scholarly journals Human Herpesvirus 8 Interaction with Target Cells Involves Heparan Sulfate

Virology ◽  
2001 ◽  
Vol 282 (2) ◽  
pp. 245-255 ◽  
Author(s):  
Shaw M. Akula ◽  
Fu-Zhang Wang ◽  
Jeffrey Vieira ◽  
Bala Chandran
Keyword(s):  
Heparan Sulfate ◽  
Human Herpesvirus ◽  
Human Herpesvirus 8 ◽  
Target Cells ◽  
Herpesvirus 8

scholarly journals Human Herpesvirus 8 Envelope Glycoprotein K8.1A Interaction with the Target Cells Involves Heparan Sulfate

2001 ◽  
Vol 75 (16) ◽  
pp. 7517-7527 ◽  
Author(s):  
Fu-Zhang Wang ◽  
Shaw M. Akula ◽  
Naranatt P. Pramod ◽  
Ling Zeng ◽  
Bala Chandran
Keyword(s):  
Cell Surface ◽  
Heparan Sulfate ◽  
Human Herpesvirus ◽  
Human Herpesvirus 8 ◽  
Target Cells ◽  
Dependent Manner ◽  
Unique Region ◽  
Herpesvirus 8 ◽  
Agarose Beads ◽  
Virion Envelope

ABSTRACT Human herpesvirus-8 (HHV-8) or Kaposi's sarcoma-associated herpesvirus K8.1 gene encodes for two immunogenic glycoproteins, gpK8.1A and gpK8.1B, originating from spliced messages. The 228-amino-acid (aa) gpK8.1A is the predominant form associated with the virion envelope, consisting of a 167-aa region identical to gpK8.1B and a 61-aa unique region (L. Zhu, V. Puri, and B. Chandran, Virology 262:237–249, 1999). HHV-8 has a broad in vivo and in vitro cellular tropism, and our studies showed that this may be in part due to HHV-8's interaction with the ubiquitous host cell surface molecule, heparan sulfate (HS). Since HHV-8 K8.1 gene is positionally colinear to the Epstein-Barr virus (EBV) gene encoding the gp350/gp220 protein involved in EBV binding to the target cells, gpK8.1A's ability to interact with the target cells was examined. The gpK8.1A without the transmembrane and carboxyl domains (ΔTMgpK8.1A) was expressed in a baculovirus system and purified. Radiolabeled purified ΔTMgpK8.1A protein bound to the target cells, which was blocked by unlabeled ΔTMgpK8.1A. Unlabeled ΔTMgpK8.1A blocked the binding of [3H]thymidine-labeled purified HHV-8 to the target cells. Binding of radiolabeled ΔTMgpK8.1A to the target cells was inhibited in a dose-dependent manner by soluble heparin, a glycosaminoglycan (GAG) closely related to HS, but not by other GAGs such as chondroitin sulfate A and C, N-acetyl heparin and de-N-sulfated heparin. Cell surface absorbed ΔTMgpK8.1A was displaced by soluble heparin. Radiolabeled ΔTMgpK8.1A also bound to HS expressing Chinese hamster ovary (CHO-K1) cells, and binding to mutant CHO cell lines deficient in HS was significantly reduced. The ΔTMgpK8.1A specifically bound to heparin-agarose beads, which was inhibited by HS and heparin, but not by other GAGs. Virion envelope-associated gpK8.1A was specifically precipitated by heparin-agarose beads. These findings suggest that gpK8.1A interaction with target cells involves cell surface HS-like moieties, and HHV-8 interaction with HS could be in part mediated by virion envelope-associated gpK8.1A.

