LncRNA B230334C09Rik通过miR-543-3p靶向调节GLT-1参与帕金森病发病的机制研究

The regulatory effect of lncRNA and miRNA in the occurrence of diseases has been increasingly valued, however, the research on the regulatory molecular mechanism of non-coding RNA in PD is still relatively scarce. Our previous study found that the expression of miR-543-3p was increased, and miR-543-3p down-regulates the expression of target molecule GLT-1 in PD mice. We also identify the sponge molecule of miR-543-3p, lncRNA B230334C09Rik. We found that inhibition of miR-543-3p can significantly improve pathological abnormalities in PD mice. So we hypothesize that the abnormality of lncRNA B230334C09Rik--miR-543-3p--GLT-1 axis plays a certain pathogenic effect in the pathogenesis of PD, which aggravates the neurotoxic effect of glutamate and promotes neuronal degeneration. In this project we will identify the important lncRNA, miRNA in the pathogenesis of PD, reveal the roles and mechanisms of LncRNA B230334C09Rik and miR-543-3p in the pathogenesis of PD from the molecular level, cellular level and overall level. We will explore the potential of miR-543-3p as therapeutic targets and early diagnostic markers. This study will provide a new approach for the treatment, prevention and drug development of PD.

LncRNA、miRNA在疾病发生中的调控效应越来越受到重视，但是目前PD中对非编码RNA的调控分子机制研究还相对匮乏。我们前期研究发现PD小鼠miR-543-3p表达增高，并下调了靶分子GLT-1表达，鉴定了miR-543-3p的海绵分子lncRNA B230334C09Rik，抑制miR-543-3p可以显著改善PD小鼠的病理学异常。据此推测：PD发病中lncRNA B230334C09Rik--miR-543-3p--GLT-1轴的异常发挥了一定的致病效应，加重了谷氨酸兴奋性神经毒性效应，促进了神经元退行性变化。本项目将鉴定PD发病中重要lncRNA、miRNA，从分子水平、细胞水平、整体水平揭示LncRNA B230334C09Rik、miR-543-3p在PD发病中的作用与机制，探讨miR-543-3p作为治疗靶标及早期诊断标志物的可能性。本项目将为PD防治和药物研发提供新线索。

本项目通过全转录组测序筛选了PD模型中差异表达的lncRNA、miRNA、circRNA和mRNA。鉴定了差异表达非编码RNA的靶基因，揭示了差异表达非编码RNA在多巴胺能神经元退行性变化中的作用及分子机制。我们发现miR-543-3p、miR-30a-5p、miR-186-3p、miR-186-5p在PD模型中高表达。miR-543-3p直接靶向GLT-1基因下调GLT-1表达加剧谷氨酸兴奋性毒性效应参与PD发病。抑制miR-543-3p可以显著提高PD小鼠黑质多巴胺能神经元TH表达水平，减少神经元凋亡，改善PD小鼠的运动功能障碍。我们发现miR-30a-5p增加GLT-1的泛素化降解，抑制miR-30a-5p可以减少GLT-1的泛素化降解，缓解谷氨酸的兴奋性毒性效应，保护多巴胺能神经元，改善PD小鼠运动功能障碍。我们发现转录因子RUNX3与miR-186-3p的启动子区结合，激活miR-186-3p的表达，miR-186-3p表达水平升高诱导多巴胺能神经元凋亡。miR-186-3p靶向结合slc6a3/igf1r基因3’UTR区，抑制slc6a3/igf1r基因蛋白表达。过表达IGF1R/DAT蛋白后，能够减少多巴胺能神经元凋亡，增加多巴胺水平，表明RUNX3-miR-186-3p-slc6a3/igf1r调控轴异常导致多巴胺能神经元凋亡参与了PD的发病。PD模型中miR-186-5p显著升高诱导多巴胺能神经元凋亡，敲低miR-186-5p可以抑制神经元凋亡。miR-186-5p与pik3ca基因3’UTR区靶向结合，下调pik3ca基因蛋白表达，抑制PI3K/AKT/mTOR通路诱导凋亡，过表达PIK3CA可以抑制凋亡，表明miR-186-5p-pik3ca调控轴异常参与了PD发病。本研究鉴定了PD发病中差异表达的非编码RNA，揭示了非编码RNA在PD发病过程中的作用及其分子机制，为PD的防治和药物研发提供了新线索。