Nef is an accessory protein encoded by human and simian immunodeficiency viruses (HIV and SIV), and is critical for viral pathogenicity in vivo.The structure of Nef has been resolved and the major cellular activities of Nef are generally described as down-regulation of cell surface molecules, enhancement of virus infectivity and regulation of cell signaling and activation. Macrophages represent a key target of HIV-1 infection and may contribute significantly to viral pathogenesis by facilitating viral propagation, maintaining a viral reservoir and regulating viral replication. During HIV-1 infection, various cytokines and chemokines are induced for viral advantages more than for host defense. We have previously demonstrated that HIV-1 Nef regulates the release of chemokines, MIP-1α and MIP-1ß, from infected macrophages and have proposed that this may enhance conditions for viral replication by promoting recruitment of substrate lymphocytes to sites of infection (1). However, the molecular basis for this Nef activity remains to be defined. The main goals of this thesis are to identify the functional motif in Nef that is responsible for chemokine induction in macrophages and to elucidate the relevance of this motif to other Nef functions. Using a mutagenesis approach, we have eventually identified a novel motif (KEK) that regulates chemokine production in infected macrophages after we excluded several previously described Nef motifs. This motif is conserved in both HIV-1 and SIV Nef proteins. Mutations in this domain abrogated MIP-1ß induction as well as the Nef-dependent release of other secretory factors by macrophages. However, disruption of this motif did not affect other Nef-ascribed activities such as CD4 and MHC-I down-regulation. In addition, we have determined the involvement of viral Env proteins in Nef-induced chemokine production. Distinct signaling pathways that regulate chemokine release in macrophage will also be described. Finally, several possible roles of the KEK motif are proposed and some preliminary results of co-immunoprecipitation experiments will be presented which aim to characterize cellular proteins involved in chemokine regulation by Nef. Collectively, our studies reveal a specific determinant within Nef that is critical for chemokine release by Nef. Identification of this motif paves the way for future studies to explore the molecular machanisms of Nef-regulated cell signaling pathways. Such knowledge may point to new therapeutic strategies that interrupt Nef function and limit the course of HIV-1 infection.

Alors que d’énormes efforts sont mis de l’avant pour mettre en place des stratégies thérapeutiques contre l’infection au VIH-1, il est nécessaire de mieux cerner les déterminants viraux qui aideront à l’efficacité de celles-ci. En ce sens, une volumineuse littérature scientifique suggère que les anticorps contre le VIH-1 possédant une capacité à induire une réponse effectrice dépendante de leur portion Fc puissent jouer un rôle important dans la prévention de l’infection et dans la progression de la maladie. Cependant, peu d’information est disponible concernant les déterminants reconnus par ces anticorps et comment le virus s’en protège. Le but des travaux présentés dans cette thèse est donc d’élucider les mécanismes viraux contrôlant la reconnaissance des cellules infectées par ces anticorps capables d’induire une réponse effectrice. De par les corrélats de protection identifiés au cours de l’essai vaccinal RV144, les travaux présentés ici se concentrent sur la réponse cytotoxique dépendante des anticorps (ADCC), puisqu’il s’agit d’une réponse effectrice suggérée pour avoir joué un rôle dans la protection observée dans le RV144, seul essai vaccinal anti-VIH à avoir démontré un certain degré de protection. De plus, plusieurs anticorps capables d’induire cette réponse contre le VIH sont connus pour reconnaître les glycoprotéines de surface du virus (Env) dans une conformation dite ouverte, c’est-à-dire la conformation adoptée lors de la liaison d’Env avec son récepteur CD4 (épitopes CD4i). Nous avons mis au point deux techniques in vitro permettant d’étudier ces changements de conformation ainsi que leur impact sur la réponse ADCC. Les techniques mises au point, un ÉLISA sur base cellulaire pour mesurer les changements de conformation d’Env ainsi que la mesure de la réponse ADCC par cytométrie en flux, nous ont permis de démontrer comment le virus