Live-cell lipid biochemistry reveals a role of diacylglycerol side-chain composition for cellular lipid dynamics and protein affinities
Live-cell lipid biochemistry reveals a role of diacylglycerol side-chain composition for cellular lipid dynamics and protein affinities
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DOI：
10.1073/pnas.1912684117
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发表时间：
2020-04-07
影响因子：
11.1
通讯作者：
Nadler, Andre
中科院分区：
文献类型：
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作者：
Schuhmacher, Milena;Grasskamp, Andreas T.;Nadler, Andre
每个细胞产生数千种不同的脂质种类，但缺乏对脂质化学多样性如何促进生物信号传导的深入了解，特别是因为缺乏定量研究活细胞中脂质功能的方法。使用的例子，二酰基甘油，突出的第二信使，我们在这里调查是否脂质化学多样性可以提供一个基础，细胞信号规范。我们产生了光笼脂质探针，它允许急性操纵不同的甘油二酯种类的质膜。结合uncaging实验与数学建模，我们能够确定结合常数的二酰基甘油-蛋白质相互作用，和动力学参数的二酰基甘油transbilayer运动和营业额在定量活细胞实验。引人注目的是，我们发现，亲和力和动力学变化的数量级，由于二酰基甘油侧链组成。这些差异足以解释二酰基甘油结合蛋白的差异募集，因此，不同的下游磷酸化模式。我们的方法代表了一个普遍适用的方法，用于阐明在定量活细胞实验中的亚细胞尺度上的单个脂质物种的生物学功能。
Every cell produces thousands of distinct lipid species, but insight into how lipid chemical diversity contributes to biological signaling is lacking, particularly because of a scarcity of methods for quantitatively studying lipid function in living cells. Using the example of diacylglycerols, prominent second messengers, we here investigate whether lipid chemical diversity can provide a basis for cellular signal specification. We generated photo-caged lipid probes, which allow acute manipulation of distinct diacylglycerol species in the plasma membrane. Combining uncaging experiments with mathematical modeling, we were able to determine binding constants for diacylglycerol-protein interactions, and kinetic parameters for diacylglycerol transbilayer movement and turnover in quantitative live-cell experiments. Strikingly, we find that affinities and kinetics vary by orders of magnitude due to diacylglycerol side-chain composition. These differences are sufficient to explain differential recruitment of diacylglycerol binding proteins and, thus, differing downstream phosphorylation patterns. Our approach represents a generally applicable method for elucidating the biological function of single lipid species on subcellular scales in quantitative live-cell experiments.