Oligomerization of the human immunodeficiency virus type 1 (HIV-1) Vpu protein--a genetic, biochemical and biophysical analysis.
BACKGROUND
The human immunodeficiency virus type 1(HIV-1) is a complex retrovirus and the causative agent of acquired immunodeficiency syndrome (AIDS). The HIV-1 Vpu protein is an oligomeric integral membrane protein essential for particle release, viral load and CD4 degradation. In silico models show Vpu to form pentamers with an ion channel activity.


RESULTS
Using Vpu proteins from a primary subtype C and the pNL4-3 subtype B isolates of HIV-1, we show oligomerization of the full-length protein as well as its transmembrane (TM) domain by genetic, biochemical and biophysical methods. We also provide direct evidence of the presence of Vpu pentamers in a stable equilibrium with its monomers in vitro. This was also true for the TM domain of Vpu. Confocal microscopy localized Vpu to the endoplasmic reticulum and Golgi regions of the cell, as well as to post-Golgi vesicles. In fluorescence resonance energy transfer (FRET) experiments in live cells we show that Vpu oligomerizes in what appears to be either the Golgi region or intracellular vesicles, but not in the ER.


CONCLUSIONS
We provide here direct evidence that the TM domain, is critical for Vpu oligomerization and the most favourable channel assembly is a pentamer. The Vpu oligomerization appears to be either the Golgi region or intracellular vesicles, but not in the ER.
Hussain
Amjad
A
Virology Group, International Centre for Genetic Engineering and Biotechnology, New Delhi, India. amjadicgeb@yahoo.com
Das
Suman R
SR
Tanwar
Charu
C
Jameel
Shahid
S
D000595:Amino Acid Sequence; D000818:Animals; D001704:Biopolymers; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry; D003001:Cloning, Molecular; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000648:ultrastructure / Q000821:virology; D031541:Fluorescence Resonance Energy Transfer; D016338:Genes, vpu; D006056:Golgi Apparatus / Q000737:chemistry / Q000648:ultrastructure / Q000821:virology; D015497:HIV-1 / Q000737:chemistry* / Q000378:metabolism; D054298:Human Immunodeficiency Virus Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007473:Ion Channels / Q000737:chemistry; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D054334:Viral Regulatory and Accessory Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D000086082:Viroporin Proteins
D001704:Biopolymers; D054298:Human Immunodeficiency Virus Proteins; D007473:Ion Channels; D011993:Recombinant Fusion Proteins; D054334:Viral Regulatory and Accessory Proteins; C514859:vpu protein, Human immunodeficiency virus 1; D000086082:Viroporin Proteins