Correction: Human Nucleoporins Promote HIV-1 Docking at the Nuclear Pore, Nuclear Import and Integration

The delivery of the HIV-1 genome into the nucleus is an indispensable step in retroviral infection of non-dividing cells, but the mechanism of HIV-1 nuclear import has been a longstanding debate due to controversial experimental evidence. It was commonly believed that the HIV-1 capsid would need to disassemble (uncoat) in the cytosol before nuclear import because the capsid is larger than the central channel of nuclear pore complexes (NPCs); however, increasing evidence demonstrates that intact, or nearly intact, HIV-1 capsid passes through the NPC to enter the nucleus. With the protection of the capsid, the HIV-1 core completes reverse transcription in the nucleus and is translocated to the integration site. Uncoating occurs while, or after, the viral genome is released near the integration site. These independent discoveries reveal a compelling new paradigm of this important step of the HIV-1 life cycle. In this review, we summarize the recent studies related to HIV-1 nuclear import, highlighting the spatial–temporal relationship between the nuclear entry of the virus core, reverse transcription, and capsid uncoating.

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Cone-shaped HIV-1 capsids are transported through intact nuclear pores

10.1101/2020.07.30.193524 ◽

2020 ◽

Cited By ~ 3

Author(s):

Vojtech Zila ◽

Erica Margiotta ◽

Beata Turonova ◽

Thorsten G. Müller ◽

Christian E. Zimmerli ◽

...

Keyword(s):

Nuclear Import ◽

Light And Electron Microscopy ◽

Productive Infection ◽

Nuclear Pore ◽

Major Health ◽

Replication Complex ◽

Empty Capsid ◽

Immunodeficiency Virus ◽

Subtomogram Averaging ◽

Hiv 1

AbstractHuman immunodeficiency virus (HIV-1) remains a major health threat. Viral capsid uncoating and nuclear import of the viral genome are critical for productive infection. The size of the HIV-1 capsid is generally believed to exceed the diameter of the nuclear pore complex (NPC), indicating that capsid uncoating has to occur prior to nuclear import. Here, we combined correlative light and electron microscopy with subtomogram averaging to capture the structural status of reverse transcription-competent HIV-1 complexes in infected T cells. We demonstrate that the diameter of the NPC in cellulo is sufficient for the import of apparently intact, coneshaped capsids. Subsequent to nuclear import, we detected disrupted and empty capsid fragments, indicating that uncoating of the replication complex occurs by breaking the capsid open, and not by disassembly into individual subunits. Our data directly visualize a key step in HIV-1 replication and enhance our mechanistic understanding of the viral life cycle.

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Author response: HIV-1 nuclear import in macrophages is regulated by CPSF6-capsid interactions at the nuclear pore complex

10.7554/elife.41800.031 ◽

2018 ◽

Author(s):

David Alejandro Bejarano ◽

Ke Peng ◽

Vibor Laketa ◽

Kathleen Börner ◽

K Laurence Jost ◽

...

Keyword(s):

Nuclear Pore Complex ◽

Nuclear Import ◽

Author Response ◽

Nuclear Pore ◽

Hiv 1

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A DNA-origami nuclear pore mimic reveals nuclear entry mechanisms of HIV-1 capsid

10.1101/2020.08.10.245522 ◽

2020 ◽

Author(s):

Qi Shen ◽

Chaoyi Xu ◽

Sooin Jang ◽

Qiancheng Xiong ◽

Swapnil C. Devarkar ◽

...

Keyword(s):

Nuclear Pore Complex ◽

Nuclear Import ◽

Dna Origami ◽

Nuclear Pore ◽

Nuclear Entry ◽

Cellular Factors ◽

Immunodeficiency Virus ◽

Hiv 1 ◽

Human Immunodeficiency Virus 1

SummaryThe capsid of human immunodeficiency virus 1 (HIV-1) plays a pivotal role in viral nuclear import, but the mechanism by which the viral core passages the nuclear pore complex (NPC) is poorly understood. Here, we use DNA-origami mimics of the NPC, termed NuPODs (NucleoPorins Organized by DNA), to reveal the mechanistic underpinnings of HIV-1 capsid nuclear entry. We found that trimeric interface formed via three capsid protein hexamers is targeted by a triple-arginine (RRR) motif but not the canonical phenylalanine-glycine (FG) motif of NUP153. As NUP153 is located on the nuclear face of the NPC, this result implies that the assembled capsid must cross the NPC in vivo. This hypothesis is corroborated by our observations of tubular capsid assemblies penetrating through NUP153 NuPODs. NUP153 prefers to bind highly curved capsid assemblies including those found at the tips of viral cores, thereby facilitating capsid insertion into the NPC. Furthermore, a balance of capsid stabilization by NUP153 and deformation by CPSF6, along with other cellular factors, may allow for the intact capsid to pass NPCs of various sizes. The NuPOD system serves as a unique tool for unraveling the previously elusive mechanisms of nuclear import of HIV-1 and other viruses.

