RRC ID 70555
Author Fujita T, Yokoyama T, Shirouzu M, Taguchi H, Ito T, Iwasaki S.
Title The landscape of translational stall sites in bacteria revealed by monosome and disome profiling.
Journal RNA
Abstract Ribosome pauses are associated with various cotranslational events and determine the fate of mRNAs and proteins. Thus, the identification of precise pause sites across the transcriptome is desirable; however, the landscape of ribosome pauses in bacteria remains ambiguous. Here, we harness monosome and disome (or collided ribosome) profiling strategies to survey ribosome pause sites in Escherichia coli. Compared to eukaryotes, ribosome collisions in bacteria showed remarkable differences: a low frequency of disomes at stop codons, collisions occurring immediately after 70S assembly on start codons, and shorter queues of ribosomes trailing upstream. The pause sites corresponded with the biochemical validation by integrated nascent chain profiling (iNP) to detect polypeptidyl-tRNA, an elongation intermediate. Moreover, the subset of those sites showed puromycin resistance, presenting slow peptidyl transfer. Among the identified sites, the ribosome pause at Asn586 of ycbZ was validated by biochemical reporter assay, tRNA sequencing (tRNA-seq), and cryo-electron microscopy (cryo-EM) experiments. Our results provide a useful resource for ribosome stalling sites in bacteria.
Volume 28(3)
Pages 290-302
Published 2022-3-1
DOI 10.1261/rna.078188.120
PII rna.078188.120
PMID 34906996
PMC PMC8848927
MeSH Codon, Terminator Escherichia coli Escherichia coli Proteins / genetics Escherichia coli Proteins / metabolism Peptide Chain Elongation, Translational* Peptide Chain Termination, Translational* RNA, Transfer / genetics RNA, Transfer / metabolism Ribosomes / metabolism*
IF 4.32
Prokaryotes E. coli