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Abstract— DNA replication in UV‐irradiated Chinese hamster V‐79 cells was analyzed by measuring the incorporation of 14C‐thymidine into acid precipitable form and by DNA fiber autoradiography. As expected, UV exposure resulted in a rapid deceleration in the rate of thymidine incorporation, reaching a minimum rate 60–75 min following exposure to 2–10Jm‐2. After an additional 1–2h the rate of thymidine incorporation began to recover slowly, approaching the control rate within 10h following exposure to 10 Jm‐2 or less. The mechanism of the inhibition and recovery in thymidine incorporation was examined by DNA fiber autoradiography. The results indicate that UV radiation produces lesions in DNA which temporarily block chain growth of DNA that is synthesized during the first 5 h after exposure. Within 10 h after exposure, the lesions have either been repaired or modified (or the replicative enzymes altered) so that DNA chain growth is no longer impeded.

Treatment of isolated bone cells with adenosine increased cyclic 3',5'-AMP levels markedly but failed to modify total ATP levels. 100 JIM-500 HM adenosine elicited maximal (17-fold), and 5 M*M minimal ( + 68%), effects. Equivalent concentrations of 2'-AMP or 5'-AMP were less active than adenosine; and adenine, guanosine and uridine were inactive. The effect of adenosine peaked in 15-45 sec, gradually diminished with more prolonged treatment, and disappeared almost completely when treated cells were washed and incubated for an additional 30 sec in adenosinefree medium. These results suggest that adenosine acts at or near the cell surface to stimulate adenylate cydase.Adenosine and either sodium fluoride, epinephrine, or PTH increased bone cell cyclic 3',5'-AMP levels synergistically; and a physiological concentration of PTH (1 ng/ml) was highly effective in the presence of adenosine. Simultaneous treatment with theophylline decreased the effect of adenosine, yet theophylline did not inhibit the cellular accumulation of adenosine-8-14 C. Hence, adenosine may also modify the responsiveness of adenylate cyclase to various stimuli, inhibit cyclic 3',5'-AMP phosphodiesterase activity, or both. {Endocrinology 94: 148, 1974)

Adenosine rapidly stimulated adenylate cyclase activity but did not modify cyclic AMP degradation when added to a particulate fraction prepared from isolated bone cells. The effect of adenosine was one-half maximal at 5-10 micronM, and was not mimicked by 5' AMP, inosine, guanosine, uridine, adenine, or ribose. Basal and adenosine-stimulated adenylate cyclase activites were directly proportional to the concentration of particulate protein in the assay system. Theophylline decreased the degree to which adenosine stimulated adenylate cyclase activity, whereas another phosphodiesterase inhibitor, RO-20-1724, failed to iiinfluence the effect of adenosine. Adenosine itself, and not a metabolite of adenosine is the stimulator of adenylate cyclase, since it was neither phosphorylated nor deaminated appreciably by the particulate fraction. The particulate fraction did not convert substrate ATP to adenosine in amounts sufficient to enhance adenylate cyclase. The stimulatory effect of adenosine was maximal at 1.2 mM Mg2+, declined with increases in the Mg2+ concentration, and was replaced by inhibition at 20 mM Mg2+. At 2.4 mM Mg2+, basal adenylate cyclase activity peaked at 1.1 mM ATP, and was inhibited by higher ATP concentrations. The magnitude of adenosine stimulation was greater at inhibitory concentrations of ATP than at concentrations which yielded maximum activity. The results suggest that the previously demonstrated ability of adenosine to increase cyclic 3'5' AMP levels in intact bone cells stems from its effect on adenylate cyclase. Adenosine may act by modifying the regulatory nfluence of free Mg2+, uncomplexed ATP, (or both), on adenylate cyclase. Theophylline appears to interfere with the action of adenosine by a mechanism which is distinct from its capacity to inhibit cyclic AMP phosphodiesterase activity. (Endocrinology 99:901,1976)

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