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Intracellular processing of the paramyxovirus F protein: critical role of the predicted amphipathic alpha helix adjacent to the fusion domain.

ABSTRACT: At a nonpermissive temperature, the group D temperature-sensitive mutants of Newcastle disease virus strain Australia-Victoria (AV) are defective in plaque formation, in inducing infected cells to fuse, and in incorporating the cleaved fusion glycoprotein, F1 + F2, into virus particles. In this study, the F protein of AV, expressed in chicken embryo cells, was able to complement these mutants in a plaque assay, identifying the F gene as the gene containing the group D temperature-sensitive lesions. The F genes of mutants D1, D2, and D3 were found to contain single mutations relative to the AV sequence, clustered within a predicted amphipathic alpha helix (AAH) adjacent to the hydrophobic amino terminus of F1. These mutant F proteins were inefficiently processed at the permissive temperature, a problem that was exacerbated at the nonpermissive temperature. Surprisingly, the AV F protein was also found to be partially temperature sensitive in processing. Its AAH is predicted to contain a break in the helix close to the D mutation sites, which are themselves predicted to further weaken the helix at this point. Interestingly, six revertants of the group D mutants were found to have an additional lesion in the AAH, repairing both the AV and mutant helices, resulting in a predicted perfect helix. The F protein of these revertants had overcome both the processing defects of the mutants and the temperature sensitivity of AV, indicating that the AAH region is critical for F protein processing. The lesions of a second group of revertants were localized within F2, suggesting an interaction with the F1 AAH region containing the original lesion.

SUBMITTER: Wang C

PROVIDER: S-EPMC241219 | biostudies-other | 1992 Jul

REPOSITORIES: biostudies-other

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Intracellular processing of the paramyxovirus F protein: critical role of the predicted amphipathic alpha helix adjacent to the fusion domain.

Wang C C Raghu G G Morrison T T Peeples M E ME

Journal of virology 19920701 7

At a nonpermissive temperature, the group D temperature-sensitive mutants of Newcastle disease virus strain Australia-Victoria (AV) are defective in plaque formation, in inducing infected cells to fuse, and in incorporating the cleaved fusion glycoprotein, F1 + F2, into virus particles. In this study, the F protein of AV, expressed in chicken embryo cells, was able to complement these mutants in a plaque assay, identifying the F gene as the gene containing the group D temperature-sensitive lesio ...[more]

PMID: 1602539

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Project description:Enveloped viruses utilize surface glycoproteins to bind and fuse with a target cell membrane. The zoonotic Hendra virus (HeV), a member of the family Paramyxoviridae, utilizes the attachment protein (G) and the fusion protein (F) to perform these critical functions. Upon triggering, the trimeric F protein undergoes a large, irreversible conformation change to drive membrane fusion. Previously, we have shown that the transmembrane (TM) domain of the F protein, separate from the rest of the protein, is present in a monomer-trimer equilibrium. This TM-TM association contributes to the stability of the prefusion form of the protein, supporting a role for TM-TM interactions in the control of F protein conformational changes. To determine the impact of disrupting TM-TM interactions, constructs expressing the HeV F TM with limited flanking sequences were synthesized. Coexpression of these constructs with HeV F resulted in dramatic reductions in the stability of F protein expression and fusion activity. In contrast, no effects were observed when the HeV F TM constructs were coexpressed with the nonhomologous parainfluenza virus 5 (PIV5) fusion protein, indicating a requirement for specific interactions. To further examine this, a TM peptide homologous to the PIV5 F TM domain was synthesized. Addition of the peptide prior to infection inhibited infection with PIV5 but did not significantly affect infection with human metapneumovirus, a related virus. These results indicate that targeted disruption of TM-TM interactions significantly impact viral fusion protein stability and function, presenting these interactions as a novel target for antiviral development.IMPORTANCE Enveloped viruses require virus-cell membrane fusion to release the viral genome and replicate. The viral fusion protein triggers from the pre- to the postfusion conformation, an essentially irreversible change, to drive membrane fusion. We found that small proteins containing the TM and a limited flanking region homologous to the fusion protein of the zoonotic Hendra virus reduced protein expression and fusion activity. The introduction of exogenous TM peptides may displace a TM domain, disrupting native TM-TM interactions and globally destabilizing the fusion protein. Supporting this hypothesis, we showed that a sequence-specific transmembrane peptide dramatically reduced viral infection in another enveloped virus model, suggesting a broader inhibitory mechanism. Viral fusion protein TM-TM interactions are important for protein function, and disruption of these interactions dramatically reduces protein stability.

Dataset Information

Intracellular processing of the paramyxovirus F protein: critical role of the predicted amphipathic alpha helix adjacent to the fusion domain.

Publications

Intracellular processing of the paramyxovirus F protein: critical role of the predicted amphipathic alpha helix adjacent to the fusion domain.

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