empêche l’exposition des épitopes d’Env CD4i. L’activité simultanée des protéines accessoires virales Nef et Vpu sur le retrait du récepteur CD4 de la surface des cellules infectées et l’inhibition du facteur de restriction Tétherine / BST-2 par Vpu contrôlent à la fois les niveaux d’Env et de CD4 à la surface cellulaire et donc modulent l’interaction Env-CD4 et ultimement la susceptibilité à la réponse ADCC contre les épitopes CD4i reconnus par des anticorps hautement prévalents lors de l’infection au VIH. Également, nous démontrons comment de petits composés mimant la liaison de CD4 sur Env sont capables de forcer l’exposition des épitopes CD4i, même en présence des protéines Nef et Vpu, et donc d’augmenter la susceptibilité des cellules infectées à la réponse ADCC. Une autre découverte présentée ici est la démonstration que la portion soluble d’Env produite par les cellules infectées peut interagir avec le récepteur CD4 des cellules non-infectées avoisinantes et induire leur reconnaissance et élimination par la réponse ADCC contre Env. Somme toute, la modulation de la réponse ADCC par l’interaction Env–CD4 représente un important pilier de la relation hôte – pathogène du VIH-1 de la perspective des réponses Fc-dépendantes. Les travaux présentés dans cette thèse ont le potentiel d’être utilisés dans l’élaboration de nouvelles stratégies antivirales tout en élargissant les connaissances fondamentales de cette interaction hôte – pathogène. / While huge efforts are put forth to develop therapeutic strategies against HIV-1 infection, it is necessary to better understand the viral determinants that will help the effectiveness of these approaches. Moreover, a voluminous scientific literature suggests that antibodies against HIV-1 that have the ability to induce an Fc-mediated effector response can play an important role in the prevention and control of the disease. However, little information is available regarding the determinants recognized by these antibodies and how the virus protects itself from this response. The aim of the work presented in this thesis is therefore to better elucidate the viral mechanisms controlling recognition of infected cells by antibodies capable of inducing effector responses. In regards to the correlates of protection identified in the RV144 vaccine trial, the work presented here focuses on antibody-dependant cellular cytotoxicity (ADCC), since this effector response was suggested to have played a role in the protection observed in the RV144 trial. In addition, many antibodies that induce this response against HIV are known to recognize the virus surface glycoprotein (Env) in an open conformation, that is to say, the conformation adopted by the binding of Env with the CD4 receptor (CD4i epitopes). We have developed two in vitro techniques to study these conformational changes and their impact on ADCC responses. The techniques developed, a cell-based ELISA to measure Env conformational changes and the measure of ADCC responses by flow cytometry, allowed us to demonstrate how the virus prevents exposure of Env CD4i epitopes. The simultaneous activity of viral accessory proteins Vpu and Nef on the removal of CD4 from the surface of infected cells and the Vpu-mediated inhibition of the restriction factor Tetherin / BST-2 control both Env and CD4 levels at the cell surface, thus modulating Env-CD4 interaction. This ultimately results in a decrease in the susceptibility of infected cells to ADCC responses against CD4i epitopes recognized by antibodies that are highly prevalent during HIV infection. Also, we demonstrate how using small compounds mimicking the CD4 binding of Env forces the exposure of CD4i epitopes, even in the presence of Nef and Vpu proteins, and therefore increases the susceptibility to ADCC responses against infected cells. Another discovery is presented here that demonstrates how the soluble portion of Env produced by infected cells can interact with the CD4 receptor on the bystanders non-infected cells and induce their recognition and elimination by ADCC responses against Env. Overall, the modulation of ADCC responses by Env-CD4 interaction is an important pillar of HIV-1 host – pathogen interaction from the perspective of Fc effector functions. The work presented in this thesis has the potential to be used in the development of new antiviral strategies while expanding the fundamental understanding of HIV-1 host - pathogen interactions.

