Retinol dehydrogenase 11 (RDH11) is a microsomal short-chain dehydrogenase/reductase that recognizes all-trans– and cis–retinoids as substrates and prefers NADPH as a cofactor. Previous work has suggested that RDH11 contributes to the oxidation of 11-cis–retinol to 11-cis–retinaldehyde during the visual cycle in the eye’s retinal pigment epithelium. However, the role of RDH11 in metabolism of all-trans–retinoids remains obscure. Here, we report that microsomes isolated from the testes and livers of Rdh11/ mice fed a regular diet exhibited a 3- and 1.7-fold lower rate of all-trans–retinaldehyde conversion to all-trans–retinol, respectively, than the microsomes of WT littermates. Testes and livers of Rdh11/ mice fed a vitamin A– deficient diet had 35% lower levels of all-trans–retinol than those of WT mice. Furthermore, the conversion of -carotene to retinol via retinaldehyde as an intermediate appeared to be impaired in the testes of Rdh11//retinol-binding protein 4/ (Rbp4/) mice, which lack circulating holo RBP4 and rely on dietary supplementation with -carotene for maintenance of their retinoid stores. Together, these results indicate that in mouse testis and liver, RDH11 functions as an all-trans–retinaldehyde reductase essential for the maintenance of physiological levels of all-trans–retinol under reduced vitamin A availability.