Cloning and characterization of a brain-specific cationic amino acid transporter.
rCAT3 (rat cationic amino acid transporter 3), a cDNA that encodes a novel member of the murine CAT family was isolated. The protein encoded by rCAT3 contained 619 amino acids, 53-58% of which were identical with those of the murine CAT family proteins previously described (mouse CAT1, CAT2a, CAT2b, and rat CAT1). Transient expression of rCAT3 and L-[14C]arginine incorporation experiments in COS7 cells verified a high affinity system y+ transporter activity of rCAT3. First, rCAT3-mediated L-[14C]arginine incorporation was time-dependent and saturable with half-saturation constant (Km) values of 103 +/- 12 microM (mean +/- S.E., n = 3). Second, the incorporation was specific for cationic amino acids as evidenced from the inhibition by L-arginine, L-lysine, and L-ornithine. Third, neither sodium nor chloride ions in the extracellular medium were required for the activity. Fourth, the incorporation was inhibited by high potassium-induced membrane depolarization. On Northern blot using RNAs from various rat tissues, the expression of rCAT3 mRNA was restricted to the brain. These results indicated a role of rCAT3 in the system y+ transporter activity in the nervous tissue.
Hosokawa
H
H
Department of Pharmacology, Faculty of Medicine, Graduate School of Human and Environmental Studies, Kyoto University, Kyoto 606, Japan.
Sawamura
T
T
Kobayashi
S
S
Ninomiya
H
H
Miwa
S
S
Masaki
T
T
D000595:Amino Acid Sequence; D026923:Amino Acid Transport Systems, Basic; D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D001120:Arginine / Q000378:metabolism; D001483:Base Sequence; D001692:Biological Transport; D015152:Blotting, Northern; D001923:Brain Chemistry; D019556:COS Cells; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics*; D003001:Cloning, Molecular; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008239:Lysine / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D009952:Ornithine / Q000378:metabolism; D011188:Potassium / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats
D026923:Amino Acid Transport Systems, Basic; D000596:Amino Acids; D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D012333:RNA, Messenger; C105348:Slc7a3 protein, rat; D001120:Arginine; D009952:Ornithine; D019001:Nitric Oxide Synthase; D008239:Lysine; D011188:Potassium