A FRET sensor enables quantitative measurements of membrane charges in live cells
A FRET sensor enables quantitative measurements of membrane charges in live cells
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DOI：
10.1038/nbt.3828
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发表时间：
2017-04-01
影响因子：
46.9
通讯作者：
Gaus, Katharina
中科院分区：
文献类型：
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作者：
Ma, Yuanqing;Yamamoto, Yui;Gaus, Katharina
膜电荷在蛋白质运输和信号传导中具有关键作用。然而，细胞膜的有效静电势的定量仍然具有挑战性。我们开发了一种荧光膜电荷传感器（MCS），通过福斯特共振能量转移（FRET）报告活细胞膜电荷的变化。MCS永久附着于质膜的内小叶，并对静电势的变化显示出线性、可逆和快速的响应。该传感器可以监测改变静电势的各种细胞处理，例如带电脂质的掺入和重新分布以及胞质离子浓度的改变。将传感器应用于T细胞生物学，我们用它来识别免疫突触中的带电膜结构域。此外，我们发现静电相互作用阻止了T细胞受体的自发磷酸化，并有助于T细胞中信号簇的形成。
Membrane charge has a critical role in protein trafficking and signaling. However, quantification of the effective electrostatic potential of cellular membranes has remained challenging. We developed a fluorescence membrane charge sensor (MCS) that reports changes in the membrane charge of live cells via Forster resonance energy transfer (FRET). MCS is permanently attached to the inner leaflet of the plasma membrane and shows a linear, reversible and fast response to changes of the electrostatic potential. The sensor can monitor a wide range of cellular treatments that alter the electrostatic potential, such as incorporation and redistribution of charged lipids and alterations in cytosolic ion concentration. Applying the sensor to T cell biology, we used it to identify charged membrane domains in the immunological synapse. Further, we found that electrostatic interactions prevented spontaneous phosphorylation of the T cell receptor and contributed to the formation of signaling clusters in T cells.