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New quantitative mass spectrometry approaches reveal different ADP-ribosylation phases dependent on the levels of oxidative stress


Bilan, Vera; Selevsek, Nathalie; Kistemaker, Hans A. V; Abplanalp, Jeannette; Feurer, Roxane; Filippov, Dmitri V; Hottiger, Michael O (2017). New quantitative mass spectrometry approaches reveal different ADP-ribosylation phases dependent on the levels of oxidative stress. Molecular & Cellular Proteomics, 16(5):949-958.

Abstract

Oxidative stress is a potent inducer of protein ADP-ribosylation. Although individual oxidative stress-induced ADP-ribosylated proteins have been identified, it is so far not clear to which extent different degrees of stress severity quantitatively and qualitatively alter ADP-ribosylation. Here, we investigated both quantitative and qualitative changes of the hydrogen peroxide (H2O2)-induced ADP-ribosylome using a label-free shotgun quantification and a parallel reaction monitoring (PRM) mass spectrometry approach for a selected number of identified ADP-ribosylated peptides. Although the major part of the basal HeLa ADP-ribosylome remained unchanged upon all tested H2O2 concentrations, some selected peptides change the extent of ADP-ribosylation depending on the degree of the applied oxidative stress. Low oxidative stress (i.e. 4 μm and 16 μm H2O2) caused a reduction in ADP-ribosylation of modified proteins detected under untreated conditions. In contrast, mid to strong oxidative stress (62 μm to 1 mm H2O2) induced a significant increase in ADP-ribosylation of oxidative stress-targeted proteins. The application of the PRM approach to SKOV3 and A2780, ovarian cancer cells displaying different sensitivities to PARP inhibitors, revealed that the basal and the H2O2-induced ADP-ribosylomes of SKOV3 and A2780 differed significantly and that the sensitivity to PARP inhibitors correlated with the level of ARTD1 expression in these cells. Overall, this new PRM-MS approach has proven to be sensitive in monitoring alterations of the ADP-ribosylome and has revealed unexpected alterations in proteins ADP-ribosylation depending on the degree of oxidative stress.

Abstract

Oxidative stress is a potent inducer of protein ADP-ribosylation. Although individual oxidative stress-induced ADP-ribosylated proteins have been identified, it is so far not clear to which extent different degrees of stress severity quantitatively and qualitatively alter ADP-ribosylation. Here, we investigated both quantitative and qualitative changes of the hydrogen peroxide (H2O2)-induced ADP-ribosylome using a label-free shotgun quantification and a parallel reaction monitoring (PRM) mass spectrometry approach for a selected number of identified ADP-ribosylated peptides. Although the major part of the basal HeLa ADP-ribosylome remained unchanged upon all tested H2O2 concentrations, some selected peptides change the extent of ADP-ribosylation depending on the degree of the applied oxidative stress. Low oxidative stress (i.e. 4 μm and 16 μm H2O2) caused a reduction in ADP-ribosylation of modified proteins detected under untreated conditions. In contrast, mid to strong oxidative stress (62 μm to 1 mm H2O2) induced a significant increase in ADP-ribosylation of oxidative stress-targeted proteins. The application of the PRM approach to SKOV3 and A2780, ovarian cancer cells displaying different sensitivities to PARP inhibitors, revealed that the basal and the H2O2-induced ADP-ribosylomes of SKOV3 and A2780 differed significantly and that the sensitivity to PARP inhibitors correlated with the level of ARTD1 expression in these cells. Overall, this new PRM-MS approach has proven to be sensitive in monitoring alterations of the ADP-ribosylome and has revealed unexpected alterations in proteins ADP-ribosylation depending on the degree of oxidative stress.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:05 Vetsuisse Faculty > Department of Molecular Mechanisms of Disease
07 Faculty of Science > Department of Molecular Mechanisms of Disease
Dewey Decimal Classification:570 Life sciences; biology
Language:English
Date:2017
Deposited On:08 Aug 2017 14:55
Last Modified:08 Aug 2017 14:55
Publisher:American Society for Biochemistry and Molecular Biology
ISSN:1535-9476
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1074/mcp.O116.065623
PubMed ID:28325851

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