Absolute quantification of translational regulation and burden using combined sequencing approaches

Thomas E. Gorochowski, Irina Chelysheva, Mette Eriksen, Priyanka Nair, Steen Pedersen, Zoya Ignatova

Research output: Contribution to journalArticle (Academic Journal)peer-review

15 Citations (Scopus)
210 Downloads (Pure)

Abstract

Translation of mRNAs into proteins is a key cellular process. Ribosome binding sites and stop codons provide signals to initiate and terminate translation, while stable secondary mRNA structures can induce translational recoding events. Fluorescent proteins are commonly used to characterize such elements but require the modification of a part's natural context and allow only a few parameters to be monitored concurrently. Here, we combine Ribo‐seq with quantitative RNA‐seq to measure at nucleotide resolution and in absolute units the performance of elements controlling transcriptional and translational processes during protein synthesis. We simultaneously measure 779 translation initiation rates and 750 translation termination efficiencies across the Escherichia coli transcriptome, in addition to translational frameshifting induced at a stable RNA pseudoknot structure. By analyzing the transcriptional and translational response, we discover that sequestered ribosomes at the pseudoknot contribute to a σ32‐mediated stress response, codon‐specific pausing, and a drop in translation initiation rates across the cell. Our work demonstrates the power of integrating global approaches toward a comprehensive and quantitative understanding of gene regulation and burden in living cells.
Original languageEnglish
Article numbere8719
Pages (from-to)e8719
Number of pages15
JournalMolecular Systems Biology
Volume15
Issue number5
DOIs
Publication statusPublished - 3 May 2019

Structured keywords

  • BrisSynBio
  • Bristol BioDesign Institute

Keywords

  • genetic circuits
  • Ribo‐seq
  • RNA‐seq
  • transcription
  • translation

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