Cloning and functional expression of a mouse liver organic cation transporter

Abstract

Hepatic uptake of organic cations is essential for the metabolism and secretion of numerous endobiotics and drugs. Several hepatic organic cation transporters have been kinetically defined, yet have not been isolated or cloned. We have isolated a complementary DNA (cDNA) from both murine liver and kidney cDNA libraries (mOct1/ Slc22a1), and have functionally expressed it in Xenopus laevis oocytes. Although mOct1/Slc22a1 is homologous to previously cloned rat and human organic cation transporters, organic cation transport kinetics differed markedly. The liver and kidneys have critical roles in the secretion of drugs and endobiotics into the bile and urine. Hepatic elimination of many organic cationic drugs involves uptake across the basolateral surface of the hepatocyte, hepatic biotransformation, and canalicular secretion. 1 Previous studies in the intact rat, isolated perfused rat liver, and isolated hepatocytes show that the hepatobiliary transport of organic cations is carrier-mediated. [2][3][4] In addition, the organic cation choline is an essential nutrient and precursor for the synthesis of phospholipids. Hepatic choline levels are approximately 100-fold greater than blood concentrations, and de novo choline synthesis is minimal, suggesting the existence of a transport mechanism for hepatic choline uptake. 5,6 Several organic cation transporters have been kinetically defined on the plasma membrane of hepatocytes and in lysosomes. Functional studies in rat liver plasma membrane vesicles have shown at least two distinct organic cation:H ϩ exchangers, and two electrogenic organic cation transporters. [7][8][9][10][11][12][13] The complementary DNA (cDNA) encoding for the rat electrogenic polyspecific transporter (OCT1) has been isolated using functional expression cloning in Xenopus laevis oocytes, and assaying n-methyl-nicotinamide (NMN)-inhibitable uptake of the model quaternary organic cation tetraethylammonium (TEA). 14 A human homologue of rat OCT1 was recently cloned that transports 1-methyl-4-phenylpyridinium (MPP ϩ ), and uptake is also inhibited by the organic cation NMN. 15 In this study, we have isolated a murine cDNA (mOct1/ Slc22a1) from both liver and kidney libraries, and have functionally expressed it in X. laevis oocytes. Although mOct1/Slc22a1 is homologous with OCT1, transport kinetics differed markedly. mOct1/Slc22a1 transports several organic cations, but uptake is increased by inside:outside proton gradients, is unaffected by membrane potential differences, and is not inhibited by NMN. These data indicate that this cloned cDNA is a member of the hepatic and renal organic cation transporter family.

Metabolomics reveals an essential role for peroxisome proliferator-activated receptor α in bile acid homeostasis

“…Several studies have shown that Oct2 plays an important role in renal secretion of creatinine in humans and mice ( 30,31 ). Additionally, methylnicotinamide, used as a model substrate for studying organic cation transport, can be regulated by Oct1 and Oct2 ( 32,33 ). In vivo, nicotinamide 1-oxide and methylnicotinamide are derived from nicotinamide.…”

Polyspecific Organic Cation Transporters: Structure, Function, Physiological Roles, and Biopharmaceutical Implications

“…Like most members of the SLC22 family, the organic cation and carnitine transporters have a predicted membrane topology that comprises 12 a-helical transmembrane domains (TMDs), an intracellular N-terminus, a large glycosylated extracellular loop between TMDs 1 and 2, a large intracellular loop with phosphorylation sites between TMDs 6 and 7, and an intracellular C-terminus. OCT1 has been cloned from human, rat, mouse and rabbit (13)(14)(15)(16)(17), OCT2 from human (14,15), rat (18), mouse (19), rabbit (16) and pig (20) and OCT3 from human, rat and mouse (21)(22)(23). A functional active isoform of rat OCT1 (rOCT1A) missing the first TMDs and the large extracellular loop (24) and a functional active isoform of human OCT2 (hOCT2B) lacking the last three TMDs have been cloned (25).…”

Properties and regulation of organic cation transport in freshly isolated mouse proximal tubules analyzed with a fluorescence reader-based method

“…These three isoforms, cloned from rat, mouse, and man, mediate OC transport in intestine, liver, brain, and kidney. Renal expression of OCTs is species-and isoform-specific, e.g., in rat kidney, both OCT1 and OCT2 are highly expressed, whereas OCT2 is the predominant isoform in human kidney [14,15,19,29,30,32].…”