HDAC6 as a new restriction factor with anti-replicative activity against HIV-1

The emergence of resistance to anti-retroviral drugs, which is the most common cause of treatment failure, and the existence of sanctuaries of latency, replication and spread of HIV-1 in the body, preventing the removal of integrated provirus, pose a major obstacle to the eradication of the virus in the fight against AIDS. This is why it is urgent to find new molecular and cellular targets against HIV-1, necessary for the development of new therapeutic strategies. Previous studies from our research group demonstrate a good correlation between the level of HDAC6 expression in target cells and the control exerted by the enzyme on HIV-1 replication (Valenzuela-Fernandez, A., et al., 2005). However, the mechanism through which HDAC6 hinders viral replication, once the virus has entered the cell, remains unknown. In this framework, this research project seeks to identify and characterize the mechanisms of the anti-HIV-1 activity exerted by HDAC6.

Deep functional analysis of HIV-1 envelope glycoproteins isolated from individuals showing extreme responses to infection

The extent of CD4 T cell depletion and the progression to AIDS is highly variable among HIV infected individuals due to both host and viral factors. The cytopathic potential associated to the HIV envelope glycoprotein has been suggested as one of the most relevant factors in this context. However, the evaluation of the function of the viral envelope has been a technical challenge and so far, except for the viral tropism, we lack experimental models with clinically contrasted relevance. Therefore, the main aim of this project is to define new tools of for the analysis of HIV Envelope function that describe clinically relevant parameters. We plan to analyze a collection of envelopes (n=27) partially characterized and isolated from HIV infected individuals with extreme responses to infection: rapid progressors and viremec non progressors. The parameters to be analyzed are: fusogenicity in primary cells (both CD4 T cells and macrophages), dependence and affinity for CD4, ability to form virological synapses, to recruit receptors and to signalize through them, and finally senesitivity to the inhibition of cellular cofactors (Arf6 and moesin). The different parameters will be correlated and analyzed in the context of clinical progression.

Regulation of HIV-1 fusion, entry and infection by controlling pore fusion formation by alteration of plasma membrane dynamics: involvement of Arf6 and F-actin disrupter proteins

Objective: The main aim of our research project consists in the elucidation of the functional involvement of the membrane traffic process for the recovery of the plasma membrane, as well as the integrity of cortical F-actin cytoskeleton that could occur at sites of HIV-1/cell contact. We seek for the identification and characterization of new molecular targets, which would allow the development of alternative therapeutical strategies against AIDS, by blocking HIV-1 infection at the early steps of attachment, fusion and viral entry. Experimental procedures: Real time study of the attachment, fusion and HIV-1 entry processes, of fluorescent HIV-1 viral particles defectives for replication, in permissive cells were Arf6- and gelsolin-related functions were modified, and by using dynamic fluorescent microscopy techniques: by Total Internal Reflection Fluorescence Microscopy (TIRFM).

KEY WORDS: Arf6, PIP2, gelsolin/cofilin/ERM, F-actin, pore fusion formation, regulation of HIV-1 infection, TIRFM.