OPTIMIZATION OF AN OPTOGENETICALLY ACTIVATED INITIATOR OF APOPTOSIS

Abstract

Optogenetic tools provide a window into the world of cell signaling pathways and protein-protein interactions with a precision and elegance that other biological probes can struggle to achieve. The creation of optogenetic switches for the control of cell death pathways can provide insight into the mechanisms of apoptosis and form a basis for non-invasive, next generation therapeutic strategies. Previously, we have employed Cryptochrome 2 (Cry2)/CIB, a blue light photoreceptor protein - protein dimerization module from A. thaliana in conjunction with BAX, an OMM targeting pro-apoptotic protein, for light-mediated initiation of mitochondrial outer membrane permeabilization (MOMP) and downstream apoptosis. In this work, we are further developing our light activated Cry2-BAX system (henceforth referred to as "OptoBAX") for "one click" initiation of the BAX-mediated apoptotic cascade. We also report results of experimental efforts to optimize our optogenetic switch to reduce light-independent cell death (dark activation), and to enhance experimental control of our switch by manipulating photophysical properties associated with the Cry2/CIB interaction.