The circadian protein CLOCK regulates cell metabolism via the mitochondrial carrier SLC25A10
The circadian protein CLOCK regulates cell metabolism via the mitochondrial carrier SLC25A10
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昼夜节律蛋白CLOCK通过线粒体载体SLC25A10调节细胞代谢
DOI：
10.1016/j.bbamcr.2019.03.016
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发表时间：
2019-08-01
影响因子：
5.1
通讯作者：
Lu, Chao
中科院分区：
文献类型：
--
作者：
Cai, Tingting;Hua, Bingxuan;Lu, Chao
生理功能和代谢调节是哺乳动物生物钟控制的最重要的输出。线粒体呼吸和ROS产生呈现节律性活动。线粒体载体负责线粒体物质的转移，对线粒体新陈代谢至关重要。Clock(Circadian Locomotor Output Cycle Kaput)是第一个在哺乳动物中发现的核心昼夜节律基因。然而，时钟蛋白是否可以通过线粒体载体调节线粒体功能尚不清楚。在这里，我们证明了Clock可以与线粒体载体SLC25A10结合。为了进一步分析，我们利用CRISPR/Cas9基因编辑技术建立了SLc25a10(-/-)-Hepal-6细胞系。Slc25a10(-/-)--Hepal-6细胞表现出葡萄糖稳态紊乱，氧化应激水平增加，电子传递链受损。接下来，利用免疫沉淀实验，我们发现SLC25A10中的43-84和169-210氨基酸是响应时钟结合的关键位点。最后，在SLc25a10(-/-)-Hepal-6细胞中强制表达野生型SLC25A10可以弥补SLC25A10的缺失，使糖代谢降低、严重的氧化应激和受损的电子传递链得到恢复。此外，一个突变的SLc25a10在两个关键位点发生了变化，没有表现出救援效果。综上所述，我们发现了一种新的蛋白质-蛋白质相互作用机制，在该机制中，Clock可以通过线粒体膜转运蛋白SLC25A10直接调节细胞代谢。我们的研究可能会对生物钟和线粒体代谢之间的关系提供一些新的见解。
Physiological function and metabolic regulation are the most important outputs of circadian clock controls in mammals. Mitochondrial respiration and ROS production show rhythmic activity. Mitochondrial carriers, which are responsible for mitochondrial substance transfer, are vital for mitochondrial metabolism. Clock (Circadian Locomotor Output Cycles Kaput) is the first core circadian gene identified in mammalian animals. However, whether CLOCK protein can regulate mitochondrial functions via mitochondrial carriers is unclear. Here, we showed that CLOCK can bind to the mitochondrial carrier SLC25A10. For further analysis, we established a Slc25a10(-/-)-Hepal-6 cell line using CRISPR/Cas9 gene-editing technology. Slc25a10(-/-)-Hepal-6 cells showed disordered glucose homeostasis, increased oxidative stress levels, and damaged electron transport chains. Next, using an immunoprecipitation assay, we found that amino acids 43-84 and 169-210 in SLC25A10 are key sites that respond to CLOCK binding. Finally, forced expression of wild-type SLC25A10 in Slc25a10(-/-)-Hepal-6 cells could compensate for the loss of SLC25A10; the decreased glucose metabolism, severe oxidative stress and damaged electron transport chain were recovered. In addition, a mutant Slc25a10 with changes in two key sites did not show a rescue effect. In conclusion, we identified a new protein-protein interaction mechanism in which CLOCK can directly regulate cell metabolism via the mitochondrial membrane transporter SLC25A10. Our study might provide some new insights into the relationship between circadian clock and mitochondrial metabolism.