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Journal of Virology, July 2008, p. 6788-6797, Vol. 82, No. 14
0022-538X/08/$08.00+0     doi:10.1128/JVI.00213-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Mutations in the Spacer Peptide and Adjoining Sequences in Rous Sarcoma Virus Gag Lead to Tubular Budding {triangledown}

Paul W. Keller,1 Marc C. Johnson,2 and Volker M. Vogt1*

Department of Molecular Biology and Genetics, Biotechnology Building, Cornell University, Ithaca, New York 14853,1 Department of Molecular Microbiology and Immunology, Life Science Center, University of Missouri Medical School, Columbia, Missouri 652112

Received 30 January 2008/ Accepted 24 April 2008

All orthoretroviruses encode a single structural protein, Gag, which is necessary and sufficient for the assembly and budding of enveloped virus-like particles from the cell. The Gag proteins of Rous sarcoma virus (RSV) and human immunodeficiency virus type 1 (HIV-1) contain a short spacer peptide (SP or SP1, respectively) separating the capsid (CA) and nucleocapsid (NC) domains. SP or SP1 and the residues immediately upstream are known to be critical for proper assembly. Using mutagenesis and electron microscopy analysis of insect cells or chicken cells overexpressing RSV Gag, we defined the SP assembly domain to include the last 8 residues of CA, all 12 residues of SP, and the first 4 residues of NC. Five- or two-amino acid glycine-rich insertions or substitutions in this critical region uniformly resulted in the budding of abnormal, long tubular particles. The equivalent SP1-containing HIV-1 Gag sequence was unable to functionally replace the RSV sequence in supporting normal RSV spherical assembly. According to secondary structure predictions, RSV and HIV-1 SP/SP1 and adjoining residues may form an alpha helix, and what is likely the functionally equivalent sequence in murine leukemia virus Gag has been inferred by mutational analysis to form an amphipathic alpha helix. However, our alanine insertion mutagenesis did not provide evidence for an amphipathic helix in RSV Gag. Taken together, these results define a short assembly domain between the folded portions of CA and NC, which is essential for formation of the immature Gag shell.

* Corresponding author. Mailing address: Department of Molecular Biology and Genetics, Biotechnology Bldg., Cornell University, Ithaca, NY 14853. Phone: (607) 255 2443. Fax: (607) 255 2428. E-mail: vmv1{at}cornell.edu

{triangledown} Published ahead of print on 30 April 2008.

Journal of Virology, July 2008, p. 6788-6797, Vol. 82, No. 14
0022-538X/08/$08.00+0     doi:10.1128/JVI.00213-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Heslin, D. J., Murcia, P., Arnaud, F., Van Doorslaer, K., Palmarini, M., Lenz, J. (2009). A Single Amino Acid Substitution in a Segment of the CA Protein within Gag That Has Similarity to Human Immunodeficiency Virus Type 1 Blocks Infectivity of a Human Endogenous Retrovirus K Provirus in the Human Genome. J. Virol. 83: 1105-1114 [Abstract] [Full Text]  


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