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

On dispose de deux notions de positivité pour les (1,1)-classes de cohomologies d'une variété complexe: l'effectivité numérique, induite par la positivité au sens de Lelong des formes différentielles, et la pseudoeffectivité, plus faible, induite par celle des courants. Dans une première partie, nous construisons des obstructions locales à l'effectivité numérique d'une classe pseudoeffective, ce qui permet de la décomposer en une partie nef en codimension un et un diviseur exceptionnel. Dans un second temps, nous nous intéressons au volume d'un fibré en droites, qui est un invariant mesurant sa positivité. Nous en donnons une interprétation en terme de géométrie différentielle, et nous montrons comment ce volume s'inscrit dans une théorie de l'``intersection mobile'', qui ne conserve que les parties numériquement effectives des classes de cohomologie. Finalement, nous étudions le cas des surfaces et des variétés hyperkähleriennes, pour lesquelles la géométrie de la forme d'intersection permet une description plus détaillée de ces constructions.

En la present Tesi s'ha estudiat l'efecte de diversos activadors de trimetilfosfina en diferents reaccions i s'ha fet l'esforç d'esbrinar els mecanismes de reacció. Així s'han pogut racionalitzar els comportaments dels activadors, cosa que ha permès de desenvolupar nous mètodes de síntesi o millorar-ne d'altres.En el capítol 1.2 s'ha optimitzat la transformació de nitroalcans secundaris en cetones. Els millors activadors per dur a terme la reacció reacció són el disulfur de 4-terc-butiltiofenil i l'N-(fenilseleno)ftalimida. La reacció té lloc llavors de manera suau, catalítica i sense que es generi cap subproducte pudent. L'N-(fenilseleno)ftalimida millora els rendiments de les reaccions que impliquen grups nitro impedits.L'estudi mecanístic de la reacció va aportar nous coneixements que ens van permetre variar l'activador i el nombre d'equivalents de trimetilfosfina per convertir a voluntat els nitroalcans secundaris en imines o en N-sulfenilimines. En el capítol 1.3 s'ha descrit el primer mètode de conversió directa de nitroalcans secundaris o cetooximes en N-fenilsulfenilimines. En la majoria dels casos, els rendiments són superiors al 90% i els possibles centres estereogènics en α al grup C=NSPh no es veuen afectats durant la reacció.Malauradament, el mètode descrit en el capítol 1.2 produeix la racemització total dels centres en α al grup CHNO2. Per tal de solucionar-ho, en el capítol 1.4 s'ha estudiat la hidròlisi de les N-sulfenilimines obtingudes en el capítol 1.3 catalitzada per àcids de Lewis. El AuBr3 ha estat la única sal metàl·lica capaç de promoure la hidròlisi a un pH neutre. A més, es pot realitzar la reacció de formació d'N-sulfenilimines i la hidròlisi amb AuBr3 de manera consecutiva, tot obtenint les cetones corresponents sense afectació dels centres estereogènics en α al grup CHNO2 de partida.En la segona part de la present Tesi doctoral, s'ha estudiat l'acoblament directe entre àcids carboxílics i azides mitjançat per trimetilfosfina. S'han emprat diferents activadors (PhSSPh, PySSPy, 4-PySS-4-Py, PhSeSePh, PySeSePy), alguns dels quals acceleren la reacció de manera significativa. S'ha proposat un mecanisme i s'ha reinvestigat la presència d'una reacció paràsita que explica perfectament la diferència tan gran d'activitat entre activadors aparentment tan semblants. Aquesta metòdica s'ha emprat en un pas clau de la síntesi de les fluvirucines B1-B5 i alguns anàlegs, en el nostre grup de recerca. / Different trimethylphosphine activators have been explored in different reactions, with a special effort to understand reaction mechanisms. It was possible to rationalize the different behavior and performance of the activators and to develop new synthetic methodologies or to improve the existing ones.In chapter 1.2 the transformation of secondary nitroalkanes to ketones was optimized. The best activators to carry out this reaction are 