Immunoelectron Microscopic Studies on Haemagglutinin and Haemolysin of Measles Virus in Infected HEp2 Cells

Armstrong, Mervyn; Fraser, Kenneth; Dermott, Evelyn; Shirodaria, P. V.

doi:10.1099/0022-1317-59-1-187

Cited by 4 publications

(6 citation statements)

References 13 publications

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“…8B. Interestingly, an early electron microscopy study suggests prominent spikes on the measles virus surface to correspond to the attachment protein, whereas the fusion function was considered to reside closer to the virus membrane (62). Consistent with this, a recent electron cryomicroscopy analysis of hPIV5 particles concludes that defined glycoprotein spikes, previously observed in electron microscopy studies of paramyxovirus particles (11), correspond in the case of F to the post-fusion conformation and thus represent a product of premature F refolding (63).…”

Section: Discussionmentioning

confidence: 99%

Functional Interaction between Paramyxovirus Fusion and Attachment Proteins

Lee

Prussia

Paal

et al. 2008

Journal of Biological Chemistry

140

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Paramyxovirinae envelope glycoproteins constitute a premier model to dissect how specific and dynamic interactions in multisubunit membrane protein complexes can control deep-seated conformational rearrangements. However, individual residues that determine reciprocal specificity of the viral attachment and fusion (F) proteins have not been identified. We have developed an assay based on a pair of canine distemper virus (CDV) F proteins (strains Onderstepoort (ODP) and Lederle) that share approximately 95% identity but differ in their ability to form functional complexes with the measles virus (MV) attachment protein (H). Characterization of CDV F chimeras and mutagenesis reveals four residues in CDV F-ODP (positions 164, 219, 233, and 317) required for productive interaction with MV H. Mutating these residues to the Lederle type disrupts triggering of F-ODP by MV H without affecting functionality when co-expressed with CDV H. Co-immunoprecipitation shows a stronger physical interaction of F-ODP than F-Lederle with MV H. Mutagenesis of MV F highlights the MV residues homologous to CDV F residues 233 and 317 as determinants for physical glycoprotein interaction and fusion activity under homotypic conditions. In assay reversal, the introduction of sections of the CDV H stalk into MV H shows a five-residue fragment (residues 110-114) to mediate specificity for CDV F-Lederle. All of the MV H stalk chimeras are surface-expressed, show hemadsorption activity, and trigger MV F. Combining the five-residue H chimera with the CDV F-ODP quadruple mutant partially restores activity, indicating that the residues identified in either glycoprotein contribute interdependently to the formation of functional complexes. Their localization in structural models of F and H suggests that placement in particular of F residue 233 in close proximity to the 110-114 region of H is structurally conceivable.

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Section: Discussionmentioning

confidence: 99%

Functional Interaction between Paramyxovirus Fusion and Attachment Proteins

Lee

Prussia

Paal

et al. 2008

Journal of Biological Chemistry

140

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“…Immunofluorescent staining of unfixed, uninfected cells showed that the an tiserum had a high-titer cellular antibody which was greatly reduced by absorption with uninfected Vero cells, but any reduction that might have been seen on unfixed infect ed cells was masked by the presence of hightiter antibody to the measles virus hemag glutinin and hemolysin. To prevent the im mune response to these antigens, the hemag glutinin spikes were removed from the vir ions and the hemolysin antigen was destroy ed, using trypsin and acetone treatment [9][10][11] before inoculation. This also ensured that any cellular antigens sticking to the sur face of the virions or caught in clumps of virus were removed [3].…”

Section: Discussionmentioning

confidence: 99%

“…3) but were absent after unfixed, uninfected, Vero cell absorption (lane 2). Lane 4 shows the relative position of the measles-virusspecific 78K glycoprotein precipitated using a monospecific anti-HA serum [9]. …”

