Upregulation of the Nrf2 antioxidant pathway characterizes the transition from productive to latent infection in CD4+ T cells

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CAGE-Seq Reveals that HIV-1 Latent Infection Does Not Trigger Unique Cellular Responses in a Jurkat T Cell Model

The cure for HIV-1 is currently stalled by our inability to specifically identify and target latently infected cells. HIV-1 viral RNA/DNA or viral proteins are recognized by cellular mechanisms and induce interferon responses in virus-producing cells, but changes in latently infected cells remain unknown. HIVGKO contains a green fluorescent protein (GFP) reporter under the HIV-1 promoter and a monomeric Kusabira orange 2 (mKO2) reporter under the internal elongation factor alpha (EF1α) promoter. This viral construct enables direct identification of both productively and latently HIV-1-infected cells. In this study, we aim to identify specific cellular transcriptional responses triggered by HIV-1 entry and integration using cap analysis of gene expression (CAGE). We deep sequenced CAGE tags in non-infected and latently and productively infected cells and compared their differentially expressed transcription start site (TSS) profiles. Virus-producing cells had differentially expressed TSSs related to T-cell activation and apoptosis compared to those of non-infected cells or latently infected cells. Surprisingly, latently infected cells had only 33 differentially expressed TSSs compared to those of non-infected cells. Among these, SPP1 and APOE were downregulated in latently infected cells. SPP1 or APOE knockdown in Jurkat T cells increased susceptibility to HIVGKO infection, suggesting that they have antiviral properties. Components of the phosphatidylinositol 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) pathway, MLST8, 4EBP, and RPS6, were significant TSSs in productively infected cells, and S6 kinase (S6K) phosphorylation was increased compared to that in latently infected cells, suggesting that mTOR pathway activity plays a role in establishing the latent reservoir. These findings indicate that HIV-1 entry and integration do not trigger unique transcriptional responses when infection becomes latent

Assisted evolution enables HIV-1 to overcome a high trim5α-imposed genetic barrier to rhesus macaque tropism

Diversification of antiretroviral factors during host evolution has erected formidable barriers to cross-species retrovirus transmission. This phenomenon likely protects humans from infection by many modern retroviruses, but it has also impaired the development of primate models of HIV-1 infection. Indeed, rhesus macaques are resistant to HIV-1, in part due to restriction imposed by the TRIM5α protein (rhTRIM5α). Initially, we attempted to derive rhTRIM5α-resistant HIV-1 strains using two strategies. First, HIV-1 was passaged in engineered human cells expressing rhTRIM5α. Second, a library of randomly mutagenized capsid protein (CA) sequences was screened for mutations that reduced rhTRIM5α sensitivity. Both approaches identified several individual mutations in CA that reduced rhTRIM5α sensitivity. However, neither approach yielded mutants that were fully resistant, perhaps because the locations of the mutations suggested that TRIM5α recognizes multiple determinants on the capsid surface. Moreover, even though additive effects of various CA mutations on HIV-1 resistance to rhTRIM5α were observed, combinations that gave full resistance were highly detrimental to fitness. Therefore, we employed an 'assisted evolution' approach in which individual CA mutations that reduced rhTRIM5α sensitivity without fitness penalties were randomly assorted in a library of viral clones containing synthetic CA sequences. Subsequent passage of the viral library in rhTRIM5α-expressing cells resulted in the selection of individual viral species that were fully fit and resistant to rhTRIM5α. These viruses encoded combinations of five mutations in CA that conferred complete or near complete resistance to the disruptive effects of rhTRIM5α on incoming viral cores, by abolishing recognition of the viral capsid. Importantly, HIV-1 variants encoding these CA substitutions and SIVmac239 Vif replicated efficiently in primary rhesus macaque lymphocytes. These findings demonstrate that rhTRIM5α is difficult to but not impossible to evade, and doing so should facilitate the development of primate models of HIV-1 infection

Il ruolo del recettore dei glucocorticoidi del NAcc nei comportamenti legati allo stress. Dissezione genetica via trasduzione virale.

