Extrusion-modulated DnaA activity oscillations coordinate DNA replication with biomass growth

Area of Science:

• Microbiology
• Molecular Biology
• Genetics

Background:

• DNA replication control is crucial for bacterial growth.
• Existing models of DNA replication initiation in Escherichia coli, based on DnaA activity oscillations, lack direct experimental evidence and fail to explain the decoupling from dnaA expression.

Purpose of the Study:

• To establish a direct link between DnaA activity oscillations and DNA replication initiation.
• To investigate the regulatory mechanisms controlling DnaA activity independently of dnaA expression.
• To propose and validate a novel model for replication initiation.

Main Methods:

• Monitoring DnaA activity oscillations under various growth conditions and genetic perturbations.
• Analyzing the relationship between DnaA activity peaks and replication initiation timing.
• Investigating the role of DNA-binding proteins, specifically H-NS, in modulating DnaA activity and replication timing.

Main Results:

• Demonstrated robust oscillations in DnaA activity that precisely peak at replication initiation in Escherichia coli.
• Showed that DnaA activity oscillations persist independently of dnaA transcription levels.
• Proposed and provided evidence for an extrusion model where DNA-binding proteins regulate DnaA activity in response to biomass-DNA imbalance.
• Confirmed that perturbing H-NS affects DnaA activity and replication timing.

Conclusions:

• DnaA activity oscillations are a direct and robust trigger for DNA replication initiation in Escherichia coli.
• Replication initiation is regulated by mechanisms modulating DnaA activity independently of its expression levels.
• The proposed extrusion model offers a new framework for understanding DNA replication control in bacteria.