scholarly journals Molecular dissection of an inhibitor targeting the HIV integrase dependent preintegration complex nuclear import

2018 ◽  
Vol 21 (1) ◽  
pp. e12953 ◽  
Author(s):  
Kylie M. Wagstaff ◽  
Stephen Headey ◽  
Sushama Telwatte ◽  
David Tyssen ◽  
Anna C. Hearps ◽  
...  
Keyword(s):  
Nuclear Import ◽  
Hiv Integrase ◽  
Molecular Dissection ◽  
Preintegration Complex

scholarly journals Importin α3 Interacts with HIV-1 Integrase and Contributes to HIV-1 Nuclear Import and Replication

2010 ◽  
Vol 84 (17) ◽  
pp. 8650-8663 ◽  
Author(s):  
Zhujun Ao ◽  
Kallesh Danappa Jayappa ◽  
Binchen Wang ◽  
Yingfeng Zheng ◽  
Sam Kung ◽  
...  
Keyword(s):  
T Cells ◽  
Nuclear Import ◽  
Expression System ◽  
293T Cell ◽  
Cellular Protein ◽  
Pcr Analysis ◽  
Hiv Integrase ◽  
Preintegration Complex ◽  
Importin Α ◽  
Hiv 1

ABSTRACT HIV-1 employs the cellular nuclear import machinery to actively transport its preintegration complex (PIC) into the nucleus for integration of the viral DNA. Several viral karyophilic proteins and cellular import factors have been suggested to contribute to HIV-1 PIC nuclear import and replication. However, how HIV interacts with different cellular machineries to ensure efficient nuclear import of its preintegration complex in dividing and nondividing cells is still not fully understood. In this study, we have investigated different importin α (Impα) family members for their impacts on HIV-1 replication, and we demonstrate that short hairpin RNA (shRNA)-mediated Impα3 knockdown (KD) significantly impaired HIV infection in HeLa cells, CD4+ C8166 T cells, and primary macrophages. Moreover, quantitative real-time PCR analysis revealed that Impα3-KD resulted in significantly reduced levels of viral 2-long-terminal repeat (2-LTR) circles but had no effect on HIV reverse transcription. All of these data indicate an important role for Impα3 in HIV nuclear import. In an attempt to understand how Impα3 participates in HIV nuclear import and replication, we first demonstrated that the HIV-1 karyophilic protein integrase (IN) was able to interact with Impα3 both in a 293T cell expression system and in HIV-infected CD4+ C8166 T cells. Deletion analysis suggested that a region (amino acids [aa] 250 to 270) in the C-terminal domain of IN is involved in this viral-cellular protein interaction. Overall, this study demonstrates for the first time that Impα3 is an HIV integrase-interacting cofactor that is required for efficient HIV-1 nuclear import and replication in both dividing and nondividing cells.

scholarly journals Factors that mold the nuclear landscape of HIV-1 integration

2020 ◽  
Author(s):  
Gregory J Bedwell ◽  
Alan N Engelman
Keyword(s):  
Nuclear Import ◽  
Integration Site ◽  
Preintegration Complex ◽  
Retroviral Integration ◽  
Host Interactions ◽  
Aids Pandemic ◽  
Nuclear Environment ◽  
Site Targeting ◽  
Viral Dna Integration ◽  
Hiv 1

Abstract The integration of retroviral reverse transcripts into the chromatin of the cells that they infect is required for virus replication. Retroviral integration has far-reaching consequences, from perpetuating deadly human diseases to molding metazoan evolution. The lentivirus human immunodeficiency virus 1 (HIV-1), which is the causative agent of the AIDS pandemic, efficiently infects interphase cells due to the active nuclear import of its preintegration complex (PIC). To enable integration, the PIC must navigate the densely-packed nuclear environment where the genome is organized into different chromatin states of varying accessibility in accordance with cellular needs. The HIV-1 capsid protein interacts with specific host factors to facilitate PIC nuclear import, while additional interactions of viral integrase, the enzyme responsible for viral DNA integration, with cellular nuclear proteins and nucleobases guide integration to specific chromosomal sites. HIV-1 integration favors transcriptionally active chromatin such as speckle-associated domains and disfavors heterochromatin including lamina-associated domains. In this review, we describe virus-host interactions that facilitate HIV-1 PIC nuclear import and integration site targeting, highlighting commonalities among factors that participate in both of these steps. We moreover discuss how the nuclear landscape influences HIV-1 integration site selection as well as the establishment of active versus latent virus infection.

