| 1. |
- Kretova, Miroslava, et al.
(författare)
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TGF-beta/NF1/Smad4-mediated suppression of ANT2 contributes to oxidative stress in cellular senescence
- 2014
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Ingår i: Cellular Signalling. - : Elsevier BV. - 0898-6568 .- 1873-3913. ; 26:12, s. 2903-2911
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Tidskriftsartikel (refereegranskat)abstract
- 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-beta. 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-beta-mediated suppression of ANT2 through NF1/Smad4 complexes contributes to oxidative stress and DNA damage during induction of cellular senescence.
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| 2. |
- Luciakova, Katarina, et al.
(författare)
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Growth-dependent repression of human adenine nucleotide translocator-2 (ANT2) transcription : evidence for the participation of Smad and Sp family proteins in the NF1-dependent repressor complex
- 2008
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Ingår i: Biochemical Journal. - 0264-6021 .- 1470-8728. ; 412, s. 123-130
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Tidskriftsartikel (refereegranskat)abstract
- NF1 (nuclear factor 1) binds to two upstream elements of the human ANT2 (adenine nucleotide translocator-2) promoter and actively represses expression of the gene in growth-arrested diploid skin fibroblasts [Luciakova, Barath, Poliakova, Persson and Nelson (2003) J. Biol. Chem. 278, 30624-30633]. ChIP (chromatin immunoprecipitation) and co-immunoprecipitation analyses of nuclear extracts from growth-arrested and growth-activated diploid cells demonstrate that NF1, when acting as a repressor, is part of a multimeric complex that also includes Smad and Sp-family proteins. This complex appears to be anchored to both the upstream NF1-repressor elements and the proximal promoter, Sp1-dependent activation elements in growth-arrested cells. In growth-activated cells, the repressor complex dissociates and NF1 leaves the promoter. As revealed by co-immunoprecipitation experiments, NF1-Smad4-Sp3 complexes are present in nuclear extracts only from growth-inhibited cells, suggesting that the growth-state-dependent formation of these complexes is not an ANT2 promoter-specific event. Consistent with the role of Smad proteins in the repression complex, TGF-beta (transforming growth factor-beta) can fully repress ANT2 transcription in normally growing fibroblasts. Finally, pull-down experiments of in vitro transcribed/translated NF1 isoforms by GST (glutathione transferase)-Smad and GST-Smad MH fusion proteins indicate direct physical interactions between members of the two families. These findings suggest a possible functional relationship between the NF1 and Smad proteins that has not been previously observed.
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| 3. |
- Luciakova, Katarina, et al.
(författare)
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TGF-beta signals the formation of a unique NF1/Smad4-dependent transcription repressor-complex in human diploid fibroblasts
- 2011
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 411:3, s. 648-653
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Tidskriftsartikel (refereegranskat)abstract
- We earlier reported the formation of a unique nuclear NF1/Smad complex in serum-restricted fibroblasts that acts as an NF1-dependent repressor of the human adenine nucleotide translocase-2 gene (ANT2) [K. Luciakova, G. Kollarovic, P. Barath, B.D. Nelson, Growth-dependent repression of human adenine nucleotide translocator-2 (ANT2) transcription: evidence for the participation of Smad and Sp family proteins in the NF1-dependent repressor complex, Biochem. J. 412 (2008) 123-130]. In the present study, we show that TGF-beta, like serum-restriction: (a) induces the formation of NF1/Smad repressor complexes, (b) increases binding of the complexes to the repressor elements (Go elements) in the ANT2 promoter, and (c) inhibits ANT2 expression. Repression of ANT2 by TGF-beta is eliminated by mutating the NF1 binding sites in the Go repressor elements. All of the above responses to TGF-beta are prevented by inhibitors of TGF-beta RI and MAPK p38. These inhibitors also prevent NF1/Smad4 repressor complex formation and repression of ANT2 expression in serum-restricted cells, suggesting that similar signaling pathways are initiated by TGF-beta and serum-restriction. The present finding that NF1/Smad4 repressor complexes are formed through TGF-beta signaling pathways suggests a new, but much broader, role for these complexes in the initiation or maintenance of the growth-inhibited state.
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