Cooperative binding of the class I major histocompatibility complex cytoplasmic domain and human immunodeficiency virus type 1 Nef to the endosomal AP-1 complex via its mu subunit.
Human immunodeficiency virus type 1 Nef provides immune evasion by decreasing the expression of major histocompatibility complex class I (MHC-I) at the surfaces of infected cells. The endosomal clathrin adaptor protein complex AP-1 is a key cellular cofactor for this activity, and it is recruited to the MHC-I cytoplasmic domain (CD) in the presence of Nef by an uncharacterized mechanism. To determine the molecular basis of this recruitment, we used an MHC-I CD-Nef fusion protein to represent the MHC-I CD/Nef complex during protein interaction assays. The MHC-I CD had no intrinsic ability to bind AP-1, but it conferred binding activity when fused to Nef. This activity was independent of the canonical leucine-based AP-binding motif in Nef; it required residue Y320 in the MHC-I CD and residues E62-65 and P78 in Nef, and it involved the mu but not the gamma/sigma subunits of AP-1. The impaired binding of mutants encoding substitutions of E62-65 or P78 in Nef was rescued by replacing the Y320SQA sequence in the MHC-I CD with YSQL, suggesting that Nef allows the YSQA sequence to act as if it were a canonical mu-binding motif. These data identify the mu subunit of AP-1 (mu1) as the key target of the MHC-I CD/Nef complex, and they indicate that both Y320 in the MHC-I CD and E62-65 in Nef interact directly with mu1. The data support a cooperative binding model in which Nef functions as a clathrin-associated sorting protein that allows recognition of an incomplete, tyrosine-based mu-binding signal in the MHC-I CD by AP-1.
University of California-San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0679, USA.
Colleen M
CM
Noviello
Serge
S
Benichou
John C
JC
Guatelli
D020816:Amino Acid Motifs; D019943:Amino Acid Substitution / Q000235:genetics; D002460:Cell Line; D015497:HIV-1 / Q000276:immunology*; D015395:Histocompatibility Antigens Class I / Q000378:metabolism*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000378:metabolism; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D054311:nef Gene Products, Human Immunodeficiency Virus / Q000378:metabolism*
D015395:Histocompatibility Antigens Class I; D050505:Mutant Proteins; D018808:Transcription Factor AP-1; D054311:nef Gene Products, Human Immunodeficiency Virus; C514837:nef protein, Human immunodeficiency virus 1