RIP140通过抑制SIRT5表达介导心肌细胞能量代谢紊乱（英文）

Repression of SIRT5 is Involved in RIP140-mediated Metabolic Dysregulation in Cardiomyocytes

【目的】探讨去琥珀酰化酶Sirtuin5(SIRT5)对受体相互作用蛋白140(RIP140)诱导的心肌线粒体能量代谢紊乱的影响。【方法】利用腺病毒感染心肌细胞诱导RIP140过表达,利用质粒转染诱导SIRT5基因过表达,si RNA干扰敲低RIP140、SIRT5表达;q RT-PCR和Western blot检测SIRT5、RIP140的表达变化;q RT-PCR检测线粒体编码基因的表达情况,TMRE染色法评估线粒体膜电位,试剂盒检测细胞耗氧和ATP生成情况。所有数据均来自至少3次的独立实验。【结果】过表达RIP140明显抑制SIRT5的表达(P<0.05);敲低内源性RIP140能增加SIRT5的表达(P<0.05)。过表达RIP140亦能诱导线粒体蛋白高度琥珀酰化,抑制SIRT5的去琥珀酰化酶活性。除此,RIP140能诱导线粒体功能紊乱和能量代谢损伤作用,表现为线粒体编码基因的表达抑制(P<0.05),线粒体膜电位降低(P<0.05),细胞耗氧和ATP合成减少(P<0.05),而过表达SIRT5能抑制RIP140介导的上述能量代谢损伤(P<0.05)。此外,SIRT5受RIP140的负性调控作用依赖于过氧化物增殖体激活受体α(PPARα)。【结论】RIP140通过抑制SIRT5诱导心肌细胞线粒体功能紊乱和能量代谢损伤。

【Objective】 To investigate the effect of desuccinylase Sirtuin5(SIRT5) on receptor-interacting protein 140(RIP140)-mediated metabolic dysfunction in cardiomyocytes.【Methods】RIP140 was overexpressed by Adenovirus infection and SIRT5 was overexpressed by plasmid transfection. RIP140 and SIRT5 were knocked down by si RNA interference. The expression of RIP140 and SIRT5 were measured by q RT-PCR and western blot. The transcription levels of mitochondrial DNA-encoded genes were detected by q RT-PCR. Mitochondrial membrane potential was detected by tetramethylrhodamine ethyl ester(TMRE)fluorescence anlysis. Cellular oxygen consumption and ATP production were investigated by assay kits. All data are from at least three independent experiments.【Results】RIP140 overexpression significantly attenuated SIRT5 expression(P<0.05),whereas knockdown of endogenous RIP140 elevated SIRT5 expression(P<0.05)in cardiomyocytes. Superabundant RIP140 also induced hypersuccinylation of mitochondrial proteins,suggesting RIP140 could repress the desuccinylase activity of SIRT5. Moreover,SIRT5 overexpression reversed RIP140-mediated mitochondrial dysfunction and energy metabolic impairment,such as repression of mitochondrial DNA-encoded genes(P<0.05),decrease of mitochondrial membrane potential(P<0.05),as well as reduction of cellular oxygen consumption(P<0.05)and ATP production(P<0.05). Furthermore,the regulation of RIP140 on SIRT5 was dependent on the peroxisome proliferatoractivated receptor α(PPARα)in cardiomyocytes.【Conclusion】RIP140 induces mitochondrial dysfunction and metabolic impairment through repression of SIRT5 in cardiomyocytes.