1887

Abstract

The primary human immunodeficiency virus type 1 (HIV-1) Nef mutant F12-HIVNef is characterized by three rare amino acid substitutions, GE, VL and EG. It was reported previously that the expression of F12-HIVNef in the context of the highly productive NL4-3 HIV-1 strain blocks virus replication at the level of virus assembly and/or release by a mechanism depending on the presence of the CD4 intracytoplasmic tail. Here, it is reported that NL4-3 HIV-1 strains expressing F12-HIV alleles that were back-mutated in each amino acid substitution readily replicated in CD4 cells. Attempting to correlate possible functional alterations with antiviral effects, both F12-HIVNef and its back mutants were tested in terms of well-characterized markers of Nef expression. Both F12-HIVNef and its GE back mutant did not down-regulate CD4 as the consequence of a greatly reduced rate of CD4 internalization. On the other hand, F12-HIVNef as well as the EG and LV back mutants failed to activate the p62 Nef-associated kinase (p62NAK). Thus, only F12-HIVNef was defective in both accelerated rates of CD4 internalization and p62NAK activation, whereas at least one Nef function was restored in all of the back mutants. Infection of cells expressing Nef-resistant CD4 molecules with HIV-1 strains encoding F12-HIVNef back mutants showed that both the lack of accelerated CD4 endocytosis and an, as yet, still unidentified function are required for the F12-HIVNef inhibitory phenotype. These results provide a detailed functional analysis of the F12-HIV allele and support the idea that both CD4 accelerated internalization and p62NAK activation are part of the essential steps in the virus replication cycle.

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2001-11-01
2024-03-28
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References

  1. Aiken C., Konner J., Landau N. R., Lenburg M. E., Trono D. 1994; Nef induces CD4 endocytosis: requirement for a critical dileucine motif in the membrane-proximal CD4 cytoplasmic domain. Cell 75:853–864
    [Google Scholar]
  2. Arora V. K., Molina R. P., Foster J. L., Blakemore J. L., Chernoff J., Fredericksen B. L., Garcia J. V. 2000; Lentivirus Nef specifically activates Pak2. Journal of Virology 74:11081–11087
    [Google Scholar]
  3. Bell I., Ashman C., Maughan J., Hooker E., Cook F., Reinhart T. A. 1998; Association of simian immunodeficiency virus Nef with the T-cell receptor (TCR) ζ chain leads to TCR down-modulation. Journal of General Virology 79:2717–2727
    [Google Scholar]
  4. Brambilla A., Turchetto L., Gatti A., Bovolenta C., Veglia F., Santagostino E., Gringeri A., Clementi M., Poli G., Bagnarelli P., Vicenzi E. 1999; Defective nef alleles in a cohort of hemophiliacs with progressing and nonprogressing HIV-1 infection. Virology 259:349–368
    [Google Scholar]
  5. Bryant M. L., Ratner L., Duronio R. J., Kishore N. S., Devadas B., Adams S. P., Grodon J. I. 1991; Incorporation of 12-methoxydodecanoate into the human immunodeficiency virus 1 Gag polyprotein precursor inhibits its proteolytic processing and virus production in a chronically infected human lymphoid cell line. Proceedings of the National Academy of Sciences, USA 88:2055–2059
    [Google Scholar]
  6. Carlini F., Federico M., Equestre M., Ricci S., Ratti G., Zibai Q., Verani P., Rossi G. B. 1992; Sequence analysis of an HIV-1 proviral DNA from a non-producer chronically infected Hut-78 cellular clone. Journal of Virological Diseases 1:40–55
