Logo of jvirolPermissionsJournals.ASM.orgJournalJV ArticleJournal InfoAuthorsReviewers
J Virol. 1997 Jul; 71(7): 4892–4903.
PMCID: PMC191719

Inactivation of the human immunodeficiency virus type 1 inhibitory elements allows Rev-independent expression of Gag and Gag/protease and particle formation.


The expression of gag, pol, and env of human immunodeficiency virus type 1 (HIV-1) depends on the presence of the viral Rev protein. This dependence is, at least in part, due to the presence of negatively acting sequences (inhibitory or instability elements [INS]) located within unspliced and partially spliced mRNAs. The positive interaction of Rev with the Rev-responsive element in these mRNAs counteracts the negative effects of the inhibitory sequences. Here, we demonstrate that in addition to the previously identified INS1 within p17gag, several other INS elements exist within the gag/pol region of HIV-1. These elements act independently of each other and were eliminated by mutagenesis after the introduction of multiple point mutations not affecting the coding region, leading to constitutive high levels of Gag expression. Expression vectors containing an intact or nearly intact p55gag region allowed the production of immature viral particles in mammalian cells in the absence of any other HIV proteins. The introduction of additional mutations in the protease region allowed efficient production of Gag/protease, which resulted in processing of the Pr55gag precursor and production of mature Gag particles with a lentivirus-like conical-core structure. The elimination of a newly identified INS element within pol and the previously identified CRS located within int was accomplished by the same methodology. Sequence comparisons of the identified inhibitory elements revealed no apparent homologies and demonstrated that these sequences are not splice sites. These results demonstrate that the elimination of INS elements leads to efficient expression of HIV-1 mRNAs in the absence of Rev or any posttranscriptional activating mechanisms.

Full Text

The Full Text of this article is available as a PDF (1.0M).

