Department of Biochemistry, Pavillon de Recherche Appliquée Sur le Cancer, Université de Sherbrooke, Sherbrooke, Québec, Canada.
RNA Biol. 2021 Sep;18(9):1339-1353. doi: 10.1080/15476286.2020.1847866. Epub 2020 Dec 11.
RNAs are highly regulated at the post-transcriptional level in neurodegenerative diseases and just a few mutations can significantly affect the fate of neuronal cells. To date, the impact of G-quadruplex (G4) regulation in neurodegenerative diseases like Parkinson's disease (PD) has not been analysed. In this study, potential G4s located in deregulated genes related to the nervous system were initially identified and were found to be significantly enriched. Several G4 sequences found in the 5' untranslated regions (5'UTR) of mRNAs associated with Parkinson's disease were demonstrated to in fact fold by biochemical assays. Subcloning of the full-length 5'UTRs of these candidates upstream of a luciferase reporter system led to the demonstration that the G4s of both Parkin RBR E3 Ubiquitin Protein Ligase (PRKN) and Vacuolar Protein Sorting-Associated Protein 35 (VPS35) significantly repressed the translation of both genes in SH-SY5Y cells. Subsequently, a strategy of using label-free RNA affinity purification assays with either of these two G4 sequences as bait isolated the Guanine Nucleotide-Binding Protein-Like 1 (GNL1). The latter was shown to have a higher affinity for the G4 sequences than for their mutated version. This study sheds light on new RNA G-quadruplexes located in genes dysregulated in Parkinson disease and a new G4-binding protein, GNL1.
RNAs 在神经退行性疾病中高度受到转录后水平的调控，少数突变就可以显著影响神经元细胞的命运。迄今为止，G-四链体（G4）在神经退行性疾病如帕金森病（PD）中的调控作用尚未得到分析。在这项研究中，最初鉴定了与神经系统相关的失调基因中潜在的 G4，发现它们明显富集。通过生化测定证实，与帕金森病相关的 mRNA 的 5'非翻译区（5'UTR）中发现的几个 G4 序列实际上可以折叠。将这些候选物的全长 5'UTR 亚克隆到荧光素酶报告系统的上游，证明 Parkin RBR E3 泛素蛋白连接酶（PRKN）和液泡蛋白分选相关蛋白 35（VPS35）的 G4 显著抑制了 SH-SY5Y 细胞中这两个基因的翻译。随后，使用这两个 G4 序列之一作为诱饵的无标记 RNA 亲和纯化测定的策略分离出鸟嘌呤核苷酸结合蛋白样 1（GNL1）。结果表明，后者与 G4 序列的亲和力高于其突变版本。本研究揭示了位于帕金森病失调基因中的新的 RNA G-四链体和新的 G4 结合蛋白 GNL1。
