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Apoptotic DNA degradation into oligonucleosomal fragments, but not apoptotic nuclear morphology, relies on a cytosolic pool of DFF40/CAD endonuclease
Iglesias Guimarais, Victoria (Universitat Autònoma de Barcelona. Institut de Neurociències)
Gil Guiñon, Estel (Universitat Autònoma de Barcelona. Institut de Neurociències)
Gabernet, Gisela (Universitat Autònoma de Barcelona. Institut de Neurociències)
Garcia i Belinchón, Maria Mercè (Universitat Autònoma de Barcelona. Institut de Neurociències)
Sánchez-Osuna, María (Universitat Autònoma de Barcelona. Institut de Neurociències)
Casanelles Abella, Elisenda (Universitat Autònoma de Barcelona. Institut de Neurociències)
Comella i Carnicé, Joan Xavier, 1963- (Vall d'Hebron Institut de Recerca)
Yuste Mateos, Víctor J. (Víctor José) (Universitat Autònoma de Barcelona. Departament de Bioquímica i Biologia Molecular)

Fecha: 2012
Resumen: Apoptotic cell death is characterized by nuclear fragmentation and oligonucleosomal DNA degradation, mediated by the caspase-dependent specific activation of DFF40/CAD endonuclease. Here, we describe how, upon apoptotic stimuli, SK-N-AS human neuroblastoma-derived cells show apoptotic nuclear morphology without displaying concomitant internucleosomal DNA fragmentation. Cytotoxicity afforded after staurosporine treatment is comparable with that obtained in SH-SY5Y cells, which exhibit a complete apoptotic phenotype. SK-N-AS cell death is a caspase-dependent process that can be impaired by the pan-caspase inhibitor q-VD-OPh. The endogenous inhibitor of DFF40/CAD, ICAD, is correctly processed, and dff40/cad cDNA sequence does not reveal mutations altering its amino acid composition. Biochemical approaches show that both SH-SY5Y and SK-N-AS resting cells express comparable levels of DFF40/CAD. However, the endonuclease is poorly expressed in the cytosolic fraction of healthy SK-N-AS cells. Despite this differential subcellular distribution of DFF40/CAD, we find no differences in the subcellular localization of both pro-caspase-3 and ICAD between the analyzed cell lines. After staurosporine treatment, the preferential processing of ICAD in the cytosolic fraction allows the translocation of DFF40/CAD from this fraction to a chromatin-enriched one. Therefore, the low levels of cytosolic DFF40/CAD detected in SK-N-AS cells determine the absence of DNA laddering after staurosporine treatment. In these cells DFF40/CAD cytosolic levels can be restored by the overexpression of their own endonuclease, which is sufficient to make them proficient at degrading their chromatin into oligonucleosome-size fragments after staurosporine treatment. Altogether, the cytosolic levels of DFF40/CAD are determinants in achieving a complete apoptotic phenotype, including oligonucleosomal DNA degradation.
Nota: Número d'acord de subvenció MICINN/FEDER/BFU2008-01328
Nota: Número d'acord de subvenció MICINN/FEDER/TRA2009-0185
Nota: Número d'acord de subvenció MICINN/FEDER/SAF2010-19953
Nota: Número d'acord de subvenció CIBERNED/CB06/05/0042
Nota: Número d'acord de subvenció AGAUR/SGR2009-346
Derechos: Tots els drets reservats.
Lengua: Anglès
Documento: article ; recerca ; publishedVersion
Materia: Apoptosis ; Caspase ; Cell death ; Neuroblastoma ; Subcellular fractionation ; DFF40/CAD ; Oligonucleosomal DNA degradation
Publicado en: Journal of biological chemistry, Vol. 287, Num. 10 (Mar 2012) , p. 7766-7779, ISSN 1083-351X

DOI: 10.1074/jbc.M111.290718
PMID: 22253444

18 p, 6.6 MB

El registro aparece en las colecciones:
Documentos de investigación > Documentos de los grupos de investigación de la UAB > Centros y grupos de investigación (producción científica) > Ciencias de la salud y biociencias > Institut de Neurociències (INc)
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 Registro creado el 2015-09-21, última modificación el 2020-11-21

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