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Molecular Cell Nov 2021ADP-ribose (ADPr) readers are essential components of ADP-ribosylation signaling, which regulates genome maintenance and immunity. The identification and discrimination...
ADP-ribose (ADPr) readers are essential components of ADP-ribosylation signaling, which regulates genome maintenance and immunity. The identification and discrimination between monoADPr (MAR) and polyADPr (PAR) readers is difficult because of a lack of suitable affinity-enrichment reagents. We synthesized well-defined ADPr probes and used these for affinity purifications combined with relative and absolute quantitative mass spectrometry to generate proteome-wide MAR and PAR interactomes, including determination of apparent binding affinities. Among the main findings, MAR and PAR readers regulate various common and distinct processes, such as the DNA-damage response, cellular metabolism, RNA trafficking, and transcription. We monitored the dynamics of PAR interactions upon induction of oxidative DNA damage and uncovered the mechanistic connections between ubiquitin signaling and ADP-ribosylation. Taken together, chemical biology enables exploration of MAR and PAR readers using interaction proteomics. Furthermore, the generated MAR and PAR interaction maps significantly expand our current understanding of ADPr signaling.
Topics: ADP-Ribosylation; Adenosine Diphosphate; Adenosine Diphosphate Ribose; Allosteric Site; Animals; Antibodies, Monoclonal; Binding Sites; Biotinylation; Cell Communication; DNA Damage; Genetic Techniques; HeLa Cells; Humans; Mass Spectrometry; Mice; Protein Binding; Protein Processing, Post-Translational; Proteome; Proteomics; Signal Transduction; Ubiquitin; Ubiquitin-Protein Ligases
PubMed: 34551281
DOI: 10.1016/j.molcel.2021.08.037 -
Genes & Development Mar 2020Poly-adenosine diphosphate-ribose polymerases (PARPs) promote ADP-ribosylation, a highly conserved, fundamental posttranslational modification (PTM). PARP catalytic... (Review)
Review
Poly-adenosine diphosphate-ribose polymerases (PARPs) promote ADP-ribosylation, a highly conserved, fundamental posttranslational modification (PTM). PARP catalytic domains transfer the ADP-ribose moiety from NAD to amino acid residues of target proteins, leading to mono- or poly-ADP-ribosylation (MARylation or PARylation). This PTM regulates various key biological and pathological processes. In this review, we focus on the roles of the PARP family members in inflammation and host-pathogen interactions. Here we give an overview the current understanding of the mechanisms by which PARPs promote or suppress proinflammatory activation of macrophages, and various roles PARPs play in virus infections. We also demonstrate how innovative technologies, such as proteomics and systems biology, help to advance this research field and describe unanswered questions.
Topics: ADP-Ribosylation; Host-Pathogen Interactions; Humans; Inflammation; Macrophages; Poly(ADP-ribose) Polymerases; Proteomics; Research; Systems Biology; Virus Diseases
PubMed: 32029454
DOI: 10.1101/gad.334425.119 -
Chemical Reviews Jul 2023Biomolecular condensates are reversible compartments that form through a process called phase separation. Post-translational modifications like ADP-ribosylation can... (Review)
Review
Biomolecular condensates are reversible compartments that form through a process called phase separation. Post-translational modifications like ADP-ribosylation can nucleate the formation of these condensates by accelerating the self-association of proteins. Poly(ADP-ribose) (PAR) chains are remarkably transient modifications with turnover rates on the order of minutes, yet they can be required for the formation of granules in response to oxidative stress, DNA damage, and other stimuli. Moreover, accumulation of PAR is linked with adverse phase transitions in neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. In this review, we provide a primer on how PAR is synthesized and regulated, the diverse structures and chemistries of ADP-ribosylation modifications, and protein-PAR interactions. We review substantial progress in recent efforts to determine the molecular mechanism of PAR-mediated phase separation, and we further delineate how inhibitors of PAR polymerases may be effective treatments for neurodegenerative pathologies. Finally, we highlight the need for rigorous biochemical interrogation of ADP-ribosylation in vivo and in vitro to clarify the exact pathway from PARylation to condensate formation.
