Inefficient Formation of a Complex among CXCR4, CD4 and gp120 in U937 Clones Resistant to X4 gp120–gp41-Mediated Fusion

Norwood, David; Feng, Yanru; Moriuchi, Masako; Jones-Trower, Agnes; Stantchev, Tzanko S.; Moriuchi, Hiroyuki; Broder, Christopher C.; Dimitrov, Dimiter S.

doi:10.1006/exmp.1999.2299

Cited by 14 publications

(14 citation statements)

References 20 publications

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“…This result was expected, given that this MAb was obtained as a result of immunization with a peptide equivalent to the N terminus of CXCR4 (57). In contrast, the reciprocal chimera (2444) was recognized by the entire panel of MAbs save for the N-terminal MAb 4G10.…”

Section: Generation and Characterization Of Mabs To Cxcr4mentioning

confidence: 70%

“…We believe that this is due to conformational heterogeneity for several reasons. First, the entire panel of MAbs bound to conformation-dependent epitopes, since solubilization of CXCR4 with a variety of detergents prevented immunoprecipitation of this receptor by all of the MAbs, except for 4G10, which recognizes a linear determinant in the N-terminal domain (57). Second, the known posttranslational modifications of CXCR4 (N-linked glycosylation and sulfation of the N-terminal domain) had little or no effect on antibody reactivity (3,8,23).…”

Section: Discussionmentioning

confidence: 99%

“…Coexpression of CD4 with CXCR4 in 293T cells did not affect MAb reactivity with CXCR4 (data not shown). In addition to MAbs 1, 2, 8, 12, 16, 17, and 18, we also used 12G5, a conformationdependent MAb that recognizes a determinant in the first and second extracellular loops of CXCR4 (6,22), and MAb 4G10, which was generated against the N-terminal domain of CXCR4 (57). The specificity of the newly developed MAbs was also confirmed by treating Jurkat cells with the CXCR4 inhibitor T22 ( Fig.…”

Section: Generation and Characterization Of Mabs To Cxcr4mentioning

confidence: 99%

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Antigenically Distinct Conformations of CXCR4

Baribaud

Edwards

Sharron

et al. 2001

J Virol

144

156

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The major human immunodeficiency virus type 1 (HIV-1) coreceptors are the chemokine receptors CCR5 and CXCR4. The patterns of expression of the major coreceptors and their use by HIV-1 strains largely explain viral tropism at the level of entry. However, while virus infection is dependent upon the presence of CD4 and an appropriate coreceptor, it can be influenced by a number of factors, including receptor concentration, affinity between envelope gp120 and receptors, and potentially receptor conformation. Indeed, seven-transmembrane domain receptors, such as CCR5, can exhibit conformational heterogeneity, although the significance for virus infection is uncertain. Using a panel of monoclonal antibodies (MAbs) to CXCR4, we found that CXCR4 on both primary and transformed T cells as well as on primary B cells exhibited considerable conformational heterogeneity. The conformational heterogeneity of CXCR4 explains the cell-type-dependent ability of CXCR4 antibodies to block chemotaxis to stromal cell-derived factor 1␣ and to inhibit HIV-1 infection. In addition, the MAb most commonly used to study CXCR4 expression, 12G5, recognizes only a subpopulation of CXCR4 molecules on all primary cell types analyzed. As a result, CXCR4 concentrations on these important cell types have been underestimated to date. Finally, while the factors responsible for altering CXCR4 conformation are not known, we found that they do not involve CXCR4 glycosylation, sulfation of the N-terminal domain of CXCR4, or pertussis toxin-sensitive G-protein coupling. The fact that this important HIV-1 coreceptor exists in multiple conformations could have implications for viral entry and for the development of receptor antagonists.The discovery of the receptors used by human immunodeficiency virus type 1 (HIV-1) to infect cells, coupled with a greater understanding of the membrane fusion-inducing conformational changes undergone by the viral envelope protein (Env) upon receptor binding, has identified several promising drug and vaccine targets (reviewed in reference 12). The viral Env protein binds cell surface CD4 with a high affinity, resulting in conformational changes that enable Env to bind a coreceptor (32,54,56). Coreceptor binding is thought to trigger the ability of Env to mediate fusion between the viral and cellular membranes. The major HIV-1 coreceptors are the chemokine receptors CCR5 and CXCR4 (reviewed in reference 11). The R5 virus strains that use CCR5 with CD4 to infect cells are largely responsible for virus transmission and are typically macrophage tropic. The accrual of mutations in Env can lead to X4 virus strains that use CXCR4 in place of CCR5 or R5X4 virus strains that can use both receptors. While X4 virus strains do not always evolve in infected individuals, their emergence is a harbinger of progression to AIDS (51, 52).While virus infection is dependent upon the presence of CD4 and an appropriate coreceptor, it can be influenced by receptor concentration (21,29,30,43,48), affinity between Env and receptors (28), and potenti...

