Natural tuning of restriction endonuclease synthesis by cluster of rare arginine codons

Iwona Mruk,Agata Witczak,Tadeusz Kaczorowski

doi:10.1038/s41598-019-42311-w

Abstract

Restriction–modification (R-M) systems are highly widespread among bacteria and archaea, and they appear to play a pivotal role in modulating horizontal gene transfer, as well as in protecting the host organism against viruses and other invasive DNA particles. Type II R-M systems specify two independent enzymes: a restriction endonuclease (REase) and protective DNA methyltransferase (MTase). If the cell is to survive, the counteracting activities as toxin and antitoxin, must be finely balanced in vivo. The molecular basis of this regulatory process remains unclear and current searches for regulatory elements in R-M modules are focused mainly at the transcription step. In this report, we show new aspects of REase control that are linked to translation. We used the EcoVIII R-M system as a model. Both, the REase and MTase genes for this R-M system contain an unusually high number of rare arginine codons (AGA and AGG) when compared to the rest of the E. coli K-12 genome. Clusters of these codons near the N-terminus of the REase greatly affect the translational efficiency. Changing these to higher frequency codons for E. coli (CGC) improves the REase synthesis, making the R-M system more potent to defend its host against bacteriophages. However, this improved efficiency in synthesis reduces host fitness due to increased autorestriction. We hypothesize that expression of the endonuclease gene can be modulated depending on the host genetic context and we propose a novel post-transcriptional mode of R–M system regulation that alleviates the potential lethal action of the restriction enzyme.

Highlights

Restriction-modification (R-M) systems are highly widespread among prokaryotes[1,2,3]
This led us to focus on the codon usage for a few R-M systems isolated from E. coli
We found a significant difference in the overall GC content of the R-M system unit compared to its natural carrier - plasmid pEC156 (49.3%) and to the E. coli genome (50.8%)

Summary

Introduction

Restriction-modification (R-M) systems are highly widespread among prokaryotes[1,2,3]. The REase cleaves at the recognition sequence while the MTase adds methyl group to modify such sites, to make them protected from the action of the cognate REase[4] This way the genomic DNA is marked as the self, in contrast to any foreign DNA entering the cell that lacks such modification (non-self) and become the target for degradation by host REase. R-M systems must have a time-dependent control that initially favours the expression and action of the MTase if the newly introduced gene are to be acquired by the new host[19,20,21,22] The details underlying these molecular processes are unclear, though it seems that genetic feedback loops play a central role. We hypothesise that the expression of the endonuclease is modulated by host genetic context, and we propose a novel, post-transcriptional mode of R–M system regulation that may help alleviate the lethality of unbalanced restriction enzyme gene expression

Methods

Results

Conclusion