Human Herpesvirus-8 (HHV-8/KSHV) Induces Reactive Oxygen Species in Endothelial Cells That Facilitate Virus Infection.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 605-605 ◽  
Author(s):  
Jian Feng Wang ◽  
Xuefeng Zhang ◽  
Bala Chandran ◽  
Jerome E. Groopman
Keyword(s):  
Reactive Oxygen Species ◽  
Endothelial Cells ◽  
Virus Entry ◽  
Reactive Oxygen ◽  
Human Herpesvirus ◽  
Human Herpesvirus 8 ◽  
Ros Generation ◽  
Target Cells ◽  
Oxygen Species ◽  
Herpesvirus 8

Abstract Reactive oxygen species (ROS) can activate replication of certain viruses, induce the production of various inflammatory mediators and play a critical role in carcinogenesis and tumor development. Kaposi’s sarcoma (KS) is the most prevalent HIV-associated cancer and is caused by infection with human herpesvirus-8 (HHV-8/KSHV). KS tissue has been reported to possess increased levels of ROS. We studied if ROS generation is related to HHV-8 infection and its role in virus entry into endothelial cells. Incubation of dermal microvascular endothelial cells (DMECs) with highly purified HHV-8 induced rapid increases in the production of intracellular hydrogen peroxide (H2O2), one of the major forms of ROS. Intracellular H2O2 was also induced by the treatment of DMECs with the HHV-8 envelope glycoprotein B (gB). The gB protein possesses an RGD motif, binds to the integrin molecules, alpha3 and beta1, and is a major mediator of virus entry into target cells. To test if it was integrin ligation that induces the production of ROS, we treated DMECs with fibronectin or laminin, the respective natural ligands for alpha3 and beta1 integrins. We observed a similar induction of intracellular ROS in DMECs by either matrix protein. These results indicated that the HHV-8-induced production of ROS was, at least in part, mediated by stimulation of integrins through the RGD-containing viral gB protein. ROS have recently been shown to function as second messengers in cellular signaling. To assess at which steps of cell signaling ROS may be functioning, we studied the signaling cascade in DMECs activated by the HHV-8 gB protein. Previous studies have shown that HHV-8, through gB/integrin interaction, induces cytoskeletal rearrangement and activates focal adhesion kinase (FAK), Src kinase and Akt, which are critical for virus entry into the target cells. We found that the activation of FAK, c-Src or Akt by this viral protein was inhibited by pretreatment with N-acetyl-L-cysteine (NAC), a potent thiol antioxidant. These results suggested that generation of ROS was involved in HHV-8-triggered signaling. We next examined if a change in ROS production modulated HHV-8 virus entry. We used green fluorescent protein (GFP)-labeled HHV-8 at a multiplicity of infection of 5–6, and quantitated the infection by fluorescence analysis of the DMECs. Short term exposure to low concentrations of H2O2 enhanced HHV-8 infection in DMECs, while treatment with NAC significantly decreased infection. These data indicated that ROS generation participated in HHV-8-mediated signaling and entry into target cells. Our study demonstrates a novel role of ROS in virus pathogenesis and provides a framework for the development of novel antioxidant strategies in AIDS-KS treatment.

scholarly journals Human Herpesvirus 8 Envelope-Associated Glycoprotein B Interacts with Heparan Sulfate-like Moieties

Virology ◽  
2001 ◽  
Vol 284 (2) ◽  
pp. 235-249 ◽  
Author(s):  
Shaw M. Akula ◽  
Naranatt P. Pramod ◽  
Fu-Zhang Wang ◽  
Bala Chandran
Keyword(s):  
Heparan Sulfate ◽  
Human Herpesvirus ◽  
Human Herpesvirus 8 ◽  
Glycoprotein B ◽  
Herpesvirus 8

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus (Human Herpesvirus 8) Infection of Human Fibroblast Cells Occurs through Endocytosis

2003 ◽  
Vol 77 (14) ◽  
pp. 7978-7990 ◽  
Author(s):  
Shaw M. Akula ◽  
Pramod P. Naranatt ◽  
Neelam-Sharma Walia ◽  
Fu-Zhang Wang ◽  
Barbara Fegley ◽  
...  
Keyword(s):  
Kaposi's Sarcoma ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Human Herpesvirus 8 ◽  
Target Cells ◽  
Fibroblast Cells ◽  
Coated Pits ◽  
Herpesvirus 8 ◽  
Endocytic Vesicles

