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 Previous Article | Table of Contents | Next Article 
Blood, Vol. 93 No. 9 (May 1), 1999:
pp. 3053-3063
Anticancer Drugs Induce Caspase-8/FLICE Activation and Apoptosis in the
Absence of CD95 Receptor/Ligand Interaction
By
Sebastian Wesselborg,
Ingo H. Engels,
Evi Rossmann,
Marek Los, and
Klaus Schulze-Osthoff
From the Department of Internal Medicine I,
Eberhard-Karls-University, Tübingen, Germany.
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ABSTRACT |
Proteases of the caspase family are the critical executioners of
apoptosis. Their activation has been mainly studied upon triggering of
death receptors, such as CD95 (Fas/APO-1) and tumor necrosis factor-R1,
which recruit caspase-8/FLICE as the most proximal effector to the
receptor complex. Because apoptosis induced by anticancer drugs has
been proposed to involve CD95/CD95 ligand interaction, we investigated
the mechanism of caspase activation by daunorubicin, doxorubicin,
etoposide, and mitomycin C. In Jurkat leukemic T cells, all drugs
induced apoptosis and the cleavage of procaspase-8 to its active p18
subunit. However, cells resistant to CD95 were equally susceptible to
anticancer drugs and activated caspase-8 with a similar kinetic and
dose response as CD95-sensitive cells. The broad caspase inhibitor
benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone prevented apoptosis
and caspase-8 activation in response to CD95 and drug treatment,
whereas a neutralizing CD95 decoy as well as a dominant-negative FADD
construct selectively abrogated CD95, but not drug-induced effects. A
potent activation of caspase-8 was also induced by cycloheximide,
indicating that it was independent of protein synthesis. Our data,
therefore, show that (1) anticancer drug-induced apoptosis does not
require de novo synthesis of death ligands or CD95 interaction, and (2)
that caspase-8 can be activated in the absence of a death receptor signaling.
© 1999 by The American Society of Hematology.
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INTRODUCTION |
CHEMOTHERAPEUTIC AGENTS kill cancer cells
by multiple mechanisms, including intercalation into DNA, inhibition of
DNA replication, cell membrane damage, or free radical
generation.1,2 Although the primary intracellular targets
of drug action are rather distinct, it has become evident that
drug-induced cytotoxicity ultimately converges on a common pathway,
causing apoptosis. Cells exposed to anticancer drugs display apoptotic
alterations, such as cell shrinkage, chromatin condensation, and
internucleosomal DNA fragmentation.3 A close link between
apoptosis and the mechanism of drug action has been demonstrated by the
involvement of similar genetic components. Overexpression of Bcl-2
proteins can confer drug resistance in transfected tumor
cells.4-6 A number of investigations exposed a critical
role of the tumor suppressor p53 in apoptosis after drug
treatment.5,7,8 Finally, it has been recently shown that
drug-induced cytotoxicity involves proteases of the caspase family,
because specific inhibitors of caspases prevented cell death after
treatment with different anticancer agents.9-12
Caspases are currently considered as the central executioners of many,
if not all, apoptotic pathways. In mammalian cells, at least 12 different caspases exist, which are cysteine proteases that cleave
their substrates after aspartate residues.13-15 Caspases are synthesized as inactive proenzymes that are proteolytically processed to form an active tetrameric complex composed of two heterodimeric subunits of about 10 and 20 kD. The activation mechanism of caspases is currently unknown and is an area of intense research. Activation may proceed through autoproteolysis of the precursor, through mutual processing, or by still undefined processes. An increasing number of proteins have been found to be cleaved by caspases, and for some of them an apoptotic function has been proposed.13-16 Among different substrates are enzymes
involved in genome function, such as the DNA repair enzyme
poly(adenosine diphosphate-ribose)polymerase (PARP) and
DNA-dependent protein kinase (DNA-PK), or regulators of
the cell cycle, including retinoblastoma protein, the p53 regulator
MDM-2, MEKK, and protein kinase C- . Substrates of the nucleus and
cytoskeleton include lamins, Gas2, gelsolin, and fodrin. Furthermore,
it has been found that DNA cleavage is triggered upon caspase-mediated
degradation of the inhibitory subunit of a novel endonuclease,
designated caspase-activated DNase.17
It is currently unknown why there are so many different caspases in
mammalian cells. The most intensively studied member is caspase-3,
which is activated by multiple apoptotic stimuli. Depletion of
caspase-3 through homologous recombination results in excessive accumulation of neuronal cells caused by a lack of apoptosis in the
brain, whereas it has no affect in other tissues, indicating that
caspase-3 is redundant in many cell types.18 The
physiologic relevance of the other members of the caspase family is
also not well defined. Cells express more than one of these proteases, but whether all are functionally required for a single apoptotic pathway remains obscure. Current knowledge indicates that individual caspases have distinct substrate specificities, inhibitor profiles, and
abilities to process each other. These findings suggest that caspases
form a hierarchical network which, similar to the complement system,
may function as an amplifier for a given apoptotic stimulus.
