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DNA damage triggers a phosphorylation-based signaling cascade known as the DNA damage response. p19INK4d, a member of the INK4 family of CDK4/6 inhibitors, has been reported to participate in the DNA damage response promoting DNA repair and cell survival. Here, we provide mechanistic insight into the activation mechanism of p19INK4d linked to the response to DNA damage. Results showed that p19INK4d becomes phosphorylated following UV radiation, b-amyloid peptide and cisplatin treatments. ATM-Chk2/ATR-Chk1 signaling pathways were found to be differentially involved in p19INK4d phosphorylation depending on the type of DNA damage. Two sequential phosphorylation events at serine 76 and threonine 141 were identified using p19INK4d single-point mutants in metabolic labeling assays with 32P-orthophosphate. CDK2 and PKA were found to participate in p19INK4d phosphorylation process and that they would mediate serine 76 and threonine 141 modifications respectively. Nuclear translocation of p19INK4d induced by DNA damage was shown to be dependent on serine 76 phosphorylation. Most importantly, both phosphorylation sites were found to be crucial for p19INK4d function in DNA repair and cell survival. In contrast, serine 76 and threonine 141 were dispensable for CDK4/6 inhibition highlighting the independence of p19INK4d functions, in agreement with our previous findings. These results constitute the first description of the activation mechanism of p19INK4d in response to genotoxic stress and demonstrate the functional relevance of this activation following DNA damage. © 2012 Marazita et al.


Documento: Artículo
Título:CDK2 and PKA mediated-sequential phosphorylation is critical for p19INK4d function in the DNA damage response
Autor:Marazita, M.C.; Florencia Ogara, M.; Sonzogni, S.V.; Martí, M.; Dusetti, N.J.; Pignataro, O.P.; Cánepa, E.T.
Filiación:Laboratorio de Biología Molecular, Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad de Buenos Aires, Argentina
INSERM, U624 Stress Cellulaire, Marseille, France
Laboratorio de Endocrinología Molecular y Transducción de señales, Instituto de Biología y Medicina Experimental-CONICET, Ciudad de Buenos Aires, Argentina
Palabras clave:amyloid beta protein; ATM protein; ATR protein; checkpoint kinase 1; checkpoint kinase 2; cisplatin; cyclic AMP dependent protein kinase; cyclin dependent kinase 2; cyclin dependent kinase 4; cyclin dependent kinase 6; cyclin dependent kinase inhibitor 2D; phosphate p 32; serine; threonine; amyloid beta protein; CDK2 protein, human; cisplatin; cyclic AMP dependent protein kinase; cyclin dependent kinase 2; cyclin dependent kinase inhibitor 2D; DNA; article; cell nucleus; cell survival; conformational transition; controlled study; DNA damage; DNA repair; point mutation; protein phosphorylation; signal transduction; ultraviolet radiation; cell cycle; cell line; cell strain HEK293; DNA damage; DNA repair; drug effect; gene expression regulation; genetics; human; metabolism; mutation; phosphorylation; protein transport; radiation exposure; Amyloid beta-Peptides; Cell Cycle; Cell Line; Cell Survival; Cisplatin; Cyclic AMP-Dependent Protein Kinases; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase Inhibitor p19; DNA; DNA Damage; DNA Repair; Gene Expression Regulation; HEK293 Cells; Humans; Mutation; Phosphorylation; Protein Transport; Signal Transduction; Ultraviolet Rays
Título revista:PLoS ONE
Título revista abreviado:PLoS ONE
CAS:amyloid beta protein, 109770-29-8; checkpoint kinase 2, 244634-79-5; cisplatin, 15663-27-1, 26035-31-4, 96081-74-2; cyclin dependent kinase 2, 141349-86-2; cyclin dependent kinase 4, 147014-97-9; phosphate p 32, 8027-28-9; serine, 56-45-1, 6898-95-9; threonine, 36676-50-3, 72-19-5; DNA, 9007-49-2; Amyloid beta-Peptides; CDK2 protein, human,; Cisplatin, 15663-27-1; Cyclic AMP-Dependent Protein Kinases,; Cyclin-Dependent Kinase 2,; Cyclin-Dependent Kinase Inhibitor p19; DNA, 9007-49-2


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---------- APA ----------
Marazita, M.C., Florencia Ogara, M., Sonzogni, S.V., Martí, M., Dusetti, N.J., Pignataro, O.P. & Cánepa, E.T. (2012) . CDK2 and PKA mediated-sequential phosphorylation is critical for p19INK4d function in the DNA damage response. PLoS ONE, 7(4).
---------- CHICAGO ----------
Marazita, M.C., Florencia Ogara, M., Sonzogni, S.V., Martí, M., Dusetti, N.J., Pignataro, O.P., et al. "CDK2 and PKA mediated-sequential phosphorylation is critical for p19INK4d function in the DNA damage response" . PLoS ONE 7, no. 4 (2012).
---------- MLA ----------
Marazita, M.C., Florencia Ogara, M., Sonzogni, S.V., Martí, M., Dusetti, N.J., Pignataro, O.P., et al. "CDK2 and PKA mediated-sequential phosphorylation is critical for p19INK4d function in the DNA damage response" . PLoS ONE, vol. 7, no. 4, 2012.
---------- VANCOUVER ----------
Marazita, M.C., Florencia Ogara, M., Sonzogni, S.V., Martí, M., Dusetti, N.J., Pignataro, O.P., et al. CDK2 and PKA mediated-sequential phosphorylation is critical for p19INK4d function in the DNA damage response. PLoS ONE. 2012;7(4).