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Journal of Virology, March 2003, p. 3785-3798, Vol. 77, No. 6
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.6.3785-3798.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Infectivity of a Human Respiratory Syncytial Virus Lacking the SH, G, and F Proteins Is Efficiently Mediated by the Vesicular Stomatitis Virus G Protein

University of Alabama School of Medicine, Birmingham, Alabama 35294

Received 4 October 2002/ Accepted 11 November 2002

To examine the requirements of the human respiratory syncytial virus (HRSV) SH (small hydrophobic), G (attachment), and F (fusion) proteins for virus infectivity and morphology, we used the prototype A2 strain of HRSV to generate a series of cDNAs from which (i) the SH open reading frame (ORF), (ii) the SH and G ORFs, or (iii) the SH, G, and F ORFs were deleted. Each deleted ORF was replaced as follows: the SH ORF was replaced with that of green fluorescent protein; the G ORF was replaced with that of G^vsv, a chimeric glycoprotein consisting of the vesicular stomatitis Indiana virus (VSIV) G protein ecto- and transmembrane domains coupled to the HRSV F cytoplasmic tail; and the F ORF was replaced with that of marker protein ß-glucuronidase. The number of genes and the intergenic junctions in the constructs were kept as found in A2 virus in order to maintain authentic levels of transcription. Infectious viruses were recovered from all three engineered cDNAs and designated RSsh, RSsh,g/G^vsv, and RSsh,g,f/G^vsv, respectively. Low-pH-induced syncytium formation was observed in cells infected with viruses RSSH,G/G^vsv and RSSH,G,F/G^vsv, indicating that G^vsv was expressed and functional. Neutralization of infectivity by anti-VSIV G antibodies and inhibition of entry by ammonium chloride showed that RSSH,G,F/G^vsv infectivity was mediated by G^vsv and that an acidification step was required for entry into the host cell, similar to VSIV virions. All three engineered viruses displayed growth kinetics and virus yields similar to a wild-type A2 virus, both in Vero and HEp-2 cells. Abundant virus-induced filaments were observed at the surface of cells infected with each of the three engineered viruses or with virus A2, indicating that neither the SH and G proteins nor the F protein ecto- and transmembrane domains were required for the formation of these structures. This is the first report of the recovery of an infectious HRSV lacking a fusion protein of the Paramyxoviridae family and of manipulation of the HRSV entry pathway via incorporation of a nonparamyxoviral transmembrane glycoprotein.

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