Tra2β对RAGE基因可变剪接的调控及对RAGE介导的Aβ神经毒性的影响

The receptor for advanced glycation endproducts (RAGE) gene expresses an important membrane receptor that mediates the neurotoxicity of Aβ in the brain. In addition, by alternative splicing, the gene also expresses another soluble protein isoform of RAGE (esRAGE, endogenous secretary RAGE) that is an antagonist against the function of RAGE and reduces the Aβ-mediated neurotoxicity. An imbalanced esRAGE /RAGE ratio is known to be closely related with Alzheimer’s disease (AD) pathogenesis. Thus, it is hypothesized that restoring the esRAGE/RAGE ratio to normal by interfering the alternative splicing of RAGE gene may decrease the neurotoxicity of Aβ and would provide a new effective strategy in AD prevention and treatment. In preliminary studies, we demonstrated that Tra2β plays an important role in regulating the alternative splicing of RAGE gene and the esRAGE/RAGE ratio. In addition, we also detected the significantly decreased expression of Tra2β proteins in the cell model of AD pathogenesis, and in brain autopsy slices of AD patient, which is consistent with the report showing the abnormal high expression of Tra2β in human postmortem brain tissue of AD patients. These results imply that Tra2β is probably involved in AD pathogenesis through regulating the alternative splicing of RAGE gene, supporting our hypothesis. The aim of this project is to examine the hypothesis by investigating the molecular mechanism underlying the regulation of alternative splicing of the RAGE gene by Tra2β from multiple aspects, including Tra2β over-expression or silence, characterization of nucleic acid sequences critical for Tra2β to bind with RAGE transcripts. Furthermore, we will study the effect of the Tra2β-regulated alternative splicing of the RAGE gene in early AD pathogenesis using the cell model of AD pathogenesis. The results will bring new data to the elucidation of AD pathogenesis and provide a basis for exploring the possibility to develop a new strategy for the prevention and treatment of AD.

RAGE是介导脑内Aβ神经毒性的膜受体，而RAGE基因经可变剪接产生的另一亚型—可溶性分泌型esRAGE能拮抗RAGE的功能。研究显示，esRAGE/RAGE比例失衡与阿尔茨海默病（AD）密切关联。通过干预RAGE基因可变剪接而降低Aβ神经毒性,可能在AD防治中起作用。但RAGE剪接机制尚不清楚。我们前期研究显示，Tra2β参与RAGE可变剪接并上调esRAGE/RAGE比例。在AD脑和细胞模型中，检测到Tra2β表达下调，提示Tra2β可能通过调控RAGE剪接而参与AD发生。因此，本项目将从Tra2β的表达、RAGE基因上的Tra2β高亲和序列方面，研究Tra2β对RAGE剪接的调控机制；并利用模拟AD早期病理的细胞模型，通过干预Tra2β表达，分析Tra2β对RAGE剪接的调控及其对AD早期病理过程中Aβ神经毒性的影响。

RAGE是介导脑内Aβ神经毒性的膜受体，而RAGE基因经可变剪接产生的另一亚型—可溶性分泌型esRAGE能拮抗RAGE的功能。研究显示，esRAGE/RAGE比例失衡与阿尔茨海默病（AD）密切关联。干预RAGE基因可变剪接,可能在AD防治中起作用。但RAGE剪接机制尚不清楚。本项目的主要研究目标是，探索前体mRNA剪接因子Tra2β对RAGE基因可变剪接的调控及其在AD早期病理发生中的作用。主要进展包括：（1）剪接因子Tra2β和hnRNP A1参与了RAGE 的剪接调控：我们在纯化的RAGE 前体mRNA剪接小体中，检测到Tra2β和hnRNP A1蛋白；在细胞中过量表达和抑制表达Tra2β和hnRNP A1，检测到mRAGE/esRAGE的剪接比例显著改变，且hnRNP A1 与Tra2β功能相反。提示这两种蛋白均可调控 RAGE的可变剪接。（2）葡萄糖代谢障碍可能通过调控Tra2β和hnRNP A1的表达来影响mRAGE/esRAGE剪接：利用葡萄糖代谢障碍模型模拟AD早期的神经元损伤，检测到mRAGE/esRAGE比例的变化趋势与AD脑内一致，以及Tra2β表达量的降低和hnRNP A1表达量的增加。利用过量表达Tra2β和抑制表达hnRNP A1干预Tra2β和hnRNP A1表达的变化，则由糖代谢障碍引起的mRAGE/esRAGE 剪接的改变也被减弱。提示葡萄糖代谢障碍可能通过调控Tra2β和hnRNP A1的表达来影响mRAGE/esRAGE剪接。（3）在AD病人血样的单个核细胞（PBMCs）中，检测到mRAGE/esRAGE 和 hnRNP A1/ Tra2β-1比例的同步改变，提示在AD病人PBMCs细胞中，可能同样存在Tra2β和hnRNP A1对 mRAGE/esRAGE剪接的调控。（4）鉴于前体mRNA剪接调控在脑疾病中的重要功能，我们还拓展了神经功能相关基因的剪接调控在脑缺血性神经元损伤中的功能的研究: 我们发现，脑缺血后，剪接蛋白NSSR1在星型胶质细胞中表达的显著上调，可以降低脑缺血引起的神经元损伤，其作用可能是通过调控星型胶质细胞中NCAM-L1和 CREB的前体mRNA可变剪接来完成的。.相关研究结果作为通讯作者已录用论文1篇（Glia，2015，in press），IF 5.5.另有1篇正在J Neurochem投稿修回中。