A conserved dileucine motif mediates clathrin and AP-2-dependent endocytosis of the HIV-1 envelope protein.
During the assembly of enveloped viruses viral and cellular components essential for infectious particles must colocalize at specific membrane locations. For the human and simian immunodeficiency viruses (HIV and SIV), sorting of the viral envelope proteins (Env) to assembly sites is directed by trafficking signals located in the cytoplasmic domain of the transmembrane protein gp41 (TM). A membrane proximal conserved GYxxØ motif mediates endocytosis through interaction with the clathrin adaptor AP-2. However, experiments with SIV(mac239) Env indicate the presence of additional signals. Here we show that a conserved C-terminal dileucine in HIV(HxB2) also mediates endocytosis. Biochemical and morphological assays demonstrate that the C-terminal dileucine motif mediates internalization as efficiently as the GYxxØ motif and that both must be removed to prevent Env internalization. RNAi experiments show that depletion of the clathrin adaptor AP-2 leads to increased plasma membrane expression of HIV Env and that this adaptor is required for efficient internalization mediated by both signals. The redundancy of conserved endocytosis signals and the role of the SIV(mac239) Env GYxxØ motif in SIV pathogenesis, suggest that these motifs have functions in addition to endocytosis, possibly related to Env delivery to the site of viral assembly and/or incorporation into budding virions.
Byland
Rahel
R
Cell Biology Unit, MRC-Laboratory for Molecular Cell Biology and Department of Biochemistry and Molecular Biology, University College London, London WC1E 6BT, United Kingdom.
Vance
Patricia J
PJ
Hoxie
James A
JA
Marsh
Mark
M
D033962:Adaptor Protein Complex 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D015704:CD4 Antigens / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002966:Clathrin / Q000378:metabolism*; D017124:Conserved Sequence; D004705:Endocytosis; D015686:Gene Products, env / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D015497:HIV-1; D006367:HeLa Cells; D006801:Humans; D007930:Leucine / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism
D033962:Adaptor Protein Complex 2; D015704:CD4 Antigens; D002966:Clathrin; D015686:Gene Products, env; D011993:Recombinant Fusion Proteins; D007930:Leucine