    [Google Scholar]
  7. Chowers M. Y., Spina C. A., Kwoh T. J., Fitch N. J. S., Richman D. D., Guatelli J. C. 1994; Optimal infectivity in vitro of human immunodeficiency virus type 1 requires an intact nef gene. Journal of Virology 68:2906–2914
    [Google Scholar]
  8. Cohen G. B., Gandhi R. T., Davis D. M., Mandelboim O., Chen B. K., Strominger J. L., Baltimore D. 1999; The selective downregulation of class I major histocompatibility complex proteins by HIV-1 protects HIV-infected cells from NK cells. Immunity 10:661–671
    [Google Scholar]
  9. Collins K. L., Chen B. K., Kalams S. A., Walker B. D., Baltimore D. 1998; HIV-1 Nef protein protects infected primary cells against killing by cytotoxic T lymphocytes. Nature 391:397–401
    [Google Scholar]
  10. Craig H. M., Pandori M. W., Guatelli J. C. 1998; Interaction of HIV-1 Nef with the cellular dileucine-based sorting pathway is required for CD4 down-regulation and optimal viral infectivity. Proceedings of the National Academy of Sciences, USA 95:11229–11234
    [Google Scholar]
  11. D’Aloja P., Olivetta E., Bona R., Nappi F., Pedacchia D., Pugliese K., Ferrari G., Verani P., Federico M. 1998; gag , vif , and nef genes contribute to the homologous viral interference induced by a nonproducer human immunodeficiency virus type 1 (HIV-1) variant: identification of novel HIV–1-inhibiting viral protein mutants. Journal of Virology 72:4308–4319
    [Google Scholar]
  12. Fackler O. T., Luo W., Geyer M., Alberts A. S., Peterlin B. M. 1999; Activation of Vav by Nef induces cytoskeletal rearrangements and downstream effector functions. Molecular Cell 3:729–739
    [Google Scholar]
  13. Fackler O. T., Lu X., Frost J. A. A., Geyer M., Jiang B., Luo W., Abo A., Alberts A. S., Peterlin B. M. 2000; p21-activated kinase 1 plays a critical role in cellular activation by Nef. Molecular and Cellular Biology 20:2619–2627
    [Google Scholar]
  14. Federico M., Taddeo B., Carlini F., Nappi F., Verani P., Rossi G. B. 1993; A recombinant retrovirus carrying a non-producer human immunodeficiency virus (HIV) type 1 variant induces resistance to superinfecting HIV. Journal of General Virology 74:2099–2110
    [Google Scholar]
  15. Franken P., Arold S., Padilla A., Bodeus M., Hoh F., Strub M. P., Boyer M., Jullien M., Benarous R., Dumas C. 1997; HIV-1 Nef protein: purification, crystallization, and preliminary X-ray diffraction studies. Protein Science 6:2681–2683
    [Google Scholar]
  16. Greenberg M. E., Bronson S., Lock M., Neumann M., Pavlakis G. N., Skowronski J. 1997; Co-localization of HIV-1 Nef with the AP-2 adaptor protein complex correlates with Nef-induced CD4 down-regulation. EMBO Journal 16:6964–6976
    [Google Scholar]
  17. Greenberg M. E., Iafrate A. J., Skowronski J. 1998; The SH3 domain-binding surface and an acidic motif in HIV-1 Nef regulate trafficking of class I MHC complexes. EMBO Journal 17:2777–2789
    [Google Scholar]
  18. Grzesiek S., Bax A., Hu J. S., Kaufman J., Palmer I., Stahl S. J., Tjandra N., Wingfield P. T. 1997; Refined solution structure and backbone dynamics of HIV-1 Nef. Protein Science 6:1248–1263
    [Google Scholar]
  19. Khan I. H., Sawai E. T., Antonio E., Weber C. J., Mandell C. P., Montbriand P., Luciw P. A. 1998; Role of the SH3-ligand domain of simian immunodeficiency virus Nef in interaction with Nef-associated kinase and simian AIDS in rhesus macaques. Journal of Virology 72:5820–5830
    [Google Scholar]
