Search tips
Search criteria 


Logo of jvirolPermissionsJournals.ASM.orgJournalJV ArticleJournal InfoAuthorsReviewers
J Virol. 1993 August; 67(8): 4639–4650.
PMCID: PMC237849

Analysis of human immunodeficiency virus type 1 nef gene sequences present in vivo.


The nef genes of the human immunodeficiency viruses type 1 and 2 (HIV-1 and HIV-2) and the related simian immunodeficiency viruses (SIVs) encode a protein (Nef) whose role in virus replication and cytopathicity remains uncertain. As an attempt to elucidate the function of nef, we characterized the nucleotide and corresponding protein sequences of naturally occurring nef genes obtained from several HIV-1-infected individuals. A consensus Nef sequence was derived and used to identify several features that were highly conserved among the Nef sequences. These features included a nearly invariant myristylation signal, regions of sequence polymorphism and variable duplication, a region with an acidic charge, a (Pxx)4 repeat sequence, and a potential protein kinase C phosphorylation site. Clustering of premature stop codons at position 124 was noted in 6 of the 54 Nef sequences. Further analysis revealed four stretches of residues that were highly conserved not only among the patient-derived HIV-1 Nef sequences, but also among the Nef sequences of HIV-2 and the SIVs, suggesting that Nef proteins expressed by these retroviruses are functionally equivalent. The "Nef-defining" sequences were used to evaluate the sequence alignments of known proteins reported to share sequence similarity with Nef sequences and to conduct additional computer-based searches for similar protein sequences. A gene encoding the consensus Nef sequence was also generated. This gene encodes a full-length Nef protein that should be a valuable tool in further studies of Nef function.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (2.6M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Adachi A, Gendelman HE, Koenig S, Folks T, Willey R, Rabson A, Martin MA. Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone. J Virol. 1986 Aug;59(2):284–291. [PMC free article] [PubMed]
  • Allan JS, Coligan JE, Lee TH, McLane MF, Kanki PJ, Groopman JE, Essex M. A new HTLV-III/LAV encoded antigen detected by antibodies from AIDS patients. Science. 1985 Nov 15;230(4727):810–813. [PubMed]
  • Anderson S, Shugars DC, Swanstrom R, Garcia JV. Nef from primary isolates of human immunodeficiency virus type 1 suppresses surface CD4 expression in human and mouse T cells. J Virol. 1993 Aug;67(8):4923–4931. [PMC free article] [PubMed]
  • Balfe P, Simmonds P, Ludlam CA, Bishop JO, Brown AJ. Concurrent evolution of human immunodeficiency virus type 1 in patients infected from the same source: rate of sequence change and low frequency of inactivating mutations. J Virol. 1990 Dec;64(12):6221–6233. [PMC free article] [PubMed]
  • Barker WC, Dayhoff MO. Viral src gene products are related to the catalytic chain of mammalian cAMP-dependent protein kinase. Proc Natl Acad Sci U S A. 1982 May;79(9):2836–2839. [PubMed]
  • Blumberg BM, Epstein LG, Saito Y, Chen D, Sharer LR, Anand R. Human immunodeficiency virus type 1 nef quasispecies in pathological tissue. J Virol. 1992 Sep;66(9):5256–5264. [PMC free article] [PubMed]
  • Burch HB, Nagy EV, Lukes YG, Cai WY, Wartofsky L, Burman KD. Nucleotide and amino acid homology between the human thyrotropin receptor and the HIV-1 Nef protein: identification and functional analysis. Biochem Biophys Res Commun. 1991 Nov 27;181(1):498–505. [PubMed]
  • Burger H, Weiser B, Flaherty K, Gulla J, Nguyen PN, Gibbs RA. Evolution of human immunodeficiency virus type 1 nucleotide sequence diversity among close contacts. Proc Natl Acad Sci U S A. 1991 Dec 15;88(24):11236–11240. [PubMed]
