HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 VPU PROTEIN INDUCES DEGRADATION OF CHIMERIC ENVELOPE GLYCOPROTEINS BEARING THE CYTOPLASMIC AND ANCHOR DOMAINS OF CD4 - ROLE OF THE CYTOPLASMIC DOMAIN IN VPU-INDUCED DEGRADATION IN THE ENDOPLASMIC-RETICULUM

The human immunodeficiency virus type 1 (HIV-1) Vpu protein is a trans membrane phosphoprotein which induces rapid degradation of CD4 in the endoplasmic reticulum (ER). To identify sequences in CD4 for Vpu-induc ed degradation, we generated four chimeric envelope glycoproteins havi ng the ectodomain of HIV-1 gp160, the anchor domain of CD4, and 38, 25 , 24, and 18 amino acids (aa) of the CD4 cytoplasmic domain. Using the vaccinia virus-T7 RNA polymerase expression system, we analyzed the e xpression of chimeric proteins in the presence and absence of Vpu. In singly transfected cells, the chimeric envelope glycoproteins having 3 8, 24, and 18 aa of the CD4 cytoplasmic domain were endoproteolyticall y cleaved and biologically active in the fusion of HeLa CD4+ cells. Ho wever, one of the chimeras having 25 aa of the CD4 cytoplasmic tail wa s retained in the ER using the transmembrane ER retention signal and w as defective in membrane fusion. Furthermore, biochemical analyses of the coexpressing cells revealed that the Vpu protein induced degradati on of the envelope glycoproteins having 38, 25, and 24 aa of the CD4 c ytoplasmic tail and degradation occurred in the ER. Consequently, the fusion-competent glycoproteins did not induce the formation of syncyti a in HeLa CD4+ cells expressing Vpu. However, the HIV-1 gp160 and chim eric envelope glycoprotein having the membrane-proximal 18 aa of the C D4 cytoplasmic tail were stable and fusion competent in cells expressi ng Vpu. In addition, we examined the stability of CD4 molecules in the presence of Vpu. Coexpression analyses revealed that the Vpu protein induced degradation of CD4 whereas mutant CD4 having the membrane-prox imal 18 aa of the cytoplasmic domain was relatively stable in the pres ence of Vpu. Taken together, these studies have elucidated that the Vp u protein requires sequences or sequence determinants in the cytoplasm ic domain of CD4 to induce degradation of the glycoproteins in the cel l.