BackgroundCold environments pose serious threats on human health, with increased risk for myocardial infarction, stroke, frostbite, and hypothermia. Acquired cold acclimation is required to minimize cold-induced injures and to improve metabolic health. However, the underlying mechanisms still remains to be fully elucidated. ObjectiveWe aimed to identify critical amino acids involved in cold acclimation and unmask regulatory mechanisms. MethodsA total of twenty male participants were recruited and followed up after 3 months’ natural cold exposure. Cold-induced vasodilation (CIVD) tests and clinical biochemical analysis were performed at baseline and after 3-months cold exposure, whilst blood samples were collected, and plasma amino acids analysis was performed using targeted metabolomics. To further confirm the effect of lysine on cold tolerance and explain the latent mechanism, mice were challenged with chronic cold exposure for 7 days with lysine supplement, then core and local surface temperature as well as thermogenesis activity were detected. ResultsContinuous cold exposure shortened the CIVD onset time and increased the average finger temperature. Levels of the plasma amino acids lysine and glycine were decreased in both humans and mice. Venn analysis from three datasets revealed that lysine was the only significantly changed plasma amino which strongly correlated with the altered CIVD. Moreover, mice sustained a relatively higher core temperature and surface temperature in the back, tail and paws upon lysine supplementation. Furthermore, lysine supplementation increased the level of histone H3K18cr and promoted the gene and protein expression of Cpt1a, Cpt2 and Cyp27a1 in liver. ConclusionOur work identified lysine as a critical amino acid for the remodeling hepatic histone crotonylation and contributing to cold acclimation.