Ji Yuhao, Heidari Ali, Nzigou Mombo Brice, Wegner Seraphine V
Institute of Physiological Chemistry and Pathobiochemistry, University of Münster 48149 Münster Germany
Chem Sci. 2023 Dec 11;15(3):1027-1038. doi: 10.1039/d3sc04815b. eCollection 2024 Jan 17.
Optogenetics has opened new possibilities in the remote control of diverse cellular functions with high spatiotemporal precision using light. However, delivering light to optically non-transparent systems remains a challenge. Here, we describe the photoactivation of light-oxygen-voltage-sensing domains (LOV domains) with generated light from a chemiluminescence reaction between luminol and HO. This activation is possible due to the spectral overlap between the blue chemiluminescence emission and the absorption bands of the flavin chromophore in LOV domains. All four LOV domain proteins with diverse backgrounds and structures (iLID, BcLOV4, nMagHigh/pMagHigh, and VVDHigh) were photoactivated by chemiluminescence as demonstrated using a bead aggregation assay. The photoactivation with chemiluminescence required a critical light-output below which the LOV domains reversed back to their dark state with protein characteristic kinetics. Furthermore, spatially confined chemiluminescence produced inside giant unilamellar vesicles (GUVs) was able to photoactivate proteins both on the membrane and in solution, leading to the recruitment of the corresponding proteins to the GUV membrane. Finally, we showed that reactive oxygen species produced by neutrophil like cells can be converted into sufficient chemiluminescence to recruit the photoswitchable protein BcLOV4-mCherry from solution to the cell membrane. The findings highlight the utility of chemiluminescence as an endogenous light source for optogenetic applications, offering new possibilities for studying cellular processes in optically non-transparent systems.
光遗传学为利用光以高时空精度远程控制多种细胞功能开辟了新的可能性。然而，将光传递到光学不透明系统仍然是一个挑战。在此，我们描述了利用鲁米诺与过氧化氢之间的化学发光反应产生的光对光-氧-电压传感结构域（LOV结构域）进行光激活。这种激活是可能的，因为蓝色化学发光发射与LOV结构域中黄素发色团的吸收带之间存在光谱重叠。使用珠聚集试验证明，所有四种具有不同背景和结构的LOV结构域蛋白（iLID、BcLOV4、nMagHigh/pMagHigh和VVDHigh）都通过化学发光被光激活。化学发光的光激活需要一个临界光输出，低于该临界光输出，LOV结构域会以蛋白质特征动力学恢复到其黑暗状态。此外，在巨型单层囊泡（GUV）内部产生的空间受限化学发光能够光激活膜上和溶液中的蛋白质，导致相应蛋白质募集到GUV膜上。最后，我们表明，嗜中性粒细胞样细胞产生的活性氧可以转化为足够的化学发光，以将可光开关蛋白BcLOV4-mCherry从溶液募集到细胞膜上。这些发现突出了化学发光作为光遗传学应用的内源性光源的实用性，为研究光学不透明系统中的细胞过程提供了新的可能性。
