microRNA调节线粒体融合蛋白OPA1参与化疗药物诱导细胞凋亡的机制

miRNA is highly conserved endogenous single-strand non-coding RNA in eukaryotes,which serves the pivotal functions of regulating gene expression,development, anti-virus, hematopoiesis, proliferation, apoptosis, and and lipometabolism.Latest sudies have shown that miRNAs play key roles in regulating proliferation,differentiation and apoptosis.We have screened the potential targets in liver cancer that response to sorafenib,a chemotherapeutic drug-induced apoptosis. We found that OPA1 can function as a critical regulator in mitochondria fusion and fission, proliferation and apoptosis. OPA1 may be a tumor suppressor in liver cancer to regulate tumorigensis. Our data have shown that there is a significant differentiation for OPA1 protein expression between normal liver tissues and malignant liver tissues.OPA1 also is involved in regulating the sensitivity of tumor cells to chemtherapeutic drugs. Recently, we got some primary data that suggested that opa1 is regulated by a number of miRNAs. In this project, we will study how miRNAs regulate anti-tumor drugs-induced OPA1 down-regulation and mitochondria fragmentation and apoptosis. We will study how miRNAs affects mitochondria dynamics to regulate cancer cell proliferation,differentiation and apoptosis. Our study will provide more information of working mechanisms of miRNA in anti-tumor chemodrug-induced apoptosis via mito-proteins.

MicroRNA（miRNA）是一类内源性，具有调控功能的非编码RNA。miRNA参与维持包括发育、病毒防御、造血、器官形成和脂肪代谢等生理过程。最近研究表明miRNA在诱导细胞增殖、分化和凋亡中有非常关键的作用。在研究化疗药物治疗肝癌靶点时，我们筛选到一个敏感因子OPA1，它能够通过调控肝癌细胞线粒体分裂与融合来影响细胞增殖和凋亡；调节肿瘤细胞对化疗药物敏感性应答。研究发现OPA1在正常肝脏组织和癌变肝组织表达呈显著性差异。OPA1可能与肝癌的发生、发展和治疗密切相关。我们的最新实验结果表明，opa1基因受到多个miRNA调控。我们将进一步验证OPA1特异性miRNA的存在；在细胞和动物个体水平上研究化疗药物如何对miRNA进行调节，miRNA如何通过OPA1诱导线粒体损伤。这些工作的开展将为我们了解miRNA调节线粒体动态变化及其对细胞程序性死亡分子调控机制提供新思路。

结题摘要：.本研究利用体外培养人肝癌细胞为模型，在分子生物学水平上对miRNA调控OPA1参与化疗药物诱导的细胞凋亡进行了比较详细的探讨。初步阐明了miRNA27能够介导化疗药物Sorafenib诱导的细胞凋亡：1.体外Luciferase Report检测了miRNA-27,128和miRNA203三种miRNA，结果只有miRNA27能够与opa1的3’UTR结合，抑制opa1表达；miRNA27点突变抑制了miRNA27正常功能的发挥。2.细胞水平实验证明Sorafenib处理肝癌细胞能够上调miRNA27表达水平和抑制opa1蛋白表达。与luciferase report结果一致的是另外两种miRNA即miRNA128和203均没有明显变化。3. 细胞内实验表明miRNA27高表达（over-expression）能够抑制OPA1，增强Sorafenib诱导的细胞凋亡效应。4.miRNA27过表达能够增加Sorafenib诱导的肝癌细胞凋亡，这种效应能够被OPA1过表达降低。上述实验结果已经初步证实了我们提出的miRNA通过线粒体分裂融合蛋白OPA1调控化疗药物诱导的细胞凋亡。