scholarly journals Human Immunodeficiency Virus Type 1 Enters Brain Microvascular Endothelia by Macropinocytosis Dependent on Lipid Rafts and the Mitogen-Activated Protein Kinase Signaling Pathway

2002 ◽  
Vol 76 (13) ◽  
pp. 6689-6700 ◽  
Author(s):  
Nancy Q. Liu ◽  
Albert S. Lossinsky ◽  
Waldemar Popik ◽  
Xia Li ◽  
Chandrasekhar Gujuluva ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Endothelial Cells ◽  
Protein Kinase ◽  
Virus Entry ◽  
Mitogen Activated Protein Kinase ◽  
Cytoplasmic Vesicles ◽  
Immunodeficiency Virus ◽  
Mitogen Activated Protein ◽  
Hiv 1

ABSTRACT Brain microvascular endothelial cells (BMVECs) present an incomplete barrier to human immunodeficiency virus type 1 (HIV-1) neuroinvasion. In order to clarify the mechanisms of HIV-1 invasion, we have examined HIV-1 uptake and transcellular penetration in an in vitro BMVEC model. No evidence of productive infection was observed by luciferase, PCR, and reverse transcriptase assays. Approximately 1% of viral RNA and 1% of infectious virus penetrated the BMVEC barrier without disruption of tight junctions. The virus upregulated ICAM-1 on plasma membranes and in cytoplasmic vesiculotubular structures. HIV-1 virions were entangled by microvilli and were taken into cytoplasmic vesicles through surface invaginations without fusion of the virus envelope with the plasma membrane. Subsequently, the cytoplasmic vesicles fused with lysosomes, the virions were lysed, and the vesicles diminished in size. Upon cell entry, HIV-1 colocalized with cholera toxin B, which targets lipid raft-associated GM1 ganglioside. Cholesterol-extracting agents, cyclodextrin and nystatin, and polyanion heparin significantly inhibited virus entry. Anti-CD4 had no effect and the chemokine AOP-RANTES had only a slight inhibitory effect on virus entry. HIV-1 activated the mitogen-activated protein kinase (MAPK) pathway, and inhibition of MAPK/Erk kinase inhibited virus entry. Entry was also blocked by dimethylamiloride, indicating that HIV-1 enters endothelial cells by macropinocytosis. Therefore, HIV-1 penetrates BMVECs in ICAM-1-lined macropinosomes by a mechanism involving lipid rafts, MAPK signaling, and glycosylaminoglycans, while CD4 and chemokine receptors play limited roles in this process.

scholarly journals Interaction of the CC-Chemokine RANTES with Glycosaminoglycans Activates a p44/p42 Mitogen-Activated Protein Kinase-Dependent Signaling Pathway and Enhances Human Immunodeficiency Virus Type 1 Infectivity

2002 ◽  
Vol 76 (5) ◽  
pp. 2245-2254 ◽  
Author(s):  
Theresa Li-Yun Chang ◽  
Cynthia J. Gordon ◽  
Branka Roscic-Mrkic ◽  
Christine Power ◽  
Amanda E. I. Proudfoot ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Protein Kinase ◽  
Cell Surface ◽  
Signaling Pathway ◽  
Human Immunodeficiency Virus Type ◽  
Mitogen Activated Protein Kinase ◽  
Immunodeficiency Virus ◽  
Mitogen Activated Protein ◽  
Hiv 1

ABSTRACT The interaction of the CC-chemokine RANTES with its cell surface receptors transduces multiple intracellular signals: low concentrations of RANTES (1 to 10 nM) stimulate G-protein-coupled receptor (GPCR) activity, and higher concentrations (1 μM) activate a phosphotyrosine kinase (PTK)-dependent pathway. Here, we show that the higher RANTES concentrations induce rapid tyrosine phosphorylation of multiple proteins. Several src-family kinases (Fyn, Hck, Src) are activated, as is the focal adhesion kinase p125 FAK and, eventually, members of the p44/p42 mitogen-activated protein kinase (MAPK) family. This PTK signaling pathway can be activated independently of known seven-transmembrane GPCRs for RANTES because it occurs in cells that lack any such RANTES receptors. Instead, activation of the PTK signaling pathway is dependent on the expression of glycosaminoglycans (GAGs) on the cell surface, in that it could not be activated by RANTES in GAG-deficient cells. We have previously demonstrated that RANTES can both enhance and inhibit infection of cells with human immunodeficiency virus type 1 (HIV-1). Here we show that activation of both PTK and MAPK is involved in the enhancement of HIV-1 infectivity caused by RANTES in cells that lack GPCRs for RANTES but which express GAGs.

scholarly journals Plasma Membrane-Targeted Raf Kinase Activates NF-κB and Human Immunodeficiency Virus Type 1 Replication in T Lymphocytes

