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Journal of Virology, March 2008, p. 2106-2119, Vol. 82, No. 5
0022-538X/08/$08.00+0     doi:10.1128/JVI.02337-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Molecular Characterization of Feline Immunodeficiency Virus Budding

Benjamin G. Luttge,¹ Miranda Shehu-Xhilaga,^1, Dimiter G. Demirov,^1, Catherine S. Adamson,¹ Ferri Soheilian,² Kunio Nagashima,² Andrew G. Stephen,³ Robert J. Fisher,³ and Eric O. Freed^1*

Virus-Cell Interaction Section, HIV Drug Resistance Program, National Cancer Institute at Frederick, Frederick, Maryland 21702-1201,¹ Image Analysis Laboratory, Advanced Technology Program, SAIC-Frederick, National Cancer Institute at Frederick, Frederick, Maryland 21702-1201,² Protein Chemistry Laboratory, Advanced Technology Program, SAIC-Frederick, National Cancer Institute at Frederick, Frederick, Maryland 21702-1201³

Received 29 October 2007/ Accepted 11 December 2007

Infection of domestic cats with feline immunodeficiency virus (FIV) is an important model system for studying human immunodeficiency virus type 1 (HIV-1) infection due to numerous similarities in pathogenesis induced by these two lentiviruses. However, many molecular aspects of FIV replication remain poorly understood. It is well established that retroviruses use short peptide motifs in Gag, known as late domains, to usurp cellular endosomal sorting machinery and promote virus release from infected cells. For example, the Pro-Thr/Ser-Ala-Pro [P(T/S)AP] motif of HIV-1 Gag interacts directly with Tsg101, a component of the endosomal sorting complex required for transport I (ESCRT-I). A Tyr-Pro-Asp-Leu (YPDL) motif in equine infectious anemia virus (EIAV), and a related sequence in HIV-1, bind the endosomal sorting factor Alix. In this study we sought to identify and characterize FIV late domain(s) and elucidate cellular machinery involved in FIV release. We determined that mutagenesis of a PSAP motif in FIV Gag, small interfering RNA-mediated knockdown of Tsg101 expression, and overexpression of a P(T/S)AP-binding fragment of Tsg101 (TSG-5') each inhibited FIV release. We also observed direct binding of FIV Gag peptides to Tsg101. In contrast, mutagenesis of a potential Alix-binding motif in FIV Gag did not affect FIV release. Similarly, expression of the HIV-1/EIAV Gag-binding domain of Alix (Alix-V) did not disrupt FIV budding, and FIV Gag peptides showed no affinity for Alix-V. Our data demonstrate that FIV relies predominantly on a Tsg101-binding PSAP motif in the C terminus of Gag to promote virus release in HeLa cells, and this budding mechanism is highly conserved in feline cells.

Published ahead of print on 19 December 2007.

Present address: Department of Medicine, Monash University, Alfred Campus, 85 Commerical Rd., Prahran 3181, Victoria, Australia.

Present address: Institute of Molecular Virology, Von Esmarch Str. 56, 48149 Muenster, Germany.