Human immunodeficiency virus type-1 (HIV-1) Pr55gag virus-like particles are potent activators of human monocytes

ABSTRACT We demonstrate that a genetically engineered human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) composed mainly of p66 or p51 subunits can be incorporated into virus-like particles (VLPs) when coexpressed with HIV-1 Pr55 gag . VLP-associated RT exhibited a detergent-resistant association with immature cores during sucrose gradient equilibrium centrifugation, suggesting that RT is incorporated into VLPs. However, RT that retains downstream integrase (IN) is severely inhibited in terms of incorporation into VLPs. Results from immunofluorescence tests reveal that RT-IN is primarily localized at the perinuclear area and exhibits poor colocalization with Gag. IN removal leads to a redistribution of RT throughout the cytoplasm and improved RT incorporation into VLPs. Similar results were observed for RT-IN in which alanine was substituted for 186-Lys-Arg-Lys-188 residues of the IN putative nuclear localization signal, suggesting that IN karyophilic properties may partly account for the inhibitory effect of IN on RT incorporation. Although the membrane-binding capacity of RT was markedly reduced compared to that of wild-type Gag or Gag-Pol, the correlation of membrane-binding ability with particle incorporation efficiency was incomplete. Furthermore, we observed that membrane-binding-defective myristylation-minus RT can be packaged into VLPs at the same level as its normal myristylated counterpart. This suggests that the incorporation of RT into VLPs is independent of membrane affinity but very dependent on RT-Gag interaction. Results from a genetic analysis suggest that the Gag-interacting regions of RT mainly reside in the thumb subdomain and that the RT-binding domains of Gag are located in the matrix (MA) and p6 regions.

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Baculovirus-Derived Human Immunodeficiency Virus Type 1 Virus-Like Particles Activate Dendritic Cells and Induce Ex Vivo T-Cell Responses

Journal of Virology ◽

10.1128/jvi.00050-06 ◽

2006 ◽

Vol 80 (18) ◽

pp. 9134-9143 ◽

Cited By ~ 88

Author(s):

L. Buonaguro ◽

M. L. Tornesello ◽

M. Tagliamonte ◽

R. C. Gallo ◽

L. X. Wang ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Dendritic Cells ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Ex Vivo ◽

Secondary Response ◽

Virus Like Particles ◽

Immunodeficiency Virus ◽

Hiv 1

ABSTRACT We have recently developed a candidate human immunodeficiency virus type 1 (HIV-1) vaccine model based on HIV-1 Pr55 gag virus-like particles (HIV-VLPs), produced in a baculovirus expression system and presenting a gp120 molecule from a Ugandan HIV-1 isolate of clade A (HIV-VLPAs). The HIV-VLPAs show the induction in BALB/c mice of systemic and mucosal neutralizing antibodies as well as cytotoxic T lymphocytes, by intraperitoneal as well as intranasal administration. In the present article, the effects of the baculovirus-expressed HIV-VLPs on human immature monocyte-derived dendritic cells (MDDCs) have been evaluated. The HIV-VLPs efficiently induce maturation and activation of MDDCs and are incorporated into MDDCs preferentially via an actin-dependent macropinocytosis and endocytosis. The HIV-VLP-activated MDDCs show enhanced Th1- and Th2-specific cytokine production, and the effects of HIV-VLPs on MDDCs are not mediated through Toll-like receptors 2 and 4 (TLR2 and -4) signaling. Finally, HIV-VLP-loaded MDDCs are able to induce a primary and secondary response in autologous human CD4+ T cells in an ex vivo immunization assay. Our results on the interaction and processing of baculovirus HIV-VLPs by MDDCs give an insight into the mechanisms underlying the immune response induced by HIV-VLPAs in vivo.

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Induction of Systemic and Mucosal Cross-Clade Neutralizing Antibodies in BALB/c Mice Immunized with Human Immunodeficiency Virus Type 1 Clade A Virus-Like Particles Administered by Different Routes of Inoculation

Journal of Virology ◽

10.1128/jvi.79.11.7059-7067.2005 ◽

2005 ◽

Vol 79 (11) ◽

pp. 7059-7067 ◽

Cited By ~ 62

Author(s):

