Inhibition of measles virus replication and RNA synthesis by actinomycin D

Measles virus replication and RNA synthesis in Vero cells are inhibited by actinomycin at concentrations which inhibit cellular RNA synthesis. Drug present from the 2nd to the 24th hr post infection inhibited infectivity but not hemagglutinating activity or cell fusion. Infectivity was much less sensitive to drug added during the second 24-hr period, and 52S RNA was labeled and incorporated into virions during this later time interval.     

Temperature-dependent actinomycin D effect on RNA synthesis during synchronous development in Neurospora crassa

Actinomycin D at a concentration of 5 g/ml of medium inhibited DNA-dependent RNA synthesis by 92% at 35 C, 42% at 30 C, and 28% at 25 C in Neurospora crassa. This concentration also inhibited the development of conidiophores and conidia at 35 C, but not at 30 or 25 C. Mycelia which were induced to synchronous development formed conidiophores in 2.5 hr and conidia in 4.5 hr at 35 C in the absence of drug additives. Addition of actinomycin D to synchronously developing mycelia at zero time and at 0.5-hr intervals thereafter at 35 C indicated that RNA synthesis required for conidiophores occurred before 0.5 hr and for conidia before 2 hr. Addition of cycloheximide at the same times to another synchronous mycelial series at 35 C indicated that protein synthesis required for conidiophores occurred before 2 hr and for conidia before 3.5–4 hr.This work was supported in part by U.S. Public Health Service Training Grant 1 TO1 GM 01968 01.     

Inhibitory effects of actinomycin D and cycloheximide on neuronal death in adult Manduca sexta

A decline in circulating 20-hydroxyecdysone permits the emergence of the adult Manduca sexta moth; this endocrine signal also triggers the death of approximately half of the neurons in the unfused abdominal ganglia of the moth central nervous system. This programmed death of neurons was markedly reduced by treatment with either actinomysin D (an RNA synthesis inhibitor) or cycloheximide (a protein synthesis inhibitor). Similar results were found after addition of these agents to ventral nerve cord cultures. The effectiveness of these treatments in delaying or blocking neuronal death depended upon their time of administration relative to the normal time of post-emergence death in the particular motoneuron under study: late-dying neurons, for example, could still be saved by these treatments even after early-dying neurons had already initiated degeneration. In both intact moths and cultured ventral nerve cords, the ability of actinomycin D to prevent neuronal death waned at the same time at which replacement of the steroid hormone could no longer block neuronal death. This suggests that the steroid commitment point represents the time at which the genes that mediate cell death are transcribed. Cycloheximide remained effective in delaying or blocking neuronal death until shortly before the onset of degeneration, suggesting that ongoing protein synthesis is essential for the initiation of the degeneration response. 1994 John Wiley & Sons, Inc.     

Effect of vitamin E an actinomycin D on protein and RNA synthesis in rat liver

In vitro experiments showed that RNA synthesis intensity in the rat liver with E-hypovitaminosis decreases considerably while the level of the labelled precursors incorporation into protein does not differ from the norm. Under conditions of E-hypovitaminosis the inhibitory effect of actinomycin D on the RNA synthesis is pronounced more clearly as compared to the norm. In the case of the E-hypovitaminous rate liver chyme preincubation with alpha-tocopherol there is no inhibitory effect of actinomycin D on the protein biosynthesis.     

Effects of actinomycin D on ribosomal RNA and protein synthesis as revealed by high resolution autoradiography.

Incorporation of tritiated amino acids and uridine was studied in untreated and actinomycin D treated HeLa cells by high resolution autoradiography. Results showed a non-selective inhibition of protein synthesis by actinomycin, as measured by the decrease in radioactive amino acid uptake. When cells pretreated with actinomycin D were incubated with radioactive amino acids and uridine, amino acid uptake in the nucleolus still occurred, while uridine uptake was almost completely eliminated. These findings suggest that in the absence of ribosomal RNA precursor synthesis, nucleolar protein synthesis continues to some extent, and that this protein is transported to the nucleolus.     

The effect of actinomycin D on RNA synthesis and nucleolar ultrastructure in the liver of rats treated with fluorouracil

Summary Rats were given cytidine-³H and 10 min later 50 mg fluorouracil. They were killed after 25 hours. Actinomycin D was given at various times before sacrifice. The collapse of the nucleolus and the segregation of its components, seen in rats sacrificed one hour after administration of actinomycin D only, was prevented by prior treatment with fluorouracil. In rats treated with fluorouracil and given actinomycin 12 or 20 hours prior to death, there was a more or less pronounced collapse of the nucleolus but no typical segregation of its components. Radioautographs of livers from untreated rats or rats given actinomycin only at the times mentioned, and killed 25 hours after administration of cytidine-³H, were labelled mainly over the cytoplasm. Radioautographs from rats, treated with fluorouracil only, or fluorouracil plus actinomycin, showed labelling over the nucleoli, but depressed labelling over the cytoplasm. Biochemical analysis of RNA labelling showed high ribosomal peaks in untreated rats and rats treated with actinomycin only. Rats treated with fluorouracil, or fluorouracil plus actinomycin showed no labelling of the 29S and 18S ribosomal peaks. The results indicate that fluorouracil blocks or delays the formation of ribosomal RNA and that the inhibition, at least in part, takes place in the nucleolus.This work was supported by grants from the Swedish Medical Research Council (Project K68-12X-623-04), the Swedish Cancer Society (Project 6831), the Medical Faculty of Uppsala and the Swedish Society for Medical Research.     

Sequence-specific inhibition of RNA elongation by actinomycin D

We have developed a method to localize specific sites where RNA elongation is arrested due to DNA-bound ligands. The method was used to determine apparent binding sites for actinomycin D. We have found 14 strong RNA hindrance sites along nucleotide sequence of T7 and D111 T7 DNA of 380 nucleotides full length under low actinomycin D concentration conditions. Nucleotide sequence of all the sites is described by general formula XGCY where X ‡ G and Y ‡ C.