When newly synthesized RNA was pulse-labeled by addition of [5-3H]uridine, the incorporation being terminated after 30 sec by addition of an antibiotic inhibitor of RNA synthesis, significant incorporation of label into acid-insoluble material other than RNA occurred. When correction for this effect was made, the parameters of decay of pulse-labeled RNA revealed with actinomycin D at 10 µg/ml were significantly lower than those previously reported: a mean half-life of 36 ± 3 sec and a mean stable fraction of 39 ± 4% were found. Streptolydigin, echinomycin, and triostins A and C yielded (corrected) curves for decay of pulse-labeled RNA similar to those seen using actinomycin. With streptolydigin and echinomycin the parameters of decay were independent of antibiotic concentration over a wide range and were comparable with the parameters quoted for actinomycin. Similar values were also given by the triostin antibiotics tested at 5 µg/ml. Echinomycin appeared to be approximately 4-5 times more potent than actinomycin D, judged by the relative concentrations required to halt incorporation of the precursor and reveal decay of pulselabeled RNA. Direct measurements, using incorporation of [methyl-3H]thymidine, showed that both actinomycin and echinomycin inhibit DNA synthesis in Bacillus megaterium. Streptolydigin had no inhibitory action. Actinomycin, echinomycin, and streptolydigin all caused degradation of polysomes in protoplasts of B. megaterium. With each antibiotic the time to 50% decay was approximately 5 min, in agreement with earlier estimates using actinomycin. A large discrepancy between the apparent half-lives of decay of pulse-labeled RNA and of polysomes therefore was seen with all antibiotics tested. The results attest to the usefulness of studies on the decay of pulse-labeled RNA as a means of investigating the specificity, rapidity of action, and relative potency of antibiotics which inhibit RNA synthesis.
ACKNOWLEDGMENTS We thank Mr. N. F. Totty for technical assistance, and Drs. E. Cundliffe and D. J. W. Burns for their advice and help in the experiments on breakdown of polysomes. We are especially grateful to Drs. G. B. Whitfield, Jr., H. Bickel, K. Scheibli, and H. Otsuka for generous supplies of the antibiotics.
- Copyright ©, 1974, by Academic Press, Inc.