels. The resultant protein product, α-Syn 112, a variant of α-Syn with 112 amino acids, is recognized for augmenting α-Syn aggregation. Notably, this regulatory effect minimally impacts the levels of full-length SNCA140 mRNA, as evidenced by qRT-PCR. Additionally, we observed a correlation between the disease-associated allele and miR-4697-3p with increased cell death, substantiated by assessments including cell viability assays, alterations in cell morphology, and TUNEL assays.
Our research reveals that the disease-associated allele of rs329648 is linked to higher levels of miR-4697-3p. This increase in miR-4697-3p leads to elevated SNCA112 mRNA levels, consequently promoting the formation of α-Syn aggregates. Furthermore, miR-4697-3p appears to play a role in increased cell death, potentially contributing to the pathogenesis of PD.
帕金森病（PD）是一种常见的神经退行性疾病，具有多方面的遗传基础。全基因组关联研究（GWAS）在确定与PD易感性相关的基因变异方面发挥了关键作用。当前研究旨在深入探究先前GWAS中鉴定出的与PD相关的单核苷酸多态性（SNP）rs329648影响PD发病机制的分子机制。
利用CRISPR/Cas9介导的基因组编辑机制，我们证明了rs329648的疾病相关等位基因与分化的SH-SY5Y细胞中miR-4697-3p表达增加有关。我们发现miR-4697-3p有助于α-突触核蛋白（α-Syn）高分子量复合物的形成，这表明α-Syn聚集体形成，蛋白质免疫印迹分析证明了这一点。此外，我们的研究表明miR-4697-3p可提高SNCA112 mRNA水平。产生的蛋白质产物α-Syn 112是一种含有112个氨基酸的α-Syn变体，已知可增强α-Syn聚集。值得注意的是，定量逆转录聚合酶链反应证明，这种调节作用对全长SNCA140 mRNA水平的影响最小。此外，我们观察到疾病相关等位基因和miR-4697-3p与细胞死亡增加之间存在相关性，包括细胞活力测定、细胞形态改变和TUNEL测定等评估证实了这一点。
我们的研究表明，rs329648的疾病相关等位基因与更高水平的miR-4697-3p有关。miR-4697-3p的这种增加导致SNCA112 mRNA水平升高，从而促进α-Syn聚集体的形成。此外，miR-4697-3p似乎在增加细胞死亡中起作用，可能有助于PD的发病机制。
