Ribosomes slide on lysine-encoding homopolymeric A stretches

Elife. 2015 Feb 19;4:e05534. doi: 10.7554/eLife.05534.

Abstract

Protein output from synonymous codons is thought to be equivalent if appropriate tRNAs are sufficiently abundant. Here we show that mRNAs encoding iterated lysine codons, AAA or AAG, differentially impact protein synthesis: insertion of iterated AAA codons into an ORF diminishes protein expression more than insertion of synonymous AAG codons. Kinetic studies in E. coli reveal that differential protein production results from pausing on consecutive AAA-lysines followed by ribosome sliding on homopolymeric A sequence. Translation in a cell-free expression system demonstrates that diminished output from AAA-codon-containing reporters results from premature translation termination on out of frame stop codons following ribosome sliding. In eukaryotes, these premature termination events target the mRNAs for Nonsense-Mediated-Decay (NMD). The finding that ribosomes slide on homopolymeric A sequences explains bioinformatic analyses indicating that consecutive AAA codons are under-represented in gene-coding sequences. Ribosome 'sliding' represents an unexpected type of ribosome movement possible during translation.

Keywords: E. coli; S. cerevisiae; biochemistry; poly(A); ribosome; sliding.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Blotting, Western
  • Codon / genetics*
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Gene Deletion
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Lysine / genetics*
  • Molecular Sequence Data
  • Poly A / genetics
  • Protein Biosynthesis / genetics*
  • RNA Helicases / genetics
  • RNA Helicases / metabolism
  • RNA Stability / genetics
  • RNA, Messenger / genetics*
  • Reverse Transcriptase Polymerase Chain Reaction
  • Ribosomes / genetics*
  • Ribosomes / metabolism
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism

Substances

  • Codon
  • Luminescent Proteins
  • RNA, Messenger
  • Saccharomyces cerevisiae Proteins
  • red fluorescent protein
  • Poly A
  • NAM7 protein, S cerevisiae
  • RNA Helicases
  • Lysine