Department of Biology, University of Missouri-St. Louis, St. Louis, Missouri, United States of America.
PLoS One. 2022 Sep 29;17(9):e0275235. doi: 10.1371/journal.pone.0275235. eCollection 2022.
Parkinson's disease is the second most common age-related, neurodegenerative disease. A small collection of genes has been linked to Parkinson's disease including LRRK2, SAT1, and SNCA, the latter of which encodes the protein alpha-synuclein that aggregates in Lewy bodies as a hallmark of the disease. Overexpression of even wild-type versions of these genes can lead to pathogenesis, yet the regulatory mechanisms that control protein production of the genes are not fully understood. Pumilio proteins belong to the highly conserved PUF family of eukaryotic RNA-binding proteins that post-transcriptionally regulate gene expression through binding conserved motifs in the 3' untranslated region (UTR) of mRNA targets known as PUF Recognition Elements (PREs). The 3'UTRs of LRRK2, SNCA and SAT1 each contain multiple putative PREs. Knockdown (KD) of the two human Pumilio homologs (Pumilio 1 and Pumilio 2) in a neurodegenerative model cell line, SH-SY5Y, resulted in increased SNCA and LRRK2 mRNA, as well as alpha-synuclein levels, suggesting these genes are normally repressed by the Pumilio proteins. Some studies have indicated a relationship between Pumilio and microRNA activities on the same target, especially when their binding sites are close together. LRRK2, SNCA, and SAT1 each contain several putative microRNA-binding sites within the 3'UTR, some of which reside near PREs. Small RNA-seq and microRNA qPCR assays were performed in both wild type and Pumilio KD SH-SY5Y cells to analyze global and differential microRNA expression. One thousand four hundred and four microRNAs were detected across wild type and Pumilio KD cells. Twenty-one microRNAs were differentially expressed between treatments, six of which were previously established to be altered in Parkinson's disease patient samples or research models. Expression of ten miRs predicted to target LRRK2 and SNCA was verified by RT-qPCR. Collectively, our results demonstrate that Pumilios and microRNAs play a multi-faceted role in regulating Parkinson's disease-associated genes.
帕金森病是第二常见的与年龄相关的神经退行性疾病。一小部分基因与帕金森病有关，包括 LRRK2、SAT1 和 SNCA，后者编码的蛋白质α-突触核蛋白在路易体中聚集，是该疾病的标志。即使是这些基因的野生型版本的过表达也可能导致发病机制，但控制基因蛋白质产生的调节机制尚不完全清楚。Pumilio 蛋白属于高度保守的真核 RNA 结合蛋白 PUF 家族，通过结合 mRNA 靶标 3'非翻译区 (UTR) 中的保守基序，即 PUF 识别元件 (PREs)，在后转录水平上调节基因表达。LRRK2、SNCA 和 SAT1 的 3'UTR 都包含多个假定的 PRE。在神经退行性模型细胞系 SH-SY5Y 中敲低 (KD) 两种人类 Pumilio 同源物 (Pumilio 1 和 Pumilio 2)，导致 SNCA 和 LRRK2 mRNA 以及α-突触核蛋白水平增加，表明这些基因通常受到 Pumilio 蛋白的抑制。一些研究表明 Pumilio 与同一靶标上的 microRNA 活性之间存在关系，尤其是当它们的结合位点靠近时。LRRK2、SNCA 和 SAT1 的 3'UTR 都包含几个假定的 microRNA 结合位点，其中一些位于 PRE 附近。在野生型和 Pumilio KD SH-SY5Y 细胞中进行了小 RNA-seq 和 microRNA qPCR 测定，以分析全局和差异 microRNA 表达。在野生型和 Pumilio KD 细胞中检测到 1404 种 microRNAs。处理之间有 21 种 microRNAs 表达差异，其中 6 种在帕金森病患者样本或研究模型中已经改变。预测靶向 LRRK2 和 SNCA 的 10 个 miRs 的表达通过 RT-qPCR 进行了验证。总的来说，我们的结果表明 Pumilios 和 microRNAs 在调节帕金森病相关基因方面发挥着多方面的作用。
