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Epstein-Barr virus binding to CD21 activates the initial viral promoter via NF-kappaB induction.

Sugano N, Chen W, Roberts ML, Cooper NR - J. Exp. Med. (1997)

Bottom Line: Epstein-Barr virus (EBV), an oncogenic human herpesvirus, binds to and infects normal human B lymphocytes via CD21, the CR2 complement receptor.Studies of the mechanisms that enable EBV to infect nonactivated, noncycling B cells provide compelling evidence for a sequence of events in which EBV binding to CD21 on purified resting human B cells rapidly activates the NF-kappaB transcription factor, which, in turn, binds to and mediates transcriptional activation of Wp, the initial viral latent gene promoter.Thus, EBV binding to its cellular receptor on resting B cells triggers an NF-kappaB-dependent intracellular signaling pathway which is required for infection.

View Article: PubMed Central - PubMed

Affiliation: Scripps Research Institute, Department of Immunology, La Jolla, California 92037, USA.

ABSTRACT
Epstein-Barr virus (EBV), an oncogenic human herpesvirus, binds to and infects normal human B lymphocytes via CD21, the CR2 complement receptor. Studies of the mechanisms that enable EBV to infect nonactivated, noncycling B cells provide compelling evidence for a sequence of events in which EBV binding to CD21 on purified resting human B cells rapidly activates the NF-kappaB transcription factor, which, in turn, binds to and mediates transcriptional activation of Wp, the initial viral latent gene promoter. Thus, EBV binding to its cellular receptor on resting B cells triggers an NF-kappaB-dependent intracellular signaling pathway which is required for infection.

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Related in: MedlinePlus

NF-κB induction by CD21 ligands. (A) NF-κB activation by EBV. Nuclear extracts from purified resting B cells incubated with B95-8 EBV  for the designated times were analyzed by EMSA for ability to bind to an NF-κB consensus probe. (B) Specificity of NF-κB binding. Homologous wild  type (wt) and mutant (mut) NF-κB probes were evaluated for ability to inhibit binding of the NF-κB consensus probe by EMSA. (C) NF-κB activation  by C3dg. Nuclear extracts from purified resting B cells incubated with microbeads coated with C3dg or BSA were evaluated for the presence of activated  NF-κB by EMSA. (D) CD21 dependence of NF-κB activation. Purified resting B cells were incubated with soluble gp105 or OKB7 for 1 h, or with the  same amounts of gp105 (EBV + gp105) or OKB7 (EBV + OKB7) for 1 h before EBV addition. Nuclear extracts were prepared 15 min after EBV addition. B cells were also incubated in BSA- or gp105-coated plastic culture wells (pl.) for 15 min. Ability to bind to an NF-κB consensus probe was evaluated by EMSA. (E) Composition of NF-κB. Nuclear extracts from purified resting B cells 30 min after EBV addition were incubated with p50, p65, c-rel  or p52 Abs to NF-κB subunits before addition of the NF-κB consensus probe and analysis by EMSA. (F, top) Effect of calphostin C on NF-κB activation. Nuclear extracts prepared 30 min after EBV addition to purified resting B cells that had been preincubated for 2 h with calphostin C (50 nM) or  buffer, were examined for NF-κB activation by EMSA. (F, bottom) Assessment of IκBα. The same samples were evaluated for IκBα by the Western blotting procedure. Control lanes (−) do not contain EBV.
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Figure 1: NF-κB induction by CD21 ligands. (A) NF-κB activation by EBV. Nuclear extracts from purified resting B cells incubated with B95-8 EBV for the designated times were analyzed by EMSA for ability to bind to an NF-κB consensus probe. (B) Specificity of NF-κB binding. Homologous wild type (wt) and mutant (mut) NF-κB probes were evaluated for ability to inhibit binding of the NF-κB consensus probe by EMSA. (C) NF-κB activation by C3dg. Nuclear extracts from purified resting B cells incubated with microbeads coated with C3dg or BSA were evaluated for the presence of activated NF-κB by EMSA. (D) CD21 dependence of NF-κB activation. Purified resting B cells were incubated with soluble gp105 or OKB7 for 1 h, or with the same amounts of gp105 (EBV + gp105) or OKB7 (EBV + OKB7) for 1 h before EBV addition. Nuclear extracts were prepared 15 min after EBV addition. B cells were also incubated in BSA- or gp105-coated plastic culture wells (pl.) for 15 min. Ability to bind to an NF-κB consensus probe was evaluated by EMSA. (E) Composition of NF-κB. Nuclear extracts from purified resting B cells 30 min after EBV addition were incubated with p50, p65, c-rel or p52 Abs to NF-κB subunits before addition of the NF-κB consensus probe and analysis by EMSA. (F, top) Effect of calphostin C on NF-κB activation. Nuclear extracts prepared 30 min after EBV addition to purified resting B cells that had been preincubated for 2 h with calphostin C (50 nM) or buffer, were examined for NF-κB activation by EMSA. (F, bottom) Assessment of IκBα. The same samples were evaluated for IκBα by the Western blotting procedure. Control lanes (−) do not contain EBV.

