Characterization of hepatitis C virus envelope glycoprotein complexes expressed by recombinant vaccinia viruses

Ralston, Robert; Thudium, Kent; Berger, Kimberly; Kuo, C.; Gervase, Barbara; Hall, John; Selby, Mark; Kuo, George; Houghton, Michael; Choo, Qui‐Lim

doi:10.1128/jvi.67.11.6753-6761.1993

Cited by 195 publications

(70 citation statements)

References 38 publications

(58 reference statements)

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“…Recombinant vaccinia viruses expressing HCV genes derived from the HCV-1 strain (genotype 1A) have previously been described. 13,28 The 2 vaccinia constructs used in this study are the vv-core/E1 encoding aa 1-340 of HCV and vv-E1/E2/NS2 encod- ing aa 136-966. Wild-type vaccinia virus (Western Reserve strain) was used as control.…”

Section: Methodsmentioning

confidence: 99%

Identification of immunodominant hepatitis C virus (HCV)-specific cytotoxic T-cell epitopes by stimulation with endogenously synthesized HCV antigens

et al. 2001

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Hepatitis C virus (HCV)-specific CD8 ؉ cytotoxic T lymphocytes (CTL) are believed to play an important role in the pathogenesis of liver cell injury and viral clearance in HCV infection. Because HCV does not efficiently infect human cells in vitro and primary infected hepatocytes cannot be used as stimulator/target cells for CTL analysis, development of efficient systems to activate and expand CTL in vitro, reproducing antigen presentation to CTL occurring during natural infection, is mandatory to study CTL activity and to define the hierarchy of immunodominance of CTL epitopes. To achieve this goal, 5 different defective adenoviruses carrying structural and nonstructural HCV genes (core, core-E1-E2, E2, NS3-NS4A, NS3-NS5A) were used to in- The hepatitis C virus (HCV) is a positive-stranded RNA virus of 9.5 Kb that belongs to the Flaviviridae family. 1 At least 400 million people are infected with HCV worldwide, and up to 85% of infected patients can become chronic carriers. 2 The HLA class I-restricted T-cell response plays an important role in the control of viral infections 3 by lysing virus-infected cells and by secreting cytokines involved in the inhibition of viral replication and in the recruitment of nonspecific inflammatory cells at the site of viral replication. 4 Although the cytotoxic T lymphocyte (CTL) response against HCV is detectable in the peripheral blood and the intrahepatic infiltrates of patients with acute and chronic HCV infection, 5-17 its role in HCV pathogenesis is still largely undefined. In particular, the inability of HCV to infect human cells in vitro has severely hampered the study of the CTL function and the development of experimental systems to mimic antigen processing occurring in vivo is urgently needed. This limitation has partially been overcome by the use of synthetic peptides to stimulate HCV-specific CTL responses. [6][7][8]10,11,[15][16][17] By this approach, however, subdominant epitopes can be strongly stimulatory, and cryptic sequences can acquire the capacity to activate a CTL response, if multiple rounds of stimulation are performed in vitro.Therefore, the hierarchy of protective responses and the balance present in vivo between T-cell populations of different specificity and functional activity may not be accurately reproduced in vitro by the peptide-stimulation method. Recombinant viruses and plasmids encoding viral genes represent a particularly promising approach to induce antibody-and cellmediated immune responses, and they have recently been used for in vivo immunization against HCV. [18][19][20][21][22][23] These vectors allow the endogenous synthesis of viral antigens to occur inside human cells, thereby creating the conditions for natural cytosolic generation of viral peptides, their transport to the lumen of the endoplasmic reticulum, binding to HLA class I molecules, and presentation on the cell surface. In this study, we describe the use of 4 adenoviruses carrying the core, core-E1-E2, E2, NS3-NS4A, or NS3-NS5A genes to stimulate cytotoxic T cells in vit...

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Section: Methodsmentioning

confidence: 99%

Identification of immunodominant hepatitis C virus (HCV)-specific cytotoxic T-cell epitopes by stimulation with endogenously synthesized HCV antigens

et al. 2001

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“…The recombinant E2 protein was produced and purified from Chinese hamster ovary cells expressing HCV (type 1a) complementary DNA (cDNA) encoding the envelope region E2 (amino acids 384-715) as previously described. 9,11,14 The recombinant E1/E2 heterodimer was produced in insect cells by recombinant baculovirus Ac827 expressing HCV (type 1b) cDNA encoding the envelope region E1/E2 (amino acids 155-810) and purified as described previously. 9,14,15 Protein concentrations were measured by BCA Protein Assay Reagent (Pierce, Rockford, IL).…”

Section: Methodsmentioning

confidence: 99%

High titers of antibodies inhibiting the binding of envelope to human cells correlate with natural resolution of chronic hepatitis C

et al. 1998

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“…An interesting difference between BVDV and HCV may reside in the nature of the linkage between E1 and E2 proteins within the dimers. While for BVDV the complex was clearly shown to be disulfide bonded both in cell lysates and mature virions [5,8], purified HCV glycoprotein complexes expressed in recombinant systems were reported to be associated mainly in a noncovalent fashion [9]. Only a fraction of E1 and E2 present in the lysates of cells infected with vaccinia virus-HCV recombinants has been shown to be associated via disulfide linkages which was suggested to be part of a nonproductive pathway leading to misfolding and aggregation [10].…”

mentioning

confidence: 99%

Role of disulfide bond formation in the folding and assembly of the envelope glycoproteins of a pestivirus

Branza‐Nichita

Lazar

Durantel

et al. 2002

Biochemical and Biophysical Research Communications

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Bovine viral diarrhea virus (BVDV) is a pestivirus member of the Flaviviridae family, closely related to, and used as a surrogate model for the hepatitis C virus. Its envelope contains the E1 and E2 glycoproteins, disulfide linked into homo- and heterodimers. In this study, we investigate the role of disulfide bond formation in the folding, assembly, and stability of BVDV glycoproteins. We provide molecular evidence that intact disulfide bonds are critical for the acquirement of a stable conformation of E2 monomers. Forcing the E2 glycoproteins to adopt a reduced conformation either co- or post-translationally before assembly into dimers, determines their misfolding and degradation by proteasome. In contrast, dimerization of E2 glycoproteins results in a conformation resistant to reducing agents and degradation. Furthermore, inhibition of the ER-alpha-mannosidase activity leads to impairment of misfolded E2 degradation, demonstrating the involvement of this enzyme in targeting viral proteins towards proteasomal degradation.

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Characterization of hepatitis C virus envelope glycoprotein complexes expressed by recombinant vaccinia viruses

Cited by 195 publications

References 38 publications

Identification of immunodominant hepatitis C virus (HCV)-specific cytotoxic T-cell epitopes by stimulation with endogenously synthesized HCV antigens

Identification of immunodominant hepatitis C virus (HCV)-specific cytotoxic T-cell epitopes by stimulation with endogenously synthesized HCV antigens

High titers of antibodies inhibiting the binding of envelope to human cells correlate with natural resolution of chronic hepatitis C

Role of disulfide bond formation in the folding and assembly of the envelope glycoproteins of a pestivirus

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