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HIV-1 capsid binds host cofactors to mediate nuclear import

Acta Crystallographica Section A Foundations and Advances ◽

10.1107/s2053273314098799 ◽

2014 ◽

Vol 70 (a1) ◽

pp. C120-C120

Author(s):

Amanda Price ◽

David Jacques ◽

Sebastian Essig ◽

Tom Elliott ◽

Upul Halambage ◽

...

Keyword(s):

Nuclear Import ◽

Binding Sites ◽

Essential Role ◽

Antiretroviral Drugs ◽

Nuclear Pore ◽

The Core ◽

Early Stages ◽

Important Target ◽

Hexameric Form ◽

Hiv 1

The capsid (CA) protein of HIV-1, which forms the core of the virus, has been shown to have an increasingly important role in the early stages of the virus lifecycle, in particular during reverse transcription and nuclear import. We recently solved the structure of a fragment of the human cofactor CPSF6 in complex with the N-terminal domain of HIV-1 CA, revealing a previously unknown interface used by the virus to recruit CPSF6, which is required for the virus to successfully complete the early stages of its lifecycle. Using a recently developed hexameric unit of CA, we have solved the structure of the CPSF6 peptide with CA in a context that more closely resembles an intact CA lattice. This has revealed that CPSF6 contacts HIV-1 CA using an additional second site only present in the hexameric form of CA. Furthermore, we have now solved the structure of a fragment of NUP153 (an HIV-1 cofactor that is integral to the nuclear pore) in complex with hexameric CA and discovered that this also forms contacts specific to hexameric CA. Moreover, the binding sites for CPSF6 and NUP153 on CA overlap at one crucial residue, which is remarkably mimicked by two drugs independently discovered to bind at this same site. Together, these data provide evidence for an essential role for CA in HIV-1 infection, and highlights CA as an important target for antiretroviral drugs.

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Analysis of the Viral Elements Required in the Nuclear Import of HIV-1 DNA

Journal of Virology ◽

10.1128/jvi.01952-09 ◽

2009 ◽

Vol 84 (2) ◽

pp. 729-739 ◽

Cited By ~ 48

Author(s):

Lise Rivière ◽

Jean-Luc Darlix ◽

Andrea Cimarelli

Keyword(s):

Nuclear Import ◽

Matrix Protein ◽

Active Phase ◽

Cell Types ◽

Target Cells ◽

Nuclear Pore ◽

Viral Dna ◽

The Matrix ◽

Central Polypurine Tract ◽

Hiv 1

ABSTRACT HIV-1 possesses an exquisite ability to infect cells independently from their cycling status by undergoing an active phase of nuclear import through the nuclear pore. This property has been ascribed to the presence of karyophilic elements present in viral nucleoprotein complexes, such as the matrix protein (MA); Vpr; the integrase (IN); and a cis-acting structure present in the newly synthesized DNA, the DNA flap. However, their role in nuclear import remains controversial at best. In the present study, we carried out a comprehensive analysis of the role of these elements in nuclear import in a comparison between several primary cell types, including stimulated lymphocytes, macrophages, and dendritic cells. We show that despite the fact that none of these elements is absolutely required for nuclear import, disruption of the central polypurine tract-central termination sequence (cPPT-CTS) clearly affects the kinetics of viral DNA entry into the nucleus. This effect is independent of the cell cycle status of the target cells and is observed in cycling as well as in nondividing primary cells, suggesting that nuclear import of viral DNA may occur similarly under both conditions. Nonetheless, this study indicates that other components are utilized along with the cPPT-CTS for an efficient entry of viral DNA into the nucleus.

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Characterization of HIV-1 Vpr Nuclear Import: Analysis of Signals and Pathways

The Journal of Cell Biology ◽

10.1083/jcb.143.4.875 ◽

1998 ◽

Vol 143 (4) ◽

pp. 875-885 ◽

Cited By ~ 143

Author(s):

Yonchu Jenkins ◽

Michele McEntee ◽

Karsten Weis ◽

Warner C. Greene

Keyword(s):

