Genetic re-engineering of polyunsaturated phospholipid profile of Saccharomyces cerevisiae identifies a novel role for Cld1 in mitigating the effects of cardiolipin peroxidation.
Genetic re-engineering of polyunsaturated phospholipid profile of Saccharomyces cerevisiae identifies a novel role for Cld1 in mitigating the effects of cardiolipin peroxidation.
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DOI：
10.1016/j.bbalip.2018.06.016
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发表时间：
2018-10
期刊：
影响因子：
--
通讯作者：
Greenberg ML
中科院分区：
文献类型：
--
作者：
Lou W;Ting HC;Reynolds CA;Tyurina YY;Tyurin VA;Li Y;Ji J;Yu W;Liang Z;Stoyanovsky DA;Anthonymuthu TS;Frasso MA;Wipf P;Greenberger JS;Bayır H;Kagan VE;Greenberg ML
心磷脂（CL）是一种独特的磷脂，几乎完全定位于线粒体膜内，在那里它被合成。新合成的CL经过酰基重塑产生富含不饱和酰基的CL种。Cld1是酵母菌中唯一确定的CL特异性磷脂酶，是启动CL重塑途径所必需的。在高等真核生物中，氯化氯的过氧化作用，产生CLOX，与细胞信号事件有关，引发细胞凋亡。CLOX可以被酶水解，从而释放具有信号特性的脂质介质。我们之前的研究结果表明，CLD1表达上调是对氧化应激的反应，并且CL重塑的生理作用之一是去除过氧化的CL。为了利用强大的酵母模型来研究CLD1在CL过氧化中的功能，我们在酵母中表达H. brasiliensis Δ12-desaturase基因，然后合成PUFAs，并将其整合到CL物种中。利用LC-MS-based氧化还原磷脂组学，我们鉴定并定量了cld1Δ与WT细胞中CL和其他磷脂的分子种类。CLD1的缺失导致按时间排列的寿命、线粒体膜电位和呼吸能力的急剧下降，以及单羟基-过氧CL水平的增加，特别是在高度不饱和的CL物种中，包括四脂酰CL。此外，纯化的Cld1对CLOX表现出更高的亲和力，并且H2O2处理细胞增加了Cld1在对数生长期的表达。这些数据表明，CLD1的表达是减轻氧化应激所必需的。本研究的发现有助于我们全面了解CL重塑及其在减轻氧化应激中的作用。
Cardiolipin (CL) is a unique phospholipid localized almost exclusively within the mitochondrial membranes where it is synthesized. Newly synthesized CL undergoes acyl remodeling to produce CL species enriched with unsaturated acyl groups. Cld1 is the only identified CL-specific phospholipase in yeast and is required to initiate the CL remodeling pathway. In higher eukaryotes, peroxidation of CL, yielding CLOX, has been implicated in the cellular signaling events that initiate apoptosis. CLOX can undergo enzymatic hydrolysis, resulting in the release of lipid mediators with signaling properties. Our previous findings suggested that CLD1 expression is upregulated in response to oxidative stress, and that one of the physiological roles of CL remodeling is to remove peroxidized CL. To exploit the powerful yeast model to study functions of CLD1 in CL peroxidation, we expressed the H. brasiliensis Δ12-desaturase gene in yeast, which then synthesized PUFAs that are incorporated into CL species. Using LC-MS-based redox phospholipidomics, we identified and quantified the molecular species of CL and other phospholipids in cld1Δ vs. WT cells. Loss of CLD1 led to a dramatic decrease in chronological lifespan, mitochondrial membrane potential, and respiratory capacity, and increased levels of mono-hydroperoxy-CLs, particularly among the highly unsaturated CL species, including tetralinoleoyl-CL. In addition, purified Cld1 exhibited a higher affinity for CLOX, and treatment of cells with H2O2 increased CLD1 expression in the logarithmic growth phase. These data suggest that CLD1 expression is required to mitigate oxidative stress. The findings from this study contribute to our overall understanding of CL remodeling and its role in mitigating oxidative stress.
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