All rabbits immunized with type A influenza virions have a serum haemagglutination-inhibition antibody response biased to a single epitope in antigenic site B

Lambkin, Robert; Dimmock, Nigel J.

doi:10.1099/0022-1317-76-4-889

Cited by 20 publications

(13 citation statements)

References 33 publications

(37 reference statements)

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“…Most neutralizing Abs are directed to epitopes in the globular "head" domain of HA, the location of the sialic acid binding site. Abs neutralize infectivity by sterically blocking HA binding to its receptor and by interfering with conformational alterations required for HA-mediated fusion of viral and cellular membranes (11). Fusion occurs after the internalization of virions into endosomes, where the acidic pH triggers HA to expose its fusion peptide.…”

mentioning

confidence: 99%

Influenza A Virus Hemagglutinin Trimerization Completes Monomer Folding and Antigenicity

Magadán

Khurana

Das

et al. 2013

J Virol

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c Influenza A virus (IAV) remains an important human pathogen largely because of antigenic drift, the rapid emergence of antibody escape mutants that precludes durable vaccination. The most potent neutralizing antibodies interact with cognate epitopes in the globular "head" domain of hemagglutinin (HA), a homotrimeric glycoprotein. The H1 HA possesses five distinct regions defined by a large number of mouse monoclonal antibodies (MAbs), i.e., Ca1, Ca2, Cb, Sa, and Sb. Ca1-Ca2 sites require HA trimerization to attain full antigenicity, consistent with their locations on opposite sides of the trimer interface. Here, we show that full antigenicity of Cb and Sa sites also requires HA trimerization, as revealed by immunofluorescence microscopy of IAVinfected cells and biochemically by pulse-chase radiolabeling experiments. Surprisingly, epitope antigenicity acquired by HA trimerization persists following acid triggering of the globular domains dissociation and even after proteolytic release of monomeric heads from acid-treated HA. Thus, the requirement for HA trimerization by trimer-specific MAbs mapping to the Ca, Cb, and Sa sites is not dependent upon the bridging of adjacent monomers in the native HA trimer. Rather, complete antigenicity of HA (and, by inference, immunogenicity) requires a final folding step that accompanies its trimerization. Once this conformational change occurs, HA trimers themselves would not necessarily be required to induce a highly diverse neutralizing response to epitopes in the globular domain.T he influenza A virus (IAV) hemagglutinin (HA) glycoprotein attaches virions to target cells by binding terminal sialic acid residues on cell surface glycans (1, 2). As a prototypical homotrimeric type I integral membrane protein, HA is synthesized in the endoplasmic reticulum (ER) of infected cells and transported through the Golgi complex (GC) to the plasma membrane (PM), where it is incorporated into budding virions. A variable number (depending on the strain) of N-linked oligosaccharides are added cotranslationally as HA is extruded into the ER through the translocon and subsequently trimmed and modified extensively during HA transport to the cell surface. In addition, the cytoplasmic COOH terminus of HA is palmitoylated during its intracellular transport, likely in an early GC compartment (3).HA folding begins cotranslationally, as shown by the acquisition of intrachain disulfide bonds and by the binding of monoclonal antibodies (MAbs) specific for discontinuous epitopes within HA to nascent chains (4). Initial folding of HA monomers is likely completed shortly after chain termination (within 1 or 2 min), whereas HA trimerization occurs with a half-life of ϳ5 min (5, 6). The localization of the later process appears to be strain (and perhaps subtype) specific, with prototype H2 (7) and H3 (8) HAs trimerizing in the ER while the A/Puerto Rico/8/34 (PR8) H1 HA trimerizes in either the ER-Golgi intermediate compartment or the cis-GC (3, 6).HA is of great medical importance by virtue of its recog...

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confidence: 99%

Influenza A Virus Hemagglutinin Trimerization Completes Monomer Folding and Antigenicity

Magadán

Khurana

Das

et al. 2013

J Virol

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“…Some data show that the antibody response might focus on one epitope of HA immunized with the whole type A influenza virions in an animal model [29, 30]. And this bias can be observed more obviously after the second immunization.…”

Section: Discussionmentioning

confidence: 99%

A Candidate Vaccine against Influenza Virus Intensively Improved the Immunogenicity of a Neutralizing Epitope