4,4'-bis-tert-butyldiphenyl disulfide and N-(phenylselenenyl)phthalimide. With these improvements, the reaction takes place catalytically, under mild conditions and avoiding the generation of a stinking subproduct. N- (phenylselenenyl)phthalimide improves the yield of the reactions when sterically hindered nitro groups are used as starting material.The knowledge gained through mechanistic studies allowed us to change the activator and the number of equivalents of trimethylphosphine to transform secondary nitroalkanes to imines or N-phenylsulfenyl ketimines. In chapter 1.3 we describe the first direct conversion of secondary nitroalkanes or ketoximes to N-phenylsulfenyl ketimines. Reaction yields are generally over 90% and α stereogenic centers to the C=NSPh group are not affected during the reaction.Unfortunately, the method described in chapter 1.2 produces the total racemization of α stereogenic centers to CHNO2. To solve this problem, in chapter 1.4 we studied the hydrolysis of the N-sulfenilimines obtained in chapter 1.3, catalyzed by a Lewis acid. We proved that AuBr3 is the only metallic salt that hydrolyzes N-sulfenilimines under neutral pH conditions. Moreover, it is possible to do both reactions, the formation of N-phenylsulfenyl ketimines and its hydrolysis with AuBr3, sequentially in one pot without affecting the α stereocenters of the starting material CHNO2.Also, the direct coupling between carboxylic acids and azides mediated by trimethylphosphine has been studied. Different activators (PhSSPh, 2-PySS-2-Py, 4-PySS-4-Py, PhSeSePh, 2-PySeSe-2-Py) have been used, some of them producing a significant acceleration of the reaction rate. We propose a reaction mechanism for this coupling and the existence of a parasitic reaction. This parasitic reaction explains the huge difference in activity between activators with similar structures. In our group, this new methodology has already been used in a crucial step of the synthesis of fluvirucins B2-B5 and other analogues.

The accessory protein Nef of human immunodeficiency virus type 1 (HIV-1) has long been known to enhance the infectivity of HIV-1 progeny virions. The multipass transmembrane proteins serine incorporator 3 (SERINC3) and SERINC5 were recently identified as novel antiviral proteins that restrict HIV-1 infectivity. Nef enhances HIV-1 infectivity by removing SERINCs from the plasma membrane, which prevents their incorporation into progeny HIV-1 virions. To exploit this potent intrinsic antiretroviral factor for potential therapy development, it is critical to explore the determinants in SERINC5 that govern its downregulation by Nef and its restriction on HIV-1 infectivity. Here I report that the ability to inhibit HIV-1 infectivity is conserved among vertebrate SERINC5 proteins, whereas the sensitivity to downregulation by Nef is not. However, a Nef-resistant SERINC5 became Nef-sensitive when its intracellular loop 4 (ICL4) was replaced by that of Nef-sensitive human SERINC5. Conversely, human SERINC5 became resistant to Nef when its ICL4 was replaced by that of a Nef-resistant SERINC5. In general, ICL4 regions from SERINCs that exhibited resistance to a given Nef conferred resistance to the same Nef when transferred to a sensitive SERINC, and vice versa. I demonstrate that human SERINC5 can be modified to restrict HIV-1 infectivity even in the presence of Nef. Moreover, by generating chimeras between SERINC5 and SERINC2, which does not exhibit antiretroviral activity, I demonstrate that SERINC5’s inhibitory function, unlike the sensitivity to Nef, requires the participation of more than one region. Helix 4 and extracellular loop 5 (ECL5) of SERINC5 are both required for the potent restriction of HIV-1 infectivity. In contrast, a large amino-terminal portion of SERINC5 is not required for its antiretroviral activity of SERINC5. The determinants in ECL5 disperse throughout the loop. Furthermore, the ECL5 of SERINC5 is a hotspot region that determines the Env-dependent antiretroviral activity of SERINC5.