scholarly journals The C-Terminal Proline-Rich Tail of Human Immunodeficiency Virus Type 2 Vpx Is Necessary for Nuclear Localization of the Viral Preintegration Complex in Nondividing Cells

2000 ◽  
Vol 74 (13) ◽  
pp. 6162-6167 ◽  
Author(s):  
Heather A. Pancio ◽  
Nancy Vander Heyden ◽  
Lee Ratner
Keyword(s):  
Human Immunodeficiency Virus ◽  
Fusion Protein ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Localization ◽  
Nuclear Import ◽  
Preintegration Complex ◽  
Immunodeficiency Virus ◽  
Infected Cells

ABSTRACT Human immunodeficiency virus type 2 (HIV-2), like other lentiviruses, is capable of infecting nondividing T cells and macrophages. The present work shows that in HIV-2-infected cells, Vpx is necessary for efficient nuclear import of the preintegration complex. In agreement with this finding, the subcellular localization of a GFP-Vpx fusion protein was found to be predominantly nuclear. However, deletion of the proline-rich C-terminal 11 residues of Vpx resulted in a shift of the fusion protein to the cytoplasm. Furthermore, the same deletion in the context of the provirus resulted in a decrease in nuclear import of the preintegration complex and attenuated replication in macrophages.

scholarly journals Nuclear Import of the Preintegration Complex Is Blocked upon Infection by Human Immunodeficiency Virus Type 1 in Mouse Cells

2006 ◽  
Vol 81 (2) ◽  
pp. 677-688 ◽  
Author(s):  
Naomi Tsurutani ◽  
Jiro Yasuda ◽  
Naoki Yamamoto ◽  
Byung-Il Choi ◽  
Motohiko Kadoki ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Nuclear Localization ◽  
Reverse Transcription ◽  
Nuclear Import ◽  
Viral Dna ◽  
Preintegration Complex ◽  
Immunodeficiency Virus ◽  
Mouse Cells ◽  
Hiv 1

ABSTRACT Mouse cells do not support human immunodeficiency virus type 1 (HIV-1) replication because of host range barriers at steps including virus entry, transcription, RNA splicing, polyprotein processing, assembly, and release. The exact mechanisms for the suppression, however, are not completely understood. To elucidate further the barriers against HIV-1 replication in mouse cells, we analyzed the replication of the virus in lymphocytes from human CD4/CXCR4 transgenic mice. Although primary splenocytes and thymocytes allowed the entry and reverse transcription of HIV-1, the integration efficiency of the viral DNA was greatly reduced in these cells relative to human peripheral blood mononuclear cells, suggesting an additional block(s) before or at the point of host chromosome integration of the viral DNA. Preintegration processes were further analyzed using HIV-1 pseudotyped viruses. The reverse transcription step of HIV-1 pseudotyped with the envelope of murine leukemia virus or vesicular stomatitis virus glycoprotein was efficiently supported in both human and mouse cells, but nuclear import of the preintegration complex (PIC) of HIV-1 was blocked in mouse cells. We found that green fluorescent protein (GFP)-labeled HIV-1 integrase, which is known to be important in the nuclear localization of the PIC, could not be imported into the nucleus of mouse cells, in contrast to human cells. On the other hand, GFP-Vpr localized exclusively to the nuclei of both mouse and human cells. These observations suggest that, due to the dysfunction of integrase, the nuclear localization of PIC is suppressed in mouse cells.

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

1998 ◽  
Vol 143 (4) ◽  
pp. 875-885 ◽  
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.