L'inattivazione del gene del recettore dei glucocorticoidi nei neuroni dopaminocettivi (topi GRD1aCre) incide profondamente su alcuni comportamenti legati allo stress, tra cui le risposte comportamentali a psicostimolanti e sul ritiro sociale acquisito dopo uno stress sociale ripetute. A livello fisiologico, la mutazione del GR in neuroni postsinaptici riduce fortemente l'attività elettrica dei neuroni dopaminergici presinaptici della VTA, mostrando un ruolo del GR nella modulazione del feedback esercitata dai neuroni dopaminocettivi sui neuroni DA. Nei topi GRD1aCre, il gene GR è ricombinato in varie strutture tra cui il NACC, striato dorsale, l'amigdala e gli strati basali della corteccia, tra cui il NACC e mPFC sono noti per controllare l'attività dei neuroni DA della VTA. E 'quindi essenziale perfezionare l'identificazione delle strutture cerebrali pertinenti.Glucocorticoid Receptor gene inactivation in dopaminoceptive neurons (GRD1aCre mice) deeply affects some stress-related behaviors, including behavioral responses to psychostimulants and social withdrawal acquired after repeated social stress. At the physiological level, the mutation of GR in post-synaptic neurons deeply reduces the electrical activity of presynaptic DA neurons of the VTA, showing a role for GR in the modulation of the feedback exerted by dopaminoceptive neurons on DA neurons. In GRD1aCre mice, GR gene is recombined in several structures including the NAcc, the dorsal striatum, the amygdala and the basal layers of the cortex, among which the NAcc and the mPFC are known to control VTA DA neurons activity. It is therefore essential to refine the identification of the relevant brain structures. My objective was to identify whether the GR in the NAcc is important for the observed phenotypes. We therefore specifically inactivated GR, by stereotactic injection of AAV2 expressing the Cre recombinase and the GFP in GRloxP/loxP mice. Control animals were injecting in parallel with an AAV2 expressing only the GFP. We performed locomotor sensitization to cocaine in mutant and control animals. We observed a marked sensitization but no differences between genotypes

Social rank, dopaminergic system and stress response

L'organisation hiérarchique est la marque de nombreux vertébrés. Des associations entre rang social et état de santé sont signalées chez de nombreuses espèces incluant l’humain. Cependant, les mécanismes impliqués sont mal compris. La réponse au stress, spécifique au rang social chez de nombreux animaux, intervient dans de nombreuses pathologies psychiatriques. Ceci suggère un rôle de la réponse au stress dans le processus liant statut social et comportement. Mon travail de thèse vise à mieux comprendre les associations qui existent entre le rang social et le comportement individuel chez la souris. Mon travail vise à clarifier la direction de cette association en observant les conséquences du rang social sur le comportement et la vulnérabilité aux maladies psychiatriques, et réciproquement, en identifiant des marqueurs individuels précoces pouvant contribuer à façonner le destin social. Au niveau physiologique, je me concentre sur la réponse au stress et ses conséquences sur le système dopaminergique (DA), en tant que médiateur potentiel des phénotypes sociaux. Pour répondre à ces questions, j'ai évalué le rang social de mâles adultes C57BL/6 via des tests compétitifs. J'ai ensuite testé ces animaux pour l'anxiété, la sociabilité et les compétences cognitives, avant et après établissement hiérarchique. Nous avons enregistré l'activité électrophysiologique de cellules DA dans la région tégmentale ventrale. J'ai quantifié la libération de DA dans la voie mésocorticolimbique et testé les souris pour la dépression et la dépendance. Enfin, j’ai évalué les conséquences d’une inactivation du récepteur des glucocorticoïdes dans les cellules DAceptives pour la dominance sociale.Hierarchical organization is the hallmark of many social vertebrates. Associations between social rank and health are reported within many species, including humans. Yet, the mechanisms underlying these associations are poorly understood. The stress response, specific to the social rank in many animals, is also involved in many psychiatric pathologies. This suggests a role of stress response in the process linking social status and behavior. My PhD work aims to better understand the associations that exist between social rank and individual behavior in tetrad housed mice. My work also seeks to clarify the direction of this association by observing the consequences of social positions on behavior and vulnerability to psychiatric illnesses, and reciprocally, by identifying early individual markers that can participate in shaping social fate. At the physiological level, I am focusing on the stress response and its consequences on the dopamine (DA) system, as potential mediator for such social-related phenotypes. To address these questions, I first assessed adult male wild-type mice for social rank with precedence, territorial and competition tests. I then tested animals of different social ranks for behaviors such as anxiety, sociability and cognitive skills either before or after hierarchy establishment. We recorded the electrophysiological activity of DA cells in the ventral tegmental area. I quantified the DA release throughout the mesocorticolimbic pathway and assessed mice for depressive-like behaviors and addiction. Finally, I addressed the role of the glucocorticoids by assessing mutated mice lacking glucocorticoids receptor in DAceptive cells for social dominance