One of the best-defined apoptotic pathways is mediated by the surface
receptor CD95 (APO-1/Fas).19-21 Triggering of the receptor by its natural ligand CD95L or agonistic antibodies induces the formation of a death-inducing signaling complex (DISC), which consists
of the adapter protein FADD and FLICE/caspase-8.22-24 Complex formation is initiated through homophilic interaction of the
death domains present in the intracellular part of both CD95 and FADD.
FADD, in addition, contains a second interacting region, called the
death effector domain (DED), which couples to caspase-8 as the most
proximal element in the caspase cascade. Further downstream in the
death pathway, caspase-8 presumably triggers the proteolytic activation
of other caspases and cleavage of cellular substrates.
Although it is evident that different anticancer drugs ultimately
mediate a common apoptotic pathway through the activation of caspases,
the events occuring between primary target action of the drugs and the
activation of apoptotic effectors are unclear. Recently, it has been
proposed that drug-induced apoptosis occurs through the CD95
pathway.25-27 It has been observed that several anticancer
drugs, such as doxorubicin, methotrexate, or bleomycin induce the
upregulation of membrane CD95 and induction of CD95L expression,
followed by the subsequent autocrine or paracrine induction of
CD95-dependent apoptosis. Cell lines resistant to CD95 were found to be
insensitive to anticancer drug-induced apoptosis. Furthermore,
drug-induced apoptosis was prevented by CD95 neutralizing antibodies.
However, there are also reports indicating that antitumor drugs may
induce apoptosis by other pathways.28-32
In the present study, we investigated the mechanism of anticancer
drug-induced apoptosis and the requirement of the CD95 system. We show
that different antineoplastic drugs, such as daunorubicin, doxorubicin,
etoposide, and mitomycin C induce caspase-dependent apoptosis to a
similar extent in both CD95-sensitive and resistant leukemic T cells.
Interestingly, caspase-8 was activated in both cell types by these
drugs and also by the protein synthesis inhibitor cycloheximide. Our
data indicate that (1) anticancer drugs induce caspase-8 activation and
apoptosis independent of CD95L/receptor interaction, and that (2)
caspase-8 can be activated in the absence of receptor signals even on
inhibition of protein synthesis. Thus, these findings show that
caspase-8, which has been previously regarded as the most proximal
caspase in CD95 signaling, can be activated independently of
death-receptor interaction.
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MATERIALS AND METHODS |
Cells and reagents.
The human leukemic T-cell lines Jurkat and CEM were maintained in
RPMI-1640 supplemented with 10% heat-inactivated fetal calf serum
(FCS), 100 U of penicillin per milliliter, 0.1 mg streptomycin per
milliliter, and 10 mmol/L HEPES (all from GIBCO-BRL, Eggenstein, Germany). Cells were grown at 37°C in a 5% CO2
atmosphere and maintained in log phase. The CD95-resistant Jurkat
subline, Jurkat-R, was generated by continuous culture in the presence
of anti-CD95 monoclonal antibody (MoAb) (IgG3, 1 µg/mL;
Cell Diagnostica, Münster, Germany) for 6 months. HeLa cells
stably transfected with a chimeric construct of a dominant-negative
FADD mutant or the vector alone fused to green-fluorescent protein
(GFP) were kindly provided by Dr P. Scheurich and Dr H. Wajant
(University of Stuttgart, Stuttgart, Germany)33 and
cultivated in RPMI-1640 supplemented with 5% FCS, 10 mmol/L HEPES and
antibiotics. The CD95-neutralizing chimeric protein glutathione-S
transferase (GST) CD95, consisting of the extracellular part of CD95
fused to GST, was a gift from Dr E. Gulbins (University of
Tübingen, Tübingen, Germany) and produced in
Escherichia coli. The broad-range caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (zVAD-fmk) was purchased from Enzyme Systems (Dublin, CA). The chemotherapeutic drugs
daunorubicin, etoposide, and mitomycin C were obtained from the
clinical pharmacy (Medical Clinics, Tübingen, Germany), and doxorubicin was purchased from Sigma (Deisenhofen, Germany).
Daunorubicin and mitomycin C were dissolved in methanol and doxorubicin
and etoposide in ethanol and kept as stock solutions at  70°C.
Measurement of cell death.
