AbstractCaloric restriction (CR) is known to promote health and longevity, likely via modification of the gut microbiota (GM). However, functional and metabolic changes induced in the GM during CR are still unidentified. Here, we investigated the short- and long-term effects of CR on the rat GM using a metaproteogenomic approach. We show that a switch from ad libitum (AL) low fat diet to CR in young rats is able to induce rapid and deep changes in their GM metaproteomic profile, related to a reduction of the Firmicutes/Bacteroidetes ratio and an expansion of lactobacilli. Specifically, we observed a significant change in the expression of the microbial enzymes responsible for short-chain fatty acid biosynthesis, with CR boosting propionogenesis and limiting butyrogenesis and acetogenesis. Furthermore, these CR-induced effects were maintained up to adulthood and started to be reversed after a short-term diet change. We also found that CR alters the abundance of an array of host proteins released in stool, mainly related to epithelial barrier integrity and inflammation. Hence, our results provide thorough information about CR-induced modifications to GM and host functional activity, and might constitute the basis for novel GM-based approaches aimed at monitoring the effectiveness of dietary interventions.