线粒体蛋白ISCA2突变通过[4Fe-4S]合成缺陷介导脑病发生的机制研究

幼儿线粒体铁硫簇组装蛋白ISCA2 G77S突变引起的临床表现同呼吸链复合体中[4Fe-4S]亚基缺陷所致的线粒体脑病相似，但机制不清楚。课题组前期工作显示G77S突变选择性影响蛋白质结合[2Fe-2S]，而敲低Hela细胞ISCA2导致复合体活性降低及含[4Fe-4S]亚基下调，为此假设：①ISCA2中[2Fe-2S]的结合参与线粒体中[4Fe-4S]生成；②ISCA2缺陷能引起[4Fe-4S]合成障碍，进而影响氧化磷酸化系统最终促进脑病发生。本项目计划：①蛋白水平阐述ISCA2蛋白在铁硫簇组装中的功能；②细胞水平阐明ISCA2铁硫簇组装功能及在氧化磷酸化和能量代谢中的地位；③动物水平验证ISCA2条件性敲除是否会通过阻断铁硫簇组装影响线粒体能量代谢导致线粒体脑病。预期阐明ISCA2参与[4Fe-4S]组装及缺陷导致线粒体脑病发生的机制，从而为铁硫簇缺陷引起脑病的精准诊治提供理论基础。

Iron-sulfur clusters are involved in many important physiological processes ranging from electron transport to enzyme reaction catalyzation. Mitochondrial iron-sulfur cluster assembly protein ISCA2 G77S mutation would lead to a clinical manifestation which is similar to the mitochondria encephalopathy caused by mutation in complex-Ⅰsubunits which contain [4Fe-4S] cluster. Recently, our group conducted experiments and identified that ISCA2 protein can bind both mononuclear iron and the active [2Fe-2S] cluster. G77S mutation of ISCA2 selectively affects the [2Fe-2S] cluster binding activity of protein. In Hela cells, ISCA2 knockdown led to the decline of oxidative phosphorylation activity and down-regulation of subunits containing [4Fe-4S] cluster in respiratory chain. So we propose a hypothesis that the [2Fe-2S] cluster binding in ISCA2 is involved in the process of [4Fe-4S] cluster assembly in mitochondria. Deficiency of ISCA2 protein may lead to defect of [4Fe-4S] cluster biogenesis, which affect the function of oxidative phosphorylation (OXPHOS) and eventually contribute to encephalopathy. Our work will clarify the function of ISCA2 in the assembly of [4Fe-4S] cluster, and further analyse the importance of iron-sulfur cluster assembly function in mitochondrial oxidative phosphorylation and energy metabolism. At last, we will construct ISCA2 conditional knockout mouse model to observe whether ISCA2 knockout in nervous system would lead to mitochondria encephalopathy through the way of blocking the assembly of [4Fe-4S] cluster in mitochondrial. If the work finished, we may demonstrate the pathogenic mechanism of mitochondrial encephalopathy caused by ISCA2-mutation-mediated defect of [4Fe-4S] cluster biogenesis.