Quantitative dynamics of intracellular NMN by genetically encoded biosensor

Area of Science:

• Biochemistry
• Cell Biology
• Metabolic Engineering

Background:

• Nicotinamide mononucleotide (NMN) is a key precursor to NAD+, vital for cellular energy and aging processes.
• Existing methods cannot measure live cell NMN dynamics, hindering research into its uptake and regulation.
• Understanding NMN metabolism is critical for addressing NAD+-related pathologies and aging.

Purpose of the Study:

• To develop genetically encoded sensors for quantifying subcellular NMN in live cells.
• To investigate the mechanisms of exogenous NMN and nicotinamide riboside (NR) uptake.
• To identify regulators of intracellular NMN levels and NMN/NAD+ ratios.

Main Methods:

• Engineering NMN-responsive protein scaffolds fused to luciferase and fluorescent proteins.
• Developing genetically encoded bioluminescent and fluorescent sensors.
• Quantifying subcellular NMN dynamics in live cells and different cellular compartments.

Main Results:

• The developed sensors successfully visualized NMN dynamics and uptake mechanisms in live cells.
• Subcellular NMN levels and NMN/NAD+ ratios were measured across compartments.
• Regulation of NMN by Nudts, Pxmp2, and identification of Slc25a45 as a potential mitochondrial regulator were observed.

Conclusions:

• Genetically encoded sensors offer a powerful new tool for real-time visualization of NMN metabolism.
• This technology facilitates deeper understanding of NMN uptake, regulation, and its role in aging.
• The findings pave the way for new therapeutic strategies targeting NAD+ metabolism.