SMC-like Wadjet system prevents plasmid transfer into Clostridium cellulovorans.

Abstract

This study demonstrates the impact of a Structure Maintenance of Chromosome (SMC)-like Wadjet system on the horizontal gene transfer of plasmids by conjugation to a recipient that naturally containing such a system for the first time. A Clostridium cellulovorans mutant with dramatically improved efficiency to receive plasmid DNA by conjugation was isolated and sequenced. Three spontaneous chromosomal deletions included a type II restriction-modification system, a putative CRISPR system, and a cluster of ORFs named jetABCD encoding a putative Wadjet system. Since nearly nothing is known about the role of naturally occurring Wadjet systems in their native host bacteria, markerless chromosomal deletion of jetABCD in the C. cellulovorans wildtype strain 743B was achieved and the effect on conjugative plasmid uptake was studied. The transconjugation frequency of the jetABCD mutant was increased by about five orders of magnitude compared to wildtype C. cellulovorans recipient cells. Bioinformatic analysis of genome sequences of the Bacillota phylum revealed near-complete mutually exclusive possession of either plasmids < 40 kb or jetABCD genes, indicating high efficiency of Wadjet systems in small plasmid prevention in bacteria. Importantly, the implications of this study go beyond the case of C. cellulovorans. Our study demonstrates that the eradication of Wadjet systems can dramatically improve the uptake of recombinant plasmids and thereby enhance genetic engineering of bacterial strains of interest for biotechnological applications. KEY POINTS: • Native Wadjet system inhibits plasmid transfer by conjugation in C. cellulovorans • Deleting jetABCD increased plasmid uptake by about five orders of magnitude • Possession of Wadjet systems efficiently block plasmid maintenance in Bacillota.