ADP-Ribosylation and Antiviral Resistance in Plants.

ADP-ribosylation (ADPRylation) is a versatile posttranslational modification in eukaryotic cells which is involved in the regulation of a wide range of key biological processes, including DNA repair, cell signalling, programmed cell death, growth and development and responses to biotic and abiotic stresses. Members of the poly(ADP-ribosyl) polymerase (PARP) family play a central role in the process of ADPRylation. Protein targets can be modified by adding either a single ADP-ribose moiety (mono(ADP-ribosyl)ation; MARylation), which is catalysed by mono(ADP-ribosyl) transferases (MARTs or PARP "monoenzymes"), or targets may be decorated with chains of multiple ADP-ribose moieties (PARylation), via the activities of PARP "polyenzymes". Studies have revealed crosstalk between PARylation (and to a lesser extent, MARylation) processes in plants and plant-virus interactions, suggesting that these tight links may represent a novel factor regulating plant antiviral immunity. From this perspective, we go through the literature linking PARylation-associated processes with other plant regulation pathways controlling virus resistance. Once unraveled, these links may serve as the basis of innovative strategies to improve crop resistance to viruses under challenging environmental conditions which could mitigate yield losses.

Can molecular mimicry explain the cytokine storm of SARS-CoV-2?: An in silico approach.

PARP14 and PARP9 play a key role in macrophage immune regulation. SARS-CoV-2 is an emerging viral disease that triggers hyper-inflammation known as a cytokine storm. In this study, using in silico tools, we hypothesize about the immunological phenomena of molecular mimicry between SARS-CoV-2 Nsp3 and the human PARP14 and PARP9. The results showed an epitope of SARS-CoV-2 Nsp3 protein that contains consensus sequences for both human PARP14 and PARP9 that are antigens for MHC Classes 1 and 2, which can potentially induce an immune response against human PARP14 and PARP9; while its depletion causes a hyper-inflammatory state in SARS-CoV-2 patients.

When PARPs Meet Antiviral Innate Immunity.

The poly(ADP-ribose) polymerases (PARPs) family contains 17 members in humans, sharing a PARP domain to transfer ADP-ribose groups to target proteins to trigger ADP-ribosylation. The roles of PARPs have evolved from DNA damage repair to diverse biological processes, such as gene transcription, cellular stress response, etc. Recently, seminal studies have demonstrated the critical roles of PAPRs in antiviral innate immunity. PARPs catalyze ADP-ribosylation, a fundamental post-translational modification, using NAD+ as a substrate. ADP-ribosylation can occur either as mono- or poly-(ADP-ribosyl)ation, which is initially linked to DNA damage repair, as exemplified by PARP1. Recent advances in host antiviral immunity demonstrated that several PARPs, such as PARP9, 11, 12, 13, 14, etc., have broad-spectrum antiviral activities that are independent of their ADP-ribosylation.

Viral macrodomains: a structural and evolutionary assessment of the pharmacological potential.

Viral macrodomains possess the ability to counteract host ADP-ribosylation, a post-translational modification implicated in the creation of an antiviral environment via immune response regulation. This brought them into focus as promising therapeutic targets, albeit the close homology to some of the human macrodomains raised concerns regarding potential cross-reactivity and adverse effects for the host. Here, we evaluate the structure and function of the macrodomain of SARS-CoV-2, the causative agent of COVID-19. We show that it can antagonize ADP-ribosylation by PARP14, a cellular (ADP-ribosyl)transferase necessary for the restriction of coronaviral infections. Furthermore, our structural studies together with ligand modelling revealed the structural basis for poly(ADP-ribose) binding and hydrolysis, an emerging new aspect of viral macrodomain biology. These new insights were used in an extensive evolutionary analysis aimed at evaluating the druggability of viral macrodomains not only from the Coronaviridae but also Togaviridae and Iridoviridae genera (causing diseases such as Chikungunya and infectious spleen and kidney necrosis virus disease, respectively). We found that they contain conserved features, distinct from their human counterparts, which may be exploited during drug design.

