Vpu-dependent block to incorporation of GaLV Env into lentiviral vectors.
BACKGROUND
The gibbon ape leukemia virus (GaLV) Env protein mediates entry into a wide range of human cells and is frequently used to pseudotype retroviral vectors. However, an incompatibility exists between GaLV Env and lentiviral vectors that results in decreased steady-state levels of the mature GaLV Env in cells and prevents its incorporation into lentiviral vector particles.


RESULTS
We identified the HIV-1 Vpu protein as the major cause of the depletion in GaLV Env levels that occurs when lentiviral vector components are present. This activity of Vpu targeted the mature (cleaved) form of the GaLV Env that exists within or beyond the trans-Golgi. The activity required two conserved phospho-serines in the cytoplasmic tail of Vpu that are known to recruit beta TrCP, a substrate adaptor for an SCF E3 ubiquitin ligase complex, and could be blocked by mutation of lysine 618 in the GaLV Env tail. Moreover, the Vpu-mediated decrease of GaLV Env levels was inhibited by the lysosomal inhibitor, bafilomycin A1. Interestingly, this activity of Vpu was only observed in the presence of other lentiviral vector components.


CONCLUSIONS
Similar to the mechanism whereby Vpu targets BST-2/tetherin for degradation, these findings implicate beta-TrCP-mediated ubiquitination and the endo-lysosomal pathway in the degradation of the GaLV Env by lentiviral vector components. Possibly, the cytoplasmic tail of the GaLV Env contains features that mimic bona fide targets of Vpu, important to HIV-1 replication. Furthermore, the lack of effect of Vpu on GaLV Env in the absence of other HIV-1 proteins, suggests that a more complex interaction may exist between Vpu and its target proteins, with the additional involvement of one or more component(s) of the HIV-1 replication machinery.
Christodoulopoulos
Ilias
I
Department of Molecular Microbiology and Immunology, Keck School of Medicine, University of Southern California, Los Angeles 90033, California, USA.
Droniou-Bonzom
Magali E
ME
Oldenburg
Jill E
JE
Cannon
Paula M
PM
D002460:Cell Line; D015686:Gene Products, env / Q000037:antagonists & inhibitors* / Q000235:genetics*; D015497:HIV-1 / Q000235:genetics*; D054298:Human Immunodeficiency Virus Proteins / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D017989:Leukemia Virus, Gibbon Ape / Q000235:genetics*; D025941:Protein Interaction Mapping; D054334:Viral Regulatory and Accessory Proteins / Q000235:genetics* / Q000378:metabolism*; D044844:beta-Transducin Repeat-Containing Proteins / Q000378:metabolism; D000086082:Viroporin Proteins
D015686:Gene Products, env; D054298:Human Immunodeficiency Virus Proteins; D054334:Viral Regulatory and Accessory Proteins; D044844:beta-Transducin Repeat-Containing Proteins; C514859:vpu protein, Human immunodeficiency virus 1; D000086082:Viroporin Proteins