In olive fruits, sugars are the main soluble components providing energy and acting as precursors for olive oil biosynthesis. Large quantities of glucose, fructose and galactose are often found in olive pulp. To analyze sugar transport processes in Olea europaea, a cDNA encoding a monosaccharide transporter, designated OeMST2 (Olea europaea monosaccharide transporter 2) was cloned. An open reading frame of 1,569 bp codes for a protein of 523 amino acids and a calculated molecular weight of 57.6 kDa. The protein is homologous to other sugar transporters identified so far in higher plants. Expression of this cDNA in an hxt-null Saccharomyces cerevisiae strain deficient in glucose transport restored its capacity to grow on and to transport glucose. The encoded protein showed high affinity for D-glucose (K(m), 25 microM) and was also able to recognize D-galactose and the analogs 3-O-methyl-D-glucose and 2-deoxy-D-glucose, but not D-fructose, D-arabinose, sucrose or D-mannitol. Maximal transport activity was high at acidic pH (5.0), and the initial D-[(14)C]glucose uptake rates were strongly inhibited by the protonophore carbonyl cyanide m-chlorophenylhydrazone, confirming that OeMST2 is a H(+)/monosaccharide transporter. The expression of OeMST2 was studied during the ripening process. Transcript levels increased during fruit maturation, suggesting that OeMST2 takes part in the massive accumulation of monosaccharides in olive fruits. Monosaccharide:H(+) transport system activity and OeMST2 expression were negatively regulated by glucose in suspension-cultured cells. Glucose-mediated OeMST2 repression was impaired by mannoheptulose, suggesting the involvement of a hexokinase-dependent signaling pathway.