NAD+ augmentation restores mitophagy and limits accelerated aging in Werner syndrome

Metabolic dysfunction is a primary feature of Werner syndrome (WS), a human premature aging disease caused by mutations in the gene encoding the Werner (WRN) DNA helicase. WS patients exhibit severe metabolic phenotypes, but the underlying mechanisms are not understood, and whether the metabolic deficit can be targeted for therapeutic intervention has not been determined. Here we report impaired mitophagy and depletion of NAD+, a fundamental ubiquitous molecule, in WS patient samples and WS invertebrate models. WRN regulates transcription of a key NAD+ biosynthetic enzyme nicotinamide nucleotide adenylyltransferase 1 (NMNAT1). NAD+ repletion restores NAD+ metabolic profiles and improves mitochondrial quality through DCT-1 and ULK-1-dependent mitophagy. At the organismal level, NAD+ repletion remarkably extends lifespan and delays accelerated aging, including stem cell dysfunction, in Caenorhabditis elegans and Drosophila melanogaster models of WS. Our findings suggest that accelerated aging in WS is mediated by impaired mitochondrial function and mitophagy, and that bolstering cellular NAD+ levels counteracts WS phenotypes.


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All studies must disclose on these points even when the disclosure is negative. Sample sizes for behavioral experiments were determined by the current standard used for mice in behavioral neuroscience experiments, based on the minimal amount of mice required to detect significance with an alpha rate set at .05 in a standardly powered experiment. For imaging experiments, between 4 and 5 mice were imaged per experimental paradigm.
No data was excluded from the analysis.
To increase replication of the C. elegans-based studies, we used large amount of worms (upto 100 worms/group) and had at least two repeats. For all other experiments that were carried out, all attempts at replication were successful.
Animal/samples (mice) were assigned randomly to the various experimental groups, and mice were randomly selected for behavioral experiments.
In data collection and analysis (e.g., worm behavioral studies, worm imaging data analysis, as well as imaging and data analysis of EM), the performer(s) was blinded with experimental design. The primary fibroblast cell lines HT01 (#AG09599) and the WS01 (#AG03141) cells were acquired from Coriell Institute. The WRN-KD cells were siRNA knockdown in HT01 cells using WRN human siRNA ologo duplex (CAT#: SR322215, Origene). Briefly, siRNAs were incubated in Optimem with 4 ml RNA Interferin (siRNA transfection reagent, Polyplus) per 1 ng RNA for 15 min and added to complete media for a final concentration of 30 nM siRNA. After 3-day incubation, cells were applied for further experiments. Knockdown efficiency was examined by western blot. All other primary human fibroblasts (detailed in Table S1)