Nuclear Import ◽

Nuclear Pore ◽

Nuclear Targeting ◽

Soluble Receptors ◽

Preintegration Complex ◽

Gtp Hydrolysis ◽

Localization Signal ◽

Targeting Signals ◽

Hiv 1

While the Vpr protein of HIV-1 has been implicated in import of the viral preintegration complex across the nuclear pore complex (NPC) of nondividing cellular hosts, the mechanism by which Vpr enters the nucleus remains unknown. We now demonstrate that Vpr contains two discrete nuclear targeting signals that use two different import pathways, both of which are distinct from the classical nuclear localization signal (NLS)- and the M9-dependent pathways. Vpr import does not appear to require Ran-mediated GTP hydrolysis and persists under conditions of low energy. Competition experiments further suggest that Vpr directly engages the NPC at two discrete sites. These sites appear to form distal components of a common import pathway used by NLS- and M9-containing proteins. Together, our data suggest that Vpr bypasses many of the soluble receptors involved in import of cellular cargoes. Rather, this viral protein appears to directly access the NPC, a property that may help to ensure the capacity of HIV to replicate in nondividing cellular hosts.

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Human Nucleoporins Promote HIV-1 Docking at the Nuclear Pore, Nuclear Import and Integration

PLoS ONE ◽

10.1371/journal.pone.0046037 ◽

2012 ◽

Vol 7 (9) ◽

pp. e46037 ◽

Cited By ~ 98

Author(s):

Francesca Di Nunzio ◽

Anne Danckaert ◽

Thomas Fricke ◽

Patricio Perez ◽

Juliette Fernandez ◽

...

Keyword(s):

Nuclear Import ◽

Nuclear Pore ◽

Hiv 1

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Nup153 Unlocks the Nuclear Pore Complex for HIV-1 Nuclear Translocation in Nondividing Cells

Journal of Virology ◽

10.1128/jvi.00648-18 ◽

2018 ◽

Vol 92 (19) ◽

Cited By ~ 21

Author(s):

Cindy Buffone ◽

Alicia Martinez-Lopez ◽

Thomas Fricke ◽

Silvana Opp ◽

Marco Severgnini ◽

...

Keyword(s):

Nuclear Import ◽

Site Selection ◽

Nuclear Translocation ◽

Integration Site ◽

The State ◽

Nuclear Pore ◽

Binding Interface ◽

Nuclear Factors ◽

Hiv 1 ◽

Integration Site Selection

ABSTRACTHuman immunodeficiency virus type 1 (HIV-1) displays the unique ability to infect nondividing cells. The capsid of HIV-1 is the viral determinant for viral nuclear import. To understand the cellular factors involved in the ability of HIV-1 to infect nondividing cells, we sought to find capsid mutations that allow the virus to infect dividing but not nondividing cells. Because the interaction of capsid with the nucleoporin protein 153 (Nup153) is important for nuclear import of HIV-1, we solved new crystal structures of hexameric HIV-1 capsid in complex with a Nup153-derived peptide containing a phenylalanine-glycine repeat (FG repeat), which we used to guide structure-based mutagenesis of the capsid-binding interface. HIV-1 viruses with mutations in these capsid residues were tested for their ability to infect dividing and nondividing cells. HIV-1 viruses with capsid N57 substitutions infected dividing but not nondividing cells. Interestingly, HIV-1 viruses with N57 mutations underwent reverse transcription but not nuclear translocation. The mutant capsids also lost the ability to interact with Nup153 and CPSF6. The use of small molecules PF74 and BI-2 prevented the interaction of FG-containing nucleoporins (Nups), such as Nup153, with the HIV-1 core. Analysis of integration sites in HIV-1 viruses with N57 mutations revealed diminished integration into transcriptionally active genes in a manner resembling that of HIV-1 in CPSF6 knockout cells or that of HIV-1-N74D. The integration pattern of the N57 mutant HIV-1 can be explained by loss of capsid interaction with CPSF6, whereas capsid interaction with Nup153 is required for HIV-1 to infect nondividing cells. Additionally, the observed viral integration profiles suggested that integration site selection is a multiparameter process that depends upon nuclear factors and the state of the cellular chromatin.IMPORTANCEOne of the key advantages that distinguish lentiviruses, such as HIV-1, from all other retroviruses is its ability to infect nondividing cells. Interaction of the HIV-1 capsid with Nup153 and CPSF6 is important for nuclear entry and integration; however, the contribution of each of these proteins to nuclear import and integration is not clear. Using genetics, we demonstrated that these proteins contribute to different processes: Nup153 is essential for the HIV-1 nuclear import in nondividing cells, and CPSF6 is important for HIV-1 integration. In addition, nuclear factors such as CPSF6 and the state of the chromatin are known to be important for integration site selection; nevertheless, the preferential determinant influencing integration site selection is not known. This work demonstrates that integration site selection is a multiparameter process that depends upon nuclear factors and the state of the cellular chromatin.

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