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PMID: 9463369 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Viral protein R regulates nuclear import of the HIV-1 pre-integration complex.

The EMBO journal ·Vol. 17 ·No. 4 ·1998-02-16 ·Pages 909-17

Popov S, Rexach M, Zybarth G, Reiling N, Lee MA, Ratner L, Lane CM, Moore MS, Blobel G, Bukrinsky M

Abstract

Replication of human immunodeficiency virus type 1 (HIV-1) in non-dividing cells critically depends on import of the viral pre-integration complex into the nucleus. Genetic evidence suggests that viral protein R (Vpr) and matrix antigen (MA) are directly involved in the import process. An in vitro assay that reconstitutes nuclear import of HIV-1 pre-integration complexes in digitonin-permeabilized cells was used to demonstrate that Vpr is the key regulator of the viral nuclear import process. Mutant HIV-1 pre-integration complexes that lack Vpr failed to be imported in vitro, whereas mutants that lack a functional MA nuclear localization sequence (NLS) were only partially defective. Strikingly, the import defect of the Vpr- mutant was rescued when recombinant Vpr was re-added. In addition, import of Vpr- virus was rescued by adding the cytosol of HeLa cells, where HIV-1 replication had been shown to be Vpr-independent. In a solution binding assay, Vpr associated with karyopherin alpha, a cellular receptor for NLSs. This association increased the affinity of karyopherin alpha for basic-type NLSs, including that of MA, thus explaining the positive effect of Vpr on nuclear import of the HIV-1 pre-integration complex and BSA-NLS conjugates. These results identify the biochemical mechanism of Vpr function in transport of the viral pre-integration complex to, and across, the nuclear membrane.

MeSH Terms
Biological Transport/drug effects,genetics Cell Membrane Permeability Cell Nucleus/genetics,metabolism,virology Cell-Free System/physiology Cytoplasm/physiology Defective Viruses/genetics,metabolism Digitonin/pharmacology Gene Products, gag/physiology Gene Products, vpr/genetics,metabolism,physiology HIV Antigens/physiology HIV-1/genetics HeLa Cells Humans Nuclear Proteins/metabolism Protein Binding/genetics Viral Proteins Virus Integration alpha Karyopherins gag Gene Products, Human Immunodeficiency Virus vpr Gene Products, Human Immunodeficiency Virus
Chemicals
Gene Products, gag Gene Products, vpr HIV Antigens Nuclear Proteins Viral Proteins alpha Karyopherins gag Gene Products, Human Immunodeficiency Virus p17 protein, Human Immunodeficiency Virus Type 1 vpr Gene Products, Human Immunodeficiency Virus Digitonin
Authors & Affiliations
10 authors, click to expand affiliations / ORCID
Popov S
The Picower Institute for Medical Research, Manhasset, NY 11030, USA.
Rexach M
Zybarth G
Reiling N
Lee M A
Ratner L
Lane C M
Moore M S
Blobel G
Bukrinsky M
References (62)
62 references, click to expand
  1. Protein import into nuclei: association and dissociation reactions involving transport substrate, transport factors, and nucleoporins.
    Cell. 1995 Dec 1;83(5):683-92 PMID: 8521485
  2. Retroviral integration: structure of the initial covalent product and its precursor, and a role for the viral IN protein.
