Effects of ectodomain sequences between HR1 and HR2 of F1 protein on the specific membrane fusion in paramyxoviruses

Abstract

Objectives: To explore the effects of ectodomain sequences between HR1 and HR2 of F1 protein on the specific interaction with its homologous hemagglutinin-neuraminidase (HN) in paramyxoviruses.

Methods: Site-directed mutagenesis was used to obtain mutants containing new enzyme sites on the F genes of Newcastle disease virus (NDV) and human parainfluenza virus (hPIV), and four DNA segments located between the HR1 and HR2 (NDV F-1, hPIV F-1, NDV F-2 and hPIV F-2) were obtained by cutting mutant F genes with specific endonucleases. Gene recombination was used to get chimeric F proteins NDV-C1 and hPIV-C1 by exchanging NDV F-1 and hPIV F-1 each other, and NDV-C2 and hPIV-C2 were also obtained by the same way. All the mutants and chimeric F proteins were co-expressed with their homologous or heterologous HN proteins in eukaryocytes. The fusion functions were assayed with Giemsa staining and reporter gene method for qualitative and quantitative analyses, respectively. The cell surface expression of F proteins was assayed with fluorescence-activated cell sorter (FACS) for quantitative analysis.

Results: All the mutants of F proteins had the same functions as their relevant wild types. Chimeric F proteins NDV-C1 and hPIV-C1 had 76.34 and 65.82% of fusion activities, and NDV-C2 and hPIV-C2 had 96.25 and 93.78% of fusion activities, respectively, as compared with their relevant wild types. The analysis of FACS indicated that all the mutants and chimeric F proteins had almost the same expression efficiencies as their relevant wild types.

Conclusions: The segments of NDV F-1 and hPIV F-1 were important for their specific membrane fusion, but NDV F-2 and hPIV F-2 were not.