Inhibitory effect of UvrD and DinG on the replication of ColE1-derived plasmids in Escherichia coli

Abstract

CspA has been identified as a major cold-shock protein in Escherichia coli. CspA binds to RNAs which are abnormally folded at low temperature and then acts as an RNA chaperone unfolding those RNAs. The dramatic expression of cspA at low temperature is contributed by posttranscriptional stability and robust translatability. Interestingly, when cspA mRNA encoding a premature nonsense codon was overexpressed at low temperature, cell growth was completely inhibited. This phenotype was termed LACE (the low temperature-dependent antibiotic effect of truncated cspA expression), and this lethality resulted from exclusive stalling of most ribosomes on mutant cspA mRNAs. In a previous study, we demonstrated that overexpression of the ATP-dependent DNA helicases, UvrD and DinG, suppressed the lethality and ribosome stalling caused by mutant cspA mRNA. In the present study, we attempted to elucidate how these two DNA helicases help recover normal growth under LACE condition. Interestingly, we found that UvrD and DinG appeared to have an ability to down-regulate the replication of pUC-based high copy plasmid. In plasmid copy number tests, the amount of pUC-based plasmid encoding mutant cspA was reduced by 3–10-fold when either UvrD or DinG was expressed. Through a β-galactosidase activity assay, we also confirmed that expression of the lacZα gene inserted into the pUC-based plasmid was significantly reduced due to down-regulation of plasmid replication. Our findings imply that UvrD and DinG, known as non-replicative helicases, play a novel role in the regulation of ColE1-like plasmid replication.