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Int J Radiat Biol. 2000 Jun;76(6):841-51.

ATM protein is required for radiation-induced apoptosis and acts before mitochondrial collapse.

Author information

1
UMR 218 du Centre National de la Recherche Scientifique, LRC n 1 du Commissariat à l'Energie Atomique, Institut Curie-Recherche, Paris, France.

Abstract

PURPOSE:

To define the role of the ataxia telangiectasia (A-T) mutated gene (ATM) in activation and progress of apoptosis.

MATERIAL AND METHODS:

Three normal and three A-T EBV-transformed cell lines were studied. Following irradiation (IR), Fas activation or ceramide exposure, viability and apoptosis were measured by trypan blue dye exclusion assay and as sub-G1 cell fraction by flow cytometric analysis of propidium iodide stained cultures, respectively. Activation of caspase-3 was evaluated by immunoblot and by an in vitro activity assay on cytosolic cell extracts. To assess changes in mitochondrial potential and reactive oxygen species, cells were stained by 3,3'-dihexyloxacarbocynine iodide or hydroethidine, respectively, and scored by flow cytometry.

RESULTS:

The observations establish that A-T cells are equipped with a proficient apoptotic machinery, as demonstrated by their ability to undergo mitochondrial collapse and caspase-3 activation after Fas activation or ceramide treatment. Both treatments have a similar cytotoxic effect on normal and A-T cells. In contrast, in spite of the stronger cytotoxicity induced by IR exposure, irradiated A-T cells are unable to undergo mitochondrial collapse and caspase-3 activation.

CONCLUSIONS:

The data indicate that ATM is necessary in the initiation of molecular pathway(s) leading to IR-induced apoptosis, and suggest that increased radiosensitivity of A-T cells is more likely a direct consequence of necrotic cell death.

PMID:
10902739
DOI:
10.1080/09553000050029002
[Indexed for MEDLINE]

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