Inhibition of RNA synthesis by deoxyadenosine plus deoxycoformycin in resting lymphocytes

Deoxyadenosine has been implicated in the lymphocytopenia that occurs in immunodeficient children with an inherited deficiency of adenosine deaminase (ADA) and in leukemic patients treated with the ADA inhibitor deoxycoformycin. The recent reports of deoxyadenosine toxicity to nondividing lymphocytes indicates a challenge to the mechanism for deoxyadenosine toxicity, which involves the inhibition of ribonucleotide reductase by dATP, leading to the inhibition of DNA synthesis. This study provides evidence for the inhibition of transcription by deoxyadenosine as an alternative mechanism of toxicity. The incubation of resting peripheral blood lymphocytes with deoxyadenosine plus deoxycoformycin led to an inhibition of uridine incorporation. The extent of inhibition increased with the increasing time of incubation and concentration of deoxyadenosine. Replacement of deoxyadenosine with other nucleosides, adenosine or deoxyguanosine, had no effect, suggesting that deoxyadenosine-induced inhibition was not due to the reduced transport of uridine. Separation of DNA from RNA by differential alkaline hydrolysis showed that the reduction of uridine incorporation was primarily in the RNA fraction. The time sequence of the reduction in uridine incorporation coincided with that of the accumulation of dATP, but preceded that of ATP depletion and cell lysis. The phosphorylation of uridine into UTP was slightly reduced by deoxyadenosine, but this could not entirely account for the reduced incorporation of uridine into RNA. Finally, the direct measurement of RNA synthesis by the incorporation of UTP into isolated nuclei showed that RNA synthesis was inhibited to 88% and 41% of control values in lymphocytes preincubated with 20 microM deoxyadenosine for 3 and 15 hr, respectively. These findings demonstrate that deoxyadenosine plus deoxycoformycin inhibits RNA synthesis in resting lymphocytes.