Membrane Transport of Nucleobases, Nucleosides and Nucleotides

In mammalian cells, nucleoside transport (NT) systems of several types mediate entry and exit of nucleosides. Transporter-mediated equilibrative NT processes of broad specificity are widely distributed in cells and tissues, and Na^+ - linked, concentrative NT processes have been identified in hepatocytes and in lymphocytic cells, and in vesicles and cells from epithelia. Subtypes of NT systems are known (i) in the equilibrative transporters through major differences (2-3 orders) in sensitivity to NT inhibitors such as NBMPR (nitrobenzylthioinosine), and (ii) in Na^+ - linked transporters through distinctions in substrate specificity and Na^+ /K^+ cosubstrate specificity. While exploration of Na^+ - linked NT systems has proceeded mainly with fresh preparations from differentiated tissues, such systems are also expressed in cultured IEC-6 intestinal epithelial cells and in cultured mouse leukemia L1210 cells. Wild-type L1210 clones express three genetically distinct NT systems in which mutations have been obtained. In IEC-6 monolayers, Na^+ -linked adenosine transport is evidently regulated since this activity is expressed after confluence and a "maturation" interval, and flux levels are markedly influenced by the presence in the medium of insulin, EGF, and hydrocortisone. NBMPR, a tightly-bound, photoactivable ligand at NT sites on erythrocytes from various species has served in the identification of NT glycoproteins, and partial purification of NT glycoproteins of pig and human erythrocytes has enabled development of antibodies directed at those transporters. Polyclonal antibodies against the human transporter are in use in a Joint effort by S.A. Baldwin and coworkers (London) and C.E. Cass and coworkers (Edmonton) to obtain cDNA encoding NT proteins from an expression library prepared from BeWo cells (cultured human choriocarcinoma cells which "overexpress" the NBMPR-sensitive, equilibrative transporter). Both equilibrative and concentrative transporters for nucleobases have been recognized, but transporter phenotypes have been difficult to define. Some cells have substantial transporter-independent permeability to nucleobases and in some cells both hypoxanthine and nucleosides appear to be substrates for the same transporter. With papaverine-stop methods in the assay of nucleobase fluxes, it has become clear that acyclovir and related acyclic nucleosides are substrates for the nucleobase transporter of human erythrocytes, which is NBMPR-insensitive. Alt