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Inhibition of homologous recombination by the plasmid MucA'B complex
C Venderbure1, A Chastanet, F Boudsocq
1Institut Curie, Centre Universitaire, F-91405 Orsay, France.
Abstract:
By its functional interaction with a RecA polymer, the mutagenic UmuD'C complex possesses an antirecombination activity. We show here that MucA'B, a functional homolog of the UmuD'C complex, inhibits homologous recombination as well. In F- recipients expressing MucA'B from a Ptac promoter, Hfr x F- recombination decreased with increasing MucA'B concentrations down to 50-fold. In damage-induced pKM101-containing cells expressing MucA'B from the native promoter, recombination between a UV-damaged F lac plasmid and homologous chromosomal DNA decreased 10-fold. Overexpression of MucA'B together with UmuD'C resulted in a synergistic inhibition of recombination. RecA[UmuR] proteins, which are resistant to UmuD'C inhibition of recombination, are inhibited by MucA'B while promoting MucA'B-promoted mutagenesis efficiently. The data suggest that MucA'B and UmuD'C contact a RecA polymer at distinct sites. The MucA'B complex was more active than UmuD'C in promoting UV mutagenesis, yet it did not inhibit recombination more than UmuD'C does. The enhanced mutagenic potential of MucA'B may result from its inherent superior capacity to assist DNA polymerase in trans-lesion synthesis. In the course of this work, we found that the natural plasmid pKM101 expresses around 45,000 MucA and 13,000 MucB molecules per lexA(Def) cell devoid of LexA. These molecular Muc concentrations are far above those of the chromosomally encoded Umu counterparts. Plasmid pKM101 belongs to a family of broad-host-range conjugative plasmids. The elevated levels of the Muc proteins might be required for successful installation of pKM101-like plasmids into a variety of host cells.
Insights
The MucA
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- The UmuD'C complex interacts with RecA to inhibit recombination.
- Homologous recombination is a critical DNA repair pathway.
Purpose of the Study:
- To investigate the antirecombination activity of the MucA'B complex, a functional homolog of UmuD'C.
- To compare the mutagenic and antirecombination activities of MucA'B and UmuD'C.
Main Methods:
- Experiments were conducted in E. coli strains with varying expression levels of MucA'B and UmuD'C.
- Recombination frequencies were measured using Hfr x F- crosses and plasmid-chromosome recombination assays.
- Mutagenesis assays were performed to assess the trans-lesion synthesis capabilities.
Main Results:
- MucA'B inhibits homologous recombination in a dose-dependent manner, similar to UmuD'C.
- Overexpression of MucA'B and UmuD'C together resulted in synergistic inhibition of recombination.
- MucA'B efficiently promotes mutagenesis and inhibits RecA proteins resistant to UmuD'C inhibition, suggesting distinct interaction sites with RecA.
Conclusions:
- MucA'B, like UmuD'C, possesses antirecombination activity through interaction with RecA polymers.
- MucA'B exhibits enhanced mutagenic potential, possibly due to superior DNA polymerase assistance in trans-lesion synthesis.
- High expression levels of MucA'B from plasmid pKM101 may be crucial for plasmid establishment in diverse host cells.