  • Cammarota G, Scheirle A, Takacs B, Doran DM, Knorr R, Bannwarth W, Guardiola J, Sinigaglia F. Identification of a CD4 binding site on the beta 2 domain of HLA-DR molecules. Nature. 1992 Apr 30;356(6372):799–801. [PubMed]
  • Chou PY, Fasman GD. Empirical predictions of protein conformation. Annu Rev Biochem. 1978;47:251–276. [PubMed]
  • Cullen BR. Regulation of human immunodeficiency virus replication. Annu Rev Microbiol. 1991;45:219–250. [PubMed]
  • Daniel MD, Kirchhoff F, Czajak SC, Sehgal PK, Desrosiers RC. Protective effects of a live attenuated SIV vaccine with a deletion in the nef gene. Science. 1992 Dec 18;258(5090):1938–1941. [PubMed]
  • Delassus S, Cheynier R, Wain-Hobson S. Evolution of human immunodeficiency virus type 1 nef and long terminal repeat sequences over 4 years in vivo and in vitro. J Virol. 1991 Jan;65(1):225–231. [PMC free article] [PubMed]
  • Dever TE, Glynias MJ, Merrick WC. GTP-binding domain: three consensus sequence elements with distinct spacing. Proc Natl Acad Sci U S A. 1987 Apr;84(7):1814–1818. [PubMed]
  • Devereux J, Haeberli P, Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. [PMC free article] [PubMed]
  • Dillon PJ, Rosen CA. A rapid method for the construction of synthetic genes using the polymerase chain reaction. Biotechniques. 1990 Sep;9(3):298–300. [PubMed]
  • Eckert KA, Kunkel TA. High fidelity DNA synthesis by the Thermus aquaticus DNA polymerase. Nucleic Acids Res. 1990 Jul 11;18(13):3739–3744. [PMC free article] [PubMed]
  • Fisher AG, Ensoli B, Looney D, Rose A, Gallo RC, Saag MS, Shaw GM, Hahn BH, Wong-Staal F. Biologically diverse molecular variants within a single HIV-1 isolate. Nature. 1988 Aug 4;334(6181):444–447. [PubMed]
  • Franchini G, Robert-Guroff M, Ghrayeb J, Chang NT, Wong-Staal F. Cytoplasmic localization of the HTLV-III 3' orf protein in cultured T cells. Virology. 1986 Dec;155(2):593–599. [PubMed]
  • Garcia JV, Alfano J, Miller AD. The negative effect of human immunodeficiency virus type 1 Nef on cell surface CD4 expression is not species specific and requires the cytoplasmic domain of CD4. J Virol. 1993 Mar;67(3):1511–1516. [PMC free article] [PubMed]
  • Garcia JV, Miller AD. Serine phosphorylation-independent downregulation of cell-surface CD4 by nef. Nature. 1991 Apr 11;350(6318):508–511. [PubMed]
  • Gaynor R. Cellular transcription factors involved in the regulation of HIV-1 gene expression. AIDS. 1992 Apr;6(4):347–363. [PubMed]
  • Guy B, Kieny MP, Riviere Y, Le Peuch C, Dott K, Girard M, Montagnier L, Lecocq JP. HIV F/3' orf encodes a phosphorylated GTP-binding protein resembling an oncogene product. Nature. 1987 Nov 19;330(6145):266–269. [PubMed]
  • Guy B, Rivière Y, Dott K, Regnault A, Kieny MP. Mutational analysis of the HIV nef protein. Virology. 1990 Jun;176(2):413–425. [PubMed]
  • Hahn BH, Gonda MA, Shaw GM, Popovic M, Hoxie JA, Gallo RC, Wong-Staal F. Genomic diversity of the acquired immune deficiency syndrome virus HTLV-III: different viruses exhibit greatest divergence in their envelope genes. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4813–4817. [PubMed]
  • Harris M, Hislop S, Patsilinacos P, Neil JC. In vivo derived HIV-1 nef gene products are heterogeneous and lack detectable nucleotide binding activity. AIDS Res Hum Retroviruses. 1992 May;8(5):537–543. [PubMed]
  • Hovanessian AG. On the HIV nef gene product. Res Virol. 1992 Jan-Feb;143(1):31–34. [PubMed]
  • Kaminchik J, Bashan N, Itach A, Sarver N, Gorecki M, Panet A. Genetic characterization of human immunodeficiency virus type 1 nef gene products translated in vitro and expressed in mammalian cells. J Virol. 1991 Feb;65(2):583–588. [PMC free article] [PubMed]
