A Ratiometric Fluorescent Biosensor Reveals Dynamic Regulation of Long-Chain Fatty Acyl-CoA Esters Metabolism
A Ratiometric Fluorescent Biosensor Reveals Dynamic Regulation of Long-Chain Fatty Acyl-CoA Esters Metabolism
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比例荧光生物传感器揭示长链脂肪酰基辅酶 A 酯代谢的动态调节
DOI：
10.1002/anie.202101731
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Wang Jing
中科院分区：
文献类型：
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作者：
Wang Weibo;Wei Qingpeng;Zhang Jiayuan;Zhang Meiqi;Wang Chuchen;Qu Renyu;Wang Yuan;Yang Guangfu;Wang Jing
尽管人们越来越认识到长链脂肪酰辅酶 A 酯 (LCACoA) 的生物学影响，但由于缺乏检测活细胞中 LCACoA 的方法，我们对这些酰基辅酶 A 分子的亚细胞分布和调节功能的了解有限。在这里，我们报告了一种基因编码荧光传感器的开发，该传感器能够对活细胞和亚细胞区室中的 LCACoA 进行比率定量。我们展示了这种 FadR-cpYFP 融合“LACSer 传感器”如何经历 LCACoA 诱导的构象变化，这些变化反映在易于检测的荧光响应中，并展示了实时监测人类细胞中 LCACoA 的概念验证。随后，我们在科学研究中应用了 LACSer，调查了 ACSL 酶的破坏如何差异性地降低胞浆和线粒体 LCACoA 水平，并展示酰基辅酶 A 结合蛋白 (ACBP) 的遗传破坏如何改变线粒体 LCACoA 的积累。
Despite increasing awareness of the biological impacts of long‐chain fatty acyl‐CoA esters (LCACoAs), our knowledge about the subcellular distribution and regulatory functions of these acyl‐CoA molecules is limited by a lack of methods for detecting LCACoAs in living cells. Here, we report development of a genetically encoded fluorescent sensor that enables ratiometric quantification of LCACoAs in living cells and subcellular compartments. We demonstrate how this FadR‐cpYFP fusion “LACSer sensor” undergoes LCACoA‐induced conformational changes reflected in easily detectable fluorescence responses, and show proof‐of‐concept for real‐time monitoring of LCACoAs in human cells. Subsequently, we applied LACSer in scientific studies investigating how disruption of ACSL enzymes differentially reduces cytosolic and mitochondrial LCACoA levels, and show how genetic disruption of an acyl‐CoA binding protein (ACBP) alters mitochondrial accumulation of LCACoAs.