HIV-1 Nef induces p47phox phosphorylation leading to a rapid superoxide anion release from the U937 human monoblastic cell line

Eleonora Olivetta, Cinzia Mallozzi, Vitalba Ruggieri, Donatella Pietraforte, Maurizio Federico, Massimo Sanchez

Research output: Contribution to journalArticlepeer-review

Abstract

The Nef protein of the human immunodeficiency virus type 1 (HIV-1) plays a crucial role in AIDS pathogenesis by modifying host cell signaling pathways. We investigated the effects of Nef on the NADPH oxidase complex, a key enzyme involved in the generation of reactive oxygen species during the respiratory burst in human monocyte/macrophages. We have recently shown that the inducible expression of HIV-1 Nef in human macrophages cell line modulates in bi-phasic mode the superoxide anion release by NADPH oxidase, inducing a fast increase of the superoxide production, followed by a delayed strong inhibition mediated by Nef-induced soluble factor(s). Our study is focused on the molecular mechanisms involved in Nef-mediated activation of NADPH oxidase and superoxide anion release. Using U937 cells stably transfected with different Nef alleles, we found that both Nef membrane localization and intact SH3-binding domain are needed to induce superoxide release. The lack of effect during treatment with a specific MAPK pathway inhibitor, PD98059, demonstrated that Nef-induced superoxide release is independent of Erk1/2 phosphorylation. Furthermore, Nef induced the phosphorylation and then the translocation of the cytosolic subunit of NADPH oxidase complex p47phox to the plasma membrane. Adding the inhibitor PP2 prevented this process, evidencing the involvement of the Src family kinases on Nef-mediated NADPH oxidase activation. In addition, LY294002, a specific inhibitor of phosphoinositide 3-kinase (PI3K) inhibited both the Nef-induced p47phox phosphorylation and the superoxide anion release. These data indicate that Nef regulates the NADPH oxidase activity through the activation of the Src kinases and PI3K. J. Cell. Biochem. 106: 812-822, 2009.

Original languageEnglish
Pages (from-to)812-822
Number of pages11
JournalJournal of Cellular Biochemistry
Volume106
Issue number5
DOIs
Publication statusPublished - Apr 1 2009

Keywords

  • NADPH oxidase
  • PI3K
  • ROS
  • Src family kinases
  • U937 cell line

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

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