Ratiometric NAD⁺ Sensors Reveal Subcellular\nNAD⁺ Modulators

Abstract

Mapping NAD⁺ dynamics in live cells and human\nis essential\nfor translating NAD⁺ interventions into effective therapies.\nYet, genetically encoded NAD⁺ sensors with better specificity\nand pH resistance are still needed for the cost-effective monitoring\nof NAD⁺ in both subcellular compartments and clinical samples.\nHere, we introduce multicolor, resonance energy transfer-based NAD⁺ sensors covering nano- to millimolar concentration ranges\nfor clinical NAD⁺ measurement and subcellular NAD⁺ visualization. The sensors captured the blood NAD⁺ increase\ninduced by NMN supplementation and revealed the distinct subcellular\neffects of NAD⁺ precursors and modulators. The sensors\nthen enabled high-throughput screenings for mitochondrial and nuclear\nNAD⁺ modulators and identified α-GPC, a cognition-related\nmetabolite that induces NAD⁺ redistribution from mitochondria\nto the nucleus relative to the total adenine nucleotides, which was\nfurther confirmed by NAD⁺ FRET microscopy.