Clinical PARP inhibitors do not abrogate PARP1 exchange at DNA damage sites in vivo.

DNA breaks recruit and activate PARP1/2, which deposit poly-ADP-ribose (PAR) to recruit XRCC1-Ligase3 and other repair factors to promote DNA repair. Clinical PARP inhibitors (PARPi) extend the lifetime of damage-induced PARP1/2 foci, referred to as 'trapping'. To understand the molecular nature of 'trapping' in cells, we employed quantitative live-cell imaging and fluorescence recovery after photo-bleaching. Unexpectedly, we found that PARP1 exchanges rapidly at DNA damage sites even in the presence of clinical PARPi, suggesting the persistent foci are not caused by physical stalling. Loss of Xrcc1, a major downstream effector of PAR, also caused persistent PARP1 foci without affecting PARP1 exchange. Thus, we propose that the persistent PARP1 foci are formed by different PARP1 molecules that are continuously recruited to and exchanging at DNA lesions due to attenuated XRCC1-LIG3 recruitment and delayed DNA repair. Moreover, mutation analyses of the NAD+ interacting residues of PARP1 showed that PARP1 can be physically trapped at DNA damage sites, and identified H862 as a potential regulator for PARP1 exchange. PARP1-H862D, but not PARylation-deficient PARP1-E988K, formed stable PARP1 foci upon activation. Together, these findings uncovered the nature of persistent PARP1 foci and identified NAD+ interacting residues involved in the PARP1 exchange.

Murine Coronavirus Infection Activates the Aryl Hydrocarbon Receptor in an Indoleamine 2,3-Dioxygenase-Independent Manner, Contributing to Cytokine Modulation and Proviral TCDD-Inducible-PARP Expression.

The aryl hydrocarbon receptor (AhR) is a cytoplasmic receptor/transcription factor that modulates several cellular and immunological processes following activation by pathogen-associated stimuli, though its role during virus infection is largely unknown. Here, we show that AhR is activated in cells infected with mouse hepatitis virus (MHV), a coronavirus (CoV), and contributes to the upregulation of downstream effector TCDD-inducible poly(ADP-ribose) polymerase (TiPARP) during infection. Knockdown of TiPARP reduced viral replication and increased interferon expression, suggesting that TiPARP functions in a proviral manner during MHV infection. We also show that MHV replication induced the expression of other genes known to be downstream of AhR in macrophages and dendritic cells and in livers of infected mice. Further, we found that chemically inhibiting or activating AhR reciprocally modulated the expression levels of cytokines induced by infection, specifically, interleukin 1ß (IL-1ß), IL-10, and tumor necrosis factor alpha (TNF-α), consistent with a role for AhR activation in the host response to MHV infection. Furthermore, while indoleamine 2,3-dioxygenase (IDO1) drives AhR activation in other settings, MHV infection induced equal expression of downstream genes in wild-type (WT) and IDO1-/- macrophages, suggesting an alternative pathway of AhR activation. In summary, we show that coronaviruses elicit AhR activation by an IDO1-independent pathway, contributing to upregulation of downstream effectors, including the proviral factor TiPARP, and to modulation of cytokine gene expression, and we identify a previously unappreciated role for AhR signaling in CoV pathogenesis.IMPORTANCE Coronaviruses are a family of positive-sense RNA viruses with human and agricultural significance. Characterizing the mechanisms by which coronavirus infection dictates pathogenesis or counters the host immune response would provide targets for the development of therapeutics. Here, we show that the aryl hydrocarbon receptor (AhR) is activated in cells infected with a prototypic coronavirus, mouse hepatitis virus (MHV), resulting in the expression of several effector genes. AhR is important for modulation of the host immune response to MHV and plays a role in the expression of TiPARP, which we show is required for maximal viral replication. Taken together, our findings highlight a previously unidentified role for AhR in regulating coronavirus replication and the immune response to the virus.