Molecular cloning and functional characterization of a GABA transporter from the CNS of the cabbage looper, Trichoplusia ni.
A cDNA encoding a GABA transporter in the caterpillar Trichoplusia ni has been cloned and expressed in baculovirus-infected insect cells. The cDNA contains an ORF encoding a 608-residue protein, designated TrnGAT. Hydropathy analysis of the deduced amino acid sequence suggests 12 transmembrane domains, a structure similar to that of all other cloned Na+/Cl(-)-dependent GABA transporters. The deduced amino acid sequence shows high identity with a GABA transporter (MasGAT) expressed in the embryo of Manduca sexta. Expression of TrnGAT mRNA was detected only in the brain. Sf21 cells infected with recombinant baculovirus exhibited a 20- to 30-fold increase in [3H]GABA uptake compared to control-infected cells. Several blockers of GABA uptake were used to determine the pharmacological profile of TrnGAT. Although most similar to mammalian neuronal GABA transporter GAT-1 in its kinetic properties, stoichiometry of ionic dependence and pharmacological properties, TrnGAT may be distinguished from mammalian GAT-1 by the inability of cyclic GABA analogues, such as nipecotic acid and its derivatives, to inhibit GABA uptake by the insect protein. The unique pharmacology of TrnGAT suggests that the GABA transport system in the lepidopteran CNS could be a useful target in the future development of rapidly-acting neuroactive agents used to control agriculturally-important insects.
Gao
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Department of Zoology, University of Western Ontario, London, Canada.
McLean
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H
Caveney
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Donly
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C
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000494:pharmacology; D000818:Animals; D016367:Baculoviridae; D001483:Base Sequence; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D050485:GABA Plasma Membrane Transport Proteins; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D026901:Membrane Transport Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009036:Moths; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009420:Nervous System / Q000378:metabolism*; D027361:Organic Anion Transporters; D016133:Polymerase Chain Reaction; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D018411:Spodoptera; D014162:Transfection; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D000596:Amino Acids; D002352:Carrier Proteins; D050485:GABA Plasma Membrane Transport Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D027361:Organic Anion Transporters; D011994:Recombinant Proteins; D005680:gamma-Aminobutyric Acid; D012964:Sodium