Inhibiting translation elongation can aid genome duplication in Escherichia coli.

Kamila K Myka,Milind K Gupta,Mark S Dillingham,Caroline Meharg,Peter Mcglynn,Nigel J Savery,Michelle Hawkins,Aisha H Syeda,Robert G Lloyd

doi:10.1093/nar/gkw1254

Abstract

Conflicts between replication and transcription challenge chromosome duplication. Escherichia coli replisome movement along transcribed DNA is promoted by Rep and UvrD accessory helicases with Δrep ΔuvrD cells being inviable under rapid growth conditions. We have discovered that mutations in a tRNA gene, aspT, in an aminoacyl tRNA synthetase, AspRS, and in a translation factor needed for efficient proline–proline bond formation, EF-P, suppress Δrep ΔuvrD lethality. Thus replication-transcription conflicts can be alleviated by the partial sacrifice of a mechanism that reduces replicative barriers, namely translating ribosomes that reduce RNA polymerase backtracking. Suppression depends on RelA-directed synthesis of (p)ppGpp, a signalling molecule that reduces replication-transcription conflicts, with RelA activation requiring ribosomal pausing. Levels of (p)ppGpp in these suppressors also correlate inversely with the need for Rho activity, an RNA translocase that can bind to emerging transcripts and displace transcription complexes. These data illustrate the fine balance between different mechanisms in facilitating gene expression and genome duplication and demonstrate that accessory helicases are a major determinant of this balance. This balance is also critical for other aspects of bacterial survival: the mutations identified here increase persistence indicating that similar mutations could arise in naturally occurring bacterial populations facing antibiotic challenge.

Highlights

Cell survival from one generation to the relies on efficient and faithful replication of the genome
Escherichia coli replisome movement along transcribed DNA is promoted by Rep and UvrD accessory helicases with rep uvrD cells being inviable under rapid growth conditions
We have discovered that mutations in a tRNA gene, aspT, in an aminoacyl tRNA synthetase, AspRS, and in a translation factor needed for efficient proline–proline bond formation, elongation factor P (EF-P), suppress rep uvrD lethality

Summary

Introduction

Cell survival from one generation to the relies on efficient and faithful replication of the genome. The template for replication frequently harbours obstacles that have the potential to interfere with the progression of replisome complexes, the macromolecular machines responsible for driving genome duplication. One major challenge is the very high free energy of binding of transcription complexes that creates a need to disrupt many RNA polymerase–nucleic acid interactions as replication proceeds. Paused RNAPs can diffuse backwards along the template resulting in displacement of the transcript 3 end from the RNAP active site. This causes an inability to resume transcription, creating barriers to replication that threaten genome stability [4,5]

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Conclusion