RNA Polymerase Activity in Purified Rice Dwarf Virus

Abstract

SummaryPurified rice dwarf virus (RDV) particles catalyze incorporation of H3-UMP from H3-UTP into an acid-insoluble polynucleotide product.Optimal synthesis of the polynucleotide occurs at 35C and within a pH range of 8.5-9.0. The H3-UMP incorporation is strictly dependent on all other three ribonucleoside triphosphates and Mg++ ion, and is partially dependent on energy regenerating system. The reaction is stimulated slightly by chymotrypsin, not inhibited by actinomycin D or DNase, and strongly inhibited by RNase or pyrophosphate. The reaction proceeds for at least two hours without any detectable lag and is directly proportional to the amount of RDV particles. The polymerase activity cosediments with intact RDV particles in sucrose density gradient and little activity is associated with the empty shells of RDV. The reaction product is RNase-sensitive and is specifically hybridized with heat-denatured, unlabeled RDV-RNA but not with heat-denatured RNA from cytoplasmic polyhedrosis virus of silkworm.

In Vitro Transcription by RNA Polymerase Associated with Rice Gall Dwarf Virus

“…Three viruses, WTV, RDV and RGDV, have so far been grouped as Phytoreoviruses and the properties of the R N A polymerase associated with two of them, WTV and RDV, have been reported (Black & Knight, 1970;Reddy et al, 1977;Kodama & Suzuki, 1973). In this paper, we demonstrate that R G D V also has an R N A polymerase associated with the virus particles, and show that the properties of the enzyme are similar to those of WTV and RDV.…”

“…However, there appear to be some differences. The optimum activity of RGDV is at 25 °C whereas that of RDV is at 35 °C (Kodama & Suzuki, 1973). RGDV also appears to have much less activity per unit weight of virus particles than does RDV.…”

“…2 also shows the time course of incorporation by RDV particles. Even though the optimum temperature of activity for RDV is 35 °C (Kodama & Suzuki 1973), its activity was much higher than that of RGDV at 25 °C.…”

“…The final pellets were resuspended in 0.1 M-glycine, 0.01 M-MgCI2, adjusted to pH 7.0 with 0-1 M-NaOH. Virus concentration was estimated spectrophotometrically using A260n m = 6"0 for 1 mg/ml of virus (the value for RDV) (Kodama & Suzuki, 1973).…”

“…Rice dwarf virus (RDV) and wound tumour virus (WTV) have been reported to have the virion-associated enzyme (Kodama & Suzuki, 1973;Black & Knight, 1970).…”