Dataset Information

B cell epitope of human cytomegalovirus phosphoprotein 65 (HCMV pp65) induced anti-dsDNA antibody in BALB/c mice.

ABSTRACT:

Background

HCMV phosphoprotein 65 (HCMVpp65) is a putative immunogen that acts as an accelerator, inducing autoantibody and exacerbating autoimmune response in susceptible animals. The immunity to pp65_336-439 instigates autoimmunity, suggesting that pp65_336-439 contains crucial B cell epitope(s) for the development of nephritis. This study narrowed down the target epitope to pp65_422-439 for immunization of BALB/c mice and mapping of B cell epitope.

Methods

The target epitope pp65_422-439 reactivity and B cell epitope mapping was examined in serum from pp65_422-439-immunized mice and patients with systemic lupus erythematosus (SLE). Kidney tissue from immunized mice was examined for signs of immune complex nephritis.

Results

Anti-pp65_422-439 antibody in serum either from patients with SLE or from pp65_422-439-immunized mice exhibited cross-reactivity to several nuclear components such as double-stranded DNA (dsDNA). Moreover, the pp65_422-439-immunized mice developed initial signs of glomerulonephritis such as deposition of immunoglobulin G/M (IgG/IgM) and third complement component (C3). With B cell epitope mapping by pp65_422-439-derived decapeptides, one dominant epitope, pp65_428-437, was identified in serum from pp65_422-439-immunized mice and patients with SLE with anti-pp65_422-439 antibody. Epitope spreading from pp65_428-437 to pp65_430-439 was found in pp65_422-439-immunized mice in which we generated monoclonal antibodies to pp65_425-434 and pp65_430-439. However, dsDNA positive reactivity was exclusively observed in Crithidia luciliae stains with pp65_430-439-reactive monoclonal antibody. Additionally, we observed the amelioration of autoimmunity following the elevation of IgM targeting pp65_428-437. CONCLUSIONS: Our data suggest that pp65_428-437 may be an autoimmune or lupus-prone B cell epitope and may catalyze further epitope spreading for inducing autoantibodies in lupus-susceptible individuals.

SUBMITTER: HoHsieh A

PROVIDER: S-EPMC5359867 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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Publications

B cell epitope of human cytomegalovirus phosphoprotein 65 (HCMV pp65) induced anti-dsDNA antibody in BALB/c mice.

HoHsieh Ao A Wang Chin Man CM Wu Yeong-Jian Jan YJ Chen Albert A Chang Ming-I MI Chen Ji-Yih JY

Arthritis research & therapy 20170321 1

<h4>Background</h4>HCMV phosphoprotein 65 (HCMVpp65) is a putative immunogen that acts as an accelerator, inducing autoantibody and exacerbating autoimmune response in susceptible animals. The immunity to pp65<sub>336-439</sub> instigates autoimmunity, suggesting that pp65<sub>336-439</sub> contains crucial B cell epitope(s) for the development of nephritis. This study narrowed down the target epitope to pp65<sub>422-439</sub> for immunization of BALB/c mice and mapping of B cell epitope.<h4>Met ...[more]

PMID: 28320458

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Project description:Abstract: We have identified a cytomegalovirus virion protein capable of modulating the rapid induction of an interferon-like response in cells that follows virus binding and penetration. Functional genomics revealed a role for the major cytomegalovirus structural protein, pp65 (ppUL83), in counteracting this response. The underlying mechanism involves a differential impact of this structural protein on the regulation of interferon response factor 3 (IRF-3). In contrast, NF-{kappa}B is activated independent of pp65, and neither STAT1 nor STAT3 becomes activated by either virus. pp65 is sufficient to prevent the activation of IRF-3 when introduced alone into cells. pp65 acts by inhibiting nuclear accumulation of IRF-3 and is associated with a reduced IRF-3 phosphorylation state. Thus, this investigation shows that the major structural protein of cytomegalovirus is committed to the modulation of the IRF-3 response, a primary mediator of the type I interferon response. By subverting IRF-3, the virus escapes throwing a central alarm devoted to both immediate antiviral control and regulation of the immune response. Polyadenylated RNA from 107 confluent HFs was purified by using Oligotex (QIAGEN, Valencia, Calif.) from Trizol (Invitrogen)-extracted total RNA. This RNA was prepared, reverse transcribed, labeled with Cy3-dUTP or Cy5-dUTP (Amersham, Little Chalfont, Buckinghamshire, United Kingdom) by random primed synthesis with DNA pol I Klenow (Amersham Life Science, Inc., Cleveland, Ohio), and hybridized to spotted, human cDNA microarrays as previously described. Sequence-verified human cDNA microarrays (HE and HG series, 31,000 spots; HD51 series, 17,000 spots) were produced at Stanford. Images were collected by using a GenePix 4000B microarray scanner, manually flagged to eliminate poor spots and analyzed by GenePix Pro 2.0 (Axon, Union City, Calif.). A replicate experimental design type is where a series of replicates are performed to evaluate reproducibility or as a pilot study to determine the appropriate number of replicates for a subsequent experiments. Keywords: disease_state_design, pathogenicity_design

Project description:Human cytomegalovirus (HCMV) is the leading cause of congenital infection and is associated with a wide range of neurodevelopmental disabilities and intrauterine growth restriction. Yet our current understanding of the mechanisms modulating transplacental HCMV transmission is poor. The placenta, given its critical function in protecting the fetus, has evolved effective yet largely uncharacterized innate immune barriers against invading pathogens. Here we show that the intrinsic cellular restriction factor apolipoprotein B editing catalytic subunit-like 3A (APOBEC3A [A3A]) is profoundly upregulated following ex vivo HCMV infection in human decidual tissues-constituting the maternal aspect of the placenta. We directly demonstrated that A3A severely restricted HCMV replication upon controlled overexpression in epithelial cells, acting by a cytidine deamination mechanism to introduce hypermutations into the viral genome. Importantly, we further found that A3 editing of HCMV DNA occurs both ex vivo in HCMV-infected decidual organ cultures and in vivo in amniotic fluid samples obtained during natural congenital infection. Our results reveal a previously unexplored role for A3A as an innate anti-HCMV effector, activated by HCMV infection in the maternal-fetal interface. These findings pave the way to new insights into the potential impact of APOBEC proteins on HCMV pathogenesis.IMPORTANCE In view of the grave outcomes associated with congenital HCMV infection, there is an urgent need to better understand the innate mechanisms acting to limit transplacental viral transmission. Toward this goal, our findings reveal the role of the intrinsic cellular restriction factor A3A (which has never before been studied in the context of HCMV infection and vertical viral transmission) as a potent anti-HCMV innate barrier, activated by HCMV infection in the authentic tissues of the maternal-fetal interface. The detection of naturally occurring hypermutations in clinical amniotic fluid samples of congenitally infected fetuses further supports the idea of the occurrence of A3 editing of the viral genome in the setting of congenital HCMV infection. Given the widely differential tissue distribution characteristics and biological functions of the members of the A3 protein family, our findings should pave the way to future studies examining the potential impact of A3A as well as of other A3s on HCMV pathogenesis.

Dataset Information

B cell epitope of human cytomegalovirus phosphoprotein 65 (HCMV pp65) induced anti-dsDNA antibody in BALB/c mice.

Background

Methods

Results

Publications

B cell epitope of human cytomegalovirus phosphoprotein 65 (HCMV pp65) induced anti-dsDNA antibody in BALB/c mice.

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