title: A Genome-Wide Screen for Machinery Involved in Downregulation of MHC Class I by HIV-1 Nef.
abstract: The HIV-1-encoded protein, Nef, plays a key role in the development of AIDS. One of Nef's functions is to keep MHC class I off the surface of infected cells, a process that requires the host proteins clathrin and AP-1. To identify other proteins involved in this pathway, we carried out a genome-wide siRNA library screen on HeLa cells co-expressing HLA-A2 and an inducible form of Nef. Out of 21,121 siRNA pools, 100 were selected for further analysis, based on their ability to either inhibit or enhance downregulation of MHC-I by Nef. When cells were treated with the same siRNA pools as those used in the screen, 79% produced a similar phenotype. However, when the cells were treated with different siRNA reagents targeting the same genes, only 16% produced a similar phenotype. This indicates that most of the hits found in the original screen are likely to have been off-target, an important concern that is often not taken into account in siRNA screening studies. Nevertheless, we identified novel host factors involved in Nef-induced downregulation of MHC-I, including four genes, MIIP, CAMSAP3, SLC6A3, and KCTD19, where multiple reagents produced a strong inhibitory effect on Nef activity. Other hits slightly below our very high stringency cutoff point may also deserve further study. Thus, our dataset is a valuable resource for scientists investigating the pathogenesis of HIV.
authors.affiliation: University of Cambridge, Cambridge Institute for Medical Research, Cambridge, CB2 0XY, United Kingdom.
authors.forename: Maja K
authors.initials: MK
authors.lastname: Choma
authors.affiliation: University of Cambridge, Cambridge Institute for Medical Research, Cambridge, CB2 0XY, United Kingdom.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Lumb
authors.affiliation: University of Cambridge, Cambridge Institute for Medical Research, Cambridge, CB2 0XY, United Kingdom.
authors.forename: Patrycja
authors.initials: P
authors.lastname: Kozik
authors.affiliation: University of Cambridge, Cambridge Institute for Medical Research, Cambridge, CB2 0XY, United Kingdom.
authors.forename: Margaret S
authors.initials: MS
authors.lastname: Robinson
mesh_terms: D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D050483:Dopamine Plasma Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D055106:Genome-Wide Association Study; D015789:HLA-A2 Antigen / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D015221:Potassium Channels / Q000235:genetics / Q000378:metabolism; D054311:nef Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D050483:Dopamine Plasma Membrane Transport Proteins; D015789:HLA-A2 Antigen; D047908:Intracellular Signaling Peptides and Proteins; C480900:MIIP protein, human; D008869:Microtubule-Associated Proteins; D015221:Potassium Channels; C493214:SLC6A3 protein, human; D054311:nef Gene Products, Human Immunodeficiency Virus