Temporal and site-specific adp-ribosylation dynamics upon different genotoxic stresses

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The DNA damage response revolves around transmission of information via post-translational modifications, including reversible protein ADP-ribosylation. Here, we applied a massspectrometry-based Af1521 enrichment technology for the identification and quantification of ADPribosylation sites as a function of various DNA damage stimuli and time. In total, we detected 1681 ADP-ribosylation sites residing on 716 proteins in U2OS cells and determined their temporal dynamics after exposure to the genotoxins H2O2 and MMS. Intriguingly, we observed a widespread but low-abundance serine ADP-ribosylation response at the earliest time point, with later time points centered on increased modification of the same sites. This suggests that early serine ADPribosylation events may serve as a platform for an integrated signal response. While treatment with H2O2 and MMS induced homogenous ADP-ribosylation responses, we observed temporal differences in the ADP-ribosylation site abundances. Exposure to MMS-induced alkylating stress induced the strongest ADP-ribosylome response after 30 min, prominently modifying proteins involved in RNA processing, whereas in response to H2O2-induced oxidative stress ADP-ribosylation peaked after 60 min, mainly modifying proteins involved in DNA damage pathways. Collectively, the dynamic ADP-ribosylome presented here provides a valuable insight into the temporal cellular regulation of ADP-ribosylation in response to DNA damage.

Udgave nummer11
Antal sider16
StatusUdgivet - 2021

Bibliografisk note

Funding Information:
Funding:The work carried out in this study was in part supported by the Novo Nordisk Foundation Center for Protein Research, the Novo Nordisk Foundation (grant agreement numbers NNF14CC0001 and NNF13OC0006477), Danish Council of Independent Research (grant agreement numbers 4002-00051, 4183-00322A, 8020-00220B and 0135-00096B), and The Danish Cancer Society (grant agreement R146-A9159-16-S2). The proteomics technology applied was part of a project that has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement EPIC-XS-823839.

Publisher Copyright:
© 2021 by the authors. Licensee MDPI, Basel, Switzerland.

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