Heat-Shock Protein 70 Can Replace Viral Protein R of HIV-1 during Nuclear Import of the Viral Preintegration Complex

2000 ◽  
Vol 259 (2) ◽  
pp. 398-403 ◽  
Author(s):  
Isabelle Agostini ◽  
Serguei Popov ◽  
Jianhua Li ◽  
Larisa Dubrovsky ◽  
Tang Hao ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Nuclear Import ◽  
Viral Protein ◽  
Heat Shock Protein 70 ◽  
Preintegration Complex ◽  
Viral Protein R ◽  
Hiv 1

scholarly journals The First HxRxG Motif in Simian Immunodeficiency Virus mac239 Vpr Is Crucial for G2/M Cell Cycle Arrest

2002 ◽  
Vol 76 (22) ◽  
pp. 11704-11709 ◽  
Author(s):  
Sandra M. Mueller ◽  
Sabine M. Lang
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Simian Immunodeficiency Virus ◽  
Nuclear Import ◽  
M Cell ◽  
Preintegration Complex ◽  
Cycle Arrest ◽  
Immunodeficiency Virus ◽  
Helical Region

ABSTRACT The highly conserved Vpr protein mediates cell cycle arrest, transcriptional transactivation, and nuclear import of the preintegration complex in human immunodeficiency virus type 1. To identify functional domains in simian immunodeficiency virus (SIV) mac239 Vpr, we mutagenized selected motifs within an α-helical region and two C-terminal HxRxG motifs. All Vpr mutants located to the nucleus. Substitution of four amino acids in the α-helical domain did not interfere with cell cycle arrest, while a single substitution abolished cell cycle arrest function. Mutation of the first HxRxG motif to AxAxA also resulted in loss of cell cycle arrest, while mutation of the second motif had no effect. Interestingly, both Vpr mutants impaired in cell cycle arrest function also showed reduced transactivation of the SIV long terminal repeat, suggesting that arrest of cells at G2/M mediates or contributes to transactivation by Vpr.

scholarly journals HumanImmunodeficiency Virus Type 1 DNA Nuclear Import and Integration AreMitosis Independent in CyclingCells

2003 ◽  
Vol 77 (24) ◽  
pp. 13412-13417 ◽  
Author(s):  
Richard A. Katz ◽  
James G. Greger ◽  
Pamela Boimel ◽  
Anna Marie Skalka
Keyword(s):  
Virus Type ◽  
Nuclear Import ◽  
Replication Fork ◽  
Integration Site ◽  
Nuclear Entry ◽  
Preintegration Complex ◽  
Daughter Cells ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT An essential step in human immunodeficiency virus type 1 (HIV-1) replication is the movement of the viral preintegration complex from the cytoplasm into the nucleus. The pathway(s) and timing for HIV-1 DNA nuclear entry in cycling cells have not been established. Here, we show that if cycling cells are infected before S phase, viral DNA can be integrated prior to passage of the host DNA replication fork through the integration site, as indicated by stable inheritance in both daughter cells. We conclude that efficient nuclear entry can occur independently of mitotic nuclear disassembly in cycling cells.

scholarly journals Hsp40 Facilitates Nuclear Import of the Human Immunodeficiency Virus Type 2 Vpx-Mediated Preintegration Complex

2007 ◽  
Vol 82 (3) ◽  
pp. 1229-1237 ◽  
Author(s):  
Xiaogang Cheng ◽  
Michael Belshan ◽  
Lee Ratner
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Import ◽  
Nuclear Translocation ◽  
Down Regulation ◽  
Preintegration Complex ◽  
Immunodeficiency Virus ◽  
Interfering Rna

ABSTRACT Human immunodeficiency virus type 2 (HIV-2) Vpx is required for nuclear translocation of the viral preintegration complex (PIC) in quiescent cells. In order to decipher the mechanism of action of Vpx, a cDNA library was screened with the yeast two-hybrid assay, resulting in the identification of heat shock protein 40, Hsp40/DnaJB6, as a Vpx-interactive protein. Interaction with Vpx was confirmed by glutathione S-transferase (GST) pull-down and coimmunoprecipitation assays. Overexpression of Hsp40/DnaJB6 enhanced Vpx nuclear import, whereas overexpression of a nuclear localization mutant of Hsp40/DnaJB6 (H31Q) or down-regulation of Hsp40/DnaJB6 by small interfering RNA (siRNA) reduced the nuclear import of Vpx. Hsp40/DnaJB6 competed with the Pr55Gag precursor protein for the binding of Vpx and incorporation into virus-like particles. Overexpression of Hsp40/DnaJB6 promoted viral PIC nuclear import, whereas siRNA down-regulation of Hsp40/DnaJB6 inhibited PIC nuclear import. These results demonstrate a role for Hsp40/DnaJB6 in the regulation of HIV-2 PIC nuclear transport.

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