Production Of Genomic RNA Research Articles

Altering SARS Coronavirus Frameshift Efficiency Affects Genomic and Subgenomic RNA Production

In previous studies, differences in the amount of genomic and subgenomic RNA produced by coronaviruses with mutations in the programmed ribosomal frameshift signal of ORF1a/b were observed. It was not clear if these differences were due to changes in genomic sequence, the protein sequence or the frequency of frameshifting. Here, viruses with synonymous codon changes are shown to produce different ratios of genomic and subgenomic RNA. These findings demonstrate that the protein sequence is not the primary cause of altered genomic and subgenomic RNA production. The synonymous codon changes affect both the structure of the frameshift signal and frameshifting efficiency. Small differences in frameshifting efficiency result in dramatic differences in genomic RNA production and TCID50 suggesting that the frameshifting frequency must stay above a certain threshold for optimal virus production. The data suggest that either the RNA sequence or the ratio of viral proteins resulting from different levels of frameshifting affects viral replication.

Association of nucleocapsid polypeptides with defective RNA synthesis in a temperature-sensitive mutant of sendai virus

A temperature-sensitive mutant of Sendai virus, is 105, does not synthesize virus-specific RNA at the restrictive temperature (38°). In this study, all virus-specific polypeptides of the mutant were synthesized for several hours at this temperature in about the same amounts as in cells incubated with wild-type virus, when infection was initiated at permissive temperature. However, the major nucleocapsid polypeptide (NP) did not assemble into nucleocapsids, whereas newly synthesized P, the minor nucleocapsid polypeptide, was able to associate with preexisting nucleocapsids. Moreover, pulse-chase experiments showed that, in the mutant, P was only transiently associated with nucleocapsids, suggesting some transiently related nucleocapsid function. After a shift to the permissive temperature (30°), normal nucleocapsid formation resumed, but the production of genomic RNA and RNAs sedimenting in the region of replicative and transcriptive intermediates was substantially higher than normal. This latter finding suggests that a viral product(s) essential for RNA synthesis accumulated during incubation at the restrictive temperature. Since nucleocapsids are the structures in which viral RNA synthesis takes place, these results indicate that the is 105 gene codes for a nucleocapsid polypeptide, perhaps P or NP, that is responsible for the synthesis or regulation of viral RNA.

The effects of actinomycin D on RNA synthesis in measles virus-infected cells.

The effect of actinomycin D on the production of genomic RNA, messenger RNA and protein in measles virus-infected cells were investigated. It was found that whereas high concentrations of the drug inhibited genome synthesis, the production of messenger RNA was unaffected and in some cases enhanced. These effects were shown, at least in part, to be due to a secondary effect of the drug which caused an inhibition of virus-specific protein synthesis.