ABSTRACT Kaposi's sarcoma (KS)-associated herpesvirus or human herpesvirus 8 (HHV-8) DNA and transcripts have been detected in the B cells, macrophages, keratinocytes, and endothelial and epithelial cells of KS patients. In vitro, HHV-8 infects human B, endothelial, epithelial, and fibroblast cells, as well as animal cells, and the infection is characterized by (i) absence of lytic replication by the input virus and (ii) latent infection. For its initial binding to target cells, HHV-8 uses ubiquitous heparan sulfate molecules via its envelope-associated glycoproteins gB and gpK8.1A. HHV-8 also interacts with the α3β1 integrin via its glycoprotein gB, and virus binding studies suggest that α3β1 is one of the HHV-8 entry receptors (S. M. Akula, N. P. Pramod, F. Z. Wang, and B. Chandran, Cell 108:407-419, 2002). In this study, morphological and biochemical techniques were used to examine the entry of HHV-8 into human foreskin fibroblasts (HFF). HHV-8 was detected in coated vesicles and in large, smooth-surfaced endocytic vesicles. Fusion of viral envelope with the vesicle wall was also observed. In immune electron microscopy, anti-HHV-8 gB antibodies colocalized with virus-containing endocytic vesicles. In fluorescence microscopic analyses, transferrin was colocalized with HHV-8. HHV-8 infection was significantly inhibited by preincubation of cells with chlorpromazine HCl, which blocks endocytosis via clathrin-coated pits, but not by nystatin and cholera toxin B, which blocks endocytosis via caveolae and induces the dissociation of lipid rafts, respectively. Infection was also inhibited by blocking the acidification of endosomes by NH4Cl and bafilomycin A. Inhibition of HHV-8 open reading frame 73 gene expression by chlorpromazine HCl, bafilomycin A, and NH4Cl demonstrated that the virions in the vesicles could proceed to cause an infection. Taken together, these findings suggest that for its infectious entry into HFF, HHV-8 uses clathrin-mediated endocytosis and a low-pH intracellular environment.

scholarly journals Unraveling Viral Interleukin-6 Binding to gp130 and Activation of STAT-Signaling Pathways Independently of the Interleukin-6 Receptor

2009 ◽  
Vol 83 (10) ◽  
pp. 5117-5126 ◽  
Author(s):  
Nina Adam ◽  
Björn Rabe ◽  
Jan Suthaus ◽  
Joachim Grötzinger ◽  
Stefan Rose-John ◽  
...  
Keyword(s):  
Interleukin 6 ◽  
Human Herpesvirus ◽  
Amino Acid Sequences ◽  
The Body ◽  
Activation Mechanism ◽  
Human Herpesvirus 8 ◽  
Target Cells ◽  
Herpesvirus 8 ◽  
Interleukin 6 Receptor ◽  
Direct Binding

ABSTRACT Human herpesvirus 8 encodes a viral version of interleukin-6 (vIL-6) which shows 25% sequence homology with human IL-6. In contrast to human IL-6, which first binds to the IL-6 receptor (IL-6R) and only subsequently associates with the signal transducing receptor subunit gp130, vIL-6 has been shown to directly bind to gp130 without the need of IL-6R. As a functional consequence, vIL-6 can activate far more target cells in the body since all cells express gp130, but only cells such as hepatocytes and some leukocytes express IL-6R. We sought to understand which amino acid sequences within the vIL-6 protein were responsible for its ability to bind and activate gp130 independent of IL-6R. As a first approach, we constructed chimeric IL-6 proteins in which all known gp130 interacting sites (sites II and III) were sequentially transferred from vIL-6 into the human IL-6 protein. To our surprise, human IL-6 carrying all gp130 interacting sites from vIL-6 did not show IL-6R-independent gp130 activation. Even more surprisingly, the loop between helix B and C of vIL-6, clearly shown in the crystal structure not to be in contact with gp130, is indispensable for direct binding to and activation of gp130. This points to an IL-6R induced change of site III conformation in human IL-6, which is already preformed in vIL-6. These data indicate a novel activation mechanism of human IL-6 by the IL-6R that will be important for the construction of novel hyperactive cytokine variants.

scholarly journals Human Herpesvirus 8 Envelope Glycoprotein B Mediates Cell Adhesion via Its RGD Sequence

