Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1996 Jan 9;93(1):367–371. doi: 10.1073/pnas.93.1.367

Phosphorylation-dependent human immunodeficiency virus type 1 infection and nuclear targeting of viral DNA.

A G Bukrinskaya 1, A Ghorpade 1, N K Heinzinger 1, T E Smithgall 1, R E Lewis 1, M Stevenson 1
PMCID: PMC40239  PMID: 8552640

Abstract

In the replication of human immunodeficiency virus type 1 (HIV-1), gag MA (matrix), a major structural protein of the virus, carries out opposing targeting functions. During virus assembly, gag MA is cotranslationally myristoylated, a modification required for membrane targeting of gag polyproteins. During virus infection, however, gag MA, by virtue of a nuclear targeting signal at its N terminus, facilitates the nuclear localization of viral DNA and establishment of the provirus. We now show that phosphorylation of gag MA on tyrosine and serine prior to and during virus infection facilitates its dissociation from the membrane, thus allowing it to translocate to the nucleus. Inhibition of gag MA phosphorylation either on tyrosine or on serine prevents gag MA-mediated nuclear targeting of viral nucleic acids and impairs virus infectivity. The requirement for gag MA phosphorylation in virus infection is underscored by our finding that a serine/threonine kinase is associated with virions of HIV-1. These results reveal a novel level of regulation of primate lentivirus infectivity.

