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PARP7 is a proteotoxic stress sensor which labels proteins for degradation

ABSTRACT: ADP-ribosylation is a post-translational modification that plays a critical role in cellular stress responses. We have observed that during proteotoxic stress, cellular ADP-ribosylation increases, with ADP-ribosylated proteins accumulating in cytoplasmic foci containing ubiquitin and p62. During prolonged stress, these ADP-ribosylated proteins are transported to aggresomes and subsequently degraded via autophagy. In the absence of ubiquitination, ADP-ribosylated proteins become more prevalent and less soluble, indicating that ubiquitination is indispensable for this process. Upon inhibition of PARP7, accumulation of mono(ADP-ribosyl)ated proteins in response to proteotoxic stress is impeded. PARP7 turnover is very high under normal conditions, however, the protein stabilises following proteotoxic stress and thereby forms an ideal proteotoxic stress sensor. Finally, our findings imply that contrary to the current paradigm, perhaps not all ADP-ribosylation occurs on specific sites to regulate specific protein characteristics. Instead, it may be rather promiscuous to enable efficient protein degradation to prevent irreversible damage caused by defective proteins.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

SUBMITTER: Mirna Barsoum

LAB HEAD: Karla Feijs

PROVIDER: PXD065525 | Pride | 2025-09-01

REPOSITORIES: Pride

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	DIA-NN_results_PRIDE.zip	Other
	OR3_2024_10_25_KaFei_No_RBN1.raw	Raw
	OR3_2024_10_25_KaFei_No_RBN2.raw	Raw
	OR3_2024_10_25_KaFei_No_RBN3.raw	Raw
	OR3_2024_10_25_KaFei_No_RBN4.raw	Raw

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Transcriptional changes by RBN-2397 treatment and FRA1 knockdown in NCI-H1975 cells

Project description:PARP7 inhibitors reduce tumor growth in a cell-autonomous manner and by enhancing immune recognition through restoring nucleic acid (NA)-sensing-dependent innate immune signaling. However, the molecular targets of PARP7-mediated ADP-ribosylation, regulating cell survival and innate immune signaling, remained elusive. Here, we identified PARP7 as a nuclear and cysteine-specific mono-ART that ADP-ribosylates proteins critical for regulating gene expression, such as the AP-1 transcription factor FRA1. Upon PARP7 inhibition the loss of FRA1 ADP-ribosylation increased FRA1 degradation in a PSMC3 and proteasomal-dependent manner. To further investigate how FRA1 promotes cell survival, we investigated transcriptional changes after RBN-2397 treatment and FRA1 knockdown in the PARP7 inhibitor sensitive cell line NCI-H1975 by RNA sequencing.

2023-12-06 | GSE229674 | GEO

An androgen signaling axis uses a writer and a reader of ADP-ribosylation to control protein complex assembly

Project description:Androgen signaling through the androgen receptor (AR) regulates multiple pathways in both normal and prostate cancer cells. Androgen regulates diverse aspects of the AR life cycle, including its post-translational modification, but understanding how specific modifications influence AR activity has been mostly elusive. Here, we show that androgen regulates AR through a pathway mediated by the mono-ADP ribosyltransferase, Parp7. We show that Parp7 ADP-ribosylates AR on multiple cysteines, and that a subset of these sites mediates agonist-specific recruitment of the E3 ligase Dtx3L/Parp9. Tandem macrodomains in Parp9 selectively recognize ADP ribosylated AR, and Dtx3L/Parp9 affects expression of a subset of AR-regulated genes. Parp7, ADP-ribosylation of AR, and AR-Dtx3L/Parp9 complex assembly are inhibited by 60 Olaparib, a compound used clinically to inhibit poly-ADP-ribosyltransferases Parp1/2. Our study reveals the components of a new androgen signaling axis that uses a writer and reader of ADP-ribosylation to modulate AR activity.

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An androgen signaling axis uses a writer and a reader of ADP-ribosylation to control protein complex assembly

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Oxidative stress induced ADP-ribosylome

Project description:Oxidative stress is a potent inducer of protein ADP-ribosylation. Although the proteins modified under oxidative stress have been identified, it is not clear, whether the number of modified proteins and/or the number ADP-ribosylation sites varies with stress intensity. Here, we investigated both the qualitative and quantitative changes in the HeLa cell ADP-ribosylome induced by mild (4 - 16 uM), moderate (64 - 250 uM), or severe, but non-lethal (1 mM) H2O2 concentrations using shotgun and parallel reaction monitoring mass spectrometry (PRM-MS). After moderate and severe oxidative stress, approximately 50% of proteins not ADP-ribosylated under basal conditions were induced, while ADP-ribosylation of the other 50% already ADP-ribosylated under basal conditions remained constant. In contrast, mild oxidative stress caused a reduction in detectable ADP-ribosylation of many proteins, which were induced under more severe oxidative stress. Overall, the number of ADP-ribosylation sites modified/de-modified did not change significantly under the various degrees of oxidative stress. Moreover, we applied the PRM method to study protein ADP-ribosylation in ovarian cancer cells that display different sensitivities to PARP inhibitors. These studies revealed similar ADP-ribosylation responses following H2O2 treatment, which, however, correlated in extent with ARTD1 abundance in these cells. Overall, our new MS approaches have proven to be highly useful in monitoring cellular protein ADP-ribosylation and have revealed unexpected fluctuations in proteins ADP-ribosylation depending on the degree of oxidative stress.

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Substrate Identification Using a Chemical Genetics Approach Reveals a Role for PARP-7-Mediated MARylation in Controlling Microtubule Stability in Ovarian Cancer Cells

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PARP7-mediated ADP-ribosylation stabilizes FRA1 and restrains IRF3/IRF1-dependent apoptosis and immune signaling and in cancer cells

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