Despite recent advances in optogenetics, it remains challenging to manipulate gene expression in specific populations of neurons. We present a dual-protein switch system, Cal-Light, that translates neuronal-activity-mediated calcium signaling into gene expression in a light-dependent manner. In cultured neurons and brain slices, we show that Cal-Light drives expression of the reporter EGFP with high spatiotemporal resolution only in the presence of both blue light and calcium. Delivery of the Cal-Light components to the motor cortex of mice by viral vectors labels a subset of excitatory and inhibitory neurons related to learned lever-pressing behavior. By using Cal-Light to drive expression of the inhibitory receptor halorhodopsin (eNpHR), which responds to yellow light, we temporarily inhibit the lever-pressing behavior, confirming that the labeled neurons mediate the behavior. Thus, Cal-Light enables dissection of neural circuits underlying complex mammalian behaviors with high spatiotemporal precision.
A calcium- and light-gated switch to induce gene expression in activated neurons
Development of Cal-Light system
Schematic drawing of Cal-Light system. M13 and calmodulin proteins are fused to c-terminus and n-terminus of TEV protease (TEV-C and TEV-N), respectively. When Ca2+ arises in the cytosol, M13 and calmodulin bind to each other and subsequently TEV-C and TEV-N regain proteolytic functions. However, TEV protease cannot recognize TEVseq easily in a dark condition, because TEVseq is inserted at the c-terminus of AsLOV2 J-helix. Blue light causes a conformational change of J-helix making TEVseq unmasked. Cleaved tTA translocates to the nucleus and initiates gene expression.
Max Planck Florida Institute for Neuroscience
