Actinomycin poly

C-HAAF (N-Hydroxy-N-acetyl-2-amino-fluorene was from the ICN Hadlochemlcals Division, and -actinomycin D was from Amersham. Calf thymus DNA (Worthington), was prepared according to the procedure of Muller and Crothers (15.). Poly (dAdT)" poly (dAdT) and poly(dGdC)-poly(dCdC) were purchased from PL Blochemlcals. DNA concentrations were determined absorptlometrlcally, and expressed in basepalrs. 

The binding of actinomycin D to calf thymus DNA (0.01M, 0.1M, and 0.2M NaCl) and to poly(dGdC) poly(dGdC) (0.1M NaCl) (Figure 4) was also examined by phase partition techniques. For both DNAs, a cooperative process was seen. In the case of calf thymus DNA, the decreasing salt concentration appears to Increase the degree of cooperativity. 

Kinetics. The dissociation of actinomycin D from poly(dGdC)-poly(dGdC) Is well described by a single exponential decay (Table... 

Dissociation lifetime of actinomycin D from Poly(dGdC) Poly(dGdC) as a function of phosphate/drug ratio (T 22°C) and temperature. 

The dissociation kinetics of daunorubicin and daunorubicin actinomycin D from poly(dAdT).poly(dAdT) were studied using stopped-flow techniques. The data is summarized in Table IV. The dissociation of daunorubicin from poly(dAdT).poly(dAdT) is monophaslc with a lifetime of 0.2 sec. In the ternary complex (daunorubicin actinomycin D poly(dAdT).poly(dAdT)) the dissociation of the daunorubicin is characterized by two exponential decays, yielding lifetimes of 0.2 sec and 5 sec, A single 5 sec dissociation time was observed for actinomycin D. 

This data is consistent with a model in which daunorubicin is bound to Isolated DNA binding sites and dissociates with its normal lifetime of 0.2 sec. Whereas daunorubicin and actinomycin D molecules, residing at "adjacent sites", dissociate with a time constant of approximately 5 sec. These results are indicative of a cooperative interaction between daunorubicin and actinomycin D when the two are bound to poly(dAdT).poly(dAdT). 

Alkaline hydrolysates of mRNA from vaccinia virus (Kates, 1970) and mouse sarcoma 180 cells (Mendecki et al, 1972) show that the poly (A) tracts, 100-200 nucleotides in length, are located at the 3 -ter-minal end of the RNA molecules. Analysis of the reaction products of highly purified exoribonuclease specific for 3 -OH termini also indicates that most (and possibly all) of the poly (A) sequences are at the 3 -terminal end of the mRNA s (Molloy et al, 1972). This is supported by the fact that the time course of poly( A) labeling in polysomes indicates that this sequence is assembled after the rest of the RNA molecule has been completed (Mendecki et al, 1972). Poly (A) synthesis is sensitive to actinomycin D but to an extent less than that of the rest of the RNA molecule which further argues that the poly (A) segment is added after transcription is completed (Darnell et al, 1971b Mendecki et al, 1972). 

Yoshihara K (1972) Complete dependency of poly(ADP-ribose) synthesis on DNA and its inhibition by actinomycine D. Biochem Biophys Res Commun 47 119-125... 

Hyperthermia is known to result in the appearance of a number of heat shock proteins whose synthesis is controlled at the level of both transcription and translation [3]. Thus, we have examined for effects of cycloheximide and actinomycin D on the alteration of poly(ADP-ribose) metabolism by hyperthermia. The results of Table 2 show that the presence of cyclohexamide resulted in over 80% inhibition of accumulation of poly(ADP-ribose) following hyperthermia, but did not affect accumulation in control cells. In contrast, the presence of actinomycin D, which inhibited uridine incorporation by over 90%, had no appreciable effect (data not shown). 

Further evidence for a fundamental difference in the mechanism of poly(ADPR) accumulation following PMA and MNNG treatment derives from experiments with inhibitors of macromolecular synthesis. As shown in Fig. 2 for human fibroblasts, suppression of RNA synthesis with actinomycin D (2 /ig ml" ) reduced the PMA-induced accumulation of poly(ADPR) by 90%. A similar result was obtained with cycloheximide (5 /xg ml ) which inhibits de novo protein synthesis. In contrast, these... 

An interesting aspect of the facilitated binding of actinomycin D to poly(dA-dT) -poly(dA-dT) is the question of whether the bound actinonQ cin D would Inhibit RNA polymerase, since it has been well documented that actinomycin D very effectively inhibits RNA polymerase... 

The dissociation of actinomycin D from poly(dG) poly(dC) is characterized by two slow dissociation constants, which contrasts with the single exponential decay observed for the dissociation of actinomycin D from poly(dG-dC)-poly(dG-dC) Segments of these two polynucleotides are illustrated in Figure 6 where we note that in poly(dG) poly(dC) all... 

Amphibole (crocidol-ite and amosite), serpentine (chrysotile), wollastonite, riebeglass beads Human and rabbit mesothelial cells in vitro Oligonucleosomal DNA fragmentation, loss of membrane phospholipid asymmetry, and nuclear condensation Asbestos fibres, not control particles, induced apoptosis in mesothelial cells by all assays. Induction of apoptosis was dose-dependent for all types of asbestos, with crocidolite (5 ng/cm ) inducing 15.0 1.1 % apoptosis versus control particles <4 %. Apoptosis induced by asbestos, but not by actinomycin D, was inhibited by extracellular catalase, superoxide dismutase in the presence of catalase, hypoxia (8 % oxygen), deferoxamine, 3-aminobenzamide [an inhibitor of poly(ADP-ribosyl) polymerase], and cytochalasin B. Only catalase and cytocha-lasin B decreased fibre uptake. Broaddus et al. (1996) Escape from asbestos-induced apoptosis could allow the abnormal survival of mesothelial cells with asbestos-induced mutations... 

The notion that vaccinia virus-induced RNA may be involved in shut-off was independently proposed by Rosemond-Hombeak and Moss (1975) and Bablanian (1975). Rosemond-Hornbeak and Moss (1975) infected HeLa cell suspension cultures in the presence of 5-10 xg/ml of actinomycin D and showed that short poly(A)-rich RNAs were transcribed which sedimented a 5 S. Normal vaccinia virus transcripts sediment at 10-14 S (Becker and Joklik, 1964 Salzman et al.. 

Rosemond-Hornbeak, H., and Moss, B., 1975, Inhibition of host protein synthesis by vaccinia virus, fate of cell mRNA and synthesis of small poly(A)-rich polyribonucleotides in the presence of actinomycin-D, J, ViroL 16 34. 

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