Topics: Poly Adenosine Diphosphate Ribose; Poly(ADP-ribose) Polymerases; Biomolecular Condensates; Poly ADP Ribosylation; Protein Processing, Post-Translational
PubMed: 37115110
DOI: 10.1021/acs.chemrev.2c00851 -
Biochemical Pharmacology Sep 2019ADP-ribosylation (ADPr) is an ancient reversible modification of cellular macromolecules controlling major biological processes as diverse as DNA damage repair,... (Review)
Review
ADP-ribosylation (ADPr) is an ancient reversible modification of cellular macromolecules controlling major biological processes as diverse as DNA damage repair, transcriptional regulation, intracellular transport, immune and stress responses, cell survival and proliferation. Furthermore, enzymatic reactions of ADPr are central in the pathogenesis of many human diseases, including infectious conditions. By providing a review of ADPr signalling in bacterial systems, we highlight the relevance of this chemical modification in the pathogenesis of human diseases depending on host-pathogen interactions. The post-antibiotic era has raised the need to find alternative approaches to antibiotic administration, as major pathogens becoming resistant to antibiotics. An in-depth understanding of ADPr reactions provides the rationale for designing novel antimicrobial strategies for treatment of infectious diseases. In addition, the understanding of mechanisms of ADPr by bacterial virulence factors offers important hints to improve our knowledge on cellular processes regulated by eukaryotic homologous enzymes, which are often involved in the pathogenesis of human diseases.
Topics: ADP-Ribosylation; Animals; Anti-Infective Agents; Drug Delivery Systems; Endotoxins; Humans; Signal Transduction
PubMed: 31176616
DOI: 10.1016/j.bcp.2019.06.001 -
Molecular Cell Jul 2023Modification of nucleic acids by ADP-ribosylation is catalyzed by various ADP-ribosyltransferases, including the DarT enzyme. The latter is part of the bacterial...
Modification of nucleic acids by ADP-ribosylation is catalyzed by various ADP-ribosyltransferases, including the DarT enzyme. The latter is part of the bacterial toxin-antitoxin (TA) system DarTG, which was shown to provide control of DNA replication and bacterial growth as well as protection against bacteriophages. Two subfamilies have been identified, DarTG1 and DarTG2, which are distinguished by their associated antitoxins. While DarTG2 catalyzes reversible ADP-ribosylation of thymidine bases employing a macrodomain as antitoxin, the DNA ADP-ribosylation activity of DarTG1 and the biochemical function of its antitoxin, a NADAR domain, are as yet unknown. Using structural and biochemical approaches, we show that DarT1-NADAR is a TA system for reversible ADP-ribosylation of guanosine bases. DarT1 evolved the ability to link ADP-ribose to the guanine amino group, which is specifically hydrolyzed by NADAR. We show that guanine de-ADP-ribosylation is also conserved among eukaryotic and non-DarT-associated NADAR members, indicating a wide distribution of reversible guanine modifications beyond DarTG systems.
Topics: Guanosine; ADP-Ribosylation; ADP Ribose Transferases; Eukaryotic Cells; Antitoxins; Adenosine Diphosphate Ribose
PubMed: 37390817
DOI: 10.1016/j.molcel.2023.06.013 -
Cells Oct 2021The DNA damage response revolves around transmission of information via post-translational modifications, including reversible protein ADP-ribosylation. Here, we applied...
The DNA damage response revolves around transmission of information via post-translational modifications, including reversible protein ADP-ribosylation. Here, we applied a mass-spectrometry-based Af1521 enrichment technology for the identification and quantification of ADP-ribosylation 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 HO 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 ADP-ribosylation events may serve as a platform for an integrated signal response. While treatment with HO 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 HO-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.
Topics: ADP-Ribosylation; Cell Line, Tumor; DNA Damage; Humans; Hydrogen Peroxide; Methyl Methanesulfonate; Signal Transduction; Time Factors
PubMed: 34831150
DOI: 10.3390/cells10112927 -
Proceedings of the National Academy of... Dec 2023PARP7 was reported to promote tumor growth in a cell-autonomous manner and by repressing the antitumor immune response. Nevertheless, the molecular mechanism of how...
PARP7 was reported to promote tumor growth in a cell-autonomous manner and by repressing the antitumor immune response. Nevertheless, the molecular mechanism of how PARP7-mediated ADP-ribosylation exerts these effects in cancer cells remains elusive. Here, we identified PARP7 as a nuclear and cysteine-specific mono-ADP-ribosyltransferase that modifies targets critical for regulating transcription, including the AP-1 transcription factor FRA1. Loss of FRA1 ADP-ribosylation via PARP7 inhibition by RBN-2397 or mutation of the ADP-ribosylation site C97 increased FRA1 degradation by the proteasome via PSMC3. The reduction in FRA1 protein levels promoted IRF1- and IRF3-dependent cytokine as well as proapoptotic gene expression, culminating in CASP8-mediated apoptosis. Furthermore, high PARP7 expression was indicative of the PARP7 inhibitor response in FRA1-positive lung and breast cancer cells. Collectively, our findings highlight the connected roles of PARP7 and FRA1 and emphasize the clinical potential of PARP7 inhibitors for FRA1-driven cancers.
Topics: Humans; ADP Ribose Transferases; ADP-Ribosylation; Apoptosis; Cell Transformation, Neoplastic; Gene Expression Regulation; Interferon Regulatory Factor-1; Interferon Regulatory Factor-3; Neoplasms; Nucleoside Transport Proteins; Proto-Oncogene Proteins c-fos
PubMed: 38011562
DOI: 10.1073/pnas.2309047120 -
International Journal of Molecular... Oct 2023ADP-ribosylation is a post-translational modification of proteins that plays a key role in various cellular processes, including DNA repair. Recently, significant... (Review)
Review
ADP-ribosylation is a post-translational modification of proteins that plays a key role in various cellular processes, including DNA repair. Recently, significant progress has been made in understanding the mechanism and function of ADP-ribosylation in DNA repair. ADP-ribosylation can regulate the recruitment and activity of DNA repair proteins by facilitating protein-protein interactions and regulating protein conformations. Moreover, ADP-ribosylation can influence additional post-translational modifications (PTMs) of proteins involved in DNA repair, such as ubiquitination, methylation, acetylation, phosphorylation, and SUMOylation. The interaction between ADP-ribosylation and these additional PTMs can fine-tune the activity of DNA repair proteins and ensure the proper execution of the DNA repair process. In addition, PARP inhibitors have been developed as a promising cancer therapeutic strategy by exploiting the dependence of certain cancer types on the PARP-mediated DNA repair pathway. In this paper, we review the progress of ADP-ribosylation in DNA repair, discuss the crosstalk of ADP-ribosylation with additional PTMs in DNA repair, and summarize the progress of PARP inhibitors in cancer therapy.
Topics: Humans; Poly(ADP-ribose) Polymerases; Poly(ADP-ribose) Polymerase Inhibitors; ADP-Ribosylation; DNA Repair; Protein Processing, Post-Translational; Neoplasms; Proteins
PubMed: 37834477
DOI: 10.3390/ijms241915028 -
Cell Reports Sep 2023The timely removal of ADP-ribosylation is crucial for efficient DNA repair. However, much remains to be discovered about ADP-ribosylhydrolases. Here, we characterize the...
The timely removal of ADP-ribosylation is crucial for efficient DNA repair. However, much remains to be discovered about ADP-ribosylhydrolases. Here, we characterize the physiological role of TARG1, an ADP-ribosylhydrolase that removes aspartate/glutamate-linked ADP-ribosylation. We reveal its function in the DNA damage response and show that the loss of TARG1 sensitizes cells to inhibitors of topoisomerase II, ATR, and PARP. Furthermore, we find a PARP1-mediated synthetic lethal interaction between TARG1 and PARG, driven by the toxic accumulation of ADP-ribosylation, that induces replication stress and genomic instability. Finally, we show that histone PARylation factor 1 (HPF1) deficiency exacerbates the toxicity and genomic instability induced by excessive ADP-ribosylation, suggesting a close crosstalk between components of the serine- and aspartate/glutamate-linked ADP-ribosylation pathways. Altogether, our data identify TARG1 as a potential biomarker for the response of cancer cells to PARP and PARG inhibition and establish that the interplay of TARG1 and PARG protects cells against genomic instability.
Topics: Humans; Poly(ADP-ribose) Polymerase Inhibitors; Aspartic Acid; ADP-Ribosylation; Genomic Instability; Glutamates; Carrier Proteins; Nuclear Proteins
PubMed: 37676774
DOI: 10.1016/j.celrep.2023.113113 -
ACS Chemical Biology Nov 2021Poly(ADP-ribose) polymerases, PARPs, transfer ADP-ribose onto target proteins from nicotinamide adenine dinucleotide (NAD). Current mass spectrometric analytical methods...
Poly(ADP-ribose) polymerases, PARPs, transfer ADP-ribose onto target proteins from nicotinamide adenine dinucleotide (NAD). Current mass spectrometric analytical methods require proteolysis of target proteins, limiting the study of dynamic ADP-ribosylation on contiguous proteins. Herein, we present a matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) method that facilitates multisite analysis of ADP-ribosylation. We observe divergent ADP-ribosylation dynamics for the catalytic domains of PARPs 14 and 15, with PARP15 modifying more sites on itself (+3-4 ADP-ribose) than the closely related PARP14 protein (+1-2 ADP-ribose)─despite similar numbers of potential modification sites. We identify, for the first time, a minimal peptide fragment (18 amino-acids) that is preferentially modified by PARP14. Finally, we demonstrate through mutagenesis and chemical treatment with hydroxylamine that PARPs 14/15 prefer acidic residues. Our results highlight the utility of MALDI-TOF in the analysis of PARP target modifications and in elucidating the biochemical mechanism governing PARP target selection.
Topics: ADP-Ribosylation; Chromatography, Thin Layer; Poly(ADP-ribose) Polymerases; Protein Domains; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 34647721
DOI: 10.1021/acschembio.1c00542