Rang social, système dopaminergique et réponse au stress

Hierarchical organization is the hallmark of many social vertebrates. Associations between social rank and health are reported within many species, including humans. Yet, the mechanisms underlying these associations are poorly understood. The stress response, specific to the social rank in many animals, is also involved in many psychiatric pathologies. This suggests a role of stress response in the process linking social status and behavior. My PhD work aims to better understand the associations that exist between social rank and individual behavior in tetrad housed mice. My work also seeks to clarify the direction of this association by observing the consequences of social positions on behavior and vulnerability to psychiatric illnesses, and reciprocally, by identifying early individual markers that can participate in shaping social fate. At the physiological level, I am focusing on the stress response and its consequences on the dopamine (DA) system, as potential mediator for such social-related phenotypes. To address these questions, I first assessed adult male wild-type mice for social rank with precedence, territorial and competition tests. I then tested animals of different social ranks for behaviors such as anxiety, sociability and cognitive skills either before or after hierarchy establishment. We recorded the electrophysiological activity of DA cells in the ventral tegmental area. I quantified the DA release throughout the mesocorticolimbic pathway and assessed mice for depressive-like behaviors and addiction. Finally, I addressed the role of the glucocorticoids by assessing mutated mice lacking glucocorticoids receptor in DAceptive cells for social dominance.L'organisation hiérarchique est la marque de nombreux vertébrés. Des associations entre rang social et état de santé sont signalées chez de nombreuses espèces incluant l’humain. Cependant, les mécanismes impliqués sont mal compris. La réponse au stress, spécifique au rang social chez de nombreux animaux, intervient dans de nombreuses pathologies psychiatriques. Ceci suggère un rôle de la réponse au stress dans le processus liant statut social et comportement. Mon travail de thèse vise à mieux comprendre les associations qui existent entre le rang social et le comportement individuel chez la souris. Mon travail vise à clarifier la direction de cette association en observant les conséquences du rang social sur le comportement et la vulnérabilité aux maladies psychiatriques, et réciproquement, en identifiant des marqueurs individuels précoces pouvant contribuer à façonner le destin social. Au niveau physiologique, je me concentre sur la réponse au stress et ses conséquences sur le système dopaminergique (DA), en tant que médiateur potentiel des phénotypes sociaux. Pour répondre à ces questions, j'ai évalué le rang social de mâles adultes C57BL/6 via des tests compétitifs. J'ai ensuite testé ces animaux pour l'anxiété, la sociabilité et les compétences cognitives, avant et après établissement hiérarchique. Nous avons enregistré l'activité électrophysiologique de cellules DA dans la région tégmentale ventrale. J'ai quantifié la libération de DA dans la voie mésocorticolimbique et testé les souris pour la dépression et la dépendance. Enfin, j’ai évalué les conséquences d’une inactivation du récepteur des glucocorticoïdes dans les cellules DAceptives pour la dominance sociale

Contribution de TRIM5a humain dans le contrôle de l'infection par le virus de l'immunodéficience humaine de type 1

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Understanding HIV Latency: The Road to an HIV Cure

122 hlm. : - ; 20 cm