Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2005 Jul;79(14):9038-45.
doi: 10.1128/jvi.79.14.9038-9045.2005.

Heterologous late-domain sequences have various abilities to promote budding of human immunodeficiency virus type 1

David E Ott  1 , Lori V Coren, Tracy D Gagliardi, Kunio Nagashima
Affiliations

Heterologous late-domain sequences have various abilities to promote budding of human immunodeficiency virus type 1

David E Ott et al. J Virol. 2005 Jul.

Abstract

Retroviral late (L) domains present within Gag act in conjunction with cellular proteins to efficiently release virions from the surface of the cell. Three different critical core sequences have been identified as required elements for L-domain function: PPPY, PTAP (also PSAP), and YPDL, with different retroviruses utilizing one or two of these core sequences. The human immunodeficiency virus type 1 (HIV-1) L domain is centered around a PTAP sequence in the p6 region of Gag. To assess the ability of heterologous L-domain sequences to be functionally interchanged for those in full-length HIV-1, we produced a series of constructs that replaced PTAP-containing p6(Gag) sequences with those of PPPY- or YPDL-based L domains. While previous studies had found that L domains are interchangeable in other retroviruses, most of the sequences introduced into p6(Gag) failed to substitute for PTAP-mediated L-domain function. One exception was the 11-amino-acid p2b sequence of Rous sarcoma virus (RSV) Gag, which could fully restore HIV-1 budding, while a PPPPY sequence exchange alone did not. This suggests that the RSV L domain consists of more than simply its core L-domain sequence. The HIV-p2b chimera was as infectious as the wild type, produced normal virions, and was sensitive to proteasome inhibitors. These results show that L-domain sequences are not necessarily interchangeable. Thus, HIV-1 Gag might have a more stringent requirement for L-domain function than the other retroviruses previously studied.

PubMed Disclaimer

Figures

FIG. 1.
FIG. 1.
DNA constructs. Diagrams of the various Gags from the full-length HIV-1 constructs used in this study are presented with HIV-1 sequences denoted in black, RSV in white, and EIAV in hatched shading. Sequence changes are indicated below each diagram. Protease cleavage sites that produce the mature Gag proteins are denoted in the NL4-3 diagram by white lines in Gag.
FIG. 2.
FIG. 2.
Immunoblot analysis of virion preparations. A p24CA immunoblot of equal volumes of virions isolated from transfection supernatants is presented. Samples are identified above their respective lanes. Positions of the molecular mass standards are indicated at left, and reactive bands are identified at right. sssDNA, virus preparation isolated from a sheared salmon sperm DNA transfection.
FIG. 3.
FIG. 3.
[35S]Met/Cys metabolic radiolabeling and immunoprecipitation. (A) Phosphorimages of SDS-PAGE gels from a typical radioimmunoprecipitation analysis of metabolically labeled Gag from virion and cytoplasmic lysates are presented. The lysates examined are indicated above each image, and samples are identified above their respective lanes. Positions of the molecular mass standards are indicated at left, and reactive bands are identified at right. sssDNA, virus preparation isolated from a sheared salmon sperm DNA transfection. (B) Graphical results of release factors relative to wild-type virus from three independent experiments are presented.
FIG. 4.
FIG. 4.
Proteasome inhibitor studies. (A) Immunoblot of cells treated with or without proteasome inhibitors for 8 h is presented. Samples are identified above their respective lanes. O/N, virion preparations prepared from an 18-h harvest of supernatants from transfected cultures. Positions of the molecular mass standards are indicated at left, and reactive bands are identified at right. (B) Phosphorimages of an SDS-PAGE gel from radioimmunoprecipitation analysis of metabolically labeled Gag from virion and cytoplasmic lysates are presented. Cells were labeled for 6 h in the presence or absence of proteasome inhibitors (MG-132 and lactacystin) as indicated. The lysates examined are indicated above each image, and samples are identified above their respective lanes. Positions of the molecular mass standards are indicated at left, and reactive bands are identified at right. sssDNA, virus preparation isolated from a sheared salmon sperm DNA transfection.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Adachi, A., S. Koenig, H. E. Gendelman, D. Daugherty, S. Gattoni-Celli, A. S. Fauci, and M. A. Martin. 1987. Productive, persistent infection of human colorectal cell lines with human immunodeficiency virus. J. Virol. 61:209-213. - PMC - PubMed
    1. Bleiber, G., S. Peters, R. Martinez, D. Cmarko, P. Meylan, and A. Telenti. 2004. The central region of human immunodeficiency virus type 1 p6 protein (Gag residues S14-I31) is dispensable for the virus in vitro. J. Gen. Virol. 85:921-927. - PubMed
    1. Blot, V., F. Perugi, B. Gay, M. C. Prevost, L. Briant, F. Tangy, H. Abriel, O. Staub, M. C. Dokhelar, and C. Pique. 2004. Nedd4.1-mediated ubiquitination and subsequent recruitment of Tsg101 ensure HTLV-1 Gag trafficking towards the multivesicular body pathway prior to virus budding. J. Cell Sci. 117:2357-2367. - PubMed
    1. Chen, C., F. Li, and R. C. Montelaro. 2001. Functional roles of equine infectious anemia virus Gag p9 in viral budding and infection. J. Virol. 75:9762-9770. - PMC - PubMed
    1. Demirov, D. G., and E. O. Freed. 2004. Retrovirus budding. Virus Res. 106:87-102. - PubMed

Publication types

Substances