Development of a suitable animal model of AIDS is much needed in AIDS research to study infection and pathogenesis as well as to evaluate methods of prevention and treatment of HIV infection. Small animals such as rodents are attractive candidates for AIDS research due to the availability of various inbred and genetically engineered strains, extensive knowledge or their immune system, especially in mice, and the relative ease of breeding and maintaining animal colonies. Transgenic small animal models carrying entire HIV genome or selected genes have been instrumental to understand functions of HIV genes in vivo and their role in HIV pathogenesis. The type of cells in which HIV genes are expressed seems to be an import prerequisite for the study of HIV gene functions in transgenic mice. Mice constitutively expressing the entire HIV-1 genome or HIV-1 nef gene in CD4 + T cells and in the cells of macrophage/dendritic lineage develop an AIDS-like disease very similar to AIDS disease in humans. Similarly, expression of Nef in adult mice, using inducible system, results in the AIDS-like disease. This disease is characterized by thymic atrophy, impaired thymocyte maturation, loss of CD4+ T cells, increased activation and turnover of T cells, which can occur in the absence of lymphypenia, and non-lymphoid organ disease involving the lungs and kidneys. Susceptibility of adult mice to the pathological effects of Nef suggests that the AIDS-like disease in the constitutively expressing Nef Tg mice is not due to developmental defects caused by early expression of Nef. This model highlights the important role of Nef in HIV-1 pathogenesis. The high similarity in the disease in these Tg mice with human AIDS strongly suggest that these mice are a relevant model to study AIDS. This study further evidence that mouse cells can support functions of Nef and these Tg mice represent a unique model to study Nef functions in vivo in the context of the primary immune system. Moreover, the inducible Nef Tg model has given us the ability to control the level and time of expression of Nef which was impossible to do in the previously reported constitutive Nef Tg mouse models. These mice will be useful to study immune reconstitution since Nef expression can be turned off after withdrawal from dox.

One of the hallmarks of an infection with pathogenic HIV-1 is the elevated level of immune activation that leads to rapid progression to AIDS. Surprisingly, nonhuman primates naturally infected with SIV do not exhibit an augmented activation phenotype nor severe immunodeficiency. One of the viral components implicated in determining the state of immune activation is the accessory protein Nef which has been demonstrated to affect T cell signaling pathways from within the intracellular compartment and for Nef from SIV, to downregulate TCR surface expression. Recently, Nef from HIV-1 and SIV have been demonstrated to bind the ζ chain of the TCR which functions as the primary signaling subunit of the receptor. However, the molecular details of the Nef-TCRζ interaction as well as the role of complex formation in modulation of immune activation remain largely unknown. This thesis describes work directed at elucidating the biochemical and structural features of the Nef-TCRζ interaction and the functional consequences of complex formation relevant to T cell activation. Chapter I provides a brief introduction on HIV/SIV classification and pathogenesis with an emphasis on Nef and its pleiotropic function in T cells. Chapter II describes the biochemical characterization of the interaction of the conserved core domain of Nef proteins from HIV-1, HIV-2 and SIV with the cytoplasmic domain of TCRζ. The core domains of HIV-2 ST and SIVmac239 are demonstrated to bind the cytoplasmic domain of TCRζ at two distinct regions and with different affinities. In contrast, the core domain of HIV-1 isolate ELI Nef only binds to one region and with the weakest calculated affinity among the HIV-1, HIV-2 and SIV Nef proteins studied. In addition, both the N-terminal domain and the strong TCRζ-binding core domain of SIVmac239 Nef each are demonstrated to be necessary but not sufficient for downregulation of TCR surface expression. Chapter III describes the crystallization and structure determination methods used to solve the crystal structures of the core domain of SIVmac239 Nef in complex with two overlapping TCRζ polypeptides. Crystals of