Cloning and functional characterization of a rat renal organic cation transporter isoform (rOCT1A).
Polyspecific organic cation transporters in the renal proximal tubule mediate the secretion of many clinically used drugs as well as endogenous metabolites. Recently, two organic cation transporters (rOCT1 and rOCT2) were cloned from rat kidney. In this study, we report the cloning and functional expression of an rOCT1 isoform, rOCT1A, from rat kidney. Genomic DNA cloning and sequencing demonstrated that rOCT1A is an alternatively spliced variant of rOCT1 with a deletion of 104 base pairs near the 5'-end. The uptake of [14C]tetraethylammonium (TEA) in oocytes injected with the cRNA-encoding rOCT1A was increased 16-fold over that in water-injected oocytes (29 +/- 2.8 pmol/oocyte/h versus 1.8 +/- 0.13 pmol/oocyte/h, mean +/- S.E., p < 0.05). [14C]TEA uptake in the cRNA-injected oocytes was saturable (Km = 42 +/- 11 microM) and was inhibited significantly by organic cations, including cimetidine and N1-methylnicotinamide. The amino acid sequence was deduced from the cDNA after examination of all three reading frames. Two overlapping open reading frames were found. Studies with synthetic constructs suggest that a functional organic cation transporter is encoded by the larger open reading frame. The larger open reading frame encodes a 430-amino acid protein (termed rOCT1A) that is 92% identical to rOCT1 and 57% identical to rOCT2. From hydropathy analysis, rOCT1A is predicted to have 10 transmembrane domains with both amino and carboxyl termini intracellular. RNase protection assays demonstrate the presence of rOCT1A mRNA transcripts in rat kidney cortex, medulla, and intestine. These studies demonstrate the presence of a functional, alternatively spliced organic cation transporter (rOCT1A) in rat kidney.
Zhang
L
L
Department of Biopharmaceutical Sciences, University of California, San Francisco, California 94143, USA.
Dresser
M J
MJ
Chun
J K
JK
Babbitt
P C
PC
Giacomini
K M
KM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics / Q000502:physiology*; D003001:Cloning, Molecular; D007668:Kidney / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D027702:Organic Cation Transporter 1; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D013757:Tetraethylammonium Compounds / Q000378:metabolism; D014982:Xenopus laevis
D002352:Carrier Proteins; D008565:Membrane Proteins; D027702:Organic Cation Transporter 1; D012333:RNA, Messenger; D013757:Tetraethylammonium Compounds