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DNA replication triggered by double-stranded breaks in E. coli: dependence on homologous recombination functions
1Department of Cell Biology, University of New Mexico School of Medicine, Albuquerque 87131.
Cell
|September 23, 1994
Summary
Homologous recombination-dependent DNA replication (RDR) was shown in E. coli using lambda terminase to break a plasmid cos site. This process is influenced by the chi hotspot, SOS response, and specific mutations.
Area of Science:
- Molecular Biology
- Bacteriology
- Genetics
Background:
- Homologous recombination plays a crucial role in DNA repair and genome stability.
- Understanding DNA replication mechanisms is fundamental to molecular biology.
- Bacteriophage lambda terminase is involved in DNA packaging and processing.
Purpose of the Study:
- To demonstrate and characterize homologous recombination-dependent DNA replication (RDR) in E. coli.
- To investigate the role of lambda terminase, the chi hotspot, and various genetic mutations in RDR.
- To explore the relationship between RDR and inducible stable DNA replication.
Main Methods:
- Introduction of a double-stranded break into a lambda cos site-carrying plasmid using lambda terminase in E. coli.
- Analysis of RDR in wild-type cells, cells with the chi hotspot, SOS-induced cells, and various rec mutants (recD, recBC sbcA, recN).
- Determination of DNA synthesis modes (theta or rolling circle) and resulting products (monomers or multimers).
Main Results:
- RDR was successfully demonstrated in E. coli expressing lambda terminase, initiating replication from a double-stranded break.
- The chi hotspot facilitated RDR in normal cells, but was dispensable under SOS induction or in recD mutant cells.
- recBC sbcA mutants supported RDR, while recN mutations inhibited it in normal cells but not in SOS-induced cells.
- RDR proceeded via theta or rolling circle modes, producing circular monomers or linear multimers, respectively.
- Previously identified inducible stable DNA replication was reclassified as a specific form of RDR.
Conclusions:
- Homologous recombination-dependent DNA replication is a viable replication mechanism in E. coli, inducible by specific DNA breaks.
- The chi hotspot, SOS response, and RecBC pathway components significantly modulate RDR efficiency.
- RDR represents a broader category of DNA replication, encompassing inducible stable DNA replication.
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