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SARS-CoV-2 infection of human cells is initiated by the binding of the viral Spike protein to its cell-surface receptor ACE2. We conducted a targeted CRISPRi screen to uncover druggable pathways controlling Spike protein binding to human cells. We found that the protein BRD2 is required for ACE2 transcription in human lung epithelial cells and cardiomyocytes, and BRD2 inhibitors currently evaluated in clinical trials potently block endogenous ACE2 expression and SARS-CoV-2 infection of human cells, including those of human nasal epithelia. Moreover, pharmacological BRD2 inhibition with the drug ABBV-744 inhibited SARS-CoV-2 replication in Syrian hamsters. We also found that BRD2 controls transcription of several other genes induced upon SARS-CoV-2 infection, including the interferon response, which in turn regulates the antiviral response. Together, our results pinpoint BRD2 as a potent and essential regulator of the host response to SARS-CoV-2 infection and highlight the potential of BRD2 as a novel therapeutic target for COVID-19.

Interferon restricts SARS-CoV-2 replication in cell culture, but only a handful of Interferon Stimulated Genes with antiviral activity against SARS-CoV-2 have been identified. Here, we describe a functional CRISPR/Cas9 screen aiming at identifying SARS-CoV-2 restriction factors. We identified DAXX, a scaffold protein residing in PML nuclear bodies known to limit the replication of DNA viruses and retroviruses, as a potent inhibitor of SARS-CoV-2 and SARS-CoV replication in human cells. Basal expression of DAXX was sufficient to limit the replication of SARS-CoV-2, and DAXX over-expression further restricted infection. In contrast with most of its previously described antiviral activities, DAXX-mediated restriction of SARS-CoV-2 was independent of the SUMOylation pathway. SARS-CoV-2 infection triggered the re-localization of DAXX to cytoplasmic sites and promoted its degradation. Mechanistically, this process was mediated by the viral papain-like protease (PLpro) and the proteasome. Together, these results demonstrate that DAXX restricts SARS-CoV-2, which in turn has evolved a mechanism to counteract its action.

International audience; Genetic recombination is a major force driving the evolution of some species of positive sense RNA viruses. Recombination events occur when at least two viruses simultaneously infect the same cell, thereby giving rise to new genomes comprised of genetic sequences originating from the parental genomes. The main mechanism by which recombination occurs involves the viral polymerase that generates a chimera as it switches templates during viral replication. Various experimental systems have alluded to the existence of recombination events that are independent of viral polymerase activity. The origins and the frequency of such events remain to be elucidated to this day. Furthermore, it is not known whether non-replicative recombination yields products that are different from recombinants generated by the viral polymerase. If this is the case, then non-replicative recombination may play a unique role in the evolution of positive sense RNA viruses. Finally, the sparse data available suggest that non-replicative recombination does not necessarily involve only virus-specific sequences. It is thus possible that the non-replicative recombination observed in virus-focused studies may in fact reveal a more generalized mechanism that is non-specific to virus RNAs.

The main aims of the present study were to design a fusion protein of Leishmania major stress-inducible protein 1 (LmSTI1) and Phlebotomus papatasi SP15 (PpSP15), and to express it in the form of alphavirus packaged Self-amplifying mRNA (SAM). Two combinations, PpSP15-LmSTI1 and LmSTI1-PpSP15 fusion forms, were analyzed for folding and minimum free energies of the mRNA. Conformational studies on 3D modeled fusion and native forms were performed, and the Root-Mean-Square-distance (RMSD) of the Cα atomic coordinates were calculated. Antigenicity and stability were predicted using bioinformatics tools. The coding sequences of PpSP15-LmSTI1 fusion, PpSP15, and LmSTI1 were cloned into an alphavirus-based vector and used to produce the SAM constructs. All the subcloned constructs were then subjected to packaging in the form of viral replicon particles (VRPs),and were evaluated for their ability to infect BHK-21 cells and express the recombinant fusion proteins. The in-silico analysis indicated that the PpSP15-LmSTI1 combination could be a promising candidate based on lower folding ΔG of mRNA, higher protein antigenicity and lower instability indexes, and less conformational changes compared to the native proteins and the LmSTI1-PpSP15 fusion form. Packaged SAM encoding fusion and native antigens are used for infection of mammalian cells and for recombinant protein expression. This is the first study on in silico designing and successful packaging of an alphavirus-derived SAM in the form of the VRPs to target leishmaniasis.

Since the identification of the first enteroviruses, the classification and the nomenclature of these viruses were modified several times. Even the base of the classification was changed during the 2000s when genetic criteria superseded the historical serological criteria used to identify enteroviruses. Product of these modifications, the current classification and nomenclature are confusing for students, researchers and practitioners who discover them for the first time; coxsackieviruses A and B, echoviruses and polioviruses are gathered into different species while surprisingly, in view of the etymology, the rhinoviruses now belong the genus Enterovirus. This review aims to summarize the history of the methods and concepts that were used to elaborate the successive classifications and to feature the discoveries that led to their modifications. Mostly slight, sometimes drastic, these modifications underline the history of our knowledge about the enteroviruses and their diversity; indirectly, they highlight our profound ignorance.

