Cloning and functional expression of a human Na+ and Cl--dependent neutraland cationic amino acid transporter B0+

A Na+-dependent neutral and cationic amino acid transport system (B0+) play s an important role in many cells and tissues; however, the molecular basis for this transport system is still unknown. To identify new transporters, the expressed sequence tag database was queried, and cDNA fragments with se quence similarity to the Na+/Cl--dependent neurotransmitter transporter fam ily were identified. Based on these sequences, rapid amplification of cDNA ends of human mammary gland cDNA was used to obtain a cDNA of 4.5 kilobases (kb). The open reading frame encodes a 642-amino acid protein named amino acid transporter B0+. Human ATB(0+) (hATB(0+)) is a novel member of the Na/Cl--dependent neurotransmitter transporter family with the highest sequenc e similarity to the glycine and proline transporters. Northern blot analysi s identified transcripts of similar to 4.5 kb and similar to 2 kb in the lu ng. Another tissue survey suggests expression in the trachea, salivary glan d, mammary gland, stomach, and pituitary gland. Electrophysiology and radio labeled amino acid uptake measurements were used to functionally characteri ze the transporter expressed in Xenopus oocytes. hATB(0+) was found to tran sport both neutral and cationic amino acids, with the highest affinity for hydrophobic amino acids and the lowest affinity for proline. Amino acid tra nsport was Na+ and Cl--dependent and was attenuated in the presence of 2-am inobicyclo-[2.2.1]-heptane-2-carboxylic acid, a system B0+ inhibitor. These characteristics are consistent with system B0+ amino acid transport. Thus, hATB(0+) is the first cloned B0+ amino acid transporter.