    Proc Natl Acad Sci U S A. 1989 Apr;86(8):2525-9 PMID: 2539592
  3. Role of the karyopherin pathway in human immunodeficiency virus type 1 nuclear import.
    J Virol. 1996 Feb;70(2):1027-32 PMID: 8551560
  4. The binding site of karyopherin alpha for karyopherin beta overlaps with a nuclear localization sequence.
    Proc Natl Acad Sci U S A. 1996 Jun 25;93(13):6572-6 PMID: 8692858
  5. Molecular and functional characterization of the p62 complex, an assembly of nuclear pore complex glycoproteins.
    J Cell Biol. 1996 Aug;134(3):589-601 PMID: 8707840
  6. Importin provides a link between nuclear protein import and U snRNA export.
    Cell. 1996 Oct 4;87(1):21-32 PMID: 8858145
  7. Critical role of reverse transcriptase in the inhibitory mechanism of CNI-H0294 on HIV-1 nuclear translocation.
    Proc Natl Acad Sci U S A. 1996 Oct 15;93(21):11859-64 PMID: 8876228
  8. Identification of different roles for RanGDP and RanGTP in nuclear protein import.
    EMBO J. 1996 Oct 15;15(20):5584-94 PMID: 8896452
  9. In vivo nuclear transport kinetics in Saccharomyces cerevisiae: a role for heat shock protein 70 during targeting and translocation.
    J Cell Biol. 1996 Oct;135(2):329-39 PMID: 8896592
  10. RanBP1 stabilizes the interaction of Ran with p97 nuclear protein import.
    J Cell Biol. 1996 Nov;135(3):559-69 PMID: 8909533
  11. Nuclear import and cell cycle arrest functions of the HIV-1 Vpr protein are encoded by two separate genes in HIV-2/SIV(SM).
    EMBO J. 1996 Nov 15;15(22):6155-65 PMID: 8947037
  12. In vitro model for the nuclear transport of the hepadnavirus genome.
    J Virol. 1997 Feb;71(2):1310-6 PMID: 8995655
  13. Differential expression and sequence-specific interaction of karyopherin alpha with nuclear localization sequences.
    J Biol Chem. 1997 Feb 14;272(7):4310-5 PMID: 9020149
  14. Sequence requirements for nuclear location of simian virus 40 large-T antigen.
    Nature. 1984 Sep 6-11;311(5981):33-8 PMID: 6088992
  15. Biological and biochemical characterization of a cloned Leu-3- cell surviving infection with the acquired immune deficiency syndrome retrovirus.
    J Exp Med. 1986 Jul 1;164(1):280-90 PMID: 3014036
  16. Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone.
    J Virol. 1986 Aug;59(2):284-91 PMID: 3016298
  17. Correct integration of retroviral DNA in vitro.
    Cell. 1987 May 8;49(3):347-56 PMID: 3032450
  18. Isolation of cDNA coding for the placental protein 15 (PP15).
    Nucleic Acids Res. 1988 May 25;16(10):4721 PMID: 3380696
  19. The effects of variations in the number and sequence of targeting signals on nuclear uptake.
    J Cell Biol. 1988 Oct;107(4):1279-87 PMID: 3170630
  20. Integration of retroviral DNA.
    Curr Top Microbiol Immunol. 1990;157:19-48 PMID: 2203610
  21. Cytosolic proteins that specifically bind nuclear location signals are receptors for nuclear import.
    Cell. 1991 Sep 6;66(5):837-47 PMID: 1653647
  22. Human major HSP70 protein complements the localization and functional defects of cytoplasmic mutant SV40 T antigen in Swiss 3T3 mouse fibroblast cells.
    Genes Dev. 1991 Dec;5(12A):2235-44 PMID: 1748281
  23. Macrophage tropism determinants of human immunodeficiency virus type 1 in vivo.
    J Virol. 1992 Apr;66(4):2577-82 PMID: 1548783
  24. The transport of proteins into the nucleus requires the 70-kilodalton heat shock protein or its cytosolic cognate.
    Mol Cell Biol. 1992 May;12(5):2186-92 PMID: 1569948
  25. Dual regulation of silent and productive infection in monocytes by distinct human immunodeficiency virus type 1 determinants.
    J Virol. 1992 Jun;66(6):3925-31 PMID: 1533883
  26. The two steps of nuclear import, targeting to the nuclear envelope and translocation through the nuclear pore, require different cytosolic factors.