2003 ◽  
Vol 77 (5) ◽  
pp. 3131-3147 ◽  
Author(s):  
Fu-Zhang Wang ◽  
Shaw M. Akula ◽  
Neelam Sharma-Walia ◽  
Ling Zeng ◽  
Bala Chandran
Keyword(s):  
Cell Adhesion ◽  
Cell Surface ◽  
Focal Adhesion ◽  
Kaposi's Sarcoma ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Human Herpesvirus 8 ◽  
Target Cells ◽  
Herpesvirus 8 ◽  
Rgd Sequence

ABSTRACT Human herpesvirus 8 (HHV-8) or Kaposi's sarcoma-associated herpesvirus, implicated in the pathogenesis of Kaposi's sarcoma, utilizes heparan sulfate-like molecules to bind the target cells via its envelope-associated glycoproteins gB and gpK8.1A. HHV-8-gB possesses the Arg-Gly-Asp (RGD) motif, the minimal peptide region of many proteins known to interact with subsets of host cell surface integrins. HHV-8 utilizes α3β1 integrin as one of the receptors for its entry into the target cells via its gB interaction and induces the activation of focal adhesion kinase (FAK) (S. M. Akula, N. P. Pramod, F.-Z. Wang, and B. Chandran, Cell 108:407-419, 2002). Since FAK activation is the first step in the outside-in signaling necessary for integrin-mediated cytoskeletal rearrangements, cell adhesions, motility, and proliferation, the ability of HHV-8-gB to mediate the target cell adhesion was examined. A truncated form of gB without the transmembrane and carboxyl domains (gBΔTM) and a gBΔTM mutant (gBΔTM-RGA) with a single amino acid mutation (RGD to RGA) were expressed in a baculovirus system and purified. Radiolabeled HHV-8-gBΔTM, gBΔTM-RGA, and ΔTMgpK8.1A proteins bound to the human foreskin fibroblasts (HFFs), human dermal microvascular endothelial (HMVEC-d) cells, human B (BJAB) cells, and Chinese hamster ovary (CHO-K1) cells with equal efficiency, which was blocked by preincubation of proteins with soluble heparin. Maxisorp plate-bound gBΔTM protein induced the adhesion of HFFs and HMVEC-d and monkey kidney epithelial (CV-1) cells in a dose-dependent manner. In contrast, the gBΔTM-RGA and ΔTMgpK8.1A proteins did not mediate adhesion. Adhesion mediated by gBΔTM was blocked by the preincubation of target cells with RGD-containing peptides or by the preincubation of plate-bound gBΔTM protein with rabbit antibodies against gB peptide containing the RGD sequence. In contrast, adhesion was not blocked by the preincubation of plate-bound gBΔTM protein with heparin, suggesting that the adhesion is mediated by the RGD amino acids of gB, which is independent of the heparin-binding domain of gB. Integrin-ligand interaction is dependent on divalent cations. Adhesion induced by the gBΔTM was blocked by EDTA, thus suggesting the role of integrins in the observed adhesions. Focal adhesion components such as FAK and paxillin were activated by the binding of gBΔTM protein to the target cells but not by gBΔTM-RGA protein binding. Inhibition of FAK phosphorylation by genistein blocked gBΔTM-induced FAK activation and cell adhesion. These findings suggest that HHV-8-gB could mediate cell adhesion via its RGD motif interaction with the cell surface integrin molecules and indicate the induction of cellular signaling pathways, which may play roles in the infection of target cells and in Kaposi's sarcoma pathogenesis.

scholarly journals An unusual dependence of human herpesvirus-8 glycoproteins-induced cell-to-cell fusion on heparan sulfate

2009 ◽  
Vol 390 (3) ◽  
pp. 382-387 ◽  
Author(s):  
Vaibhav Tiwari ◽  
Nissar A. Darmani ◽  
Gerald R. Thrush ◽  
Deepak Shukla
Keyword(s):  
Heparan Sulfate ◽  
Cell Fusion ◽  
Human Herpesvirus ◽  
Human Herpesvirus 8 ◽  
Herpesvirus 8

Characterization of Kaposi sarcoma–associated herpesvirus/human herpesvirus–8 infection of human vascular endothelial cells: early events

