Translation of black beetle virus RNA and heterologous viral RNAs in cell-free lysates derived from Drosophila melanogaster

It has been shown previously that Drosophila cells infected with black beetle virus synthesize an early viral protein, protein A, a putative element of the viral RNA polymerase. Synthesis of protein A declines sharply by 6 h postinfection, whereas synthesis of viral coat protein at continues for at least 14 h. The early shutoff in protein A synthesis occurred despite the presence of equimolar proportions of the mRNAs for proteins A and ax, RNAs 1 and 2, respectively. We have now been able to mimic this translational discrimination in a cell-free protein-synthesizing system prepared from infected or uninfected Drosophila cells, thus allowing further analysis of the mechanism by which translation of RNA 1 is selectively turned off. The results revealed no evidence for control by virus-encoded proteins or by virus-induced modification of mRNAs by the cell-free system. Rather, with increasing RNA concentration, viral RNA 1 was outcompeted by its genomic partner, RNA 2. This suggests that the early shutoff in intracellular synthesis of protein A is due to decreasing ability of RNA 1 to compete for a rate-controlling translational factor(s) as the concentration of viral RNAs accumulates within the infected cell.

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Early and late functions in a bipartite RNA virus: evidence for translational control by competition between viral mRNAs

Friesen¹,

Rueckert²

1984

“…Unless otherwise stated, the samples were prepared for electrophoresis by diluting [3H]uridine-labeled RNA with an equal volume (15 ,ul) of Loening buffer containing 10 M urea, 0.05% (wt/vol) xylene cyanol, and 0.05% (wt/vol) bromophenol blue. Electrophoresis was conducted for approximately 6 h at 10 V/cm (6°C) without recirculating the electrophoresis (Loening) buffer.…”

mentioning

confidence: 99%

Black Beetle Virus: Messenger for Protein B Is a Subgenomic Viral RNA

Friesen

Rueckert

1982

Black beetle virus induces the synthesis of three new proteins, protein A (molecular weight, 104,000), protein α (molecular weight, 47,000), and protein B (molecular weight, 10,000), in infected Drosophila cells. Two of these proteins, A and α, are known to be encoded by black beetle virus RNAs 1 and 2, respectively, extracted from virions. We found that RNA extracted from infected cells directed the synthesis of all three proteins when it was added to a cell-free protein-synthesizing system. When polysomal RNA was fractionated on a sucrose density gradient, the messengers for proteins A and α cosedimented with viral RNAs 1 (22S) and 2 (15S), respectively. However, the messenger for protein B was a 9S RNA (RNA 3) not found in purified virions. Like the synthesis of viral RNAs 1 and 2, intracellular synthesis of RNA 3 was not affected by the drug actinomycin D at concentrations which blocked synthesis of host cell RNA. This indicated that RNA 3 is a virus-specific subgenomic RNA and, therefore, that protein B is a virus-encoded protein.

“…In addition, small amounts of two other proteins have been detected, including a possible capsid protein precursor (8,9,14). In vivo and in vitro protein synthesis studies have shown that the virus codes for a large protein of 110K, which is a candidate RNA polymerase, a capsid protein precursor, and an 8 to 10K polypeptide of unknown function (5,6,8,12).…”

mentioning

confidence: 99%

Two Unique RNA Species of the Nodavirus Black Beetle Virus

Clewley

Crump

Avery

et al. 1982

Two-dimensional fingerprinting of RNase T1-derived oligonucleotides of the two individual RNA segments of the Nodavirus black beetle virus indicates that each RNA species possesses a distinct nucleotide sequence. Species 1 RNA has a genome complexity of approximately 3,000 nucleotides, and species 2 RNA is composed of approximately 1,500 nucleotides. Submolar amounts of oligonucleotides apparently derived from a third virus-specific RNA were also detected in black beetle virus RNA preparations.