Identification of a caspase-9 substrate and detection of its cleavage in programmed cell death during mouse development.
The caspase family of proteases represents the main machinery by which apoptosis occurs. In vitro studies have revealed that upstream caspases are activated in response to apoptotic stimuli, and the active caspases in turn process downstream effector caspases that are involved in the destruction of cellular structure. Caspase-9 is an upstream caspase that can become active in response to cellular damage, including deprivation of growth factors and exposure to oxidative stress in vitro. Little is known, however, about how activation of caspase-9 is temporally and spatially regulated in vivo, e.g. during development. We have identified vimentin as the first example of a caspase-9 substrate that is not a downstream procaspase. Immunohistochemical analysis, using a specific antibody against the vimentin fragments generated by caspase-9, showed that caspase-9 cleaves vimentin in apoptotic cells in the embryonic nervous system and the interdigital regions. This result is consistent with observations that gene knockouts of caspase-9 and its activator, Apaf-1, result in developmental defects in these tissues. Our results show that the specific antibody is useful for in situ detection of caspase-9 activation in programmed cell death.
Nakanishi
K
K
Bioarchitect Research Group, RIKEN (Institute of Physical and Chemical Research), 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Maruyama
M
M
Shibata
T
T
Morishima
N
N
D000818:Animals; D017209:Apoptosis; D053453:Caspase 9; D020169:Caspases / Q000276:immunology / Q000378:metabolism*; D002460:Cell Line; D004789:Enzyme Activation; D006801:Humans; D020287:In Situ Nick-End Labeling; D051379:Mice; D013379:Substrate Specificity
C506433:CASP9 protein, human; C506434:Casp9 protein, mouse; D053453:Caspase 9; D020169:Caspases