A non‐proteolytic release mechanism for HMCES‐DNA‐protein crosslinks
The conserved protein HMCES crosslinks to abasic (AP) sites in ssDNA to prevent strand scission and the formation of toxic dsDNA breaks during replication. Here, we report a non‐proteolytic release mechanism for HMCES‐DNA‐protein crosslinks (DPCs), which is regulated by DNA context. In ssDNA and at ssDNA‐dsDNA junctions, HMCES‐DPCs are stable, which efficiently protects AP sites against spontaneous incisions or cleavage by APE1 endonuclease. In contrast, HMCES‐DPCs are released in dsDNA, allowing APE1 to initiate downstream repair. Mechanistically, we show that release is governed by two components. First, a conserved glutamate residue, within HMCES' active site, catalyses reversal of the crosslink. Second, affinity to the underlying DNA structure determines whether HMCES re‐crosslinks or dissociates. Our study reveals that the protective role of HMCES‐DPCs involves their controlled release upon bypass by replication forks, which restricts DPC formation to a necessary minimum.
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© 2023 The Authors. Published under the terms of the CC BY NC ND 4.0 license. This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc-nd/4.0/ License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non‐commercial and no modifications or adaptations are made.
We thank T. Fröhlich for mass spectrometry analyses of recombinant HMCES protein and T. Mackens-Kiani for help with preparing Fig 1C. Research in the lab of DRS is supported by NIH grant no. GM129422 and an award from the Shurl and Kay Curci Foundation. Research in the lab of JS is supported by the European Research Council under the European Union's Horizon 2020 research and innovation programme (grant agreement number 801750), by the Alfried Krupp Prize for Young University Teachers awarded by the Alfried Krupp von Bohlen und Halbach Foundation, the European Molecular Biology Organization (YIP4644), the Vallee Foundation and the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) (Project ID 213249687—SFB 1064 and Project-ID 393547839—SFB 1361). NIH grant GM118129 to Peter M. Burgers supported RB-B. Open Access funding enabled and organized by Projekt DEAL. Open Access funding enabled and organized by Projekt DEAL.
Open Access funding enabled and organized by Projekt DEAL.
Maximilian Donsbach: Conceptualization; investigation; writing – original draft; writing – review and editing. Sophie Dürauer: Conceptualization; investigation; writing – original draft; writing – review and editing. Florian Grünert: Investigation; writing – review and editing. Kha T Nguyen: Investigation; writing – review and editing. Richa Nigam: Investigation; writing – review and editing. Denitsa Yaneva: Investigation; writing – review and editing. Pedro Weickert: Investigation; writing – review and editing. Rachel Bezalel-Buch: Resources. Daniel R Semlow: Supervision; funding acquisition; investigation; writing – review and editing. Julian Stingele: Conceptualization; supervision; funding acquisition; writing – original draft; project administration; writing – review and editing.
Conflict of Interest
The authors declare that they have no conflict of interest.