1998 ◽  
Vol 72 (4) ◽  
pp. 2788-2794 ◽  
Author(s):  
Egbert Flory ◽  
Christoph K. Weber ◽  
Peifeng Chen ◽  
Angelika Hoffmeyer ◽  
Christian Jassoy ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
T Lymphocytes ◽  
Catalytic Domain ◽  
Mitogen Activated Protein Kinase ◽  
Raf Kinase ◽  
Immunodeficiency Virus ◽  
Mitogen Activated Protein ◽  
Hiv 1

ABSTRACT Increasing evidence points to a role of the mitogenic Ras/Raf/MEK/ERK signaling cascade in regulation of human immunodeficiency virus type 1 (HIV-1) gene expression. Stimulation of elements of this pathway leads to transactivation of the HIV-1 promoter. In particular, the NF-κB motif in the HIV long terminal repeat (LTR) represents a Raf-responsive element in fibroblasts. Regulation of the Raf kinase in T cells differs from findings with a variety of cell lines that the catalytic domain of Raf (RafΔ26–303) shows no activity. In this study, we restored the activity of the kinase in T cells by fusing its catalytic domain to the CAAX motif (-Cx) of Ras, thus targeting the enzyme to the plasma membrane. Constitutive activity of Raf was demonstrated by phosphorylation of mitogen-activated protein kinase kinase (MEK) and endogenous mitogen-activated protein kinase 1/2 (ERK1/2) in A3.01 T cells transfected with RafΔ26–303-Cx. Membrane-targeted Raf also stimulates NF-κB, as judged by κB-dependent reporter assays and enhanced NF-κB p65 binding on band shift analysis. Moreover, we found that active Raf transactivates the HIVNL4-3 LTR in A3.01 T lymphocytes and that dominant negative Raf (C4) blocked 12-O-tetradecanoylphorbol-13-acetate induced transactivation. When cotransfected with infectious HIVNL4-3 DNA, membrane-targeted Raf induces viral replication up to 10-fold over basal levels, as determined by the release of newly synthesized p24 gag protein. Our study clearly demonstrates that the activity of the catalytic domain of Raf in A3.01 T cells is dependent on its cellular localization. The functional consequences of active Raf in T lymphocytes include not only NF-κB activation and transactivation of the HIVNL4-3 LTR but also synthesis and release of HIV particles.

scholarly journals Implication of p38 mitogen-activated protein kinase isoforms (α, β, γ and δ) in CD4+ T-cell infection with human immunodeficiency virus type I

2008 ◽  
Vol 89 (7) ◽  
pp. 1661-1671 ◽  
Author(s):  
Dolores Gutierrez-Sanmartin ◽  
Eduardo Varela-Ledo ◽  
Antonio Aguilera ◽  
Susana Romero-Yuste ◽  
Patricia Romero-Jung ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Cell Death ◽  
T Cell ◽  
Protein Kinase ◽  
Cell Lines ◽  
Cell Apoptosis ◽  
Mitogen Activated Protein Kinase ◽  
Immunodeficiency Virus ◽  
Mitogen Activated Protein ◽  
Hiv 1

The CD4+ T-cell reduction characteristic of human immunodeficiency virus type 1 (HIV-1) infection is thought to result, in addition to infected T-cell death, mainly from uninfected bystander T-cell apoptosis. Nevertheless, the immunological and virological mechanisms leading to T-cell death during HIV-1 infection are not yet fully understood. In the present study, we analysed the individual implication of the p38 mitogen-activated protein kinase (MAPK) isoforms (p38α, p38β, p38γ and p38δ) during apoptosis induced by HIV-1, taking into account that HIV-1 replication is known to be blocked by p38 inhibitors. For this purpose, we used the SupT1 cell line, where death induced by HIV-1 mainly occurs by uninfected bystander cell apoptosis. A variety of SupT1-based cell lines were constructed constitutively expressing, under the control of cytomegalovirus promoter (PCMV), each dominant-negative (dn) p38 isoform and each wild-type p38 isoform as a control. An enhanced green fluorescent protein marker gene, under the control of the HIV-1 promoter, was inserted in all of them. These cell lines were infected with HIV-1 and analysed by flow cytometry. We found that survival in SupT1-based cell lines infected by HIV-1 was increased by the p38αdn, p38γdn and p38δdn isoforms, but not by the p38βdn isoform. HIV-1 replication was delayed most by p38δdn and to a lesser extent by p38αdn and p38γdn. Moreover, these three isoforms, p38αdn, p38γdn and p38δdn, reduced apoptosis induced by HIV-1. These results suggest that, in SupT1-based cell lines, p38α, p38γ and p38δ, but not p38β, are implicated in both HIV-1 induced replication and apoptosis in infected and uninfected bystander cells.

scholarly journals Role of Vif in Stability of the Human Immunodeficiency Virus Type 1 Core

2000 ◽  
Vol 74 (23) ◽  
pp. 11055-11066 ◽  
Author(s):  
Åsa Öhagen ◽  
Dana Gabuzda
Keyword(s):  
Human Immunodeficiency Virus ◽  
Dna Synthesis ◽  
Virus Entry ◽  
Wild Type ◽  
The Core ◽  
Immunodeficiency Virus ◽  
The Stability ◽  
Hiv 1

