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| 1. Molecular
Mechanism of HIV-1 Infection of Resting CD4 T cells |
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A major current focus of my laboratory is on the molecular interaction of HIV-1 with human resting CD4 T cells. We are particularly interested in HIV-1 interaction with the chemokine co-receptor CXCR4 in mediating actin cytoskeletal rearrangement required for viral intracellular migration. In the past, our studies have focused on several aspects of virus-CD4 T cell interaction at the cortical actin layer. We studied the effects of viral envelope signaling on the actin rearrangement of resting CD4 T cells. These studies led to the discovery that treatment of resting CD4 T cells with gp120 or HIV-1 triggers the polymerization and depolymerization of filamentous actin (F-actin) in the cortical region. We also identified cofilin as a molecular target of CXCR4 signaling triggered by HIV gp120 to promote the cortical actin dynamics. On the basis of our studies, we proposed a molecular model suggesting a direct involvement of the cortical actin and cofilin in viral latent infection of resting T cells (Figure 1) (1). |
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Fig 1. Model of gp120-CXCR4 signaling in mediating cofilin activation and HIV latent infection. HIV envelope binding to CXCR4 triggers membrane fusion and signaling transduction that leads to cofilin activation. Following fusion, the viral preintegration complex is directly anchored onto F-actin to facilitate reverse transcription. Subsequent actin activity triggered by cofilin activation promotes viral nuclear migration. Cofilin activation increases cortical actin dynamics and actin treadmilling, which promote the movement of the viral preintegration complex toward the center of the cell, allowing HIV to gain access to the perinuclear and nuclear regions. |
My laboratory will continue to focus on
studying molecular details of how HIV interacts with the cortical actin
cytoskeleton in human resting CD4 T cells. This is an important, yet
largely unexplored research field. Although our previous studies have
established a role of the cortical actin in HIV latent infection of
resting T cells, the molecular mechanisms are currently not defined.
Mapping these molecular events is fundamental to establishing a
framework for understanding the early events in viral infection of CD4
T cells.
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| 2.
Pre-integration of Transcription of HIV-1 DNA |
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In addition to studying HIV interaction with actin, my laboratory is currently also studying pre-integration transcription. In the past, we have demonstrated that pre-integration transcription is a normal process occurring during HIV-1 infection of CD4 T cells and macrophages (2-6). We are in the process of identifying the non-integrated, transcribing HIV DNA templates (Figure 2). |
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Figure 2. Pre-integration transcription of HIV-1 Viral transcription in the absence of integration generates all classes of viral transcripts, but only early proteins such as Tat, Rev, and Nef are synthesized at low levels. Tat and Nef can modulate cellular conditions. Viral replication does not occur without integration, but infection by a second virus can rescue the unintegrated viral genomes. |
3. Development of a Rev-dependent Lentiviral Vector Targeting HIV-1 Infection |
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My laboratory has developed a
Rev-dependent lentiviral vector to target HIV-infected macrophages and
T cells, using a bacterial toxin, anthrolysin O from Bacillus anthracis (7-9). We are in the process of
constructing a series of Rev-dependent vectors carrying a variety of
bacterial toxins and human apoptotic genes to induce the decay of viral
reservoirs in infected macrophages and T cells (Figure 3).
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Figure 3. Fluorescent
microscopy of GFP expression in macrophages infected with HIV-1 and the
Rev-dependent GFP lentiviral vector. Cells were infected with HIV-1 (AD8) and then infected with the Rev-dependent lentivirus vNL-GFP-RRE-SA. Infected cells were examined with fluorescent microscopy. The left and right panels show the bright and green fluorescent fields of the same cells. Red arrows indicate an HIV-1-infected cell expressing the GFP-protein. |
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References 1. Yoder, A., D. Yu, L. Dong, S. R. Iyer, X. Xu, J. Kelly, J. Liu, W. Wang, P. J. Vorster, L. Agulto, D. A. Stephany, J. N. Cooper, J. W. Marsh and Y. Wu. 2008. HIV-1 envelope-CXCR4 interaction activates cofilin to overcome cortical actin restriction in resting CD4 T cells. Cell, 134:782-792. 2. Wu, Y. and J. W. Marsh. 2001. Selective transcription and modulation of resting T cell activity by preintegrated HIV DNA. Science 293(5534), 1503-6. 3. Wu, Y. and J. W. Marsh. 2003. Early transcription from nonintegrated DNA in human immunodeficiency virus infection. J. Virol 77(19), 10376-10382. 4. Wu, Y. and J. Marsh. 2003. Gene transcription in HIV infection. Microbes and Infection 5,1023-27. 5. Wu, Y. 2004. HIV-1 gene expression: lessons from provirus and non-integrated DNA. Retrovirology 1:13 (highly accessed) 6. Young, J., Z. Tang, Q. Yu, D. Yu, and Y. Wu. 2008. Selective killing of HIV-1-positive macrophages and T cells by a Rev-dependent lentivirus carrying anthrolysin O from Bacillus anthracis. Retrovirology 5:36 (highly accessed) 7. Kelly, J., H. Beddall, D. Yu, J. W. Marsh and Y. Wu. 2008. Persistence of transcriptional activity from non-integrated HIV-1 DNA in macrophages. Virology 372(2), 300-12. 8. Wu, Y., M. H. Beddall and J. W. Marsh 2007. Rev-dependent indicator T cell line. Current HIV Research. 5(4), 394-402 9.
Wu, Y., M. H. Beddall and
J. W. Marsh. 2007.
Rev-dependent expression vector. Retrovirology 4:12
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National Center for Biodefense & Infectious Diseases
George Mason University
10900 University Drive
Manassas, VA 20110
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