Illuminating NAD Metabolism in Live Cells and In Vivo Using a Genetically Encoded Fluorescent Sensor.

Understanding of NAD metabolism provides many critical insights into health and diseases, yet highly sensitive and specific detection of NAD metabolism in live cells and in vivo remains difficult. Here, we present ratiometric, highly responsive genetically encoded fluorescent indicators, FiNad, for monitoring NAD dynamics in living cells and animals. FiNad sensors cover physiologically relevant NAD concentrations and sensitively respond to increases and decreases in NAD. Utilizing FiNad, we performed a head-to-head comparison study of common NAD precursors in various organisms and mapped their biochemical roles in enhancing NAD levels. Moreover, we showed that increased NAD synthesis controls morphofunctional changes of activated macrophages, and directly imaged NAD declines during aging in situ. The broad utility of the FiNad sensors will expand our mechanistic understanding of numerous NAD-associated physiological and pathological processes and facilitate screening for drug or gene candidates that affect uptake, efflux, and metabolism of this important cofactor.