Intracellular processing of the paramyxovirus F protein: critical role of the predicted amphipathic alpha helix adjacent to the fusion domain.
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.
Wang
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Department of Immunology/Microbiology, Rush-Presbyterian-St. Luke's Medical Center, Chicago, Illinois 60612.
Raghu
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Morrison
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Peeples
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ME
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D002642:Chick Embryo; D004279:DNA, Viral; D005816:Genetic Complementation Test; D015726:Giant Cells; D008969:Molecular Sequence Data; D009154:Mutation; D009522:Newcastle disease virus / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D012367:RNA, Viral / Q000235:genetics; D016415:Sequence Alignment; D014760:Viral Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014763:Viral Matrix Proteins / Q000235:genetics; D010948:Viral Plaque Assay
D004279:DNA, Viral; D012367:RNA, Viral; D014760:Viral Fusion Proteins; D014763:Viral Matrix Proteins