Nef in complex with the longer TCRζDP1 (L51-D93) polypeptide grew in a tetragonal space group but only diffracted to low resolution. In contrast, crystals of the Nefcore-TCRζA63-R80 complex grew in an orthorhombic space group and diffracted to high resolution but were nearly perfectly pseudo-merohedrally twinned thus complicating structure determination. Following identification of the twin law relating the twin domains, the structure of the Nefcore-TCRζA63-R80 complex was determined using refinement procedures that accounted for crystal twinning to 2.05 Å. The structure of the Nefcore-TCRζDP1 complex was solved to 3.7 Å from a single non-twinned crystal. The altered crystal packing induced by the shorter TCRζA63-R80polypeptide is postulated to have led to a reduction in crystal symmetry and increase in proneness to crystal twinning. Chapter IV provides a detailed analysis of the structure of the Nefcore-TCRζA63-R80 complex and demonstrates its effect on modulation of Src family kinase activity. The TCRζ polypeptide adopts an alpha helical conformation and occupies a hydrophobic crevice on Nef not shared by any of Nef’s reported interaction partners. The interaction of Nefcore with TCRζ is mediated primarily by the burial of hydrophobic residues on TCRζ (L75, L77) in a hydrophobic pocket on Nef and a salt bridge between a glutamic acid (E74) on TCRζ and a basic patch on Nef consisting of two conserved arginines (R105, R106). The TCRζ polypeptide additionally orders the N-terminus of Nefcore into a polyproline type II helix that has been described to bind the SH3 domain of Src family kinases. We demonstrate that in vitro phosphorylation of TCRζcyt by Fyn and Src is specifically augmented by HIV-1 and SIV Nefcoreand suggest that Nef-TCRζ complex formation cooperatively enhances kinase activity. Chapter V contains overall conclusions, future directions and a model illustrating the proposed role of the Nef-TCRζ interaction in immune activation modulation. The Appendices contain sequences of the proteins, gene constructs and primers used in this work.

No description available.

En l'absence d'un vaccin efficace et avec des thérapies antirétrovirales incapables d'éradiquer le virus, le VIH-1 reste un problème de santé publique mondial. Des immunothérapies à base d'anticorps sont à l'étude pour éliminer les réservoirs cellulaires, qui représentent un obstacle incontournable à la guérison du VIH-1. Les glycoprotéines d'enveloppe du VIH-1 (Env) représentent le seul antigène du virus exposé à la surface des cellules infectées et constituent donc la principale cible des anticorps. L’Env non-liée adopte sa conformation « fermée », reconnue préférentiellement par les anticorps neutralisants. L'interaction avec CD4 fait passer Env dans sa conformation « ouverte », exposant des épitopes conservés reconnus par les anticorps non-neutralisants (nnAbs) présents dans le sérum d’individus infectés par le VIH-1 (sérums VIH+). Les nnAbs peuvent éliminer les cellules infectées par la cytotoxicité cellulaire dépendante des anticorps (ADCC). Cependant, les protéines accessoires Nef et Vpu diminuent l’expression de surface de CD4 et BST-2 afin d’évader à la reconnaissance et l'élimination des cellules infectées par les nnAbs. Dans cette thèse, nous caractérisons en détail la contribution d’Env, Nef et Vpu pour échapper aux réponses humorales et explorons de nouvelles stratégies pour sensibiliser les cellules infectées à l’ADCC. Pour quantifier plus adéquatement la réponse ADCC, nous avons identifié des biais dans les tests largement utilisés, notamment pour évaluer les corrélats de protection vaccinale. Il s'agit de l'incapacité à faire la distinction entre l'élimination des cellules infectées et des cellules non-infectées, et l'utilisation de constructions virales comportant un gène rapporteur empêchant l’expression de Nef. En utilisant un nouveau marquage intracellulaire, nous avons confirmé l’effet protecteur de Nef et Vpu contre l’ADCC. Ensuite, nous avons étudié les déterminants d’Env et Vpu modulant la susceptibilité des cellules infectées à l’ADCC médiée par les nnAbs. Certaines caractéristiques structurelles d'Env modulent ses transitions conformationnelles, incluant le domaine d'association du trimère, le site de clivage de la furine et la cavité Phe43. L’altération de ces composantes augmente la sensibilité des cellules infectées à l'ADCC par les sérums VIH+. Outre l’inhibition de CD4 et