Full text

PDF
367

Images in this article

368Fig. 1
on p.368
369Fig. 2
on p.369
370Fig. 3
on p.370
370Fig. 4
on p.370

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Akiyama T., Ishida J., Nakagawa S., Ogawara H., Watanabe S., Itoh N., Shibuya M., Fukami Y. Genistein, a specific inhibitor of tyrosine-specific protein kinases. J Biol Chem. 1987 Apr 25;262(12):5592–5595. [PubMed] [Google Scholar]
  2. Bowerman B., Brown P. O., Bishop J. M., Varmus H. E. A nucleoprotein complex mediates the integration of retroviral DNA. Genes Dev. 1989 Apr;3(4):469–478. doi: 10.1101/gad.3.4.469. [DOI] [PubMed] [Google Scholar]
  3. Boyle W. J., van der Geer P., Hunter T. Phosphopeptide mapping and phosphoamino acid analysis by two-dimensional separation on thin-layer cellulose plates. Methods Enzymol. 1991;201:110–149. doi: 10.1016/0076-6879(91)01013-r. [DOI] [PubMed] [Google Scholar]
  4. Brown P. O., Bowerman B., Varmus H. E., Bishop J. M. Correct integration of retroviral DNA in vitro. Cell. 1987 May 8;49(3):347–356. doi: 10.1016/0092-8674(87)90287-x. [DOI] [PubMed] [Google Scholar]
  5. Bryant M., Ratner L. Myristoylation-dependent replication and assembly of human immunodeficiency virus 1. Proc Natl Acad Sci U S A. 1990 Jan;87(2):523–527. doi: 10.1073/pnas.87.2.523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bukrinsky M. I., Haggerty S., Dempsey M. P., Sharova N., Adzhubel A., Spitz L., Lewis P., Goldfarb D., Emerman M., Stevenson M. A nuclear localization signal within HIV-1 matrix protein that governs infection of non-dividing cells. Nature. 1993 Oct 14;365(6447):666–669. doi: 10.1038/365666a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bukrinsky M. I., Sharova N., Dempsey M. P., Stanwick T. L., Bukrinskaya A. G., Haggerty S., Stevenson M. Active nuclear import of human immunodeficiency virus type 1 preintegration complexes. Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6580–6584. doi: 10.1073/pnas.89.14.6580. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Bukrinsky M. I., Sharova N., McDonald T. L., Pushkarskaya T., Tarpley W. G., Stevenson M. 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):6125–6129. doi: 10.1073/pnas.90.13.6125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Burnette B., Yu G., Felsted R. L. Phosphorylation of HIV-1 gag proteins by protein kinase C. J Biol Chem. 1993 Apr 25;268(12):8698–8703. [PubMed] [Google Scholar]
  10. Farnet C. M., Haseltine W. A. Determination of viral proteins present in the human immunodeficiency virus type 1 preintegration complex. J Virol. 1991 Apr;65(4):1910–1915. doi: 10.1128/jvi.65.4.1910-1915.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Folks T. M., Powell D., Lightfoote M., Koenig S., Fauci A. S., Benn S., Rabson A., Daugherty D., Gendelman H. E., Hoggan M. D. 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):280–290. doi: 10.1084/jem.164.1.280. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gallay P., Swingler S., Aiken C., Trono D. HIV-1 infection of nondividing cells: C-terminal tyrosine phosphorylation of the viral matrix protein is a key regulator. Cell. 1995 Feb 10;80(3):379–388. doi: 10.1016/0092-8674(95)90488-3. [DOI] [PubMed] [Google Scholar]
  13. Gheysen D., Jacobs E., de Foresta F., Thiriart C., Francotte M., Thines D., De Wilde M. Assembly and release of HIV-1 precursor Pr55gag virus-like particles from recombinant baculovirus-infected insect cells. Cell. 1989 Oct 6;59(1):103–112. doi: 10.1016/0092-8674(89)90873-8. [DOI] [PubMed] [Google Scholar]
  14. Göttlinger H. G., Sodroski J. G., Haseltine W. A. Role of capsid precursor processing and myristoylation in morphogenesis and infectivity of human immunodeficiency virus type 1. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5781–5785. doi: 10.1073/pnas.86.15.5781. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Heinzinger N. K., Bukrinsky M. I., Haggerty S. A., Ragland A. M., Kewalramani V., Lee M. A., Gendelman H. E., Ratner L., Stevenson M., Emerman M. 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):7311–7315. doi: 10.1073/pnas.91.15.7311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hidaka H., Inagaki M., Kawamoto S., Sasaki Y. Isoquinolinesulfonamides, novel and potent inhibitors of cyclic nucleotide dependent protein kinase and protein kinase C. Biochemistry. 1984 Oct 9;23(21):5036–5041. doi: 10.1021/bi00316a032. [DOI] [PubMed] [Google Scholar]
  17. Karageorgos L., Li P., Burrell C. Characterization of HIV replication complexes early after cell-to-cell infection. AIDS Res Hum Retroviruses. 1993 Sep;9(9):817–823. doi: 10.1089/aid.1993.9.817. [DOI] [PubMed] [Google Scholar]
  18. Kimpton J., Emerman M. Detection of replication-competent and pseudotyped human immunodeficiency virus with a sensitive cell line on the basis of activation of an integrated beta-galactosidase gene. J Virol. 1992 Apr;66(4):2232–2239. doi: 10.1128/jvi.66.4.2232-2239.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lee P. P., Linial M. L. Efficient particle formation can occur if the matrix domain of human immunodeficiency virus type 1 Gag is substituted by a myristylation signal. J Virol. 1994 Oct;68(10):6644–6654. doi: 10.1128/jvi.68.10.6644-6654.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lewis P. F., Emerman M. Passage through mitosis is required for oncoretroviruses but not for the human immunodeficiency virus. J Virol. 1994 Jan;68(1):510–516. doi: 10.1128/jvi.68.1.510-516.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lewis P., Hensel M., Emerman M. Human immunodeficiency virus infection of cells arrested in the cell cycle. EMBO J. 1992 Aug;11(8):3053–3058. doi: 10.1002/j.1460-2075.1992.tb05376.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lu Y. L., Spearman P., Ratner L. Human immunodeficiency virus type 1 viral protein R localization in infected cells and virions. J Virol. 1993 Nov;67(11):6542–6550. doi: 10.1128/jvi.67.11.6542-6550.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Maru Y., Peters K. L., Afar D. E., Shibuya M., Witte O. N., Smithgall T. E. Tyrosine phosphorylation of BCR by FPS/FES protein-tyrosine kinases induces association of BCR with GRB-2/SOS. Mol Cell Biol. 1995 Feb;15(2):835–842. doi: 10.1128/mcb.15.2.835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Mervis R. J., Ahmad N., Lillehoj E. P., Raum M. G., Salazar F. H., Chan H. W., Venkatesan S. The gag gene products of human immunodeficiency virus type 1: alignment within the gag open reading frame, identification of posttranslational modifications, and evidence for alternative gag precursors. J Virol. 1988 Nov;62(11):3993–4002. doi: 10.1128/jvi.62.11.3993-4002.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Pear W. S., Nolan G. P., Scott M. L., Baltimore D. Production of high-titer helper-free retroviruses by transient transfection. Proc Natl Acad Sci U S A. 1993 Sep 15;90(18):8392–8396. doi: 10.1073/pnas.90.18.8392. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Quick J., Ware J. A., Driedger P. E. The structure and biological activities of the widely used protein kinase inhibitor, H7, differ depending on the commercial source. Biochem Biophys Res Commun. 1992 Sep 16;187(2):657–663. doi: 10.1016/0006-291x(92)91245-l. [DOI] [PubMed] [Google Scholar]
  27. Roe T., Reynolds T. C., Yu G., Brown P. O. Integration of murine leukemia virus DNA depends on mitosis. EMBO J. 1993 May;12(5):2099–2108. doi: 10.1002/j.1460-2075.1993.tb05858.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Stevenson M., Haggerty S., Lamonica C., Mann A. M., Meier C., Wasiak A. Cloning and characterization of human immunodeficiency virus type 1 variants diminished in the ability to induce syncytium-independent cytolysis. J Virol. 1990 Aug;64(8):3792–3803. doi: 10.1128/jvi.64.8.3792-3803.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Taniguchi H., Manenti S. Interaction of myristoylated alanine-rich protein kinase C substrate (MARCKS) with membrane phospholipids. J Biol Chem. 1993 May 15;268(14):9960–9963. [PubMed] [Google Scholar]
  30. Thelen M., Rosen A., Nairn A. C., Aderem A. Regulation by phosphorylation of reversible association of a myristoylated protein kinase C substrate with the plasma membrane. Nature. 1991 May 23;351(6324):320–322. doi: 10.1038/351320a0. [DOI] [PubMed] [Google Scholar]
  31. Uehara Y., Fukazawa H. Use and selectivity of herbimycin A as inhibitor of protein-tyrosine kinases. Methods Enzymol. 1991;201:370–379. doi: 10.1016/0076-6879(91)01033-x. [DOI] [PubMed] [Google Scholar]
  32. Veronese F. D., Copeland T. D., Oroszlan S., Gallo R. C., Sarngadharan M. G. Biochemical and immunological analysis of human immunodeficiency virus gag gene products p17 and p24. J Virol. 1988 Mar;62(3):795–801. doi: 10.1128/jvi.62.3.795-801.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Yu M., Summers J. Phosphorylation of the duck hepatitis B virus capsid protein associated with conformational changes in the C terminus. J Virol. 1994 May;68(5):2965–2969. doi: 10.1128/jvi.68.5.2965-2969.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. von Schwedler U., Kornbluth R. S., Trono D. 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):6992–6996. doi: 10.1073/pnas.91.15.6992. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES