Title
Ribosome Excursions during mRNA Translocation Mediate Broad Branching of Frameshift Pathways
Date Issued
26 February 2015
Access level
open access
Resource Type
journal article
Author(s)
University of California
Publisher(s)
Cell Press
Abstract
Programmed ribosomal frameshifting produces alternative proteins from a single transcript. -1 frameshifting occurs on Escherichia coli's dnaX mRNA containing a slippery sequence AAAAAAG and peripheral mRNA structural barriers. Here, we reveal hidden aspects of the frameshifting process, including its exact location on the mRNA and its timing within the translation cycle. Mass spectrometry of translated products shows that ribosomes enter the -1 frame from not one specific codon but various codons along the slippery sequence and slip by not just -1 but also -4 or +2 nucleotides. Single-ribosome translation trajectories detect distinctive codon-scale fluctuations in ribosome-mRNA displacement across the slippery sequence, representing multiple ribosomal translocation attempts during frameshifting. Flanking mRNA structural barriers mechanically stimulate the ribosome to undergo back-and-forth translocation excursions, broadly exploring reading frames. Both experiments reveal aborted translation around mutant slippery sequences, indicating that subsequent fidelity checks on newly adopted codon position base pairings lead to either resumed translation or early termination.
Start page
870
End page
881
Volume
160
Issue
5
Language
English
OCDE Knowledge area
Genética humana Bioquímica, Biología molecular
Scopus EID
2-s2.0-84923378894
PubMed ID
Source
Cell
ISSN of the container
00928674
Sponsor(s)
We thank Dr. S.B. Smith for help with the optical tweezers instrument, Dr. T. Iavarone (QB3/Chemistry Mass Spectrometry Facility) for assistance on MS data collection and analysis, Dr. L. Lancaster and Prof. H. Noller for expertise in sample preparations, Dr. C.L. Hetherington for comments on the manuscript, and members of the Tinoco and Bustamante labs for discussions. This work was supported by grants from the National Institutes of Health (GM10840 to I.T.).
Sources of information: Directorio de Producción Científica Scopus