Ding²,

Liu³

et al. 2002

Int Arch Allergy Immunol

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Background: The hemagglutinin (HA) of influenza viruses is one of the major targets of the humoral response. The role of serum antibody to HA in the protection against infection has been demonstrated by long-standing observation. In previous studies, we suggested that an epitope vaccine might be a new strategy against the virus. Methods: HA sequences of 491 H3 subtype strains from the influenza sequence database were compared and analyzed. To acquire information on the immunogenicity of the F3 epitope, F3-epitope-specific antibody levels in 81 patient sera infected with influenza virus were tested by ELISA. Based on the theory of the epitope vaccine, we designed an epitope peptide F3 (C-KAYSNCYPYDVPDY-G-KAYSNCYPYDVPDY), which contains the repeated F3 epitope KAYSNCYPYDVPDY (aa92–105) on HA (H3N2). The specificity and the titer of the antibodies induced by the epitope vaccine were determined by ELISA. The neutralizing activities of these anti-F3 antibodies were shown by inhibiting influenza virus infection of MDCK cells. Results and Conclusion: Comparison of HA sequences of 491 H3 subtype strains indicates that this epitope is highly conservative. Analysis of the sera from influenza virus-infected patients revealed a very low level of F3 epitope-specific antibodies, suggesting the poor immunogenicity of the F3 epitope on influenza virus. The epitope vaccine based on the F3 epitope induced high levels of F3 epitope-specific antibodies recognizing the epitope peptide F3 (antibody titer in antisera up to 1:25,600). Besides, the antisera could also recognize the natural HA in Western blotting. Interestingly, these antisera induced by the epitope vaccine could inhibit infection of MDCK cells by influenza virus (strain A/Wuhan/359/95) in the neutralization assay. These results suggest that the epitope vaccine can intensively increase the immunogenicity of neutralizing epitopes and may provide a new way to develop an effective vaccine against influenza virus.

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“…These viruses have closely related haemagglutinins and unrelated neuraminidases [21]. Virus was purified by differential centrifugation [20].…”

Section: Virusesmentioning

confidence: 99%

“…These were immunized by inoculation of 5i10% HAU purified A\FPV\D virus into a peripheral ear vein, except for rabbit 3A1 which received A\FPV\R in the same way (Table 1 and [21,22] for more details). Rabbits 191, 192 and 2A6 received virus inactivated with 0n1% β-propiolactone (Sigma), although A\FPV does not multiply in rabbits.…”

Section: Preparation Of Antisera To Type a Influenza Virusmentioning

confidence: 99%

“…Analysis of antisera from rabbits immunized with the same virus in a variety of routes, formulations and up to three doses showed that the HA epitope specificity in all of many antisera representing the secondary antiserum response of 12 rabbits studied was also biased to a single epitope, but in antigenic site B. The response was not genetically determined because mice of three different haplotypes and rabbits of three different breeds all behaved in the same way [20][21][22]. The natural situation can be modelled with Mabs, and escape mutants were selected without hindrance provided one Mab predominated, and the other 2 or 3 Mabs in the mixture constituted a small proportion of the total titre [20].…”

Section: Introductionmentioning

confidence: 99%

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Selection of neutralizing antibody escape mutants with type A influenza virus HA-specific polyclonal antisera: possible significance for antigenic drift

1997

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Ten antisera were produced in rabbits by two or three intravenous injections of inactivated whole influenza type A virions. All contained haemagglutination-inhibition (HI) antibody directed predominantly to an epitope in antigenic site B and, in addition, various amounts of antibodies to an epitope in site A and in site D. The ability of untreated antisera to select neutralization escape mutants was investigated by incubating virus possessing the homologous haemagglutinin with antiserum adjusted to contain anti-B epitope HI titres of 100, 1000 and 10000 HIU/ml. Virus-antiserum mixtures were inoculated into embryonated hen's eggs, and progeny virus examined without further selection. Forty percent of the antisera at a titre of 1000 HIU/ml selected neutralizing antibody escape mutants as defined by their lack of reactivity to Mab HC10 (site B), and unchanged reactivity to other Mabs to site A and site D epitopes. All escape mutant-selecting antisera had a ratio of anti-site B (HC10)-epitope antibody[ratio ]other antibodies of [ges ]2·0[ratio ]1. The antiserum with the highest ratio (7·4[ratio ]1) selected escape mutants in all eggs tested in four different experiments. No antiserum used at a titre of 10000 HIU/ml allowed multiplication of any virus. All antisera used at a titre of 100 HIU/ml permitted virus growth, but this was wild-type (wt) virus. We conclude that a predominant epitope-specific antibody response, a titre of [ges ]1000 HIU/ml, and a low absolute titre of other antibodies ([les ]500 HIU/ml) are three requirements for the selection of escape mutants. None of the antisera in this study could have selected escape mutants without an appropriate dilution factor, so the occurrence of an escape mutant-selecting antiserum in nature is likely to be a rare event.

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All rabbits immunized with type A influenza virions have a serum haemagglutination-inhibition antibody response biased to a single epitope in antigenic site B

Cited by 20 publications

References 33 publications

Influenza A Virus Hemagglutinin Trimerization Completes Monomer Folding and Antigenicity

Influenza A Virus Hemagglutinin Trimerization Completes Monomer Folding and Antigenicity

A Candidate Vaccine against Influenza Virus Intensively Improved the Immunogenicity of a Neutralizing Epitope

Selection of neutralizing antibody escape mutants with type A influenza virus HA-specific polyclonal antisera: possible significance for antigenic drift

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