The human immunodeficiency virus type 1 Vpu protein specifically binds to the cytoplasmic domain of CD4: implications for the mechanism of degradation.
We have recently demonstrated that coexpression of Vpu and CD4 in HeLa cells results in the degradation of CD4 in the endoplasmic reticulum. The sensitivity of CD4 to Vpu-mediated degradation is conferred by the presence of specific sequences located between amino acids 402 and 420 in the CD4 cytoplasmic domain. Using an in vitro translation system, we also showed that degradation of CD4 by Vpu requires the two proteins to be present in the same membrane compartment. Although these results suggest that spatial proximity between CD4 and Vpu may be critical in triggering degradation, it remains unknown whether the two molecules have the ability to interact with each other. In order to better define the mechanisms involved in CD4 degradation, we investigated the existence and functional relevance of direct interactions between CD4 and Vpu. Coimmunoprecipitation experiments showed that Vpu specifically binds to the cytoplasmic tail of CD4. This phenomenon is relevant to the mechanism of CD4 degradation since the ability of CD8/CD4 chimeric molecules and various CD4 mutants to form complexes with Vpu correlates with their sensitivity to degradation. Accordingly, we found that amino acid residues in the CD4 cytoplasmic tail previously shown to be important for degradation are necessary for Vpu binding. We further demonstrate that a deletion mutant of Vpu as well as a phosphorylation mutant, both biologically inactive with regard to CD4 degradation, retained the capacity to interact with the CD4 cytoplasmic domain. Taken together, these results indicate that Vpu binding is necessary to trigger CD4 degradation. However, the binding to target molecules is not sufficient per se to cause degradation. Interaction between CD4 and Vpu is thus likely to be an early event critical in triggering a multistep process leading to CD4 degradation.
Bour
S
S
Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases.
Schubert
U
U
Strebel
K
K
D000595:Amino Acid Sequence; D015704:CD4 Antigens / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D015497:HIV-1 / Q000378:metabolism*; D006367:HeLa Cells; D054298:Human Immunodeficiency Virus Proteins; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship; D054334:Viral Regulatory and Accessory Proteins / Q000378:metabolism*; D000086082:Viroporin Proteins
D015704:CD4 Antigens; D054298:Human Immunodeficiency Virus Proteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D054334:Viral Regulatory and Accessory Proteins; C514859:vpu protein, Human immunodeficiency virus 1; D000086082:Viroporin Proteins