NA-DEPENDENT NUCLEOSIDE TRANSPORT IN LIVER - 2 DIFFERENT ISOFORMS FROM THE SAME GENE FAMILY ARE EXPRESSED IN LIVER-CELLS()

Hepatocytes show a Na+-dependent nucleoside transport activity that is kinetically heterogeneous and consistent with the expression of at le ast two independent concentrative Na+-coupled nucleoside transport sys tems (Mercader et al. Biochem. J. 317, 835-842, 1996). So far, only a single nucleoside carrier-related cDNA (SPNT) has been isolated from l iver cells (Che et al. J. Biol. Chem. 270, 13596-13599, 1995). This cD NA presumably encodes a plasma membrane protein responsible for Na+-de pendent purine nucleoside transport activity. Thus, the liver must exp ress, at least, a second nucleoside transporter which should be pyrimi dine-preferring. Homology cloning using RT-PCR revealed that a second isoform is indeed present in liver. This second isoform turned out to be identical to the 'epithelial-specific isoform' called cNT1, which s hows in fact high specificity for pyrimidine nucleosides. Although cNT 1 mRNA is present at lower amounts than SPNT mRNA, the amounts of cNT1 protein, when measured using isoform-specific polyclonal antibodies, were even higher than the SPNT protein levels. Moreover, partially pur ified basolateral plasma membrane vesicles from liver were enriched in the SPNT but not in the cNT1 protein, which suggests that the subcell ular localization of these carrier proteins is different. SPNT and cNT 1 protein amounts in crude membrane extracts from 6 h-regenerating rat livers are higher than in the preparations from sham-operated control s (3.5- and 2-fold, respectively). These results suggest that liver pa renchymal cells express at least two different isoforms of concentrati ve nucleoside carriers, the cNT1 and SPNT proteins, which show differe ntial regulation and subcellular localization.