ABSTRACT The Vif protein of human immunodeficiency virus type 1 (HIV-1) is important for virion infectivity. Previous studies have shown thatvif-defective virions exhibit structural abnormalities in the virus core and are defective in the ability to complete proviral DNA synthesis in acutely infected cells. We developed novel assays to assess the relative stability of the core in HIV-1 virions. Using these assays, we examined the role of Vif in the stability of the HIV-1 core. The integrity of the core was examined following virion permeabilization or removal of the lipid envelope and treatment with various triggers, including S100 cytosol, deoxynucleoside triphosphates, detergents, NaCl, and buffers of different pH to mimic aspects of the uncoating and disassembly process which occurs after virus entry but preceding or during reverse transcription.vif mutant cores were more sensitive to disruption by all triggers tested than wild-type cores, as determined by endogenous reverse transcriptase (RT) assays, biochemical analyses, and electron microscopy. RT and the p7 nucleocapsid protein were released more readily from vif mutant virions than from wild-type virions, suggesting that the internal nucleocapsid is less stably packaged in the absence of Vif. Purified cores could be isolated from wild-type but not vif mutant virions by sedimentation through detergent-treated gradients. These results demonstrate that Vif increases the stability of virion cores. This may permit efficient viral DNA synthesis by preventing premature degradation or disassembly of viral nucleoprotein complexes during early events after virus entry.

scholarly journals An Unrelated Monoclonal Antibody Neutralizes Human Immunodeficiency Virus Type 1 by Binding to an Artificial Epitope Engineered in a Functionally Neutral Region of the Viral Envelope Glycoproteins

2005 ◽  
Vol 79 (9) ◽  
pp. 5616-5624 ◽  
Author(s):  
Xinping Ren ◽  
Joseph Sodroski ◽  
Xinzhen Yang
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Envelope Glycoprotein ◽  
Virus Entry ◽  
Neutralizing Antibody ◽  
Viral Envelope ◽  
Epitope Tag ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Neutralizing antibodies often recognize regions of viral envelope glycoproteins that play a role in receptor binding or other aspects of virus entry. To address whether this is a necessary feature of a neutralizing antibody, we identified the V4 region of the gp120 envelope glycoprotein of human immunodeficiency virus type 1 (HIV-1) as a sequence that is tolerant of drastic change and thus appears to play a negligible role in envelope glycoprotein function. An artificial epitope tag was inserted into the V4 region without a significant effect on virus entry or neutralization by antibodies that recognize HIV-1 envelope glycoprotein sequences. An antibody directed against the artificial epitope tag was able to neutralize the modified, but not the wild-type, HIV-1. Thus, the specific target of a neutralizing antibody need not contribute functionally to the process of virus entry.

scholarly journals The Synthetic Immunomodulator Murabutide Controls Human Immunodeficiency Virus Type 1 Replication at Multiple Levels in Macrophages and Dendritic Cells

2000 ◽  
Vol 74 (17) ◽  
pp. 7794-7802 ◽  
Author(s):  
Edith C. A. Darcissac ◽  
Marie-José Truong ◽  
Joëlle Dewulf ◽  
Yves Mouton ◽  
André Capron ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Dendritic Cells ◽  
Viral Infections ◽  
Mitogen Activated Protein Kinase ◽  
Target Cells ◽  
Suppressive Activity ◽  
Nonspecific Resistance ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Macrophages and dendritic cells are known to play an important role in the establishment and persistence of human immunodeficiency virus (HIV) infection. Besides antiretroviral therapy, several immune-based interventions are being evaluated with the aim of achieving better control of virus replication in reservoir cells. Murabutide is a safe synthetic immunomodulator presenting a capacity to enhance nonspecific resistance against viral infections and to target cells of the reticuloendothelial system. In this study, we have examined the ability of Murabutide to control HIV type 1 (HIV-1) replication in acutely infected monocyte-derived macrophages (MDMs) and dendritic cells (MDDCs). Highly significant suppression of viral replication was consistently observed in Murabutide-treated cultures of both cell types. Murabutide did not affect virus entry, reverse transcriptase activity, or early proviral DNA formation in the cytoplasm of infected cells. However, treated MDMs and MDDCs showed a dramatic reduction in nuclear viral two-long terminal repeat circular form and viral mRNA transcripts. This HIV-1-suppressive activity was not mediated by inhibiting cellular DNA synthesis or by activating p38 mitogen-activated protein kinase. Furthermore, Murabutide-stimulated cells expressed reduced CD4 and CCR5 receptors and secreted high levels of β-chemokines, although neutralization of the released chemokines did not alter the HIV-1-suppressive activity of Murabutide. These results provide evidence that a clinically acceptable immunomodulator can activate multiple effector pathways in macrophages and in dendritic cells, rendering them nonpermissive for HIV-1 replication.

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