Baggett et al. 2017 (PRJNA348358)

General Details

Title Global analysis of translation termination in E. coli using release factor manipulations
Organism
Number of Samples 19
Release Date 2016/10/13 00:00
Sequencing Types
Protocol Details

Study Links

Repository Details

SRA SRP091515
ENA SRP091515
GEO
BioProject PRJNA348358

Publication

Title
Authors Baggett NE,Zhang Y,Gross CA
Journal PLoS genetics
Publication Date 2017 Mar
Abstract Terminating protein translation accurately and efficiently is critical for both protein fidelity and ribosome recycling for continued translation. The three bacterial release factors (RFs) play key roles: RF1 and 2 recognize stop codons and terminate translation; and RF3 promotes disassociation of bound release factors. Probing release factors mutations with reporter constructs containing programmed frameshifting sequences or premature stop codons had revealed a propensity for readthrough or frameshifting at these specific sites, but their effects on translation genome-wide have not been examined. We performed ribosome profiling on a set of isogenic strains with well-characterized release factor mutations to determine how they alter translation globally. Consistent with their known defects, strains with increasingly severe release factor defects exhibit increasingly severe accumulation of ribosomes over stop codons, indicative of an increased duration of the termination/release phase of translation. Release factor mutant strains also exhibit increased occupancy in the region following the stop codon at a significant number of genes. Our global analysis revealed that, as expected, translation termination is generally efficient and accurate, but that at a significant number of genes (≥ 50) the ribosome signature after the stop codon is suggestive of translation past the stop codon. Even native E. coli K-12 exhibits the ribosome signature suggestive of protein extension, especially at UGA codons, which rely exclusively on the reduced function RF2 variant of the K-12 strain for termination. Deletion of RF3 increases the severity of the defect. We unambiguously demonstrate readthrough and frameshifting protein extensions and their further accumulation in mutant strains for a few select cases. In addition to enhancing recoding, ribosome accumulation over stop codons disrupts attenuation control of biosynthetic operons, and may alter expression of some overlapping genes. Together, these functional alterations may either augment the protein repertoire or produce deleterious proteins.
PMC PMC5373646
PMID 28301469
DOI
Run Accession Study Accession Scientific Name Cell Line Library Type Treatment GWIPS-viz Trips-Viz Reads BAM BigWig (F) BigWig (R)
SRR4421280 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421281 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421282 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421283 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421284 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421285 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421286 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421287 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421288 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421289 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421290 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421291 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421292 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421293 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421294 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421295 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421296 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421299 PRJNA348358 Escherichia coli Ribo-Seq
SRR4421300 PRJNA348358 Escherichia coli Ribo-Seq
Run Accession Study Accession Scientific Name Cell Line Library Type Treatment GWIPS-viz Trips-Viz Reads BAM BigWig (F) BigWig (R)

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