Abstract
Under anaerobic growth conditions, Escherichia coli operates a two-component signal transduction system, termed Arc, that consists of ArcB protein, a transmembrane sensor kinase and ArcA protein, the cognate response regulator. In response to low oxygen levels, autophosphorylated ArcB phosphorylates ArcA, and the resulting phosphorylated ArcA (ArcA-P) functions as a transcriptional regulator of the genes necessary to maintain anaerobic growth. Under anaerobic conditions, cells maintain a slow growth rate, suggesting that the initiation of chromosomal replication is regulated to reduce the initiation frequency. DNase I footprinting experiments revealed that ArcA-P binds to the left region of the chromosomal origin, oriC. ArcA-P did not affect the in vitro replication of plasmid DNA containing the ColE1 origin nor the in vitro replication of viral DNAs; however, ArcA-P specifically inhibited in vitro E. coli chromosomal replication. This inhibition was caused by the prevention of open complex formation, a necessary step in the initiation of chromosomal replication. Our in vitro results suggest that the Arc two-component system participates in regulating chromosomal initiation under anaerobic growth conditions.
Highlights
Most organisms, including Escherichia coli, are able to adapt to variable growth conditions and environmental changes by regulating gene expression [1, 2]
Increasing the amount of Transphosphorylation reactions (TP) mixture added to the footprinting reaction revealed that an ϳ150-bp region located at the left end of oriC, including the three 13-mer AT-rich regions, DnaA box R1, and IHF binding site, was protected from DNase I cleavage (Fig. 2)
ArcA protein phosphorylated by ArcB and ATP or by carbamyl phosphate binds the left end of oriC
Summary
Most organisms, including Escherichia coli, are able to adapt to variable growth conditions and environmental changes by regulating gene expression [1, 2]. The Arc signal transduction system consists of the ArcB and ArcA proteins, a transmembrane sensor kinase and its cognate response regulator, respectively [5, 6]. In response to oxygen deficiency or redox change, ArcB autophosphorylates in an ATP-dependent manner and converts to phosphorylated ArcB (ArcB-P) via Anaerobic conditions that induce the Arc two-component signal transduction system lead to reduction in growth rate [12]. We report that the binding of ArcA-P, phosphorylated by ArcB, to oriC results in the inhibition of chromosomal initiation This result suggests that the Arc two-component signal transduction system plays a role in the regulation of chromosomal initiation at oriC in response to oxygen deficiency
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