The Na+-dependent transport and facilitated diffusion of uridine were measured after differentiation of HL-60 leukaemia cells along the monocytic pathway by phorbol 12-myristate 13-acetate (PMA). PMA (200 ng/ml) caused a marked increase in Na+-dependent uridine transport within 48 h of exposure that was attributable to an increase in transport affinity (apparent Km values of 1.15 + 0.22 and 44 + 4.4 gM for PMA-induced and uninduced cells respectively), with no change in Vmax (0.15 + 0.02 and 0.13 + 0.01 pmol/s per ,1u of cell water for PMA-induced and uninduced cells respectively). A corresponding rapid decrease in both the rate of facilitated diffusion and the formation of uracil nucleotides occurred in PMA-induced cells. As a consequence of these changes, intracellular pools of uridine 3-4-fold greater than those in the medium were generated. A similar increase in Na+-dependent transport of adenosine, inosine, guanosine, thymidine and cytidine (Km values of 1-4 #M) was observed. The effects of PMA on the activation of the Na+-dependent uridine transporter were inhibited by staurosporine, suggesting the involvement of protein kinase C. The findings indicate that a change in the balance of the cellular mechanisms employed for nucleoside transport occurs during the monocytic differentiation of HL-60 leukaemia cells.

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The effects of pertussis toxin on the Na(+)-dependent transport of uridine were studied in HL-60 leukaemia cells induced to differentiate along the granulocytic or monocytic pathways by dimethyl sulphoxide (DMSO) or phorbol 12-myristate 13-acetate (PMA) respectively. Pertussis toxin at 50 ng/ml completely inhibited the activation of Na(+)-dependent uridine transport and consequently prevented the formation of intracellular pools of free uridine which occurs in HL-60 cells induced to differentiate by DMSO. The inhibition of Na(+)-dependent uridine transport by pertussis toxin in cells exposed to DMSO was associated with a 14-fold decrease in affinity, with no change in Vmax. Pertussis toxin, however, had no effect on Na(+)-dependent uridine transport in PMA-induced HL-60 cells. Furthermore, 500 ng of cholera toxin/ml had no effect on the Na(+)-dependent uptake of uridine in DMSO-treated HL-60 cells. These results suggest that the activation of the Na(+)-dependent transport of uridine in HL-60 cells induced to differentiate along the granulocytic pathway by DMSO is coupled to a pertussis-toxin-sensitive guanine-nucleotide binding protein (G-protein).