Abstract
Therapy-induced senescence (TIS or therapy-induced premature senescence) is a key cellular program triggered in the course of cancer radiotherapy and chemotherapy with genotoxic drugs, both in cancer cells and in normal cells, whose activation critically affects the outcome of cancer therapy. Drug-induced senescent cells undergo a permanent cell cycle arrest, acquire distinctive morphological and biochemical alterations, and an enhanced secretory ability, referred to as senescence-associated secretory phenotype (SASP). The transcription factor NF-κB acts as a master regulator of the SASP, driving the expression of senescence-associated secretome components.
Here we describe protocols for the establishment of a tetracycline-regulated cell system for the investigation of the role of NF-κB in TIS. We also describe protocols routinely used in our laboratory, to investigate TIS in this Tet-On inducible expression system. Finally, we describe techniques for the validation of TIS induction.
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te Poele RH, Okorokov AL, Jardine L, Cummings J, Joel SP (2002) DNA damage is able to induce senescence in tumor cells in vitro and in vivo. Cancer Res 62:1876–1883
Mirzayans R, Scott A, Cameron M, Murray D (2005) Induction of accelerated senescence by gamma radiation in human solid tumor-derived cell lines expressing wild-type TP53. Radiat Res 163:53–62
Demaria M, O’Leary MN, Chang J, Shao L, Liu S, Alimirah F et al (2017) Cellular senescence promotes adverse effects of chemotherapy and cancer relapse. Cancer Discov 7:165–176
Coppé JP, Patil CK, Rodier F, Sun Y, Muñoz DP, Goldstein J et al (2008) Senescence-associated secretory phenotypes reveal cell-nonautonomous functions of oncogenic RAS and the p53 tumour suppressor. PLoS Biol 6:2853–2868
Kuilman T, Peeper DS (2009) Senescence-messaging secretome: SMS-ing cellular stress. Nat Rev Cancer 9:81–94
Chien Y, Scuoppo C, Wang X, Fang X, Balgley B, Bolden JE et al (2011) Control of the senescence-associated secretory phenotype by NF-κB promotes senescence and enhances chemosensitivity. Genes Dev 25:2125–2136
Crescenzi E, Pacifico F, Lavorgna A, De Palma R, D’Aiuto E, Palumbo G, Formisano S et al (2011) NF-κB-dependent cytokine secretion controls Fas expression on chemotherapy-induced premature senescent tumour cells. Oncogene 30:2707–2717
Wu PC, Wang Q, Grobman L, Chu E, Wu DY (2012) Accelerated cellular senescence in solid tumour therapy. Exp Oncol 34:298–305
Sidi R, Pasello G, Opitz I, Soltermann A, Tutic M et al (2011) Induction of senescence markers after neo-adjuvant chemotherapy of malignant pleural mesothelioma and association with clinical outcome: an exploratory analysis. Eur J Cancer 47:326–332
Kim SB, Bozeman RG, Kaisani A, Kim W, Zhang L, Richardson JA et al (2016) Radiation promotes colorectal cancer initiation and progression by inducing senescence-associated inflammatory responses. Oncogene 35:3365–3375
Iannello A, Thompson TW, Ardolino M, Lowe SW, Raulet DH (2013) p53-dependent chemokine production by senescent tumor cells supports NKG2D-dependent tumor elimination by natural killer cells. J Exp Med 210:2057–2069
Eggert T, Wolter K, Ji J, Ma C, Yevsa T, Klotz S et al (2016) Distinct functions of senescence-associated immune responses in liver tumor surveillance and tumor progression. Cancer Cell 30:533–547
Gorgoulis V, Adams PD, Alimonti A, Bennett DC, Bischof O, Bishop C et al (2019) Cellular senescence: defining a path forward. Cell 179:813–827
