Study on the molecular interactions of the HIV-1 Nef protein and the host cell endolysosomal system

Abstract

The Human Immunodeficiency Virus (HIV) is the etiologic agent of Acquired Immunodeficiency Syndrome (AIDS). It is a disease with a worldwide distribution, and it is estimated that there are currently 35 million people infected with the virus. During the life cycle, HIV promotes several changes in host cell physiology to optimize virus replication and scape innate defense mechanisms. The fast progression of HIV-1 infection in humans and animal models is closely linked to the function of the accessory protein Nef. Among the several actions of Nef, the most prominent is the downregulation of surface proteins important in the immune response, such as the CD4 receptor. It is now clear that Nef's action results in degradation of this receptor in lysosomes, but the molecular mechanisms involved are not fully elucidated. Nef forms a tripartite complex with the adaptor protein AP-2 and the cytosolic tail of CD4 in clathrin-coated pits, internalizing CD4 and directing it to lysosomal degradation. Our previous studies have shown that targeting of CD4 to lysosomes by Nef involves the entry of the receptor into the multivesicular body (MVBs) pathway via an atypical mechanism that despite not requiring cargo ubiquitination, depends on the action of the ESCRT (Endosomal Sorting Complexes Required of Transport) machinery. We also show that this depends on the interaction of Nef with Alix, and ESCRT accessory protein. Here, we propose to further dissect this mechanism by investigating whether interaction of Alix with TSG101 (ESCRT-I) and/or CHMP4 (ESCRT-III) is required for downregulation of CD4. Furthermore, we will investigate whether the adaptor protein AP-5 physically interacts with Nef and if it plays a role in CD4 degradation. Because our previous results were obtained in HeLa cells, we also propose to perform most of the experiments in CD4+ T cells, the natural target of HIV infection. A better understanding of these processes wills novel aspects of the mechanism by which Nef subverts the host cell endolisossomal system components, generating important information for better know the mechanisms of this virus pathogenesis. In addition, this study has the potential to contribute the understanding of primordial process regulation on intracellular trafficking of transmembrane proteins. (AU)