Sequential Operation of Ceramide Synthesis and ICE Cascade in CPT-11-Initiated Apoptotic Death Signaling

Suzuki, Atsushi; Iwasaki, Masaru; Kato, Michiyuki; Wagai, Nobuhiko

doi:10.1006/excr.1997.3498

Cited by 71 publications

(40 citation statements)

References 27 publications

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“…We also performed OLP loading procedure (Suzuki et al, 1997a) to further examine the direct e ect of ILP. When GST-ILP was loaded into HepG2 cells, the resistance to Fas-mediated apoptosis was encountered in the presence of actinomycin D. In contrast, GSTloaded HepG2 cells showed the Fas-mediated apoptosis in the presence of actinomycin D, same as intact HepG2 (Figure 4b).…”

Section: Caspase 3-inactivation By Ilpmentioning

confidence: 99%

“…To examine whether this complex formation is essential for the resistance to Fas-mediated apoptosis, we loaded GSTp21 into HepG2 cells with the OLP loading procedure (Suzuki et al, 1997a). When GST-p21 was loaded into HepG2 cells, the resistance to Fas-mediated apoptosis was encountered even in the presence of actinomycin D (Figure 6a and b).…”

Section: Procaspase 3/p21 Complex In Hepg2 Cellsmentioning

confidence: 99%

“…For the loading of GST-ILP or GST-p21 proteins into HepG2 cells, the OLP loading procedure (Williams and Henkart, 1994) was used for experiments with some modi®cations (Suzuki et al, 1997a). After the loading of GST-ILP or GST-p21 into HepG2 cells, cells were reacted with 1 mg/ml Fas Ab for 24 h in the presence of 0.5 mg/ml actinomycin D and death induction was examined by Hoechst 33342 staining analysis.…”

Section: Loading Of Gst-ilp or Gst-p21 Into Hepg2 Cellsmentioning

confidence: 99%

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Resistance to Fas-mediated apoptosis: activation of Caspase 3 is regulated by cell cycle regulator p21WAF1 and IAP gene family ILP

Suzuki¹,

Tsutomi²,

Akahane³

et al. 1998

Oncogene

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322

277

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The death receptor Fas transduces apoptotic death signaling mediated by caspases. In the present study, human hepatoma HepG2 cells showed the Fas-mediated apoptosis mediated by caspase, especially caspase 3, only in the presence of actinomycin D. Interestingly, cytosolic proteins extracted from intact HepG2 cells induced caspase 3 inactivation. Our results reveal that this inactivation was triggered by the direct inhibition of activated caspase 3 by IAP gene family ILP. In addition, a 53 kDa protein was co-immunoprecipitated with antihuman caspase 3 antibody from intact HepG2 cells. This protein was a complex-protein of procaspase 3 and the cell cycle regulator p21 WAF1 (p21). P21 bound to only procaspase 3, but not to activated caspase 3. We also demonstrate that p21 protein-loaded HepG2 cells resist to Fas-mediated apoptosis even in the presence of actinomycin D. Here we report that caspase 3 inactivation for the resistance to Fas-mediated apoptosis is induced by a procaspase 3/p21 complex formation and direct inhibition of activated caspase 3 by ILP.

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Section: Caspase 3-inactivation By Ilpmentioning

confidence: 99%

Section: Procaspase 3/p21 Complex In Hepg2 Cellsmentioning

confidence: 99%

Section: Loading Of Gst-ilp or Gst-p21 Into Hepg2 Cellsmentioning

confidence: 99%

See 1 more Smart Citation

Resistance to Fas-mediated apoptosis: activation of Caspase 3 is regulated by cell cycle regulator p21WAF1 and IAP gene family ILP

Suzuki¹,

Tsutomi²,

Akahane³

et al. 1998

Oncogene

Self Cite

322

277

View full text Add to dashboard Cite

show abstract

“…For the loading of GST-p21 proteins into HepG2 cells, the OLP loading procedure (Williams and Henkart, 1994) was used with some modi®cations (Suzuki et al, 1997a). After the loading of GST-p21 into HepG2 cells, cells were incubated with 1 mg/ml Fas Ab for 24 h in the presence of 0.5 mg/ml actinomycin D, and death induction was examined by Hoechst 33342 staining analysis.…”

Section: Loading Of Gst-p21 Into Hepg2 Cellsmentioning

confidence: 99%

Caspase 3 inactivation to suppress Fas-mediated apoptosis: identification of binding domain with p21 and ILP and inactivation machinery by p21

Suzuki¹,

Tsutomi²,

et al. 1999

Self Cite

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The death mediator caspase acts as the dominant regulator during cell death induction. The CPP32 subfamily, including caspase 3 (CPP32/Yama/Apopain), is essential for the cell death signaling. We recently reported that activation of caspase 3 is regulated by complex formation with p21 or ILP. In the present study, we investigated the binding domain with p21 and ILP to further characterize the caspase 3 inactivation machinery. Our results show that caspase 3 contains p21 binding domain in the N-terminus and ILP binding domain in the active site. Further, the caspase 3 binding domain in p21 was independent of the Cdk-or PCNAbinding domain. We also found caspase 3 protection by p21 from the p3-site cleavage serineproteinase contributes to the suppression machinery. Here, we propose the caspase 3 inactivation system by p21 and ILP as new essential system in the regulation of cell death.

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“…6,38 Hoechst 33342 and PI were purchased from Molecular Probes, Inc. (OR, USA). After treatment, cells were stained with the mixture of Hoechst 33342 and PI, and then observed by fluorescence microscopy.…”

Section: Assay Of Cell Viabilitymentioning

confidence: 99%

Procaspase 3/p21 complex formation to resist Fas-mediated cell death is initiated as a result of the phosphorylation of p21 by protein kinase A

et al. 2000

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Caspase 3 is an essential factor for Fas-mediated cell death and exists endogenously in cells where its activation is suppressed by p21 and ILP. Inside the cell, procaspase 3 interacts with p21 on mitochondria. In the present study, we investigated the molecular basis for procaspase 3/p21 complex formation. During Fas-mediated cell death, mitochondria are damaged, accompanied by decreased mitochondrial membrane-potential and decreased intracellular ATP levels. This mitochondrial damage occurs before an estrangement of the procaspase 3/p21 complex, and we demonstrate that intracellular ATP-deprivation also initiates an estrangement of procaspase 3/p21 complex formation and accelerates Fas-mediated cell death. In addition, our current results revealed that the phosphorylated p21 by PKA interacts with procaspase 3. Here, we report that the mitochondrial role, especially for ATP synthesis, and PKA are necessary for the procaspase 3/p21 complex formation to resist Fas-mediated cell death. Cell Death and Differentiation (2000) 7, 721 ± 728.

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Sequential Operation of Ceramide Synthesis and ICE Cascade in CPT-11-Initiated Apoptotic Death Signaling

Cited by 71 publications

References 27 publications

Resistance to Fas-mediated apoptosis: activation of Caspase 3 is regulated by cell cycle regulator p21WAF1 and IAP gene family ILP

Resistance to Fas-mediated apoptosis: activation of Caspase 3 is regulated by cell cycle regulator p21WAF1 and IAP gene family ILP

Caspase 3 inactivation to suppress Fas-mediated apoptosis: identification of binding domain with p21 and ILP and inactivation machinery by p21

Procaspase 3/p21 complex formation to resist Fas-mediated cell death is initiated as a result of the phosphorylation of p21 by protein kinase A

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