Hypersensitivity of mtDNA-depleted cells to staurosporine-induced apoptosis: roles of Bcl-2 downregulation and cathepsin B.
We show that mitochondrial DNA (mtDNA)-depleted 143B cells are hypersensitive to staurosporine-induced cell death as evidenced by a more pronounced DNA fragmentation, a stronger activation of caspase-3, an enhanced poly(ADP-ribose) polymerase-1 (PARP-1) cleavage, and a more dramatic cytosolic release of cytochrome c. We also show that B-cell CLL/lymphoma-2 (Bcl-2), B-cell lymphoma extra large (Bcl-X(L)), and myeloid cell leukemia-1 (Mcl-1) are constitutively less abundant in mtDNA-depleted cells, that the inhibition of Bcl-2 and Bcl-X(L) can sensitize the parental cell line to staurosporine-induced apoptosis, and that overexpression of Bcl-2 or Bcl-X(L) can prevent the activation of caspase-3 in ρ(0)143B cells treated with staurosporine. Moreover, the inactivation of cathepsin B with CA074-Me significantly reduced cytochrome c release, caspase-3 activation, PARP-1 cleavage, and DNA fragmentation in mtDNA-depleted cells, whereas the pan-caspase inhibitor failed to completely prevent PARP-1 cleavage and DNA fragmentation in these cells, suggesting that caspase-independent mechanisms are responsible for cell death even if caspases are activated. Finally, we show that cathepsin B is released in the cytosol of ρ(0) cells in response to staurosporine, suggesting that the absence of mitochondrial activity leads to a facilitated permeabilization of lysosomal membranes in response to staurosporine.
Laboratory of Biochemistry and Cell Biology, Faculty of Sciences, University of Namur, 61 rue de Bruxelles, Namur, Belgium.
Guillaume
G
Rommelaere
Sébastien
S
Michel
Ludovic
L
Mercy
Antoine
A
Fattaccioli
Catherine
C
Demazy
Noelle
N
Ninane
Andrée
A
Houbion
Patricia
P
Renard
Thierry
T
Arnould
D017209:Apoptosis / Q000235:genetics*; D053148:Caspase 3 / Q000378:metabolism; D002401:Cathepsin B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D045744:Cell Line, Tumor; D045304:Cytochromes c / Q000378:metabolism; D053938:DNA Fragmentation; D004272:DNA, Mitochondrial / Q000235:genetics*; D004151:Dipeptides / Q000494:pharmacology; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D019311:Staurosporine / Q000494:pharmacology*; D051020:bcl-X Protein / Q000378:metabolism
C400541:CA 074 methyl ester; D004272:DNA, Mitochondrial; D004151:Dipeptides; D004791:Enzyme Inhibitors; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019253:Proto-Oncogene Proteins c-bcl-2; D051020:bcl-X Protein; D045304:Cytochromes c; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D053148:Caspase 3; D002401:Cathepsin B; D019311:Staurosporine