Section: Radioimmunoprécipitation and Pagementioning

confidence: 99%

“…This virus, like most lipid-containing viruses, buds from the surface of the infected cell [8]. The major antigens on the surface of the virion are the hemagglutinin, situated on the spikes projecting from the surface of the en velope, and the hemolysin, lying on or near the surface of the envelope [9]. As treatment with trypsin removes the spikes from measles virions [9,10] and hemolysin antigen is de stroyed by acetone treatment [11], we com bined these methods to prepare measles virus particles with envelopes from which the ma jor viral surface antigens had been removed or rendered inactive, thus leaving other anti gens in the envelope accessible to the im mune system of an inoculated animal.…”

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confidence: 99%

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Recognition of Host-Membrane Antigens in the Envelope of Measles Virions

Armstrong¹,

Fraser²,

Shirodaria³

1984

Intervirology

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Trypsin- and acetone-treated virions from either of two strains of measles virus grown in Vero cells stimulated the production in guinea pigs of (i) virus-specific antibodies to polypeptide (P) of molecular weight 70,000 (70K) and to the portion of the HA spike embedded in the viral envelope, but not to M protein, and (ii) antibodies to two host cell membrane antigens which were identified as glycoproteins with molecular weights of 108K and 104K. These host cell antigens were present in increased amounts in infected cells and were intimately associated with the virus. Untreated measles virions grown in Vero cells also stimulated the production of antibody to the 108K glycoprotein. The host polypeptides were less antigenic in virus derived from HEp2 cells, which apparently contained less of these antigens.

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“…Immunoelectron microscopic studies have suggested that the antigenic determinants for the haemagglutinin (HA) of measles virus reside on trypsin-sensitive spikes, which protrude from the membrane of infected cells (Armstrong et al, 1982). The production of measles virus HAspecific monoclonal antibodies has facilitated the characterization of these antigenic determinants (McFarlin et al, 1980;Birrer et al, 1981 ;Trudgett et al, 1981 ;Togashi et al, 1981 ;Giraudon & Wild, 1981 ;Bohn et al, 1982).…”

Section: Introductionmentioning

confidence: 99%

Biological Activity of a Monoclonal Antibody to a Measles Virus Haemagglutinin Epitope Detected Late in Infection

Kramer

Cremer

1984

Journal of General Virology

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SUMMARYA hybrid cell line secreting monoclonal antibody 2H9c (mAb 2H9c) with measles virus haemagglutinin (HA) specificity was produced. The mAb 2H9c was non-reactive in haemagglutination inhibition and neutralization assays. A protein of apparent mol. wt. 79000 was immune-precipitated using a Triton X-100/SDS/sodium deoxycholate detergent buffer and two proteins with mol. wt. of 79000 and 69000 were immuneprecipitated using 1% Nonidet P40/sodium deoxycholate buffer in SDS-PAGE assays. Similar results were obtained with other anti-HA monoclonal antibodies, supporting the assumption that mAb 2H9c was directed against the HA polypeptide. The indirect immunofluorescence (IFA) staining pattern of acetone-fixed infected cells with mAb 2H9c differed substantially from that with other HA-specific monoclonal antibodies. Kinetic studies revealed that the reactivity of mAb 2H9c lagged behind other HAspecific monoclonal antibodies by 18 to 36 h post-infection in IFA assays. This suggests that mAb 2H9c may be directed against a binding site that arises late in infection, possibly as a result of a conformational alteration of the HA polypeptide.

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Immunoelectron Microscopic Studies on Haemagglutinin and Haemolysin of Measles Virus in Infected HEp2 Cells

Cited by 4 publications

References 13 publications

Functional Interaction between Paramyxovirus Fusion and Attachment Proteins

Functional Interaction between Paramyxovirus Fusion and Attachment Proteins

Recognition of Host-Membrane Antigens in the Envelope of Measles Virions

Biological Activity of a Monoclonal Antibody to a Measles Virus Haemagglutinin Epitope Detected Late in Infection

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