Feliciello, Isidoro; Ljubić, Sven; Đermić, Edyta; Ivanković, Siniša; Zahradka, Davor; Đermić, Damir (2024) Single-strand DNA-binding protein suppresses illegitimate recombination in Escherichia coli, acting in synergy with RecQ helicase. Scientific Reports, 14 . ISSN 2045-2322
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Abstract
Single-strand DNA-binding proteins SSB/RPA are ubiquitous and essential proteins that bind ssDNA in bacteria/eukaryotes and coordinate DNA metabolic processes such as replication, repair, and recombination. SSB protects ssDNA from degradation by nucleases, while also facilitating/regulating the activity of multiple partner proteins involved in DNA processes. Using Spi− assay, which detects aberrantly excised λ prophage from the E. coli chromosome as a measure of illegitimate recombination (IR) occurrence, we have shown that SSB inhibits IR in several DSB resection pathways. The conditional ssb-1 mutation produced a higher IR increase at the nonpermissive temperature than the recQ inactivation. A double ssb-1 recQ mutant had an even higher level of IR, while showing reduced homologous recombination (HR). Remarkably, the ssb gene overexpression complemented recQ deficiency in suppressing IR, indicating that the SSB function is epistatic to RecQ. Overproduced truncated SSBΔC8 protein, which binds to ssDNA, but does not interact with partner proteins, only partially complemented recQ and ssb-1 mutations, while causing an IR increase in otherwise wild-type bacteria, suggesting that ssDNA binding of SSB is required but not sufficient for effective IR inhibition, which rather entails interaction with RecQ and likely some other protein(s). Our results depict SSB as the main genome caretaker in E. coli, which facilitates HR while inhibiting IR. In enabling high-fidelity DSB repair under physiological conditions SSB is assisted by RecQ helicase, whose activity it controls. Conversely, an excess of SSB renders RecQ redundant for IR suppression.
| Item Type: | Article | ||||||||
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| Uncontrolled Keywords: | SSB protein; Genome stability; Truncated SSB; SSB overproduction; λ Spi⎻ assay | ||||||||
| Subjects: | NATURAL SCIENCES > Biology > Microbiology | ||||||||
| Divisions: | Division of Molecular Biology Division of Molecular Medicine |
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| Depositing User: | Ana Zečević | ||||||||
| Date Deposited: | 05 Sep 2024 09:20 | ||||||||
| URI: | http://fulir.irb.hr/id/eprint/9062 | ||||||||
| DOI: | 10.1038/s41598-024-70817-5 |
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