  20. Lama J., Mangasarian A., Trono D. 1999; Cell-surface expression of CD4 reduces HIV-1 infectivity by blocking Env incorporation in a Nef- and Vpu-inhibitable manner. Current Biology 9:622–631
    [Google Scholar]
  21. Lee C. H., Saksela K., Mirza U. A., Chait B. T., Kuriyan J. 1996; Crystal structure of the conserved core of HIV-1 Nef complexed with a Src family SH3 domain. Cell 85:931–942
    [Google Scholar]
  22. Le Gall S., Erdtmann L., Benichou S., Berlioz-Torrent C., Liu L., Benarous R., Heard J. M., Schwartz O. 1998; Nef interacts with the mu subunit of clathrin adaptor complexes and reveals a cryptic sorting signal in MHC I molecules. Immunity 8:483–495
    [Google Scholar]
  23. Lu X., Wu X., Plemenitas A., Yu H., Sawai E. T., Abo A., Peterlin B. M. 1996; CDC42 and Rac1 are implicated in the activation of the Nef-associated kinase and replication of HIV-1. Current Biology 6:1677–1684
    [Google Scholar]
  24. Lu X., Yu H., Brodsky F. M., Peterlin B. M. 1998; Interactions between HIV-1 Nef and vacuolar ATPase facilitate the internalization of CD4. Immunity 8:647–656
    [Google Scholar]
  25. Luo T., Livingston R. A., Garcia J. V. 1997; Infectivity enhancement by human immunodeficiency virus type 1 Nef is independent of its association with a cellular serine/threonine kinase. Journal of Virology 71:9524–9530
    [Google Scholar]
  26. Mangasarian A., Foti M., Aiken C., Chin D., Carpentier J. L., Trono D. 1997; The HIV-1 Nef protein acts as a connector with sorting pathways in the Golgi and at the plasma membrane. Immunity 6:67–77
    [Google Scholar]
  27. Mangasarian A., Piguet V., Wang J. K., Chen Y. L., Trono D. 1999; Nef-induced CD4 and major histocompatibility complex class I (MHC-I) down-regulation are governed by distinct determinants: N-terminal α-helix and proline repeat of Nef selectively regulate MHC-I trafficking. Journal of Virology 73:1964–1973
    [Google Scholar]
  28. Manninen A., Hiipakka M., Vihinen M., Lu W., Mayer B. J., Saksela K. 1998; SH3-domain binding function of HIV-1 Nef is required for association with a PAK-related kinase. Virology 250:273–282
    [Google Scholar]
  29. Marsh M., Pelchen-Matthews A. 1996; Endocytic and exocytic regulation of CD4 expression and function. Current Topics in Microbiology and Immunology 205:107–135
    [Google Scholar]
  30. Miller M., Warmerdam M. T., Gaston I., Greene W. C., Feinberg M. B. 1994; The human immunodeficiency virus-1 nef gene product: a positive factor for viral infection and replication in primary lymphocytes and macrophages. Journal of Experimental Medicine 179:101–113
    [Google Scholar]
  31. Nunn M. F., Marsh J. W. 1996; Human immunodeficiency virus type 1 Nef associates with a member of the p21-activated kinase family. Journal of Virology 70:6157–6161
    [Google Scholar]
  32. Olivetta E., Pugliese K., Bona R., D’Aloja P., Ferrantelli F., Santarcangelo A. C., Mattia G., Verani P., Federico M. 2000; cis expression of the F12 human immunodeficiency virus (HIV) Nef allele transforms the highly productive NL4-3 HIV type 1 to a replication-defective strain: involvement of both Env gp41 and CD4 intracytoplasmic tails. Journal of Virology 74:483–492
    [Google Scholar]
  33. Palm G. J., Zdanov A., Gaitanaris G. A., Stauber R., Pavlakis G. N., Wlodawer A. 1997; The structural basis for spectral variation in green fluorescent protein. Nature Structural Biology 4:361–365
    [Google Scholar]
