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The FEBS Journal Dec 2022ADP-ribosylation, a modification of proteins, nucleic acids, and metabolites, confers broad functions, including roles in stress responses elicited, for example, by DNA...
ADP-ribosylation, a modification of proteins, nucleic acids, and metabolites, confers broad functions, including roles in stress responses elicited, for example, by DNA damage and viral infection and is involved in intra- and extracellular signaling, chromatin and transcriptional regulation, protein biosynthesis, and cell death. ADP-ribosylation is catalyzed by ADP-ribosyltransferases (ARTs), which transfer ADP-ribose from NAD onto substrates. The modification, which occurs as mono- or poly-ADP-ribosylation, is reversible due to the action of different ADP-ribosylhydrolases. Importantly, inhibitors of ARTs are approved or are being developed for clinical use. Moreover, ADP-ribosylhydrolases are being assessed as therapeutic targets, foremost as antiviral drugs and for oncological indications. Due to the development of novel reagents and major technological advances that allow the study of ADP-ribosylation in unprecedented detail, an increasing number of cellular processes and pathways are being identified that are regulated by ADP-ribosylation. In addition, characterization of biochemical and structural aspects of the ARTs and their catalytic activities have expanded our understanding of this protein family. This increased knowledge requires that a common nomenclature be used to describe the relevant enzymes. Therefore, in this viewpoint, we propose an updated and broadly supported nomenclature for mammalian ARTs that will facilitate future discussions when addressing the biochemistry and biology of ADP-ribosylation. This is combined with a brief description of the main functions of mammalian ARTs to illustrate the increasing diversity of mono- and poly-ADP-ribose mediated cellular processes.
Topics: ADP Ribose Transferases; Protein Biosynthesis; Adenosine Diphosphate Ribose; Adenosine Diphosphate
PubMed: 34323016
DOI: 10.1111/febs.16142 -
Cellular and Molecular Life Sciences :... Feb 2023Replication of viruses requires interaction with host cell factors and repression of innate immunity. Recent findings suggest that a subset of intracellular...
Replication of viruses requires interaction with host cell factors and repression of innate immunity. Recent findings suggest that a subset of intracellular mono-ADP-ribosylating PARPs, which are induced by type I interferons, possess antiviral activity. Moreover, certain RNA viruses, including Chikungunya virus (CHIKV), encode mono-ADP-ribosylhydrolases. Together, this suggests a role for mono-ADP-ribosylation (MARylation) in host-virus conflicts, but the relevant substrates have not been identified. We addressed which PARP restricts CHIKV replication and identified PARP10 and PARP12. For PARP10, this restriction was dependent on catalytic activity. Replication requires processing of the non-structural polyprotein nsP1-4 by the protease located in nsP2 and the assembly of the four individual nsP1-nsP4 into a functional replication complex. PARP10 and PARP12 inhibited the production of nsP3, indicating a defect in polyprotein processing. The nsP3 protein encodes a macrodomain with de-MARylation activity, which is essential for replication. In support for MARylation affecting polyprotein processing, de-MARylation defective CHIKV replicons revealed reduced production of nsP2 and nsP3. We hypothesized that MARylation regulates the proteolytic function of nsP2. Indeed, we found that nsP2 is MARylated by PARP10 and, as a consequence, its proteolytic activity was inhibited. NsP3-dependent de-MARylation reactivated the protease. Hence, we propose that PARP10-mediated MARylation prevents polyprotein processing and consequently virus replication. Together, our findings provide a mechanistic explanation for the role of the viral MAR hydrolase in CHIKV replication.
Topics: ADP-Ribosylation; Chikungunya virus; Peptide Hydrolases; Polyproteins; Viral Nonstructural Proteins; Virus Replication; Poly(ADP-ribose) Polymerases
PubMed: 36840772
DOI: 10.1007/s00018-023-04717-8 -
Nucleic Acids Research Oct 2021ADP-ribosylation is a modification that targets a variety of macromolecules and regulates a diverse array of important cellular processes. ADP-ribosylation is catalysed...
ADP-ribosylation is a modification that targets a variety of macromolecules and regulates a diverse array of important cellular processes. ADP-ribosylation is catalysed by ADP-ribosyltransferases and reversed by ADP-ribosylhydrolases. Recently, an ADP-ribosyltransferase toxin termed 'DarT' from bacteria, which is distantly related to human PARPs, was shown to modify thymidine in single-stranded DNA in a sequence specific manner. The antitoxin of DarT is the macrodomain containing ADP-ribosylhydrolase DarG, which shares striking structural homology with the human ADP-ribosylhydrolase TARG1. Here, we show that TARG1, like DarG, can reverse thymidine-linked DNA ADP-ribosylation. We find that TARG1-deficient human cells are extremely sensitive to DNA ADP-ribosylation. Furthermore, we also demonstrate the first detection of reversible ADP-ribosylation on genomic DNA in vivo from human cells. Collectively, our results elucidate the impact of DNA ADP-ribosylation in human cells and provides a molecular toolkit for future studies into this largely unknown facet of ADP-ribosylation.
