Optogenetic Control of Store-Operated Calcium Channels

Store-operated Ca²⁺ entry mediated by STIM1 and ORAI1 constitutes one of the major Ca²⁺ entry routes in mammalian cells. Physiologically, the molecular choreography of STIM1–ORAI1 coupling is initiated by endoplasmic reticulum (ER) Ca²⁺ store depletion with subsequent oligomerization of the STIM1 ER-luminal domain, followed by its redistribution towards the plasma membrane to gate ORAI1 channels. Over the past five years, we have engineered a series of genetically-encoded Ca²⁺ actuators (GECAs) derived from STIM and ORAI to enable optical control of Ca²⁺ signaling with unprecedented spatial and temporal resolution. STIM1 is an ER Ca²⁺ sensor that oligomerizes and conformational switch in response to store depletion, overcoming autoinhibition and forming clusters that then couples to ORAI channel. Two general engineering approaches have been applied to confer photosensitivity to STIM1: i) fusion of a photosensory module with SOAR/CAD to recapitulate reversible CC1-SOAR/CAD intramolecular trapping (Opto-CRAC-V1); ii) replacing the STIM1 luminal domain and the transmembrane domain with a light-inducible oligomerization domain (Opto-CRAC-V2). These tools enable light-dependent STIM-ORAI coupling to elicit Ca²⁺ influx through highly Ca²⁺-selective CRAC channels in both excitable and non-excitable cells.

Meanwhile, we designed a light-operated Ca²⁺ channel (designated LOCa) by inserting a plant-derived photosensory module into the intracellular loop of an engineered ORAI1 channel. LOCa displays biophysical features reminiscent of the ORAI1 channel, which enables precise optical control over Ca²⁺ signals and hallmark Ca²⁺-dependent physiological responses. Compared to traditional pharmacological or genetic tools, GECAs rival by high precision, rapid reversibility, facile tunability and non-invasiveness. These GECAs offer new non-invasive means to probe the structure-function relations of calcium channels and significant advances toward the use of optogenetics for immunomodulation and neuromodulation.