SAG/ROC2/Rbx2 is a novel activator protein-1 target that promotes c-Jun degradation and inhibits 12-O-tetradecanoylphorbol-13-acetate-induced neoplastic transformation

Cancer Res. 2007 Apr 15;67(8):3616-25. doi: 10.1158/0008-5472.CAN-06-4020.

Abstract

SAG (sensitive to apoptosis gene) was first identified as a stress-responsive protein that, when overexpressed, inhibited apoptosis both in vitro and in vivo. SAG was later found to be the second family member of ROC1 or Rbx1, a RING component of SCF and DCX E3 ubiquitin ligases. We report here that SAG/ROC2/Rbx2 is a novel transcriptional target of activator protein-1 (AP-1). AP-1 bound both in vitro and in vivo to two consensus binding sites in a 1.3-kb region of the mouse SAG promoter. The SAG promoter activity, as measured by luciferase reporter assay, was dependent on these sites. Consistently, endogenous SAG is induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) with an induction time course following the c-Jun induction in both mouse epidermal JB6-Cl.41 and human 293 cells. TPA-mediated SAG induction was significantly reduced in JB6-Cl.41 cells overexpressing a dominant-negative c-Jun, indicating a requirement of c-Jun/AP-1. On the other hand, SAG seemed to modulate the c-Jun levels. When overexpressed, SAG remarkably reduced both basal and TPA-induced c-Jun levels, whereas SAG small interfering RNA (siRNA) silencing increased substantially the levels of both basal and TPA-induced c-Jun. Consistently, SAG siRNA silencing reduced c-Jun polyubiquitination and blocked c-Jun degradation induced by Fbw7, an F-box protein of SCF E3 ubiquitin ligase. Finally, SAG overexpression inhibited, whereas SAG siRNA silencing enhanced, respectively, the TPA-induced neoplastic transformation in JB6-Cl.41 preneoplastic model. Thus, AP-1/SAG establishes an autofeedback loop, in which on induction by AP-1, SAG promotes c-Jun ubiquitination and degradation, thus inhibiting tumor-promoting activity of AP-1.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • Carrier Proteins / metabolism*
  • Cell Transformation, Neoplastic / chemically induced
  • Cell Transformation, Neoplastic / drug effects
  • Cell Transformation, Neoplastic / genetics
  • Cell Transformation, Neoplastic / metabolism*
  • Cloning, Molecular
  • Consensus Sequence
  • Doublecortin Protein
  • Gene Expression Regulation, Neoplastic
  • Gene Silencing
  • HeLa Cells
  • Humans
  • Promoter Regions, Genetic
  • Proto-Oncogene Proteins c-jun / metabolism*
  • RNA-Binding Proteins / biosynthesis
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism*
  • Tetradecanoylphorbol Acetate / antagonists & inhibitors
  • Tetradecanoylphorbol Acetate / pharmacology
  • Transcription Factor AP-1 / metabolism*
  • Ubiquitin-Protein Ligases
  • Ubiquitins / metabolism

Substances

  • Carrier Proteins
  • Dcx protein, mouse
  • Doublecortin Protein
  • Proto-Oncogene Proteins c-jun
  • RNA-Binding Proteins
  • Transcription Factor AP-1
  • Ubiquitins
  • RNF7 protein, human
  • Ubiquitin-Protein Ligases
  • Tetradecanoylphorbol Acetate