Mentions: Nuclear extracts prepared from highly purified resting (nonactivated) human tonsil B cells incubated with EBV for varying periods of time at 37°C were assessed for ability to bind to an NF-κB consensus probe in gel shift assays. Nuclear NF-κB–binding activity increased rapidly from low constitutive levels to reach peak values 15–30 min after EBV addition (Fig. 1 A). Activation was inhibited by the unlabeled wild-type oligonucleotide, but not by a mutant oligonucleotide, demonstrating specificity (Fig. 1 B). After a modest decline, EBV-induced NF-κB–binding activity increased 24 h after EBV addition, likely due to the actions of EBNA 2 and latent membrane protein 1, two EBV latent genes expressed in the first days after infection (13, 14) which activate NF-κB (15, 16). Similar studies with purified tonsil B cells from >10 individuals with two EBV strains (Akata and B95-8) gave comparable results. C3dg-coated microbeads gave the same pattern of rapid NF-κB activation (Fig. 1 C) but without the second increase 24 h later.


Epstein-Barr virus binding to CD21 activates the initial viral promoter via NF-kappaB induction.

Sugano N, Chen W, Roberts ML, Cooper NR - J. Exp. Med. (1997)

NF-κB induction by CD21 ligands. (A) NF-κB activation by EBV. Nuclear extracts from purified resting B cells incubated with B95-8 EBV  for the designated times were analyzed by EMSA for ability to bind to an NF-κB consensus probe. (B) Specificity of NF-κB binding. Homologous wild  type (wt) and mutant (mut) NF-κB probes were evaluated for ability to inhibit binding of the NF-κB consensus probe by EMSA. (C) NF-κB activation  by C3dg. Nuclear extracts from purified resting B cells incubated with microbeads coated with C3dg or BSA were evaluated for the presence of activated  NF-κB by EMSA. (D) CD21 dependence of NF-κB activation. Purified resting B cells were incubated with soluble gp105 or OKB7 for 1 h, or with the  same amounts of gp105 (EBV + gp105) or OKB7 (EBV + OKB7) for 1 h before EBV addition. Nuclear extracts were prepared 15 min after EBV addition. B cells were also incubated in BSA- or gp105-coated plastic culture wells (pl.) for 15 min. Ability to bind to an NF-κB consensus probe was evaluated by EMSA. (E) Composition of NF-κB. Nuclear extracts from purified resting B cells 30 min after EBV addition were incubated with p50, p65, c-rel  or p52 Abs to NF-κB subunits before addition of the NF-κB consensus probe and analysis by EMSA. (F, top) Effect of calphostin C on NF-κB activation. Nuclear extracts prepared 30 min after EBV addition to purified resting B cells that had been preincubated for 2 h with calphostin C (50 nM) or  buffer, were examined for NF-κB activation by EMSA. (F, bottom) Assessment of IκBα. The same samples were evaluated for IκBα by the Western blotting procedure. Control lanes (−) do not contain EBV.
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Figure 1: NF-κB induction by CD21 ligands. (A) NF-κB activation by EBV. Nuclear extracts from purified resting B cells incubated with B95-8 EBV for the designated times were analyzed by EMSA for ability to bind to an NF-κB consensus probe. (B) Specificity of NF-κB binding. Homologous wild type (wt) and mutant (mut) NF-κB probes were evaluated for ability to inhibit binding of the NF-κB consensus probe by EMSA. (C) NF-κB activation by C3dg. Nuclear extracts from purified resting B cells incubated with microbeads coated with C3dg or BSA were evaluated for the presence of activated NF-κB by EMSA. (D) CD21 dependence of NF-κB activation. Purified resting B