  34. Piguet V., Chen Y. L., Mangasarian A., Foti M., Carpentier J. L., Trono D. 1998; Mechanism of Nef-induced CD4 endocytosis: Nef connects CD4 with the mu chain of adaptor complexes. EMBO Journal 17:2472–2481
    [Google Scholar]
  35. Piguet V., Gu F., Foti M., Demaurex N., Gruenberg J., Carpentier J. L., Trono D. 1999; Nef-induced CD4 degradation: a diacidic-based motif in Nef functions as a lysosomal targeting signal through the binding of β-COP in endosomes. Cell 97:63–73
    [Google Scholar]
  36. Renkema G. H., Manninen A., Mann D. A., Harris M., Saksela K. 1999; Identification of the Nef-associated kinase as p21-activated kinase 2. Current Biology 9:1407–1410
    [Google Scholar]
  37. Ross T. M., Oran A. E., Cullen B. R. 1999; Inhibition of HIV-1 progeny virion release by cell-surface CD4 is relieved by expression of the viral Nef protein. Current Biology 9:613–621
    [Google Scholar]
  38. Rossi G. B., Verani P., Macchi B., Federico M., Orecchia A., Nicoletti L., Buttò S., Lazzarin A., Mariani G., Ippolito G., Manzari V. 1987; Recovery of HIV-related retroviruses from Italian patients with AIDS or AIDS-related complex and from asymptomatic at-risk individuals. Annals of the New York Academy of Sciences 511:390–400
    [Google Scholar]
  39. Saksela K. 1997; HIV-1 Nef and host cell protein kinases. Frontiers of Bioscience 2:606–618
    [Google Scholar]
  40. Sawai E. T., Baur A., Struble H., Peterlin B. M., Levy J. A., Cheng-Mayer C. 1994; Human immunodeficiency virus type 1 Nef associates with a cellular serine kinase in T lymphocytes. Proceedings of the National Academy of Sciences, USA 91:1539–1543
    [Google Scholar]
  41. Sawai E. T., Baur A. S., Peterlin B. M., Levy J. A., Cheng-Mayer C. 1995; A conserved domain and membrane targeting of Nef from HIV and SIV are required for association with a cellular serine kinase activity. Journal of Biological Chemistry 270:15307–15314
    [Google Scholar]
  42. Sawai E. T., Khan I. H., Montbriand P. M., Peterlin B. M., Cheng-Mayer C., Luciw P. A. 1996; Activation of PAK by HIV and SIV Nef: importance for AIDS in rhesus macaques. Current Biology 6:1519–1527
    [Google Scholar]
  43. Schwartz O., Maréchal V., Le Gall S., Lemonnier F., Heard J. M. 1996; Endocytosis of major histocompatibility complex class I molecules is induced by the HIV-1 Nef protein. Nature Medicine 2:338–342
    [Google Scholar]
  44. Spina C. A., Kwoh T. J., Chowers M. Y., Guatelli J. C., Richman D. D. 1994; The importance of nef in the induction of human immunodeficiency virus type 1 replication from primary quiescent CD4 lymphocytes. Journal of Experimental Medicine 179:115–123
    [Google Scholar]
  45. Taddeo B., Carlini F., Verani P., Engelman A. 1996; Reversion of a human immunodeficiency virus type 1 integrase mutant at a second site restores enzyme function and virus infectivity. Journal of Virology 70:8277–8284
    [Google Scholar]
  46. Wigler M., Sweet R., Sim G. K., Wold B., Pellicer A., Lacy E., Maniatis T., Silverstein S., Awel R. 1979; Transformation of mammalian cells with genes from procaryotes and eucaryotes. Cell 16:758–777
    [Google Scholar]
  47. Xu X. N., Laffert B., Screaton G. R., Kraft M., Wolf D., Kolanus W., Mongkolsapai J., McMichael A. J., Baur A. S. 1999; Induction of Fas ligand expression by HIV involves the interaction of Nef with the T cell receptor ζ chain. Journal of Experimental Medicine 189:1489–1496
    [Google Scholar]
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