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bioRxiv · 10.1101/2020.10.27.357855

DNA damage-signaling, homologous recombination and genetic mutation induced by 5-azacytidine and DNA-protein crosslinks in Escherichia coli

Klaric, J. A.·Glass, D. J.·Perr, E. L.·Reuven, A. D.·Towne, M. J.·Lovett, S. T.
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Abstract

Covalent linkage between DNA and proteins produces highly toxic lesions and can be caused by commonly used chemotherapeutic agents, by internal and external chemicals and by radiation. In this study, using Escherichia coli, we investigate the consequences of 5-azacytidine (5-azaC), which traps covalent complexes between itself and the Dcm cytosine methyltransferase protein. DNA protein crosslink-dependent effects can be ascertained by effects that arise in wild-type but not in dcm{Delta} strains. We find that 5-azaC induces the bacterial DNA damage response and stimulates homologous recombination, a component of which is Dcm-dependent. Template-switching at an imperfect inverted repeat ("quasipalindrome", QP) is strongly enhanced by 5-azaC and this enhancement was entirely Dcm-dependent. The SOS response helps ameliorate the mutagenic effect of 5-azaC but unbalanced expression of the SOS-induced DNA polymerases, especially PolIV, stimulates QP-associated mutagenesis. In the absence of Lon protease, Dcm-dependent QP-mutagenesis is elevated, suggesting it may play a role in 5-azaC tolerance. Deletions at short tandem repeats, which occur likewise by a replication template-switch, are elevated, but only modestly, by 5-azaC. We see evidence for Dcm-dependent and-independent killing by 5-azaC in sensitive mutants, such as recA, recB, and lon; homologous recombination and deletion mutations are also stimulated in part by a Dcm-independent effect of 5-azaC. Whether this occurs by a different protein/DNA crosslink or by an alternative form of DNA damage is unknown. HighlightsO_LI5-azacytidine is broadly mutagenic and recombinogenic C_LIO_LIIn E. coli, 5-azaC promotes genetic instability through Dcm methyltransferase. C_LIO_LIThere are other, unknown lesions induced by 5-azaC besides Dcm/DNA crosslinks C_LIO_LI5-azaC induces the SOS response, protecting cells from killing and genetic instability C_LI

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Klaric, J. A., Glass, D. J., Perr, E. L., Reuven, A. D., Towne, M. J., Lovett, S. T.. 2020-10-27. DNA damage-signaling, homologous recombination and genetic mutation induced by 5-azacytidine and DNA-protein crosslinks in Escherichia coli. https://doi.org/10.1101/2020.10.27.357855

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