  • Kaplan JM, Mardon G, Bishop JM, Varmus HE. The first seven amino acids encoded by the v-src oncogene act as a myristylation signal: lysine 7 is a critical determinant. Mol Cell Biol. 1988 Jun;8(6):2435–2441. [PMC free article] [PubMed]
  • Kennelly PJ, Krebs EG. Consensus sequences as substrate specificity determinants for protein kinases and protein phosphatases. J Biol Chem. 1991 Aug 25;266(24):15555–15558. [PubMed]
  • Kestler HW, 3rd, Ringler DJ, Mori K, Panicali DL, Sehgal PK, Daniel MD, Desrosiers RC. Importance of the nef gene for maintenance of high virus loads and for development of AIDS. Cell. 1991 May 17;65(4):651–662. [PubMed]
  • Kienzle N, Bachmann M, Müller WE, Müller-Lantzsch N. Expression and cellular localization of the Nef protein from human immunodeficiency virus-1 in stably transfected B-cells. Arch Virol. 1992;124(1-2):123–132. [PubMed]
  • König R, Huang LY, Germain RN. MHC class II interaction with CD4 mediated by a region analogous to the MHC class I binding site for CD8. Nature. 1992 Apr 30;356(6372):796–798. [PubMed]
  • LaRosa GJ, Davide JP, Weinhold K, Waterbury JA, Profy AT, Lewis JA, Langlois AJ, Dreesman GR, Boswell RN, Shadduck P, et al. Conserved sequence and structural elements in the HIV-1 principal neutralizing determinant. Science. 1990 Aug 24;249(4971):932–935. [PubMed]
  • Laurent AG, Hovanessian AG, Rivière Y, Krust B, Regnault A, Montagnier L, Findeli A, Kieny MP, Guy B. Production of a non-functional nef protein in human immunodeficiency virus type 1-infected CEM cells. J Gen Virol. 1990 Oct;71(Pt 10):2273–2281. [PubMed]
  • Lochrie MA, Simon MI. G protein multiplicity in eukaryotic signal transduction systems. Biochemistry. 1988 Jul 12;27(14):4957–4965. [PubMed]
  • Luria S, Chambers I, Berg P. Expression of the type 1 human immunodeficiency virus Nef protein in T cells prevents antigen receptor-mediated induction of interleukin 2 mRNA. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5326–5330. [PubMed]
  • Martins LP, Chenciner N, Asjö B, Meyerhans A, Wain-Hobson S. Independent fluctuation of human immunodeficiency virus type 1 rev and gp41 quasispecies in vivo. J Virol. 1991 Aug;65(8):4502–4507. [PMC free article] [PubMed]
  • Maurer B, Flügel RM. The 3'-orf protein of human immunodeficiency virus 2 shows sequence homology with the bel3 gene of the human spumaretrovirus. FEBS Lett. 1987 Oct 5;222(2):286–288. [PubMed]
  • McNearney T, Hornickova Z, Markham R, Birdwell A, Arens M, Saah A, Ratner L. Relationship of human immunodeficiency virus type 1 sequence heterogeneity to stage of disease. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10247–10251. [PubMed]
  • Meyerhans A, Cheynier R, Albert J, Seth M, Kwok S, Sninsky J, Morfeldt-Månson L, Asjö B, Wain-Hobson S. Temporal fluctuations in HIV quasispecies in vivo are not reflected by sequential HIV isolations. Cell. 1989 Sep 8;58(5):901–910. [PubMed]
  • Miller AD, Buttimore C. Redesign of retrovirus packaging cell lines to avoid recombination leading to helper virus production. Mol Cell Biol. 1986 Aug;6(8):2895–2902. [PMC free article] [PubMed]
  • Miller AD, Rosman GJ. Improved retroviral vectors for gene transfer and expression. Biotechniques. 1989 Oct;7(9):980–990. [PMC free article] [PubMed]
  • Munis JR, Kornbluth RS, Guatelli JC, Richman DD. Ordered appearance of human immunodeficiency virus type 1 nucleic acids following high multiplicity infection of macrophages. J Gen Virol. 1992 Aug;73(Pt 8):1899–1906. [PubMed]
  • Niederman TM, Garcia JV, Hastings WR, Luria S, Ratner L. Human immunodeficiency virus type 1 Nef protein inhibits NF-kappa B induction in human T cells. J Virol. 1992 Oct;66(10):6213–6219. [PMC free article] [PubMed]
