1. Abstract
Genetically encoded reporters have greatly increased our understanding of biology, especially in neuroscience. While fluorescent reporters have been widely used, light delivery and phototoxicity have hindered their utility. Bioluminescence overcomes some of these challenges but requires the addition of an exogenous luciferin limiting its use. Using a modular approach we have engineered Autonomous Molecular BioluminEscent Reporter (AMBER), an indicator of membrane potential. Unlike other luciferase-luciferin bioluminescent systems AMBER encodes the genes to express both the luciferase and luciferin. AMBER is a voltage-gated luciferase coupling the functionalities of the Ciona voltage sensing domain (VSD) and bacterial luciferase, luxAB. When AMBER is co-expressed with the luciferin producing genes it reversibly switches the bioluminescent intensity as a function of membrane potential. Utilizing both biophysical and biochemical methods we show that unlike other voltage indicators AMBER modulates its enzymatic activity as a function of the membrane potential. AMBER shows a several fold increase in the luminescent (ΔL/L) output upon switching from off to on state when the cell is depolarized. In vivo expression of AMBER in C. elegans allowed detecting pharyngeal pumping action and mechanosensory neural activity from multiple worms simultaneously. Since we are able to report neural activity of multiple animals at the same time, we believe AMBER can be used in social behavior assays to elucidate the role of membrane potential underlying the behaviors.