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Originally published as Genetics Published Articles Ahead of Print on October 8, 2006.

Genetics, Vol. 174, 2137-2149, December 2006, Copyright © 2006
doi:10.1534/genetics.106.063750

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Roles of PriA Protein and Double-Strand DNA Break Repair Functions in UV-Induced Restriction Alleviation in Escherichia coli

Ivana Ivancic-Bace*,1, Ignacija Vlasic{dagger}, Gordana Cogelja-Cajo{dagger}, Krunoslav Brcic-Kostic{dagger} and Erika Salaj-Smic{dagger}

* Department of Molecular Biology, Faculty of Science, University of Zagreb and {dagger} Department of Molecular Biology, Ruder Boskovic Institute, HR-10000 Zagreb, Croatia

1 Corresponding author: Department of Molecular Biology, Faculty of Science, University of Zagreb, Horvatovac 102A, HR-10000 Zagreb, Croatia.
E-mail: ivanai{at}irb.hr

It has been widely considered that DNA modification protects the chromosome of bacteria E. coli K-12 against their own restriction–modification systems. Chromosomal DNA is protected from degradation by methylation of target sequences. However, when unmethylated target sequences are generated in the host chromosome, the endonuclease activity of the EcoKI restriction-modification enzyme is inactivated by the ClpXP protease and DNA is protected. This process is known as restriction alleviation (RA) and it can be induced by UV irradiation (UV-induced RA). It has been proposed that chromosomal unmethylated target sequences, a signal for the cell to protect its own DNA, can be generated by homologous recombination during the repair of damaged DNA. In this study, we wanted to further investigate the genetic requirements for recombination proteins involved in the generation of unmethylated target sequences. For this purpose, we monitored the alleviation of EcoKI restriction by measuring the survival of unmodified {lambda} in UV-irradiated cells. Our genetic analysis showed that UV-induced RA is dependent on the excision repair protein UvrA, the RecA-loading activity of the RecBCD enzyme, and the primosome assembly activity of the PriA helicase and is partially dependent on RecFOR proteins. On the basis of our results, we propose that unmethylated target sequences are generated at the D-loop by the strand exchange of two hemi-methylated duplex DNAs and subsequent initiation of DNA replication.






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Copyright © 2006 by the Genetics Society of America.