Topics: Adenosine Diphosphate Ribose; Bacterial Toxins; Cell Line; DNA; DNA Repair; DNA Replication; DNA, Single-Stranded; Humans; Thiolester Hydrolases; Thymidine
PubMed: 34508355
DOI: 10.1093/nar/gkab771 -
Proceedings of the National Academy of... Aug 2023PARP1 (poly-ADP ribose polymerase 1) is recruited and activated by DNA strand breaks, catalyzing the generation of poly-ADP-ribose (PAR) chains from NAD+. PAR relaxes...
PARP1 (poly-ADP ribose polymerase 1) is recruited and activated by DNA strand breaks, catalyzing the generation of poly-ADP-ribose (PAR) chains from NAD+. PAR relaxes chromatin and recruits other DNA repair factors, including XRCC1 and DNA Ligase 3, to maintain genomic stability. Here we show that, in contrast to the normal development of Parp1-null mice, heterozygous expression of catalytically inactive Parp1 (E988A, ) acts in a dominant-negative manner to disrupt murine embryogenesis. As such, all the surviving F1 mice are chimeras with mixed (neoR retention) cells that act similarly to . Pure F2 embryos were found at Mendelian ratios at the E3.5 blastocyst stage but died before E9.5. Compared to cells, genotype and expression-validated pure cells retain significant ADP-ribosylation and PARylation activities but accumulate markedly higher levels of sister chromatid exchange and mitotic bridges. Despite proficiency for homologous recombination and nonhomologous end-joining measured by reporter assays and supported by normal lymphocyte and germ cell development, cells are hypersensitive to base damages, radiation, and Topoisomerase I and II inhibition. The sensitivity of cells to base damages and Topo inhibitors exceed controls. The findings show that the enzymatically inactive PARP1 dominant negatively blocks DNA repair in selective pathways beyond wild-type PARP1 and establishes a crucial physiological difference between PARP1 inactivation vs. deletion. As a result, the expression of enzymatically inactive PARP1 from one allele is sufficient to abrogate murine embryonic development, providing a mechanism for the on-target side effect of PARP inhibitors used for cancer therapy.
Topics: Female; Pregnancy; Animals; Mice; Causality; Alleles; Genotype; Genomic Instability; ADP-Ribosylation
PubMed: 37487079
DOI: 10.1073/pnas.2301972120 -
Journal of Zhejiang University.... Jan 2021Adenosine diphosphate (ADP)-ribosylation is a unique post-translational modification that regulates many biological processes, such as DNA damage repair. During DNA... (Review)
Review
Adenosine diphosphate (ADP)-ribosylation is a unique post-translational modification that regulates many biological processes, such as DNA damage repair. During DNA repair, ADP-ribosylation needs to be reversed by ADP-ribosylhydrolases. A group of ADP-ribosylhydrolases have a catalytic domain, namely the macrodomain, which is conserved in evolution from prokaryotes to humans. Not all macrodomains remove ADP-ribosylation. One set of macrodomains loses enzymatic activity and only binds to ADP-ribose (ADPR). Here, we summarize the biological functions of these macrodomains in DNA damage repair and compare the structure of enzymatically active and inactive macrodomains. Moreover, small molecular inhibitors have been developed that target macrodomains to suppress DNA damage repair and tumor growth. Macrodomain proteins are also expressed in pathogens, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, these domains may not be directly involved in DNA damage repair in the hosts or pathogens. Instead, they play key roles in pathogen replication. Thus, by targeting macrodomains it may be possible to treat pathogen-induced diseases, such as coronavirus disease 2019 (COVID-19).
Topics: ADP-Ribosylation; COVID-19; DNA Repair; Evolution, Molecular; Humans; Models, Biological; Models, Molecular; N-Glycosyl Hydrolases; Poly(ADP-ribose) Polymerases; Protein Domains; SARS-CoV-2
PubMed: 33448184
DOI: 10.1631/jzus.B2000319 -
Nucleic Acids Research Apr 2021The functionality of DNA, RNA and proteins is altered dynamically in response to physiological and pathological cues, partly achieved by their modification. While the... (Review)
Review
The functionality of DNA, RNA and proteins is altered dynamically in response to physiological and pathological cues, partly achieved by their modification. While the modification of proteins with ADP-ribose has been well studied, nucleic acids were only recently identified as substrates for ADP-ribosylation by mammalian enzymes. RNA and DNA can be ADP-ribosylated by specific ADP-ribosyltransferases such as PARP1-3, PARP10 and tRNA 2'-phosphotransferase (TRPT1). Evidence suggests that these enzymes display different preferences towards different oligonucleotides. These reactions are reversed by ADP-ribosylhydrolases of the macrodomain and ARH families, such as MACROD1, TARG1, PARG, ARH1 and ARH3. Most findings derive from in vitro experiments using recombinant components, leaving the relevance of this modification in cells unclear. In this Survey and Summary, we provide an overview of the enzymes that ADP-ribosylate nucleic acids, the reversing hydrolases, and the substrates' requirements. Drawing on data available for other organisms, such as pierisin1 from cabbage butterflies and the bacterial toxin-antitoxin system DarT-DarG, we discuss possible functions for nucleic acid ADP-ribosylation in mammals. Hypothesized roles for nucleic acid ADP-ribosylation include functions in DNA damage repair, in antiviral immunity or as non-conventional RNA cap. Lastly, we assess various methods potentially suitable for future studies of nucleic acid ADP-ribosylation.
