Fixed-dose combination emtricitabine/tenofovir/efavirenz initiated during acute HIV infection; 96-week efficacy and durability

Demonstrate rapid initiation of co-formulated treatment during acute HIV infection (AHI) is feasible and efficacious

Étude de l'activité antivirale des protéines Myxovirus resistance 1 (MX1) contre le virus influenza A

Ma thèse a eu pour but d’élucider le mécanisme d’action des protéines antivirales Myxovirus Resistance protein 1 (MX1), des GTPases induites par l’interféron, contre le virus influenza A (IAV). Les protéines MX1 sont de puissants inhibiteurs de nombreux virus, dont IAV. Certains déterminants intrinsèques de ces protéines étaient connus comme essentiels à l’activité antivirale, telles que la capacité à hydrolyser le GTP ou à multimériser. Mais, malgré leur découverte il y a près de quarante ans, le mode d’action des protéines MX1 reste mal compris.Durant cette thèse, j’ai tout d’abord pu définir un nouvel élément clé de l’activité antivirale des protéines MX1, leur domaine amino (N)-terminal. J’ai pu montrer que les protéines MX1 possèdent une leucine extrêmement conservée dans ce domaine N-terminal, qui, lorsque mutée, abolit l’activité antivirale contre différents virus à ARN, dont IAV. Cette mutation ne perturbe pas l’activité GTPase ou la capacité des protéines MX1 à oligomériser mais dérégule la localisation subcellulaire de protéines MX1 de différentes espèces. Des prédictions structurales obtenues en utilisant AlphaFold montrent que cette leucine est orientée vers l’extérieur de la protéine et pourrait permettre l’interaction avec des partenaires cellulaires ou viraux. Cette étude structure/fonction dévoile de nouveaux éléments dans la caractérisation de ces protéines antivirales et ouvre la voie à de nouvelles stratégies pour comprendre leur mécanisme d’action.Lors de la seconde partie de ma thèse, j’ai observé pour la première fois que les protéines MX1 peuvent inhiber le virus IAV à des stades du cycle de réplication intervenant après la transcription/réplication du génome viral. En effet, nous montrons pour la première fois une inhibition au niveau de l’épissage de transcrits viraux par les protéines MX1, avec un défaut d’expression des ARN épissés du segment M et NS. En présence des protéines MX1, la nucléoprotéine (NP) virale reste dans le noyau des cellules infectées à des temps tardifs post-infection. Nous émettons l’hypothèse que le complexe d’export nucléaire viral ne peut se former dans ces conditions à cause d’un défaut d’expression de la protéine d’export nucléaire virale (NEP), produite à partir d’un ARNm viral épissé. Au-delà de ce blocage observé, dans environ un quart des cellules exprimant la protéine MX1 humaine, NP s’accumule dans des structures périnucléaires denses. Nous montrons que ceci est dû à un défaut de transport cytoplasmique des complexes ribonucléoprotéines viraux exportés du noyau, restant bloqués au niveau d’un compartiment de recyclage d'endocytose (ERC) condensé. L’imagerie en temps réel nous a permis de résoudre temporellement et dynamiquement ce processus. De plus, ces inhibitions tardives du cycle viral affectent plusieurs souches d’IAV et influenza B et sont observés dans plusieurs lingées modèles.Ces nouvelles découvertes dévoilent des aspects inattendus de la restriction antivirale conferrée par les protéines MX1 et nous amènent à repenser la biologie de ces protéines.The goal of my PhD project was to understand the mode of action of the interferon-induced antiviral GTPases Myxovirus Resistance protein 1 (MX1) proteins against influenza A virus (IAV). MX1 proteins are powerful inhibitors of many viruses including IAV. Some intrinsic determinants are known to be essential for the antiviral activity of these proteins, such as the capacity to hydrolyse GTP or to multimerize. But, despite being discovered over forty years ago, their mechanism of action remains elusive.During this thesis, I was first able to define a new key element for the antiviral activity of MX1 proteins, the N-terminal domain, that had, until now, gone unstudied. I showed that MX1 proteins possess a conserved leucine residue in their amino (N)-terminal domain, which, when mutated, abrogates antiviral activity against different RNA viruses, including IAV. This mutation did not affect the GTPase activity or oligomerization status of MX1 proteins, but affected their correct subcellular localization. AlphaFold structural predictions show that this leucine would point away from the core of the protein, suggesting that this residue could be an essential interaction hub for a potential cellular or viral factor. This structure/function study sheds new light of MX1 proteins and characterizes a new intrinsic element necessary for their antiviral activity and opens the door for new strategies to try and understand the detailed mechanism of action of these proteins.In the second part of this thesis, I report for the first time that MX1 proteins are able to inhibit later stages of IAV infection in addition to their previously defined restriction of transcription/replication steps. Indeed, we show the first evidence that IAV spliced RNAs from the M and NS segments are inhibited by MX1 proteins. Paired with the observation of an altered localization of viral nucleoprotein (NP) at late timepoints post-infection, in the nuclei of infected cells, we hypothesize that this is due to the inability of the vRNP nuclear export complex to form. This would be due to the inhibition of nuclear export protein (NEP) production, from a spliced mRNA, in the presence of MX1 proteins. Furthermore, in about a quarter of cells expressing human MX1, NP accumulated into dense structures at the perinuclear region in a subset of cells. This was further shown to be an accumulation of viral ribonucleoprotein trafficking complexes, at a condensed endocytic recycling compartment (ERC). Live imaging allowed us to temporally and dynamically characterize this phenotype. These late stage inhibitions were shown to exist for several IAV strains, for influenza B virus as well as in another cell line, suggesting that they are not artefacts of a particular strain in a particular cell line.These studies unveiled unexpected aspects to MX1 protein inhibition of IAV and will necessitate to rethink past observations through incorporation of these new findings

