The NADPH Oxidase Subunit p22 Inhibits the Function of the Tumor Suppressor Protein Tuberin

• DOI: 10.2353/ajpath.2010.090606
• Publication Date: 2010-03-20T00:51:21Z
• Authors: Karen Block; Yves Gorin; David D. New; Assaad Eid; Tomasz Chelmicki; Amanda Reed; Goutam Ghosh Chou...; Dipen J. Parekh; Hanna E. Abboud
• Keywords: 0301 basic medicine; Tumor Suppressor Proteins; NADPH Oxidases; Immunohistochemistry; Models, Biological; Gene Expression Regulation, Enzymologic; 3. Good health; 03 medical and health sciences; Cell Transformation, Neoplastic; Cell Line, Tumor; Protein Biosynthesis; Mutation; Tuberous Sclerosis Complex 2 Protein; Basic Helix-Loop-Helix Transcription Factors; Humans; Phosphorylation; Hypoxia; Carcinoma, Renal Cell

Mutations in the von Hippel-Lindau (VHL) gene give rise to renal cell carcinoma. Reactive oxygen species, generated by Nox oxidases, are involved in tumorigenesis. We have previously demonstrated that in VHL-deficient cells, p22(phox)-dependent Nox1 and Nox4 oxidases maintain hypoxia inducible factor-2alpha (HIF-2alpha) protein expression through an Akt-dependent translational pathway. Phosphorylation of tuberin, by Akt, results in its inactivation. Here we show that diphenyleneiodonium chloride, an inhibitor of Nox oxidases, and small-interfering RNA-mediated down-regulation of p22(phox) inhibit Akt-dependent phosphorylation of tuberin and stabilizes tuberin protein levels in VHL-deficient renal carcinoma cells. p22(phox)-mediated inactivation of tuberin is associated with an increase in ribosomal protein S6 kinase 1 and eukaryotic initiation factor 4E-binding protein-1 (4E-BP1) phosphorylation as well as HIF-2alpha stabilization. Importantly, we find that marked up-regulation of p22(phox) in human renal cell carcinoma correlates with increased tuberin phosphorylation, decreased tuberin protein levels, and increased phosphorylation of 4E-BP1. Our data provide the first evidence that p22(phox)-based Nox oxidases maintain HIF-2alpha protein expression through inactivation of tuberin and downstream activation of ribosomal protein S6 kinase 1/4E-BP1 pathway.