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]
  • Afonina E, Neumann M, Pavlakis GN. Preferential binding of poly(A)-binding protein 1 to an inhibitory RNA element in the human immunodeficiency virus type 1 gag mRNA. J Biol Chem. 1997 Jan 24;272(4):2307–2311. [PubMed]
  • Barker GF, Beemon K. Nonsense codons within the Rous sarcoma virus gag gene decrease the stability of unspliced viral RNA. Mol Cell Biol. 1991 May;11(5):2760–2768. [PMC free article] [PubMed]
  • Benko DM, Robinson R, Solomin L, Mellini M, Felber BK, Pavlakis GN. Binding of trans-dominant mutant Rev protein of human immunodeficiency virus type 1 to the cis-acting Rev-responsive element does not affect the fate of viral mRNA. New Biol. 1990 Dec;2(12):1111–1122. [PubMed]
  • Brighty DW, Rosenberg M. A cis-acting repressive sequence that overlaps the Rev-responsive element of human immunodeficiency virus type 1 regulates nuclear retention of env mRNAs independently of known splice signals. Proc Natl Acad Sci U S A. 1994 Aug 30;91(18):8314–8318. [PMC free article] [PubMed]
  • Chen CY, Chen TM, Shyu AB. Interplay of two functionally and structurally distinct domains of the c-fos AU-rich element specifies its mRNA-destabilizing function. Mol Cell Biol. 1994 Jan;14(1):416–426. [PMC free article] [PubMed]
  • Cochrane AW, Jones KS, Beidas S, Dillon PJ, Skalka AM, Rosen CA. Identification and characterization of intragenic sequences which repress human immunodeficiency virus structural gene expression. J Virol. 1991 Oct;65(10):5305–5313. [PMC free article] [PubMed]
  • Cullen BR. Mechanism of action of regulatory proteins encoded by complex retroviruses. Microbiol Rev. 1992 Sep;56(3):375–394. [PMC free article] [PubMed]
  • D'Agostino DM, Felber BK, Harrison JE, Pavlakis GN. The Rev protein of human immunodeficiency virus type 1 promotes polysomal association and translation of gag/pol and vpu/env mRNAs. Mol Cell Biol. 1992 Mar;12(3):1375–1386. [PMC free article] [PubMed]
  • de Wet JR, Wood KV, DeLuca M, Helinski DR, Subramani S. Firefly luciferase gene: structure and expression in mammalian cells. Mol Cell Biol. 1987 Feb;7(2):725–737. [PMC free article] [PubMed]
  • Felber BK, Hadzopoulou-Cladaras M, Cladaras C, Copeland T, Pavlakis GN. rev protein of human immunodeficiency virus type 1 affects the stability and transport of the viral mRNA. Proc Natl Acad Sci U S A. 1989 Mar;86(5):1495–1499. [PMC free article] [PubMed]
  • Fisher AG, Collalti E, Ratner L, Gallo RC, Wong-Staal F. A molecular clone of HTLV-III with biological activity. Nature. 1985 Jul 18;316(6025):262–265. [PubMed]
  • Göttlinger HG, Dorfman T, Sodroski JG, Haseltine WA. Effect of mutations affecting the p6 gag protein on human immunodeficiency virus particle release. Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3195–3199. [PMC free article] [PubMed]
  • Graham FL, van der Eb AJ. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. [PubMed]
  • Hadzopoulou-Cladaras M, Felber BK, Cladaras C, Athanassopoulos A, Tse A, Pavlakis GN. The rev (trs/art) protein of human immunodeficiency virus type 1 affects viral mRNA and protein expression via a cis-acting sequence in the env region. J Virol. 1989 Mar;63(3):1265–1274. [PMC free article] [PubMed]
  • Hentze MW. Determinants and regulation of cytoplasmic mRNA stability in eukaryotic cells. Biochim Biophys Acta. 1991 Nov 11;1090(3):281–292. [PubMed]
  • Karacostas V, Nagashima K, Gonda MA, Moss B. Human immunodeficiency virus-like particles produced by a vaccinia virus expression vector. Proc Natl Acad Sci U S A. 1989 Nov;86(22):8964–8967. [PMC free article] [PubMed]
  • Karacostas V, Wolffe EJ, Nagashima K, Gonda MA, Moss B. Overexpression of the HIV-1 gag-pol polyprotein results in intracellular activation of HIV-1 protease and inhibition of assembly and budding of virus-like particles. Virology. 1993 Apr;193(2):661–671. [PubMed]
  • Maldarelli F, Martin MA, Strebel K. Identification of posttranscriptionally active inhibitory sequences in human immunodeficiency virus type 1 RNA: novel level of gene regulation. J Virol. 1991 Nov;65(11):5732–5743. [PMC free article] [PubMed]
  • Mammano F, Ohagen A, Höglund S, Göttlinger HG. Role of the major homology region of human immunodeficiency virus type 1 in virion morphogenesis. J Virol. 1994 Aug;68(8):4927–4936. [PMC free article] [PubMed]
  • Nasioulas G, Zolotukhin AS, Tabernero C, Solomin L, Cunningham CP, Pavlakis GN, Felber BK. Elements distinct from human immunodeficiency virus type 1 splice sites are responsible for the Rev dependence of env mRNA. J Virol. 1994 May;68(5):2986–2993. [PMC free article] [PubMed]
  • Quesada-Rolander M, Mäkitalo B, Thorstensson R, Zhang YJ, Castaños-Velez E, Biberfeld G, Putkonen P. Protection against mucosal SIVsm challenge in macaques infected with a chimeric SIV that expresses HIV type 1 envelope. AIDS Res Hum Retroviruses. 1996 Jul 20;12(11):993–999. [PubMed]
  • Rabbitts PH, Forster A, Stinson MA, Rabbitts TH. Truncation of exon 1 from the c-myc gene results in prolonged c-myc mRNa stability. EMBO J. 1985 Dec 30;4(13B):3727–3733. [PMC free article] [PubMed]
  • Schwartz S, Campbell M, Nasioulas G, Harrison J, Felber BK, Pavlakis GN. Mutational inactivation of an inhibitory sequence in human immunodeficiency virus type 1 results in Rev-independent gag expression. J Virol. 1992 Dec;66(12):7176–7182. [PMC free article] [PubMed]
  • Schwartz S, Felber BK, Benko DM, Fenyö EM, Pavlakis GN. Cloning and functional analysis of multiply spliced mRNA species of human immunodeficiency virus type 1. J Virol. 1990 Jun;64(6):2519–2529. [PMC free article] [PubMed]
  • Schwartz S, Felber BK, Pavlakis GN. Distinct RNA sequences in the gag region of human immunodeficiency virus type 1 decrease RNA stability and inhibit expression in the absence of Rev protein. J Virol. 1992 Jan;66(1):150–159. [PMC free article] [PubMed]
  • Shaw G, Kamen R. A conserved AU sequence from the 3' untranslated region of GM-CSF mRNA mediates selective mRNA degradation. Cell. 1986 Aug 29;46(5):659–667. [PubMed]
  • Shyu AB, Belasco JG, Greenberg ME. Two distinct destabilizing elements in the c-fos message trigger deadenylation as a first step in rapid mRNA decay. Genes Dev. 1991 Feb;5(2):221–231. [PubMed]
  • Shyu AB, Greenberg ME, Belasco JG. The c-fos transcript is targeted for rapid decay by two distinct mRNA degradation pathways. Genes Dev. 1989 Jan;3(1):60–72. [PubMed]
  • Solomin L, Felber BK, Pavlakis GN. Different sites of interaction for Rev, Tev, and Rex proteins within the Rev-responsive element of human immunodeficiency virus type 1. J Virol. 1990 Dec;64(12):6010–6017. [PMC free article] [PubMed]
  • Vaishnav YN, Wong-Staal F. The biochemistry of AIDS. Annu Rev Biochem. 1991;60:577–630. [PubMed]
  • von Poblotzki A, Wagner R, Niedrig M, Wanner G, Wolf H, Modrow S. Identification of a region in the Pr55gag-polyprotein essential for HIV-1 particle formation. Virology. 1993 Apr;193(2):981–985. [PubMed]
  • Wisdom R, Lee W. The protein-coding region of c-myc mRNA contains a sequence that specifies rapid mRNA turnover and induction by protein synthesis inhibitors. Genes Dev. 1991 Feb;5(2):232–243. [PubMed]

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


Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...