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A reassessment of the response of the bacterial ribosome to the frameshift stimulatory signal of the human immunodeficiency virus type 1
- Source :
- RNA. 10:1225-1235
- Publication Year :
- 2004
- Publisher :
- Cold Spring Harbor Laboratory, 2004.
-
Abstract
- HIV-1 uses a programmed -1 ribosomal frameshift to produce the precursor of its enzymes. This frameshift occurs at a specific slippery sequence followed by a stimulatory signal, which was recently shown to be a two-stem helix, for which a three-purine bulge separates the upper and lower stems. In the present study, we investigated the response of the bacterial ribosome to this signal, using a translation system specialized for the expression of a firefly luciferase reporter. The HIV-1 frameshift region was inserted at the beginning of the coding sequence of the luciferase gene, such that its expression requires a −1 frameshift. Mutations that disrupt the upper or the lower stem of the frameshift stimulatory signal or replace the purine bulge with pyrimidines decreased the frameshift efficiency, whereas compensatory mutations that re-form both stems restored the frame-shift efficiency to near wild-type level. These mutations had the same effect in a eukaryotic translation system, which shows that the bacterial ribosome responds like the eukaryote ribosome to the HIV-1 frameshift stimulatory signal. Also, we observed, in contrast to a previous report, that a stop codon immediately 3′ to the slippery sequence does not decrease the frameshift efficiency, ruling out a proposal that the frameshift involves the deacylated-tRNA and the peptidyl-tRNA in the E and P sites of the ribosome, rather than the peptidyl-tRNA and the aminoacyl-tRNA in the P and A sites, as commonly assumed. Finally, mutations in 16S ribosomal RNA that facilitate the accommodation of the incoming aminoacyl-tRNA in the A site decreased the frameshift efficiency, which supports a previous suggestion that the frameshift occurs when the aminoacyl-tRNA occupies the A/T entry site.
- Subjects :
- Biology
Slippery sequence
Molecular biology
Ribosome
Article
Ribosomal frameshift
Stop codon
Frameshift mutation
Polar mutation
A-site
Eukaryotic translation
Genes, Reporter
RNA, Ribosomal, 16S
Mutation
Codon, Terminator
Escherichia coli
HIV-1
Nucleic Acid Conformation
RNA, Viral
Ribosomes
Molecular Biology
Subjects
Details
- ISSN :
- 14699001 and 13558382
- Volume :
- 10
- Database :
- OpenAIRE
- Journal :
- RNA
- Accession number :
- edsair.doi.dedup.....08a1adb9c60a3589bc37fcb99f7e3e1c
- Full Text :
- https://doi.org/10.1261/rna.7670704