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Remodeling of the core leads HIV-1 pre-integration complex in the nucleus of human lymphocytes.

Abstract : Retroviral replication proceeds through obligate integration of the viral DNA into the host genome. In particular, HIV-1 genome to enter the nucleus, must be led through the nuclear pore complex (NPC). During HIV-1 cytoplasmic journey, the viral core acts like a shell to protect the viral genetic material from antiviral sensors and ensure an adequate environment for the reverse transcription. However, the relatively narrow size of the nuclear pore channel requires that the HIV-1 core reshapes into a structure that fits the pore. On the other hand, the organization of the viral CA proteins that remain associated to the pre-integration complex (PIC) during and after nuclear translocation is still enigmatic. In this study, we analysed the progressive organizational changes of viral CA proteins within the cytoplasm and the nucleus by immuno-gold labelling. Furthermore, we set up a novel technology, HIV-1 ANCHOR, which enables the specific detection of the retrotranscribed DNA by fluorescence microscopy, thereby offering the opportunity to uncover the architecture of the potential HIV-1 PIC. Thus, we combined the immunoelectron microscopy and ANCHOR technologies to reveal the presence of DNA- and CA-positive complexes by correlated light- and electron microscopy (CLEM). During and after nuclear translocation, HIV-1 appears as a complex of viral DNA decorated by multiple viral CA proteins remodelled in a "pearl necklace" shape. Thus, we could describe how CA proteins reshape around the viral DNA to permit the entrance of the HIV-1 in the nucleus. This particular CA protein complex composed by the integrase and the retrotranscribed DNA leads HIV-1 genome inside the host nucleus.Our findings contribute to the understanding of the early steps of HIV-1 infection and provide new insights into the organization of HIV-1 CA proteins during and after viral nuclear entry. Of note, we are now able to visualize the viral DNA in viral complexes, opening up new perspectives for future studies on viral nuclear fate.Importance How the reverse transcribed genome reaches the host nucleus remains a main open question related to the infectious cycle of HIV-1. HIV-1 core has a size of ∼100 nm, largely exceeding that of the NPC channel (∼39 nm). Thus, a rearrangement of the viral CA proteins organization is required to achieve an effective nuclear translocation. The mechanistic of this process remains undefined due to the lack of a technology capable to visualize potential CA sub-complexes in association with the viral DNA in the nucleus of HIV-1-infected cells.By the means of state-of-the-art technologies (HIV-1 ANCHOR system combined with CLEM), our study shows that remodeled viral complexes retain multiple CA proteins but not intact core or only a single CA monomer. These viral CA complexes associated with the retrotranscribed DNA can be observed in the outer and inner side of the NE, and they represent potential PIC.Thus, our study shed light on critical early steps characterizing HIV-1 infection, thereby revealing novel, therapeutically exploitable points of intervention. Furthermore, we developed and provided a powerful tool enabling direct, specific and high-resolution visualization of intracellular and intranuclear HIV-1 subviral structures.
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Submitted on : Monday, April 20, 2020 - 4:49:32 PM
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Guillermo Blanco-Rodriguez, Anastasia Gazi, Blandine Monel, Stella Frabetti, Viviana Scoca, et al.. Remodeling of the core leads HIV-1 pre-integration complex in the nucleus of human lymphocytes.. Journal of Virology, American Society for Microbiology, 2020, [Epub ahead of print], pp.JVI.00135-20. ⟨10.1128/JVI.00135-20⟩. ⟨pasteur-02548457⟩

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