Fatty acid synthase and stearoyl-CoA desaturase-1 are conserved druggable cofactors of Old World Alphavirus genome replication

International audienc

SARS-CoV-2 Triggers an MDA-5-Dependent Interferon Response Which Is Unable To Control Replication in Lung Epithelial Cells

International audienc

A genome-wide CRISPR/Cas9 knock-out screen identifies the DEAD box RNA helicase DDX42 as a broad antiviral inhibitor

Genome-wide CRISPR/Cas9 knock-out genetic screens are powerful approaches to unravel new regulators of viral infections. With the aim of identifying new cellular inhibitors of HIV-1, we have developed a strategy in which we took advantage of the ability of type 1 interferon (IFN) to potently inhibit HIV-1 infection, in order to create a cellular environment hostile to viral replication. This approach led to the identification of the DEAD-box RNA helicase DDX42 as an intrinsic inhibitor of HIV-1. Depletion of endogenous DDX42 using siRNA or CRISPR/Cas9 knock-out increased HIV-1 infection, both in model cell lines and in physiological targets of HIV-1, primary CD4+ T cells and monocyte-derived macrophages (MDMs), and irrespectively of the IFN treatment. Similarly, the overexpression of a dominant-negative mutant of DDX42 positively impacted HIV-1 infection, whereas wild-type DDX42 overexpression potently inhibited HIV-1 infection. The positive impact of endogenous DDX42 depletion on HIV-1 infection was directly correlated to an increase in viral DNA accumulation. Interestingly, proximity ligation assays showed that DDX42, which can be mainly found in the nucleus but is also present in the cytoplasm, was in the close vicinity of HIV-1 Capsid during infection of primary monocyte-derived macrophages. Moreover, we show that DDX42 is also able to substantially decrease infection with other retroviruses and retrotransposition of long interspersed elements-1 (LINE-1). Finally, we reveal that DDX42 potently inhibits other pathogenic viruses, including Chikungunya virus and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)

Mitochondrial morphodynamics alteration induced by influenza virus infection as a new antiviral strategy.

Influenza virus infections are major public health threats due to their high rates of morbidity and mortality. Upon influenza virus entry, host cells experience modifications of endomembranes, including those used for virus trafficking and replication. Here we report that influenza virus infection modifies mitochondrial morphodynamics by promoting mitochondria elongation and altering endoplasmic reticulum-mitochondria tethering in host cells. Expression of the viral RNA recapitulates these modifications inside cells. Virus induced mitochondria hyper-elongation was promoted by fission associated protein DRP1 relocalization to the cytosol, enhancing a pro-fusion status. We show that altering mitochondrial hyper-fusion with Mito-C, a novel pro-fission compound, not only restores mitochondrial morphodynamics and endoplasmic reticulum-mitochondria contact sites but also dramatically reduces influenza replication. Finally, we demonstrate that the observed Mito-C antiviral property is directly connected with the innate immunity signaling RIG-I complex at mitochondria. Our data highlight the importance of a functional interchange between mitochondrial morphodynamics and innate immunity machineries in the context of influenza viral infection

Assessing recovery and functioning in former problem drinkers at different stages of their recovery journeys

Introduction and Aims. Many studies which assess functioning in recovering problem drinkers are limited to early recovery within inpatient or detoxification settings, or focus on relapse rates and treatment outcomes. This study assesses how functioning varies according to recovery stage and abstinence duration. Design and Methods. Fifty-three recovering problem drinkers participated from mutual aid groups or snowball recruitment. Cross-sectional interviewer-administered structured questionnaires assessed quality of life (QoL), self-esteem, self-efficacy, psychological and physical health. Participants could also self-complete the questionnaire. Results. Those in 'stable recovery' (5 or more years into recovery, n = 18) reported higher ratings of: three aspects of QoL-social relationships, psychological health, environment, as well as self-esteem (P < 0.05 for all variables) than those in 'early' (up to 5 years into recovery, n = 35). Depression was lower in 'stable recovery' (P = 0.027). Those in 'stable recovery' were more likely to live in their own home without professional support (P = 0.010) and have partners who had never been problem drinkers (P = 0.024). Overall, the continuous scores of many functioning variables correlated with abstinence duration indicating a continuous gain in functioning. Discussion and Conclusions. Although limited by sampling considerations, this paper shows a gradual growth in functioning over a prolonged recovery process, and provides positive findings that those in recovery may expect to experience improvements in many areas of life as abstinence duration increases. For two aspects of QoL-environment and social relationships-functioning reaches a level above population norms offering hope of moving to a functioning level beyond the pre-morbid state