Transport and countertransport of thymidine in ATP depleted and thymidine kinase-deficient novikoff rat hepatoma and mouse L cells: Evidence for a high Km facilitated diffusion system with wide nucleoside specificity

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Abstract
Incubation of cultured Novikoff rat hepatoma and mouse L cells in a glucose‐free basal medium containing 5 mM KCN and 5 mM iodoacetate for about 10 minutes resulted in a complete depletion of the cells of ATP. ATP‐depleted wild type cells or thymidine kinase‐deficient sublines of Novikoff or L cells took up thymidine rapidly from the medium without concentrating it intracellularly, and exhibited countertransport of thymidine. Thus uptake was by facilitated diffusion. This transport system differs from the substrate‐specific, low‐Km (0.5 μM) thymidine transport system previously described for various types of cultured cells in that it exhibits an at least 100‐fold higher Km and transports equally well various ribo‐ and deoxyribonucleosides. The results suggest that the rate‐limiting step in thymidine incorporation into the nucleotide pool by wild type cells is phosphorylation rather than transport, or that the cells possess two transport systems, a facilitated diffusion system with low substrate specificity and a second system which involves substrate phosphorylation by thymidine kinase.

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