Action of dichlorobenzimidazole riboside on RNA synthesis in L-929 and HeLa cells.

Abstract

A BSTR ACT 5,6-Dichloro-l-/~-o-ribofuranosylbenzimidazole (DRB) inhibits RNA synthesis in L-929 cells (mouse fibroblast line) and HeLa cells (human epithelioid carcinoma line) within 2 min of addition of the compound to the medium. By removing DRB from the medium, the inhibition is promptly and completely reversed after treatment of cells for as long as I h or even longer. The inhibitory effect of DRB on the overall rate of RNA synthesis is similar in L and HeLa ceils and is markedly concentration-dependent in the low dose range (5-20 #M or 1.6-6.4/~g/ml), but not at higher concentrations of DRB. At a concentration of 12 #M, DRB has a highly selective inhibitory effect on the synthesis of nuclear heterogeneous RNA in L cells. At higher concentrations, there is also inhibition of 45 S ribosomal precursor RNA synthesis, but at all concentrations the effect on heterogeneous RNA synthesis in L cells in considerably greater than that on preribosomal RNA synthesis. In HeLa cells, too, DRB has a selective effect on heterogeneous RNA synthesis, but quantitatively the selectivity of action is somewhat less pronounced. In both L and HeLa cells, the inhibition of synthesis of nuclear heterogeneous RNA is incomplete even at very high concentrations of DRB (150 ~zM). Thus, while DRB is a selective inhibitor of nuclear heterogeneous RNA synthesis, not all such RNA synthesis is sensitive to inhibition. It is proposed that messenger precursor RNA synthesis may largely be sensitive to inhibition by DRB. In short-term experiments, DRB has no effect on protein synthesis in L or HeLa cells. DRB has a slight to moderate inhibitory effect on uridine uptake into L cells and a moderate to marked effect on uptake of uridine into HeLa cells.

Synthesis of Prematurely Terminated Late Transcripts of Polyoma Virus DNA is Resistant to Inhibition by 5,6-Dichloro-1-beta-D-ribofuranosylbenzimidazole

“…Thus, polyoma virus RNA synthesis was almost completely inhibited by DRB, while cellular RNA synthesis was only partially inhibited. These results are consistent with previous findings (Granick, 1975;Tamm et al, 1976;Tamm, 1977;Fraser et al, 1978) that DRB preferentially inhibits RNA synthesis directed by RNA polymerase II, which is responsible for transcription of polyoma virus DNA (Shmookler et al, 1974). Fig.…”

“…DRB inhibits uptake of [3H]uridine into mammalian cells (Sehgal et al, 1976;Tamm et al, 1976). Measurement of the effect of DRB on RNA synthesis by incorporation of exogenous [3H]uridine must therefore be corrected for this inhibition.…”

“…The drug 5,6-dichloro-l-fl-D-ribofuranosylbenzimidazole (DRB), an analogue of purine ribonucleosides, selectively inhibits RNA synthesis directed by RNA polymerase II in a variety of cell types (Egyhazi, 1974;Granick, 1975;Tamm et al, 1976;Sehgal et al, 1976). The mechanism by which DRB inhibits RNA synthesis has been carefully studied in the case of adenovirus transcription (Fraser et al, 1978Sehgal et al, 1979).…”

DSIF, a novel transcription elongation factor that regulates RNA polymerase II processivity, is composed of human Spt4 and Spt5 homologs

“…DRB was originally discovered as an inhibitor of the synthesis of heterogeneous nuclear RNA in human, murine, avian, and insect cells (Egyhazi 1974;Granick 1975;Sehgal et al 1976a;Tamm et al 1976). Many of these studies showed that DRB inhibits the synthesis of full-length RNA transcripts by enhancing the pausing or premature termination of transcription by RNA Pol II.…”

Mechanism of action of dichloro-beta-D-ribofuranosylbenzimidazole: effect on in vitro transcription.

“…The action of DRB is selective for RNA polymerase II transcripts; RNA polymerase III 5S and tRNA-specific transcripts and the rates of RNA polymerase I-mediated rRNA synthesis are all not affected (1). Initial reports (2,3) indicated that DRB acted at or very close to the site of initiation of transcription, on the basis of incorporation of [3H]uridine into specific-size mRNAs of Balbiani rings in Chironomus (2) and of the labeling kinetics of HeLa cell mRNA precursors (3). The effect of adding DRB to HeLa cells is very rapid (<5 min).…”