title: An interdomain binding site on HIV-1 Nef interacts with PACS-1 and PACS-2 on endosomes to down-regulate MHC-I.
abstract: The human immunodeficiency virus type 1 (HIV-1) accessory protein Nef directs virus escape from immune surveillance by subverting host cell intracellular signaling and membrane traffic to down-regulate cell-surface major histocompatibility complex class I (MHC-I). The interaction of Nef with the sorting proteins PACS-1 and PACS-2 mediates key signaling and trafficking steps required for Nef-mediated MHC-I down-regulation. Little is known, however, about the molecular basis underlying the Nef-PACS interaction. Here we identify the sites on Nef and the PACS proteins required for their interaction and describe the consequences of disrupting this interaction for Nef action. A previously unidentified cargo subsite on PACS-1 and PACS-2 interacted with a bipartite site on Nef formed by the EEEE(65) acidic cluster on the N-terminal domain and W(113) in the core domain. Mutation of these sites prevented the interaction between Nef and the PACS proteins on Rab5 (PACS-2 and PACS-1)- or Rab7 (PACS-1)-positive endosomes as determined by bimolecular fluorescence complementation and caused a Nef mutant defective in PACS binding to localize to distorted endosomal compartments. Consequently, disruption of the Nef-PACS interaction repressed Nef-induced MHC-I down-regulation in peripheral blood mononuclear cells. Our results provide insight into the molecular basis of Nef action and suggest new strategies to combat HIV-1.
authors.affiliation: Vollum Institute, Oregon Health and Science University, Portland, OR 97239, USA.
authors.forename: Jimmy D
authors.initials: JD
authors.lastname: Dikeakos
authors.forename: Laurel
authors.initials: L
authors.lastname: Thomas
authors.forename: Grace
authors.initials: G
authors.lastname: Kwon
authors.forename: Johannes
authors.initials: J
authors.lastname: Elferich
authors.forename: Ujwal
authors.initials: U
authors.lastname: Shinde
authors.forename: Gary
authors.initials: G
authors.lastname: Thomas
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D001665:Binding Sites; D015536:Down-Regulation; D011992:Endosomes / Q000378:metabolism*; D045683:Furin / Q000378:metabolism; D006367:HeLa Cells; D015395:Histocompatibility Antigens Class I / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D013329:Structure-Activity Relationship; D033921:Vesicular Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D054311:nef Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D000596:Amino Acids; D015395:Histocompatibility Antigens Class I; C491914:PACS1 protein, human; C499638:PACS2 protein, human; D033921:Vesicular Transport Proteins; D054311:nef Gene Products, Human Immunodeficiency Virus; C514837:nef protein, Human immunodeficiency virus 1; D045683:Furin