title: HIV-1 Nef sequesters MHC-I intracellularly by targeting early stages of endocytosis and recycling.
abstract: A defining characteristic of HIV-1 infection is the ability of the virus to persist within the host. Specifically, MHC-I downregulation by the HIV-1 accessory protein Nef is of critical importance in preventing infected cells from cytotoxic T-cell mediated killing. Nef downregulates MHC-I by modulating the host membrane trafficking machinery, resulting in the endocytosis and eventual sequestration of MHC-I within the cell. In the current report, we utilized the intracellular protein-protein interaction reporter system, bimolecular fluorescence complementation (BiFC), in combination with super-resolution microscopy, to track the Nef/MHC-I interaction and determine its subcellular localization in cells. We demonstrate that this interaction occurs upon Nef binding the MHC-I cytoplasmic tail early during endocytosis in a Rab5-positive endosome. Disruption of early endosome regulation inhibited Nef-dependent MHC-I downregulation, demonstrating that Nef hijacks the early endosome to sequester MHC-I within the cell. Furthermore, super-resolution imaging identified that the Nef:MHC-I BiFC complex transits through both early and late endosomes before ultimately residing at the trans-Golgi network. Together we demonstrate the importance of the early stages of the endocytic network in the removal of MHC-I from the cell surface and its re-localization within the cell, which allows HIV-1 to optimally evade host immune responses.
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, Schulich School of Medicine and Dentistry, London, Ontario, Canada.
authors.forename: Brennan S
authors.initials: BS
authors.lastname: Dirk
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, Schulich School of Medicine and Dentistry, London, Ontario, Canada.
authors.forename: Emily N
authors.initials: EN
authors.lastname: Pawlak
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, Schulich School of Medicine and Dentistry, London, Ontario, Canada.
authors.forename: Aaron L
authors.initials: AL
authors.lastname: Johnson
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, Schulich School of Medicine and Dentistry, London, Ontario, Canada.
authors.forename: Logan R
authors.initials: LR
authors.lastname: Van Nynatten
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, Schulich School of Medicine and Dentistry, London, Ontario, Canada.
authors.forename: Rajesh A
authors.initials: RA
authors.lastname: Jacob
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, Schulich School of Medicine and Dentistry, London, Ontario, Canada.
authors.forename: Bryan
authors.initials: B
authors.lastname: Heit
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, Schulich School of Medicine and Dentistry, London, Ontario, Canada.
authors.forename: Jimmy D
authors.initials: JD
authors.lastname: Dikeakos
mesh_terms: D015536:Down-Regulation; D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism; D057809:HEK293 Cells; D015497:HIV-1 / Q000378:metabolism*; D006367:HeLa Cells; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008856:Microscopy, Fluorescence; D011485:Protein Binding; D054311:nef Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000378:metabolism*; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism; D021601:trans-Golgi Network / Q000378:metabolism; D000099193:rab11 GTP-Binding Proteins
chemical_list: D015395:Histocompatibility Antigens Class I; D054311:nef Gene Products, Human Immunodeficiency Virus; D020691:rab GTP-Binding Proteins; D020696:rab5 GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins; C514837:nef protein, Human immunodeficiency virus 1