  • Ratner L, Fisher A, Jagodzinski LL, Mitsuya H, Liou RS, Gallo RC, Wong-Staal F. Complete nucleotide sequences of functional clones of the AIDS virus. AIDS Res Hum Retroviruses. 1987 Spring;3(1):57–69. [PubMed]
  • Ratner L, Starcich B, Josephs SF, Hahn BH, Reddy EP, Livak KJ, Petteway SR, Jr, Pearson ML, Haseltine WA, Arya SK, et al. Polymorphism of the 3' open reading frame of the virus associated with the acquired immune deficiency syndrome, human T-lymphotropic virus type III. Nucleic Acids Res. 1985 Nov 25;13(22):8219–8229. [PMC free article] [PubMed]
  • Robert-Guroff M, Popovic M, Gartner S, Markham P, Gallo RC, Reitz MS. Structure and expression of tat-, rev-, and nef-specific transcripts of human immunodeficiency virus type 1 in infected lymphocytes and macrophages. J Virol. 1990 Jul;64(7):3391–3398. [PMC free article] [PubMed]
  • Saag MS, Hahn BH, Gibbons J, Li Y, Parks ES, Parks WP, Shaw GM. Extensive variation of human immunodeficiency virus type-1 in vivo. Nature. 1988 Aug 4;334(6181):440–444. [PubMed]
  • Samuel KP, Hodge DR, Chen YM, Papas TS. Nef proteins of the human immunodeficiency viruses (HIV-1 and HIV-2) and simian immunodeficiency virus (SIV) are structurally similar to leucine zipper transcriptional activation factors. AIDS Res Hum Retroviruses. 1991 Aug;7(8):697–706. [PubMed]
  • Samuel KP, Seth A, Konopka A, Lautenberger JA, Papas TS. The 3'-orf protein of human immunodeficiency virus shows structural homology with the phosphorylation domain of human interleukin-2 receptor and the ATP-binding site of the protein kinase family. FEBS Lett. 1987 Jun 22;218(1):81–86. [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PubMed]
  • Shinnick TM, Lerner RA, Sutcliffe JG. Nucleotide sequence of Moloney murine leukaemia virus. Nature. 1981 Oct 15;293(5833):543–548. [PubMed]
  • Simmonds P, Balfe P, Peutherer JF, Ludlam CA, Bishop JO, Brown AJ. Human immunodeficiency virus-infected individuals contain provirus in small numbers of peripheral mononuclear cells and at low copy numbers. J Virol. 1990 Feb;64(2):864–872. [PMC free article] [PubMed]
  • Staden R. Methods to define and locate patterns of motifs in sequences. Comput Appl Biosci. 1988 Mar;4(1):53–60. [PubMed]
  • Terwilliger E, Sodroski JG, Rosen CA, Haseltine WA. Effects of mutations within the 3' orf open reading frame region of human T-cell lymphotropic virus type III (HTLV-III/LAV) on replication and cytopathogenicity. J Virol. 1986 Nov;60(2):754–760. [PMC free article] [PubMed]
  • Tsunetsugu-Yokota Y, Matsuda S, Maekawa M, Saito T, Takemori T, Takebe Y. Constitutive expression of the nef gene suppresses human immunodeficiency virus type 1 (HIV-1) replication in monocytic cell lines. Virology. 1992 Dec;191(2):960–963. [PubMed]
  • Vartanian JP, Meyerhans A, Asjö B, Wain-Hobson S. Selection, recombination, and G----A hypermutation of human immunodeficiency virus type 1 genomes. J Virol. 1991 Apr;65(4):1779–1788. [PMC free article] [PubMed]
  • Vega MA, Guigó R, Smith TF. Autoimmune response in AIDS. Nature. 1990 May 3;345(6270):26–26. [PubMed]
  • Werner T, Ferroni S, Saermark T, Brack-Werner R, Banati RB, Mager R, Steinaa L, Kreutzberg GW, Erfle V. HIV-1 Nef protein exhibits structural and functional similarity to scorpion peptides interacting with K+ channels. AIDS. 1991 Nov;5(11):1301–1308. [PubMed]
  • Yu G, Felsted RL. Effect of myristoylation on p27 nef subcellular distribution and suppression of HIV-LTR transcription. Virology. 1992 Mar;187(1):46–55. [PubMed]
  • Zweig M, Samuel KP, Showalter SD, Bladen SV, DuBois GC, Lautenberger JA, Hodge DR, Papas TS. Heterogeneity of Nef proteins in cells infected with human immunodeficiency virus type 1. Virology. 1990 Nov;179(1):504–507. [PubMed]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)