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Section: Generation and Characterization Of Mabs To Cxcr4mentioning

confidence: 70%

Section: Discussionmentioning

confidence: 99%

Section: Generation and Characterization Of Mabs To Cxcr4mentioning

confidence: 99%

See 1 more Smart Citation

Antigenically Distinct Conformations of CXCR4

Baribaud

Edwards

Sharron

et al. 2001

J Virol

144

156

View full text Add to dashboard Cite

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“…It has been shown that modifying the codistribution of CD4 and coreceptor molecules at the points of virus-cell contact may alter the process of pore fusion formation (Dimitrov et al, 1999;Xiao et al, 2000), thereby regulating HIV-1 infection. However, our results indicate that the effect of HDAC6 on Env-mediated cell Western blot analysis of ␣-tubulin acetylation in PHA-activated PBLs treated with TSA (2 M), NaBut (500 M), Taxol (5 M), and Nocodazole (5 M).…”

Section: Discussionmentioning

confidence: 99%

Histone Deacetylase 6 Regulates Human Immunodeficiency Virus Type 1 Infection

Valenzuela-Fernández

Álvarez

Gordon-Alonso

et al. 2005

MBoC

115

212

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Efficient human immunodeficiency virus (HIV)-1 infection depends on multiple interactions between the viral gp41/gp120envelope (Env) proteins and cell surface receptors. However, cytoskeleton-associated proteins that modify membrane dynamics may also regulate the formation of the HIV-mediated fusion pore and hence viral infection. Because the effects of HDAC6-tubulin deacetylase on cortical ␣-tubulin regulate cell migration and immune synapse organization, we explored the possible role of HDAC6 in HIV-1-envelope-mediated cell fusion and infection. The binding of the gp120 protein to CD4؉ -permissive cells increased the level of acetylated ␣-tubulin in a CD4-dependent manner. Furthermore, overexpression of active HDAC6 inhibited the acetylation of ␣-tubulin, and remarkably, prevented HIV-1 envelopedependent cell fusion and infection without affecting the expression and codistribution of HIV-1 receptors. In contrast, knockdown of HDAC6 expression or inhibition of its tubulin deacetylase activity strongly enhanced HIV-1 infection and syncytia formation. These results demonstrate that HDAC6 plays a significant role in regulating HIV-1 infection and Env-mediated syncytia formation. INTRODUCTIONHuman immunodeficiency virus (HIV) infection is initiated by the virus binding to the cell surface after CD4 engagement and HIV-1 fusion with the plasma membrane (Stein et al., 1987;Maddon et al., 1988;McClure et al., 1988;Moore et al., 1997). The main coreceptors for HIV-1 infection have been shown to be CXCR4 and CCR5, which represented a major advance in understanding the mechanism of the HIV-1 infection (Cocchi et al., 1995;Alkhatib et al., 1996;Bleul et al., 1996;Choe et al., 1996;Deng et al., 1996;Doranz et al., 1996;Dragic et al., 1996;Feng et al., 1996). As a result, it has been proposed that the sequential binding of gp120 viral protein to CD4 and to one coreceptor induces specific conformational changes in gp41, facilitating viral fusion with cell membrane (Clapham and McKnight, 2002).The cell cytoskeleton is involved in the early events of viral infection, regulating viral penetration and genome uncoating, the movement of viral capsids, and integration of the viral genome. Accordingly, actin and microtubules are required for the efficient entry of herpes simplex type 1 and simian virus 40, respectively (Pelkmans et al., 2002;Marozin et al., 2004). It has been shown that disrupting the actin network can inhibit infection of HIV-1 and fusion with the host cell (Iyengar et al., 1998;Jernigan et al., 2000), presumably by disrupting the colocalization of CD4 and CXCR4 (Iyengar et al., 1998). In addition, the actin cytoskeleton seems to be necessary for activation of the reverse transcription complex (Bukrinskaya et al., 1998). However, little is known about the role of cytoskeleton-related enzymes in the control of HIV fusion and infection.Histone deacetylase 6 (HDAC6) is exclusively located in the cytoplasm, and it regulates the acetylation of ␣-tubulin (Hubbert et al., 2002;Matsuyama et al., 2002;Haggarty et al., 2003;Z...

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“…These data show that the bound antibodies dissociate on Rac1 inhibition, which is indicative for a conformational change of CXCR4 induced by Rac1 inhibition. To substantiate this notion, we used the amino-terminus specific CXCR4 mAb, 4G10, 21 which is a conformation-independent antibody. Interestingly, the conformation-independent CXCR4 antibody did not dissociate from its receptor after Rac1 inhibition in an antibody-feeding experiment ( Figure 3D).…”

Section: Rac1 Inhibition Causes a Conformational Change Of Cxcr4mentioning

confidence: 99%