L. Buonaguro ◽

M. L. Visciano ◽

M. L. Tornesello ◽

M. Tagliamonte ◽

B. Biryahwaho ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Neutralizing Antibodies ◽

Ex Vivo ◽

Lower Efficiency ◽

Virus Like Particles ◽

Immunodeficiency Virus ◽

Hiv 1

ABSTRACT We have recently developed a candidate human immunodeficiency virus type 1 (HIV-1) vaccine model, based on virus-like particles (VLPs) expressing gp120 from a Ugandan HIV-1 isolate of clade A (HIV-VLPAs), which shows the induction of neutralizing antibodies as well as cytotoxic T lymphocytes (CTL) in BALB/c mice by intraperitoneal (i.p.) administration. In the present study, immunization experiments based on a multiple-dose regimen have been performed with BALB/c mice to compare different routes of administration. i.p. and intranasal (i.n.), but not oral, administration induce systemic as well as mucosal (vaginal and intestinal) immunoglobulin G (IgG) and IgA responses. These immune sera exhibit >50% ex vivo neutralizing activity against both autologous and heterologous primary isolates. Furthermore, the administration of HIV-VLPAs by the i.n. immunization route induces a specific CTL activity, although at lower efficiency than the i.p. route. The HIV-VLPAs represent an efficient strategy to stimulate both arms of immunity; furthermore, the induction of specific humoral immunity at mucosal sites, which nowadays represent the main port of entry for HIV-1 infection, is of great interest. All these properties, and the possible cross-clade in vivo protection, could make these HIV-VLPAs a good candidate for a mono- and multicomponent worldwide preventive vaccine approach not restricted to high-priority regions, such as sub-Saharan countries.

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Contribution of Virus-Like Particles to the Immunogenicity of Human Immunodeficiency Virus Type 1 Gag-Derived Vaccines in Mice

Journal of Virology ◽

10.1128/jvi.79.3.1701-1712.2005 ◽

2005 ◽

Vol 79 (3) ◽

pp. 1701-1712 ◽

Cited By ~ 8

Author(s):

S. B. Justin Wong ◽

Robert F. Siliciano

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Expression Vectors ◽

Antigenic Determinants ◽

Gag Protein ◽

Virus Like Particles ◽

Immunodeficiency Virus ◽

Hiv 1

ABSTRACT The human immunodeficiency virus type 1 (HIV-1) Gag protein is a major target antigen for cytotoxic-T-lymphocyte-based vaccine strategies because of its high level of conservation. The murine model has been used extensively to evaluate potential HIV-1 vaccines. However, the biology of HIV-1 Gag is somewhat different in human and murine tissues. The ability of HIV-1 Gag to form virus-like particles (VLPs) in human cells is severely curtailed in murine cells. Hence, it is not known whether immunizing mice with expression vectors encoding HIV-1 Gag provides an accurate assessment of the immunogenicity of these candidate vaccines in primates. In this report, we made use of a chimeric Moloney murine leukemia virus (MMLV)-HIV-1 Gag in which the p17 matrix domain of HIV-1 was replaced with the p15 matrix and p12 domains from MMLV. Murine cells expressing this construct released significant amounts of VLPs. The construct preserved H-2 d -restricted antigenic determinants in the remaining portion of HIV-1 Gag, allowing immunogenicity studies to be performed with mice. We demonstrated that immunizing mice with plasmid DNA or adenoviral vectors encoding this chimeric Gag did not significantly increase the HIV-1 Gag-specific cellular or humoral immune response when compared to immunization with a myristoylation-incompetent version of the construct. Thus, the release of VLPs formed in vivo may not play a major role in the immunogenicity of vectors expressing HIV-1 Gag constructs.

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Productive human immunodeficiency virus type 1 (HIV-1) infection of nonproliferating human monocytes.

Journal of Experimental Medicine ◽

10.1084/jem.174.6.1477 ◽

1991 ◽

Vol 174 (6) ◽

pp. 1477-1482 ◽

Cited By ~ 247

Author(s):

J B Weinberg ◽

T J Matthews ◽

B R Cullen ◽

M H Malim

Keyword(s):

Human Immunodeficiency Virus ◽

T Lymphocytes ◽

Dna Synthesis ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Mononuclear Phagocytes ◽

Human Monocytes ◽

Immunodeficiency Virus ◽

Hiv 1

Human immunodeficiency virus type 1 (HIV-1) infection of T lymphocytes requires cellular proliferation and DNA synthesis. Human monocytes were shown to have low DNA synthesis rates, yet the monocytotropic BaL isolate of HIV-1 was able to infect these cells efficiently. Monocytes that were irradiated to assure no DNA synthesis could also be readily infected with HIV-1BaL. Such infections were associated with the integration of HIV-1BaL DNA into the high molecular weight, chromosomal DNA of monocytes. Thus, normal, nonproliferating monocytes differ from T lymphocytes in that a productive HIV-1 infection can occur independently of cellular DNA synthesis. These results suggest that normal nonproliferating mononuclear phagocytes, which are relatively resistant to the destructive effects of this virus, may serve as persistent and productive reservoirs for HIV-1 in vivo.

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