HIV-1 protein Nef inhibits activity of ATP-binding cassette transporter A1 by targeting endoplasmic reticulum chaperone calnexin

J Biol Chem. 2014 Oct 17;289(42):28870-84. doi: 10.1074/jbc.M114.583591. Epub 2014 Aug 28.

Abstract

HIV-infected patients are at increased risk of developing atherosclerosis, in part due to an altered high density lipoprotein profile exacerbated by down-modulation and impairment of ATP-binding cassette transporter A1 (ABCA1) activity by the HIV-1 protein Nef. However, the mechanisms of this Nef effect remain unknown. Here, we show that Nef interacts with an endoplasmic reticulum chaperone calnexin, which regulates folding and maturation of glycosylated proteins. Nef disrupted interaction between calnexin and ABCA1 but increased affinity and enhanced interaction of calnexin with HIV-1 gp160. The Nef mutant that did not bind to calnexin did not affect the calnexin-ABCA1 interaction. Interaction with calnexin was essential for functionality of ABCA1, as knockdown of calnexin blocked the ABCA1 exit from the endoplasmic reticulum, reduced ABCA1 abundance, and inhibited cholesterol efflux; the same effects were observed after Nef overexpression. However, the effects of calnexin knockdown and Nef on cholesterol efflux were not additive; in fact, the combined effect of these two factors together did not differ significantly from the effect of calnexin knockdown alone. Interestingly, gp160 and ABCA1 interacted with calnexin differently; although gp160 binding to calnexin was dependent on glycosylation, glycosylation was of little importance for the interaction between ABCA1 and calnexin. Thus, Nef regulates the activity of calnexin to stimulate its interaction with gp160 at the expense of ABCA1. This study identifies a mechanism for Nef-dependent inactivation of ABCA1 and dysregulation of cholesterol metabolism.

Keywords: Atherosclerosis; Cholesterol Regulation; Confocal Microscopy; Endoplasmic Reticulum (ER); Human Immunodeficiency Virus (HIV).

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • ATP Binding Cassette Transporter 1 / metabolism*
  • Atherosclerosis / metabolism
  • Calnexin / metabolism*
  • Cholesterol / metabolism
  • Endoplasmic Reticulum / metabolism*
  • Glycosylation
  • HEK293 Cells
  • HIV Envelope Protein gp160 / metabolism*
  • HIV-1 / metabolism
  • HeLa Cells
  • Humans
  • Protein Binding
  • Spectrometry, Mass, Electrospray Ionization
  • Tandem Mass Spectrometry
  • nef Gene Products, Human Immunodeficiency Virus / metabolism*

Substances

  • ABCA1 protein, human
  • ATP Binding Cassette Transporter 1
  • HIV Envelope Protein gp160
  • gp160 protein, Human immunodeficiency virus 1
  • nef Gene Products, Human Immunodeficiency Virus
  • nef protein, Human immunodeficiency virus 1
  • Calnexin
  • Cholesterol