BST-2, Vpu cible également NTB-A et PVR, des ligands de récepteurs activateurs des cellules NK. Cependant, la polyfonctionnalité de Vpu est compromise par l’augmentation de BST-2 par les interférons de type I (IFN-I), sensibilisant ainsi les cellules infectées aux réponses NK. En utilisant un modèle de souris humanisée, nous validons l'importance de Vpu pour échapper à la pression immunitaire des nnAbs in vivo. Enfin, nous avons exploré de nouvelles stratégies pour sensibiliser les cellules infectées à l'ADCC en modulant la conformation d’Env avec des mimétiques moléculaires de CD4 (CD4mc). Nous avons identifié des résidus bordant la cavité Phe43 modulant la sensibilité au CD4mc. L’accumulation d’Env induite par les IFN-I augmente la capacité du CD4mc à sensibiliser les cellules infectées à l'ADCC par les sérums VIH+. Globalement, cette thèse dévoile une caractérisation approfondie des déterminants viraux et cellulaires modulant la susceptibilité des cellules infectées par le VIH-1 aux réponses humorales. Une meilleure compréhension de ces mécanismes est nécessaire pour développer des nouvelles stratégies capables d’éradiquer les réservoirs viraux. / In the absence of an effective vaccine and with antiretroviral therapies unable to eradicate the virus, HIV-1 remains a global public health problem. Antibody-based immunotherapies are currently being investigated to eliminate cellular reservoirs, which represent a major obstacle towards an HIV-1 cure. HIV-1 envelope glycoproteins (Env) represent the only virus-specific antigen exposed at the surface of infected cells and therefore is the main target for antibodies. In its unliganded form, Env samples a ‘closed’ conformation, preferentially recognized by neutralizing antibodies. Interaction with CD4 drives Env into its ‘open’ conformation, exposing conserved epitopes recognized by non-neutralizing antibodies (nnAbs) present in sera from HIV-1 infected individuals (HIV+ sera). NnAbs can eliminate infected cells by antibody-dependent cellular cytotoxicity (ADCC). However, HIV-1 encodes for the accessory proteins Nef and Vpu which decrease cell surface levels of CD4 and BST-2, thus avoiding recognition and elimination of infected cells by nnAbs. In this thesis, we characterize in detail the contribution of Env, Nef, and Vpu to evade humoral responses and explore new strategies for sensitizing infected cells to ADCC. In an effort to develop a more adequate quantification of ADCC responses, we identified major biases in widely used assays, including the ones used to assess correlates of vaccine protection. These include the inability to distinguish between the elimination of infected and uninfected cells and the use of viral constructs coding for a reporter gene that prevents Nef expression. Using a novel intracellular staining, we confirmed the protective effect of Nef and Vpu against ADCC responses. Next, we studied the different Env and Vpu determinants modulating the susceptibility of infected cells to nnAbs-mediated ADCC responses. Certain Env structural features modulate its conformational transitions, including the trimer association domain, the furin cleavage site and the Phe43 cavity. Alterations of these components increase the susceptibility of HIV-1-infected cells to ADCC mediated by HIV+ sera. In addition to inhibiting CD4 and BST-2, Vpu also targets NTB-A and PVR, which act as ligands for NK cell activating receptors. However, we found that the polyfunctionality of Vpu can be compromised by the upregulation of BST-2 by type I interferons (IFN-I), thereby sensitizing infected cells to NK cell responses. Using a humanized mouse model, we validate the importance of Vpu to escape the immune pressure of nnAbs in vivo. Finally, we explored new strategies to bypass the protective effect of Vpu and Nef and sensitize HIV-1-infected cells to ADCC by modulating Env conformation using small CD4-mimetic compounds (CD4mc). We identified a network of residue lining the Phe43 cavity that modulates Env sensitivity to CD4mc. The enhanced surface expression of Env by type I IFNs boosts the ability of CD4mc to sensitize HIV-1-infected cells to ADCC by HIV+ sera. Overall, this thesis sheds light on a thorough characterization of viral and cellular determinants modulating the susceptibility of HIV-1-infected cells to humoral responses. A better understanding of these mechanisms is needed to develop new strategies able to eradicate viral reservoirs.