cells were incubated with soluble gp105 or OKB7 for 1 h, or with the same amounts of gp105 (EBV + gp105) or OKB7 (EBV + OKB7) for 1 h before EBV addition. Nuclear extracts were prepared 15 min after EBV addition. B cells were also incubated in BSA- or gp105-coated plastic culture wells (pl.) for 15 min. Ability to bind to an NF-κB consensus probe was evaluated by EMSA. (E) Composition of NF-κB. Nuclear extracts from purified resting B cells 30 min after EBV addition were incubated with p50, p65, c-rel or p52 Abs to NF-κB subunits before addition of the NF-κB consensus probe and analysis by EMSA. (F, top) Effect of calphostin C on NF-κB activation. Nuclear extracts prepared 30 min after EBV addition to purified resting B cells that had been preincubated for 2 h with calphostin C (50 nM) or buffer, were examined for NF-κB activation by EMSA. (F, bottom) Assessment of IκBα. The same samples were evaluated for IκBα by the Western blotting procedure. Control lanes (−) do not contain EBV.
Mentions: Nuclear extracts prepared from highly purified resting (nonactivated) human tonsil B cells incubated with EBV for varying periods of time at 37°C were assessed for ability to bind to an NF-κB consensus probe in gel shift assays. Nuclear NF-κB–binding activity increased rapidly from low constitutive levels to reach peak values 15–30 min after EBV addition (Fig. 1 A). Activation was inhibited by the unlabeled wild-type oligonucleotide, but not by a mutant oligonucleotide, demonstrating specificity (Fig. 1 B). After a modest decline, EBV-induced NF-κB–binding activity increased 24 h after EBV addition, likely due to the actions of EBNA 2 and latent membrane protein 1, two EBV latent genes expressed in the first days after infection (13, 14) which activate NF-κB (15, 16). Similar studies with purified tonsil B cells from >10 individuals with two EBV strains (Akata and B95-8) gave comparable results. C3dg-coated microbeads gave the same pattern of rapid NF-κB activation (Fig. 1 C) but without the second increase 24 h later.

Bottom Line: Epstein-Barr virus (EBV), an oncogenic human herpesvirus, binds to and infects normal human B lymphocytes via CD21, the CR2 complement receptor.Studies of the mechanisms that enable EBV to infect nonactivated, noncycling B cells provide compelling evidence for a sequence of events in which EBV binding to CD21 on purified resting human B cells rapidly activates the NF-kappaB transcription factor, which, in turn, binds to and mediates transcriptional activation of Wp, the initial viral latent gene promoter.Thus, EBV binding to its cellular receptor on resting B cells triggers an NF-kappaB-dependent intracellular signaling pathway which is required for infection.

View Article: PubMed Central - PubMed

Affiliation: Scripps Research Institute, Department of Immunology, La Jolla, California 92037, USA.

ABSTRACT
Epstein-Barr virus (EBV), an oncogenic human herpesvirus, binds to and infects normal human B lymphocytes via CD21, the CR2 complement receptor. Studies of the mechanisms that enable EBV to infect nonactivated, noncycling B cells provide compelling evidence for a sequence of events in which EBV binding to CD21 on purified resting human B cells rapidly activates the NF-kappaB transcription factor, which, in turn, binds to and mediates transcriptional activation of Wp, the initial viral latent gene promoter. Thus, EBV binding to its cellular receptor on resting B cells triggers an NF-kappaB-dependent intracellular signaling pathway which is required for infection.

Show MeSH
Related in: MedlinePlus