基于miRNAs和谷氨酸神经递质系统的精神分裂症分子致病机制研究

Our previous study has found that several candidate gene of schizophrenia identified in the Han Chinese population were predicted to be targeted by microRNA137(miRNA137) which surpassed the genome-wide significance threshold. Among the genes, ZNF804A has been proved to be a target gene for miR137. Accordingly, we propose to systematically study the mechanism of miRNAs in schizophrenia pathogenesis. Firstly, differential expression profiles of miRNAs and mRNAs are built respectively by Solexa deep-sequencing technology and Agilent G4845A microarray. Secondly, we validate some miRNAs and mRNAs which show significant difference between cases and controls in a large sample size by Q-PCR and screen the significantly differential expression miRNAs as key miRNAs. Then, we predicted the target genes of key miRNAs based on the differential expression of mRNAs and bioinformatic analysis. Thirdly, we tested and verified target genes of the key miRNAs by biological method in HEK293 and SK-N-BE cell lines. Next, we study the upstream regulation of the key miRNAs using report gene, ChIP, etc. Finally, the expression of the target genes will be down-regulated and up-regulated in the schizophrenia rat model and cell model respectively to illustrate the mechanism of the target genes involved in the glutamate neurotransmitter system in schizophrenia pathogenesis. In summary, our research may help to understand the gene expression regulation, the mechanism of miRNAs and the hypothesis of altered glutamate neurotransmission implicated in schizophrenia, and provide new clue and valuable insight into the molecular basis of the disease.

我们前期鉴定的数个精神分裂症候选基因，经预测为具有全基因组关联水平的miRNA137的靶基因，其中ZNF804A已经被证实。鉴此，本项目拟系统地研究基于miRNAs的疾病致病机制。首先通过Solexa测序和芯片技术建立首发患者和正常对照的全基因组miRNAs和mRNAs的差异表达谱。其次，运用生物信息学和Q-PCR等方法，在扩大的样本中筛选差异表达的关键miRNAs及其可能的靶基因。第三，对关键miRNAs的靶基因进行生物学方法验证，并利用报告基因和ChIP等方法，明确关键miRNAs的靶基因及上游调控机制。最后，构建疾病大鼠模型和细胞模型，通过体内体外下调和上调靶基因的表达，阐明靶基因参与谷氨酸神经递质系统的作用机制。综上所述，此项研究将有助于拓展对基因表达调控、miRNAs作用机制以及谷氨酸神经递质系统缺陷学说的认识，为精神分裂症分子发病机制的研究提供重要的新线索和新思路。

基于课题组前期对精神分裂症候选基因的研究发现，结合当前全基因组关联研究的新进展。本项目以RNA为研究对象，首先建立精神分裂症患者和正常对照的全基因组miRNA表达差异谱和转录组基因表达差异谱；其次，在扩大的研究样本中进行miRNAs、mRNAs和lncRNAs的表达分析和验证，并运用生物信息学手段筛选出关键miRNAs（miR-137，miR-148b-3p，miR-19a，miR-181b，miR-195/497，miR-30e等），mRNAs（EFNB2，EGR3，ZNF804A等），以及lncRNAs （MALAT1，MIAT，LUCAT1，AMMECR1等）；第三，对关键的miRNAs及其靶基因进行相互作用验证和功能分析；最后，对重要靶基因，通过神经细胞模型和实验动物模型，鉴定并验证与该靶基因密切相关的神经递质系统成员，以期阐明miRNA及其靶基因在精神分裂症致病机制中的作用。迄今为止，课题组已完成项目的主要研究内容，并已在国内外重要学术刊物上发表论文12篇，其中SCI收录的学术论文8篇。同时，课题组主要成员获陕西高等学校科学技术奖一等奖1项。本项目的完成不仅拓展对基因表达调控、非编码RNA作用机制以及疾病相关神经递质系统功能缺陷的认识，而且为精神分裂症分子发病机制的研究提供重要的新线索和新思路。