Oxidative stress and persistent activation of DNA damage response (DDR) are causally involved in the development of cellular senescence, a phenomenon implicated in fundamental (patho)physiological processes such as aging, fetal development and tumorigenesis. Here, we report that adenine nucleotide translocase-2 (ANT2) is consistently down-regulated in all three major forms of cellular senescence: replicative, oncogene-induced and drug-induced, in both normal and cancerous human cells. We previously reported formation of novel NF1/Smad transcription repressor complexes in growth-arrested fibroblasts. Here we show that such complexes form in senescent cells. Mechanistically, binding of the NF1/Smad complexes to the NF1-dependent repressor elements in the ANT2 gene promoter repressed ANT2 expression. Etoposide-induced formation of these complexes and repression of ANT2 were relatively late events co-incident with production and secretion of, and dependent on, TGF-β. siRNA-mediated knock-down of ANT2 in proliferating cells resulted in increased levels of reactive oxygen species (ROS) and activation of the DDR. Knock-down of ANT2, together with etoposide treatment, further intensified ROS production and DNA damage signaling, leading to enhanced apoptosis. Together, our data show that TGF-β-mediated suppression of ANT2 through NF1/Smad4 complexes contributes to oxidative stress and DNA damage during induction of cellular senescence.

• MeSH
• Cell Nucleus drug effects   metabolism   MeSH
• Cell Line MeSH
• Cytoprotection drug effects   MeSH
• Down-Regulation drug effects   MeSH
• Etoposide pharmacology   MeSH
• Humans MeSH
• Mutation MeSH
• Oxidative Stress * drug effects   MeSH
• DNA Damage MeSH
• Promoter Regions, Genetic MeSH
• Smad4 Protein metabolism   MeSH
• Repressor Proteins metabolism   MeSH
• Cellular Senescence * drug effects   MeSH
• Transforming Growth Factor beta metabolism   MeSH
• NFI Transcription Factors metabolism   MeSH
• Adenine Nucleotide Translocator 2 genetics   metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

One of the most significant insults that jeopardize cardiomyocyte homeostasis is a surge of reactive oxygen species (ROS) in the failing myocardium. Early growth response factor-1 (Egr-1) has been found to act as a transcriptional regulator in multiple biological processes known to exert deleterious effects on cardiomyocytes. We thus investigated the signaling pathways involved in its regulation by H2O2. Egr-1 mRNA levels were found to be maximally induced after 2 h in H2O2-treated H9c2 cells. Egr-1 respective response at the protein level, was found to be maximally induced after 2 h of treatment with 200 µM H2O2, remaining elevated for 6 h, and declining thereafter. H2O2- induced upregulation of Egr-1 mRNA and protein levels was ablated in the presence of agents inhibiting ERKs pathway (PD98059) and JNKs (SP600125, AS601245). Immunofluorescent experiments revealed H2O2-induced Egr-1 nuclear sequestration to be also ERK- and JNK-dependent. Overall, our results show for the first time the fundamental role of ERKs and JNKs in regulating Egr-1 response to H2O2 treatment in cardiac cells at multiple levels: mRNA, protein and subcellular distribution. Nevertheless, further studies are required to elucidate the specific physiological role of Egr-1 regarding the modulation of gene expression and determination of cell fate.

• MeSH
• Active Transport, Cell Nucleus MeSH
• Cell Nucleus enzymology   drug effects   MeSH
• Cell Line MeSH
• Time Factors MeSH
• Extracellular Signal-Regulated MAP Kinases antagonists & inhibitors   metabolism   MeSH
• Financing, Organized MeSH
• Fluorescent Antibody Technique MeSH
• Protein Kinase Inhibitors pharmacology   MeSH
• JNK Mitogen-Activated Protein Kinases antagonists & inhibitors   metabolism   MeSH
• Myocytes, Cardiac enzymology   drug effects   MeSH
• Rats MeSH
• RNA, Messenger metabolism   MeSH
• Hydrogen Peroxide pharmacology   MeSH
• Early Growth Response Protein 1 genetics   metabolism   MeSH
• Up-Regulation MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Animals MeSH