    Cell. 1992 Jun 12;69(6):939-50 PMID: 1606616
  27. Active nuclear import of human immunodeficiency virus type 1 preintegration complexes.
    Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6580-4 PMID: 1631159
  28. Human immunodeficiency virus infection of cells arrested in the cell cycle.
    EMBO J. 1992 Aug;11(8):3053-8 PMID: 1322294
  29. 70-kDa heat-shock cognate protein colocalizes with karyophilic proteins into the nucleus during their transport in vitro.
    Exp Cell Res. 1993 May;206(1):134-42 PMID: 8482354
  30. Antisense phosphorothioate oligodeoxynucleotides targeted to the vpr gene inhibit human immunodeficiency virus type 1 replication in primary human macrophages.
    J Virol. 1993 Jul;67(7):4409-14 PMID: 8510229
  31. Association of integrase, matrix, and reverse transcriptase antigens of human immunodeficiency virus type 1 with viral nucleic acids following acute infection.
    Proc Natl Acad Sci U S A. 1993 Jul 1;90(13):6125-9 PMID: 7687060
  32. Human immunodeficiency virus type 1 viral protein R localization in infected cells and virions.
    J Virol. 1993 Nov;67(11):6542-50 PMID: 8411357
  33. Human immunodeficiency virus type 1 2-LTR circles reside in a nucleoprotein complex which is different from the preintegration complex.
    J Virol. 1993 Nov;67(11):6863-5 PMID: 8411390
  34. The GTP-binding protein Ran/TC4 is required for protein import into the nucleus.
    Nature. 1993 Oct 14;365(6447):661-3 PMID: 8413630
  35. A nuclear localization signal within HIV-1 matrix protein that governs infection of non-dividing cells.
    Nature. 1993 Oct 14;365(6447):666-9 PMID: 8105392
  36. Inhibition of nuclear protein import by nonhydrolyzable analogues of GTP and identification of the small GTPase Ran/TC4 as an essential transport factor.
    J Cell Biol. 1993 Dec;123(6 Pt 2):1649-59 PMID: 8276887
  37. Signal- and energy-dependent nuclear transport of SV40 Vp3 by isolated nuclei. Establishment of a filtration assay for nuclear protein import.
    J Biol Chem. 1994 Feb 18;269(7):4910-6 PMID: 8106464
  38. HIV-1 infection of non-dividing cells.
    Nature. 1994 May 12;369(6476):107-8 PMID: 8192816
  39. Distinct effects in primary macrophages and lymphocytes of the human immunodeficiency virus type 1 accessory genes vpr, vpu, and nef: mutational analysis of a primary HIV-1 isolate.
    Virology. 1994 May 1;200(2):623-31 PMID: 8178448
  40. Biochemical mechanism of HIV-I Vpr function. Specific interaction with a cellular protein.
    J Biol Chem. 1994 Jun 3;269(22):15577-82 PMID: 8195203
  41. The nuclear localization signal of the matrix protein of human immunodeficiency virus type 1 allows the establishment of infection in macrophages and quiescent T lymphocytes.
    Proc Natl Acad Sci U S A. 1994 Jul 19;91(15):6992-6 PMID: 8041734
  42. The Vpr protein of human immunodeficiency virus type 1 influences nuclear localization of viral nucleic acids in nondividing host cells.
    Proc Natl Acad Sci U S A. 1994 Jul 19;91(15):7311-5 PMID: 8041786
  43. Vpr is required for efficient replication of human immunodeficiency virus type-1 in mononuclear phagocytes.
    Virology. 1995 Feb 1;206(2):935-44 PMID: 7531918
  44. Identification of a protein complex that is required for nuclear protein import and mediates docking of import substrate to distinct nucleoporins.
    Proc Natl Acad Sci U S A. 1995 Feb 28;92(5):1769-73 PMID: 7878057
  45. Separate domains of the Ran GTPase interact with different factors to regulate nuclear protein import and RNA processing.
    Mol Cell Biol. 1995 Apr;15(4):2117-24 PMID: 7891706
  46. Previously identified protein of uncertain function is karyopherin alpha and together with karyopherin beta docks import substrate at nuclear pore complexes.
    Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):2008-11 PMID: 7892216
  47. A karyophilic protein forms a stable complex with cytoplasmic components prior to nuclear pore binding.
    J Biol Chem. 1995 Apr 14;270(15):8559-65 PMID: 7721756
  48. Role of the basic domain of human immunodeficiency virus type 1 matrix in macrophage infection.
    J Virol. 1995 Jun;69(6):3949-54 PMID: 7745752
  49. Identification of hSRP1 alpha as a functional receptor for nuclear localization sequences.
    Science. 1995 May 19;268(5213):1049-53 PMID: 7754385
  50. Mammalian karyopherin alpha 1 beta and alpha 2 beta heterodimers: alpha 1 or alpha 2 subunit binds nuclear localization signal and beta subunit interacts with peptide repeat-containing nucleoporins.
    Proc Natl Acad Sci U S A. 1995 Jul 3;92(14):6532-6 PMID: 7604027
  51. Sequence and characterization of cytoplasmic nuclear protein import factor p97.
    J Cell Biol. 1995 Jul;130(2):265-74 PMID: 7615630
  52. Two different subunits of importin cooperate to recognize nuclear localization signals and bind them to the nuclear envelope.
    Curr Biol. 1995 Apr 1;5(4):383-92 PMID: 7627554
  53. The essential yeast nucleoporin NUP159 is located on the cytoplasmic side of the nuclear pore complex and serves in karyopherin-mediated binding of transport substrate.
    J Biol Chem. 1995 Aug 11;270(32):19017-21 PMID: 7642562
  54. Mutagenic analysis of human immunodeficiency virus type 1 Vpr: role of a predicted N-terminal alpha-helical structure in Vpr nuclear localization and virion incorporation.
    J Virol. 1995 Nov;69(11):7032-44 PMID: 7474123
  55. Mutational analysis of cell cycle arrest, nuclear localization and virion packaging of human immunodeficiency virus type 1 Vpr.
    J Virol. 1995 Dec;69(12):7909-16 PMID: 7494303
  56. Nuclear localization signal of HIV-1 as a novel target for therapeutic intervention.
    Mol Med. 1995 Jan;1(2):217-30 PMID: 8529100
  57. Human immunodeficiency virus type 1 preintegration complexes: studies of organization and composition.
    J Virol. 1997 Jul;71(7):5382-90 PMID: 9188609
  58. Cell proliferation is not required for productive HIV-1 infection of macrophages.
    Virology. 1997 Jun 9;232(2):379-84 PMID: 9191852
  59. Nuclear import, virion incorporation, and cell cycle arrest/differentiation are mediated by distinct functional domains of human immunodeficiency virus type 1 Vpr.
    J Virol. 1997 Sep;71(9):6339-47 PMID: 9261351
  60. HIV-1 infection of nondividing cells through the recognition of integrase by the importin/karyopherin pathway.
    Proc Natl Acad Sci U S A. 1997 Sep 2;94(18):9825-30 PMID: 9275210
  61. HIV-1 infection of non-dividing cells: evidence that the amino-terminal basic region of the viral matrix protein is important for Gag processing but not for post-entry nuclear import.
    EMBO J. 1997 Aug 1;16(15):4531-9 PMID: 9303297
  62. The GLFG repetitive region of the nucleoporin Nup116p interacts with Kap95p, an essential yeast nuclear import factor.
    J Cell Biol. 1995 Dec;131(6 Pt 2):1699-713 PMID: 8557738
Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
DOI
10.1093/emboj/17.4.909
Published
1998-02-16
Pages
909-17
Language
English
Region
England
NLM ID
8208664
PMCID
PMC1170440
Subset
IM
Grants
NIAID NIH HHS · AI 33776 · United States
NIAID NIH HHS · AI 36071 · United States
External Links
PubMed source DOI: 10.1093/emboj/17.4.909 PMC Full Text View journal details
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