Blood ◽  
2002 ◽  
Vol 100 (3) ◽  
pp. 888-896 ◽  
Author(s):  
Bruce J. Dezube ◽  
Maria Zambela ◽  
David R. Sage ◽  
Jian-Feng Wang ◽  
Joyce D. Fingeroth
Keyword(s):  
Endothelial Cells ◽  
Culture System ◽  
Vascular Endothelial Cells ◽  
Human Herpesvirus ◽  
Kaposi Sarcoma ◽  
Human Herpesvirus 8 ◽  
Target Cells ◽  
Reading Frame ◽  
Herpesvirus 8 ◽  
Virus Attachment

Abstract Kaposi sarcoma–associated herpesvirus (KSHV)/human herpesvirus-8 (HHV-8) is causally associated with Kaposi sarcoma (KS). The absence of a cell culture system that effectively reproduces the composite mechanisms governing initiation and maintenance of HHV-8 infection (lytic and latent) in KS endothelial cells, however, has left important questions unanswered. Here, we report a culture system in which the earliest events that accompany HHV-8 infection could be surveyed in primary endothelial cells. Binding of HHV-8 to microvascular dermal endothelial cells (MVDECs) was directly compared with other primary target cells implicated in HHV-8–associated diseases. Virus attachment, fusion, internalization and transport within MVDECs was monitored by electron microscopy. Studies of genome configuration revealed that rapid circularization of the viral DNA occurred on entry, though by 72 hours after infection linear DNAs accumulated and early as well as late lytic RNAs (T1.1, K8.1) could be detected. The latency transcripts (LT1/LT2) were first detected on day 8, demonstrating that both lytic and latent infection were initiated. Although most lytic transcripts accrued until passage, open-reading frame–74 RNAs fluctuated with a fixed periodicity, suggesting that early replication after infection of MVDECs was synchronous.

scholarly journals Cell Surface Heparan Sulfate Is a Receptor for Human Herpesvirus 8 and Interacts with Envelope Glycoprotein K8.1

2001 ◽  
Vol 75 (23) ◽  
pp. 11583-11593 ◽  
Author(s):  
Alexander Birkmann ◽  
Kerstin Mahr ◽  
Armin Ensser ◽  
Svenja Yağuboğlu ◽  
Fritz Titgemeyer ◽  
...  
Keyword(s):  
Cell Surface ◽  
Heparan Sulfate ◽  
Target Cell ◽  
Envelope Glycoprotein ◽  
Mammalian Cells ◽  
Mutant Cell ◽  
Human Herpesvirus ◽  
Human Herpesvirus 8 ◽  
Cell Recognition ◽  
Herpesvirus 8

ABSTRACT An immunodominant envelope glycoprotein is encoded by the human herpesvirus 8 (HHV-8) (also termed Kaposi's sarcoma-associated herpesvirus) K8.1 gene. The functional role of glycoprotein K8.1 is unknown, and recognizable sequence homology to K8.1 is not detectable in the genomes of most other closely related gammaherpesviruses, such as herpesvirus saimiri or Epstein-Barr virus. In search for a possible function for K8.1, we expressed the ectodomain of K8.1 fused to the Fc part of human immunoglobulin G1 (K8.1ΔTMFc). K8.1ΔTMFc specifically bound to the surface of cells expressing glycosaminoglycans but not to mutant cell lines negative for the expression of heparan sulfate proteoglycans. Binding of K8.1ΔTMFc to mammalian cells could be blocked by heparin. Interestingly, the infection of primary human endothelial cells by HHV-8 could also be blocked by similar concentrations of heparin. The specificity and affinity of these interactions were then determined by surface plasmon resonance measurements using immobilized heparin and soluble K8.1. This revealed that K8.1 binds to heparin with an affinity comparable to that of glycoproteins B and C of herpes simplex virus, which are known to be involved in target cell recognition by binding to cell surface proteoglycans, especially heparan sulfate. We conclude that cell surface glycosaminoglycans play a crucial role in HHV-8 target cell recognition and that HHV-8 envelope protein K8.1 is at least one of the proteins involved.

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