A Genome-Wide Screen for Machinery Involved in Downregulation of MHC Class I by HIV-1 Nef

PLoS One. 2015 Oct 14;10(10):e0140404. doi: 10.1371/journal.pone.0140404. eCollection 2015.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism
  • Dopamine Plasma Membrane Transport Proteins / genetics
  • Dopamine Plasma Membrane Transport Proteins / metabolism
  • Down-Regulation*
  • Genome-Wide Association Study
  • HLA-A2 Antigen / genetics
  • HLA-A2 Antigen / metabolism*
  • HeLa Cells
  • Humans
  • Intracellular Signaling Peptides and Proteins
  • Microtubule-Associated Proteins / genetics
  • Microtubule-Associated Proteins / metabolism
  • Potassium Channels / genetics
  • Potassium Channels / metabolism
  • nef Gene Products, Human Immunodeficiency Virus / genetics
  • nef Gene Products, Human Immunodeficiency Virus / metabolism*

Substances

  • Carrier Proteins
  • Dopamine Plasma Membrane Transport Proteins
  • HLA-A2 Antigen
  • IIp45 protein, human
  • Intracellular Signaling Peptides and Proteins
  • Microtubule-Associated Proteins
  • Potassium Channels
  • SLC6A3 protein, human
  • nef Gene Products, Human Immunodeficiency Virus