Nutlin-3 induces BCL2A1 expression by activating ELK1 through the mitochondrial p53-ROS-ERK1/2 pathway

Int J Oncol. 2014 Aug;45(2):675-82. doi: 10.3892/ijo.2014.2463. Epub 2014 May 26.

Abstract

Nutlin-3 which occupies the p53 binding pocket in HDM2, has been reported to activate apoptosis through both the transcriptional activity-dependent and -independent programs of p53. Transcription-independent apoptosis by nutlin-3 is triggered by p53 which is translocated to mitochondria. However, we previously demonstrated that the nutlin-3-induced mitochondrial translocation of p53 stimulates ERK1/2 activation, an anti-apoptosis signal, via mitochondrial ROS generation. We report on how nutlin-3-stimulated ERK1/2 activity inhibits p53-induced apoptosis. Among the anti-apoptotic BCL2 family proteins, BCL2A1 expression was increased by nutlin-3 at both the mRNA and protein levels, and this increase was prevented by the inhibition of ERK1/2. TEMPO, a ROS scavenger, and PFT-μ , a blocker of the mitochondrial translocation of p53, also inhibited BCL2A1 expression as well as ERK1/2 phosphorylation. In addition, nutlin-3 stimulated phosphorylation of ELK1, which was prevented by all compounds that inhibited nutlin-3-induced ERK1/2 such as U0126, PFT-μ and TEMPO. Moreover, an increase in BCL2A1 expression was weakened by the knockdown of ELK1. Finally, nutlin-3-induced apoptosis was found to be potentiated by the knockdown of BCL2A1, as demonstrated by an increase of in hypo-diploidic cells and Annexin V-positive cells. Parallel to the increase in apoptotic cells, the knockdown of BCL2A1 augmented the cleavage of poly(ADP-ribose) polymerase-1. It is noteworthy that the augmented levels of apoptosis induced by the knockdown of BCL2A1 were comparable to those of apoptosis induced by U0126. Collectively, these results suggest that nutlin-3-activated ERK1/2 may stimulate the transcription of BCL2A1 via the activation of ELK1, and BCL2A1 expression may contribute to the inhibitory effect of ERK1/2 on nutlin-3-induced apoptosis, thereby constituting a negative feedback loop of p53-induced apoptosis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis / physiology*
  • Cell Line, Tumor
  • Enzyme Activation / drug effects
  • Flow Cytometry
  • Humans
  • Imidazoles / pharmacology*
  • Immunoblotting
  • MAP Kinase Signaling System / physiology
  • Minor Histocompatibility Antigens
  • Mitochondria / drug effects
  • Mitochondria / metabolism
  • Osteosarcoma / metabolism
  • Piperazines / pharmacology*
  • Proto-Oncogene Proteins c-bcl-2 / biosynthesis*
  • RNA, Small Interfering
  • Reactive Oxygen Species / metabolism
  • Real-Time Polymerase Chain Reaction
  • Reverse Transcriptase Polymerase Chain Reaction
  • Signal Transduction / drug effects
  • Signal Transduction / physiology*
  • Transfection
  • Tumor Suppressor Protein p53 / metabolism
  • ets-Domain Protein Elk-1 / metabolism*

Substances

  • BCL2-related protein A1
  • ELK1 protein, human
  • Imidazoles
  • Minor Histocompatibility Antigens
  • Piperazines
  • Proto-Oncogene Proteins c-bcl-2
  • RNA, Small Interfering
  • Reactive Oxygen Species
  • TP53 protein, human
  • Tumor Suppressor Protein p53
  • ets-Domain Protein Elk-1
  • nutlin 3