Topics: ADP Ribose Transferases; ADP-Ribosylation; Animals; Bacteria; DNA; Humans; RNA
PubMed: 33693930
DOI: 10.1093/nar/gkab136 -
SLAS Discovery : Advancing Life... Jan 2021ADP-ribosylation is a post-translational modification involved in the regulation of many vital cellular processes. This posttranslational modification is carried out by...
ADP-ribosylation is a post-translational modification involved in the regulation of many vital cellular processes. This posttranslational modification is carried out by ADP-ribosyltransferases converting β-NAD into nicotinamide and a protein-linked ADP-ribosyl group or a chain of PAR. The reverse reaction, release of ADP-ribose from the acceptor molecule, is catalyzed by ADP-ribosylhydrolases. Several hydrolases contain a macrodomain fold, and activities of human macrodomain protein modules vary from reading or erasing mono- and poly-ADP-ribosylation. Macrodomains have been linked to diseases such as cancer, making them potential drug targets. Discovery of inhibitors requires robust biochemical tools mostly lacking for hydrolases, and here we describe an inhibitor screening assay against mono-ADP-ribosylhydrolyzing enzymes. The activity-based assay uses an α-NAD, anomer of β-NAD, which is accepted as a substrate by MacroD1, MacroD2, and ARH3 due to its resemblance to the protein-linked ADP-ribose. The amount of α-NAD present after hydrolysis is measured by chemically converting it on a microtiter plate to a fluorescent compound. We optimized the assay for MacroD2 and performed a proof-of-concept compound screening. Three compounds were identified as screening hits with micromolar potency. However, further characterization of the compounds identified them as protein destabilizers, excluding further follow-up studies. Validation and screening demonstrated the usability of the in vitro assay for MacroD2, and we also demonstrate the applicability of the assay as a tool for other human ADP-ribosylhydrolases.
Topics: ADP-Ribosylation; Biological Assay; Carboxylic Ester Hydrolases; Enzyme Activation; Humans; NAD; Protein Processing, Post-Translational
PubMed: 32527186
DOI: 10.1177/2472555220928911 -
Experimental & Molecular Medicine Dec 2022PARPs play fundamental roles in multiple DNA damage recognition and repair pathways. Persistent nuclear PARP activation causes cellular NAD depletion and exacerbates...
PARPs play fundamental roles in multiple DNA damage recognition and repair pathways. Persistent nuclear PARP activation causes cellular NAD depletion and exacerbates cellular aging. However, very little is known about mitochondrial PARP (mtPARP) and poly ADP-ribosylation (PARylation). The existence of mtPARP is controversial, and the biological roles of mtPARP-induced mitochondrial PARylation are unclear. Here, we demonstrate the presence of PARP1 and PARylation in purified mitochondria. The addition of the PARP1 substrate NAD to isolated mitochondria induced PARylation, which was suppressed by treatment with the inhibitor olaparib. Mitochondrial PARylation was also evaluated by enzymatic labeling of terminal ADP-ribose (ELTA). To further confirm the presence of mtPARP1, we evaluated mitochondrial nucleoid PARylation by ADP ribose-chromatin affinity purification (ADPr-ChAP) and PARP1 chromatin immunoprecipitation (ChIP). We observed that NAD stimulated PARylation and TFAM occupancy on the mtDNA regulatory region D-loop, inducing mtDNA transcription. These findings suggest that PARP1 is integrally involved in mitochondrial PARylation and that NAD-dependent mtPARP1 activity contributes to mtDNA transcriptional regulation.
Topics: Poly ADP Ribosylation; NAD; Poly(ADP-ribose) Polymerase Inhibitors; Mitochondria; DNA, Mitochondrial
PubMed: 36473936
DOI: 10.1038/s12276-022-00894-x -
Viruses Mar 2021The poly-adenosine diphosphate (ADP)-ribose polymerases (PARPs) are responsible for ADP-ribosylation, a reversible post-translational modification involved in many... (Review)
Review
The poly-adenosine diphosphate (ADP)-ribose polymerases (PARPs) are responsible for ADP-ribosylation, a reversible post-translational modification involved in many cellular processes including DNA damage repair, chromatin remodeling, regulation of translation and cell death. In addition to these physiological functions, recent studies have highlighted the role of PARPs in host defenses against viruses, either by direct antiviral activity, targeting certain steps of virus replication cycle, or indirect antiviral activity, via modulation of the innate immune response. This review focuses on the antiviral activity of PARPs, as well as strategies developed by viruses to escape their action.
Topics: ADP-Ribosylation; Cell Death; Host Microbial Interactions; Humans; Immunity, Innate; Immunomodulation; Poly(ADP-ribose) Polymerases; Protein Processing, Post-Translational; Viruses
PubMed: 33808354
DOI: 10.3390/v13040582 -
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