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Journal of Virology, April 2007, p. 3685-3692, Vol. 81, No. 8
0022-538X/07/$08.00+0     doi:10.1128/JVI.01538-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Mutation in the Glycosylated Gag Protein of Murine Leukemia Virus Results in Reduced In Vivo Infectivity and a Novel Defect in Viral Budding or Release

Audrey Low, Shoibal Datta, Yurii Kuznetsov, Sohail Jahid, Nayantara Kothari, Alexander McPherson, and Hung Fan^*

Department of Molecular Biology and Biochemistry and Cancer Research Institute, University of California, Irvine, California 92697-3905

Received 18 July 2006/ Accepted 23 January 2007

All gammaretroviruses, including murine leukemia viruses (MuLVs), feline leukemia viruses, and gibbon-ape leukemia virus, encode an alternate, glycosylated form of Gag polyprotein (glyco-Gag or gPr80^gag) in addition to the polyprotein precursor of the viral capsid proteins (Pr65^gag). gPr80^gag is translated from an upstream in-frame CUG initiation codon, in contrast to the AUG codon used for Pr65^gag. The role of glyco-Gag in MuLV replication has been unclear, since gPr80^gag-negative Moloney MuLV (M-MuLV) mutants are replication competent in vitro and pathogenic in vivo. However, reversion to the wild type is frequently observed in vivo. In these experiments, in vivo inoculation of a gPr80^gag mutant, Ab-X-M-MuLV, showed substantially lower (2 log) initial infectivity in newborn NIH Swiss mice than that of wild-type virus, and revertants to the wild type could be detected by PCR cloning and DNA sequencing as early as 15 days postinfection. Atomic force microscopy of Ab-X-M-MuLV-infected producer cells or of the PA317 amphotropic MuLV-based vector packaging line (also gPr80^gag negative) revealed the presence of tube-like viral structures on the cell surface. In contrast, wild-type virus-infected cells showed the typical spherical, 145-nm particles observed previously. Expression of gPr80^gag in PA317 cells converted the tube-like structures to typical spherical particles. PA317 cells expressing gPr80^gag produced 5- to 10-fold more infectious vector or viral particles as well. Metabolic labeling studies indicated that this reflected enhanced virus particle release rather than increased viral protein synthesis. These results indicate that gPr80^gag is important for M-MuLV replication in vivo and in vitro and that the protein may be involved in a late step in viral budding or release.

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* Corresponding author. Mailing address: Department of Molecular Biology and Biochemistry and Cancer Research Institute, University of California, Irvine, CA 92697-3905. Phone: (949) 824-5554. Fax: (949) 824-4023. E-mail: hyfan{at}uci.edu

Published ahead of print on 31 January 2007.

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Journal of Virology, April 2007, p. 3685-3692, Vol. 81, No. 8
0022-538X/07/$08.00+0     doi:10.1128/JVI.01538-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

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