School of Biomedical Engineering, Shanghai Jiao Tong University, Shanghai, 200030, China.
School of life science, the University of Science and Technology of China, Hefei, Anhui, 230026, China.
Cell Res. 2021 Jul;31(7):758-772. doi: 10.1038/s41422-020-00463-9. Epub 2021 Jan 19.
Ca channels are essential to cell birth, life, and death. They can be externally activated by optogenetic tools, but this requires robust introduction of exogenous optogenetic genes for expression of photosensitive proteins in biological systems. Here we present femtoSOC, a method for direct control of Ca channels solely by ultrafast laser without the need for optogenetic tools or any other exogenous reagents. Specifically, by focusing and scanning wavelength-tuned low-power femtosecond laser pulses on the plasma membrane for multiphoton excitation, we directly induced Ca influx in cultured cells. Mechanistic study reveals that photoexcited flavins covalently bind cysteine residues in Orai1 via thioether bonds, which facilitates Orai1 polymerization to form store-operated calcium channels (SOCs) independently of STIM1, a protein generally participating in SOC formation, enabling all-optical activation of Ca influx and downstream signaling pathways. Moreover, we used femtoSOC to demonstrate direct neural activation both in brain slices in vitro and in intact brains of living mice in vivo in a spatiotemporal-specific manner, indicating potential utility of femtoSOC.
钙通道对于细胞的生、死至关重要。它们可以通过光遗传学工具外部激活，但这需要在生物系统中表达光敏蛋白的外源光遗传学基因的强大引入。在这里，我们提出了 femtoSOC，这是一种仅通过超快激光直接控制 Ca 通道的方法，而不需要光遗传学工具或任何其他外源试剂。具体来说，通过将调谐波长的低功率飞秒激光脉冲聚焦并扫描到质膜上进行多光子激发，我们直接诱导了培养细胞中的 Ca 内流。机制研究表明，光激发的黄素通过硫醚键共价结合到 Orai1 的半胱氨酸残基上，这有助于 Orai1 聚合形成储存操作的钙通道 (SOC)，而无需通常参与 SOC 形成的 STIM1，从而能够以全光学方式激活 Ca 内流和下游信号通路。此外，我们使用 femtoSOC 以时空特异性的方式在体外脑切片和体内活鼠完整大脑中直接进行神经激活，表明 femtoSOC 具有潜在的应用价值。
