Association of Vpu-binding protein with microtubules and Vpu-dependent redistribution of HIV-1 Gag protein.
The efficient exit of HIV-1 particles from cells requires the action of the viral encoded protein Vpu. Vpu-binding protein (Ubp) is a cellular protein that interacts with both Vpu and the major structural component of the viral capsid (Gag) and appears to affect the efficiency of particle exit. Elucidation of the function of Ubp and characterization of the spatial distribution of Ubp may provide information pertinent to understanding the role of Ubp in virus replication. To investigate the subcellular location of Ubp, and to see whether Vpu affects the intracellular distribution of Gag, we carried out immunofluorescence localization in conjunction with confocal microscopy. Based on this analysis Ubp is present in both the nucleus and the cytoplasm. In the cytoplasm, Ubp appeared to be associated with microtubules as evidenced by cofluorescence with tubulin in the absence and in the presence of colchicine. However, cytoskeletal isolation and detergent extraction of cells resulted in association of Ubp with the soluble fractions, indicating that Ubp is not in tight association with microtubules. Moreover, flotation gradient analysis demonstrated that Ubp is cytoplasmic and not stably associated with the plasma membrane. Interestingly, expression of Vpu in cells resulted in redistribution of both Ubp and Gag to a location near the periphery of the cell. The effect of Vpu on both Ubp and Gag protein has implications for Vpu-mediated particle exit from cells.
Handley
M A
MA
McArdle Laboratory for Cancer Research, University of Wisconsin, Madison, WI 53706, USA.
Paddock
S
S
Dall
A
A
Panganiban
A T
AT
D002352:Carrier Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D015683:Gene Products, gag / Q000378:metabolism*; D015497:HIV-1 / Q000378:metabolism*; D006367:HeLa Cells; D054298:Human Immunodeficiency Virus Proteins; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D018832:Molecular Chaperones; D054334:Viral Regulatory and Accessory Proteins / Q000378:metabolism*; D000086082:Viroporin Proteins
D002352:Carrier Proteins; D015683:Gene Products, gag; D054298:Human Immunodeficiency Virus Proteins; D008869:Microtubule-Associated Proteins; D018832:Molecular Chaperones; D054334:Viral Regulatory and Accessory Proteins; C114575:SGTA protein, human; C514859:vpu protein, Human immunodeficiency virus 1; D000086082:Viroporin Proteins