A newly described coronavirus named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is the causative agent of coronavirus disease 2019 (COVID-19), has infected over 2.3 million people, led to the death of more than 160,000 individuals and caused worldwide social and economic disruption1,2. There are no antiviral drugs with proven clinical efficacy for the treatment of COVID-19, nor are there any vaccines that prevent infection with SARS-CoV-2, and efforts to develop drugs and vaccines are hampered by the limited knowledge of the molecular details of how SARS-CoV-2 infects cells. Here we cloned, tagged and expressed 26 of the 29 SARS-CoV-2 proteins in human cells and identified the human proteins that physically associated with each of the SARS-CoV-2 proteins using affinity-purification mass spectrometry, identifying 332 high-confidence protein–protein interactions between SARS-CoV-2 and human proteins. Among these, we identify 66 druggable human proteins or host factors targeted by 69 compounds (of which, 29 drugs are approved by the US Food and Drug Administration, 12 are in clinical trials and 28 are preclinical compounds). We screened a subset of these in multiple viral assays and found two sets of pharmacological agents that displayed antiviral activity: inhibitors of mRNA translation and predicted regulators of the sigma-1 and sigma-2 receptors. Further studies of these host-factor-targeting agents, including their combination with drugs that directly target viral enzymes, could lead to a therapeutic regimen to treat COVID-19. A human–SARS-CoV-2 protein interaction map highlights cellular processes that are hijacked by the virus and that can be targeted by existing drugs, including inhibitors of mRNA translation and predicted regulators of the sigma receptors.

International audience; RNA recombination is a major driving force in the evolution and genetic architecture shaping of enteroviruses. In particular, intertypic recombination is implicated in the emergence of most pathogenic circulating vaccine-derived polioviruses, which have caused numerous outbreaks of paralytic poliomyelitis worldwide. Recent experimental studies that relied on recombination cellular systems mimicking natural genetic exchanges between enteroviruses provided new insights into the molecular mechanisms of enterovirus recombination and enabled to define a new model of genetic plasticity for enteroviruses. Homologous intertypic recombinant enteroviruses that were observed in nature would be the final products of a multi-step process, during which precursor nonhomologous recombinant genomes are generated through an initial inter-genomic RNA recombination event and can then evolve into a diversity of fitter homologous recombinant genomes over subsequent intra-genomic rearrangements. Moreover, these experimental studies demonstrated that the enterovirus genome could be defined as a combination of genomic modules that can be preferentially exchanged through recombination, and enabled defining the boundaries of these recombination modules. These results provided the first experimental evidence supporting the theoretical model of enterovirus modular evolution previously elaborated from phylogenetic studies of circulating enterovirus strains. This review summarizes our current knowledge regarding the mechanisms of recombination in enteroviruses and presents a new evolutionary process that may apply to other RNA viruses

International audience; Depuis l’identification des premiers entérovirus, la classification et la nomenclature de ces virus ont été remaniées à de nombreuses reprises. Le fondement même de la classification a été modifié au cours des années 2000 lorsque les critères génétiques ont supplanté les critères sérologiques pour l’identification et la classification des entérovirus. Fruit de ces remaniements, la classification et la nomenclature actuelles sont souvent assez déroutantes pour l’étudiant, le chercheur ou le praticien qui s’intéresse pour la première fois à ces virus ; coxsackievirus A et B côtoient echovirus et poliovirus au sein de différentes espèces tandis que, de façon peu intuitive au regard de l’étymologie, les rhinovirus appartiennent désormais au genre Enterovirus. Cette revue a pour but de présenter un aperçu historique des méthodes et des concepts qui ont été utilisés pour élaborer les classifications successives de ces virus et des découvertes qui ont conduit à les réformer. Souvent subtiles, parfois profondes, les modifications apportées au cours du temps retracent l’histoire de nos connaissances concernant les entérovirus et celle de notre compréhension de la diversité qu’ils présentent ; en creux, elles révèlent surtout notre ignorance.

SARS-CoV-2 infection of human cells is initiated by the binding of the viral Spike protein to its cell-surface receptor ACE2. We conducted a targeted CRISPRi screen to uncover druggable pathways controlling Spike protein binding to human cells. We found that the protein BRD2 is required for ACE2 transcription in human lung epithelial cells and cardiomyocytes, and BRD2 inhibitors currently evaluated in clinical trials potently block endogenous ACE2 expression and SARS-CoV-2 infection of human cells, including those of human nasal epithelia. Moreover, pharmacological BRD2 inhibition with the drug ABBV-744 inhibited SARS-CoV-2 replication in Syrian hamsters. We also found that BRD2 controls transcription of several other genes induced upon SARS-CoV-2 infection, including the interferon response, which in turn regulates the antiviral response. Together, our results pinpoint BRD2 as a potent and essential regulator of the host response to SARS-CoV-2 infection and highlight the potential of BRD2 as a novel therapeutic target for COVID-19., Competing Interests: Competing Interests: JCS, JO, TM and KH are employees and shareholders of Synthego Corporation.