Dimri GP, Lee X, Basile G, Acosta M, Scott G, Roskelley C et al (1995) A biomarker that identifies senescent human cells in culture and in aging skin in vivo: distinct roles for cyclin-dependent kinases in cell cycle control. Proc Natl Acad Sci U S A 92:9363–9367
Kurz DJ, Decary S, Hong Y, Erusalimsky JD (2000) Senescence-associated (beta)-galactosidase reflects an increase in lysosomal mass during replicative ageing of human endothelial cells. J Cell Sci 113:3613–3622
Muñoz-Espín D, Rovira M, Galiana I, Giménez C, Lozano-Torres B, Paez-Ribes M et al (2018) A versatile drug delivery system targeting senescent cells. EMBO Mol Med 10(9):pii: e9355
Alimonti A, Nardella C, Chen Z, Clohessy JG, Carracedo A, Trotman LC et al (2010) A novel type of cellular senescence that can be enhanced in mouse models and human tumor xenografts to suppress prostate tumorigenesis. J Clin Invest 120:681–693
Muñoz-Espín D, Cañamero M, Maraver A, Gómez-López G, Contreras J, Murillo-Cuesta S et al (2013) Programmed cell senescence during mammalian embryonic development. Cell 155:1104–1118
Sedelnikova OA, Horikawa I, Zimonjic DB, Popescu NC, Bonner WM, Barrett JC (2004) Senescing human cells and ageing mice accumulate DNA lesions with unrepairable double-strand breaks. Nat Cell Biol 6:168–170
Rossiello F, Herbig U, Longhese MP, Fumagalli M, d’Adda di Fagagna F (2014) Irreparable telomeric DNA damage and persistent DDR signalling as a shared causative mechanism of cellular senescence and ageing. Curr Opin Genet Dev 26:89–95
Crescenzi E, Palumbo G, de Boer J, Brady HJ (2008) Ataxia telangiectasia mutated and p21CIP1 modulate cell survival of drug-induced senescent tumor cells: implications for chemotherapy. Clin Cancer Res 14:1877–1887
Gossen M, Bujard H (2001) Tetracyclines in the control of gene expression in eukaryotes. In: Nelson M, Hillen W, Greenwald RA (eds) Tetracyclines in biology, chemistry and medicine. Birkhäuser Verlag, Basel, pp 139–157
Baron U, Bujard H (2000) Tet repressor-based system for regulated gene expression in eukaryotic cells: principles and advances. Methods Enzymol 327:401–421
Berens C, Hillen W (2003) Gene regulation by tetracyclines. Constraints of resistance regulation in bacteria shape TetR for application in eukaryotes. Eur J Biochem 270:3109–3121
Baeuerle PA, Baltimore D (1988) I kappa B: a specific inhibitor of the NF-kappa B transcription factor. Science 242:540–546
Brown K, Gerstberger FS, Carlson L, Franzoso G, Siebenlist U (1995) Control of IκBα proteolysis by site-specific, signal-induced phosphorylation. Science 267:1485–1491
Traenckner EB, Pahl HL, Henkel T, Schmidt KN, Wilk S, Baeuerle PA (1995) Phosphorylation of human I kappa B-alpha on serines 32 and 36 controls I kappa B-alpha proteolysis and NF-kappa B activation in response to diverse stimuli. EMBO J 14:2876–2883
Severino J, Allen RG, Balin S, Balin A, Cristofalo VJ (2000) Is beta-galactosidase staining a marker of senescence in vitro and in vivo? Exp Cell Res 257:162–171
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Pacifico, F., Crescenzi, E., Leonardi, A. (2021). Analysis of the Contribution of NF-κB in the Regulation of Chemotherapy-Induced Cell Senescence by Establishing a Tetracycline-Regulated Cell System. In: Franzoso, G., Zazzeroni, F. (eds) NF-κB Transcription Factors. Methods in Molecular Biology, vol 2366. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1669-7_12
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DOI: https://doi.org/10.1007/978-1-0716-1669-7_12
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