For determination of cell death, 3 × 104 cells per well
were seeded in microtiter plates and treated for 24 hours with the indicated concentrations of anti-CD95 or the chemotherapeutic agents.
The leakage of fragmented DNA from apoptotic nuclei was measured by the
method of Nicoletti et al.34 Briefly, apoptotic nuclei were
prepared by lysing cells in a hypotonic lysis buffer (1% sodium
citrate, 0.1% Triton X-100, 50 µg/mL propidium iodide) and
subsequently analyzed by flow cytometry. Nuclei to the left of the 2 N
peak containing hypodiploid DNA were considered apoptotic. Phosphatidylserine externalization of apoptotic cells was visualized by
staining with annexin-V-FITC after the manufacturer's protocol (Boehringer-Mannheim, Mannheim, Germany) and subsequent analysis in a
flow cytometer by using the FSC/FL1 profile. Cell death was assessed by
the uptake of propidium iodide (2 µg/mL; Sigma) in phosphate-buffered
saline (PBS) into nonfixed cells and subsequent flow cytometric
analysis with the FSC/FL2 profile. All flow cytometry analyses were
performed on a FACScalibur (Becton Dickinson, Heidelberg, Germany) by
using CellQuest analysis software. Microscopic analysis of cell
viability was performed with nonfixed GFP-FADD-DN expressing HeLa cells
in an inverse fluorescent microscope.
Immunoblotting.
Cleavage of PARP and caspase-8 was detected by immunoblotting. In
24-well plates 1 × 106 cells were seeded and treated with
the apoptotic stimuli. After the indicated periods, cells were washed
in cold PBS and lysed in 1% Triton X-100, 50 mmol/L Tris, pH 7.6, and
150 mmol/L NaCl containing 3 µg/mL aprotinin, 3 µg/mL leupeptin, 3 µg/mL pepstatin A, and 2 mmol/L phenylmethylsulfonyl fluoride.
Subsequently, proteins were separated under reducing conditions by 8%
to 15% gradient sodium dodecyl sulfate-polyacrylamide gel
electrophoresis and electroblotted to a polyvinylidene difluoride
membrane (Amersham, Braunschweig, Germany). The loading and transfer of
equal amounts of protein was confirmed by staining the membrane with
Ponceau S. Membranes were blocked for 1 hour with 5% nonfat dry milk
powder in Tris-buffered saline (TBS: 10 mmol/L Tris-HCl pH
7.4, 100 mmol/L NaCl) and then immunoblotted for 1 hour with rabbit
anti-PARP polyclonal antibody (1:2,000; Boehringer-Mannheim) or mouse
anti-caspase-8 MoAb (1:10 dilution of a hybridoma supernatant; Cell
Diagnostica). Membranes were washed four times with TBS/0.05% Tween-20
and incubated with peroxidase-conjugated affinity-purified rabbit
antimouse Ig for 1 hour. After extensive washing, the reaction was
developed by enhanced chemiluminescent staining by using ECL reagents (Amersham).
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RESULTS |
Anticancer drug-induced apoptosis is independent of functional
CD95/CD95L interaction.
The mechanism by which antitumor drugs induce apoptosis remains
controversial. It has been previously demonstrated that antineoplastic agents, such as doxorubicin, bleomycin, and methotrexate induce the
expression of CD95L and elicit cell death by subsequent CD95 interaction.25-27 To investigate whether the inducible
interaction of CD95L with its receptor is a prerequisite for
drug-mediated apoptosis, we used the subclone Jurkat-R that had been
selected for resistance to CD95 signaling. The dose response of
drug-induced apoptosis was assessed by flow cytometric staining of
hypodiploid DNA in apoptotic nuclei. When CD95-sensitive Jurkat and
CD95-resistant Jurkat-R cells were treated with mitomycin C, both cell
lines underwent apoptosis in a similar concentration-dependent manner (Fig 1A). When the CD95 pathway was
stimulated by using agonistic anti-CD95 antibodies, cell death was
induced up to a concentration of as low as 4 ng/mL in Jurkat cells,
whereas virtually no apoptosis was observed in the CD95-resistant cell
line Jurkat-R (Fig 1B). To investigate whether chemotherapeutic drugs
other than mitomycin C induce cell death independently of CD95, both
cell lines were stimulated with daunorubicin, doxorubicin, and
etoposide (Fig 2). Similar as in the
previous experiment, all drugs induced cell death in Jurkat and
Jurkat-R cells with almost the same dose dependency.
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| Fig 1.
Effects of mitomycin C and anti-CD95 in CD95-sensitive
and resistant Jurkat cells. 3 × 104 CD95-sensitive
(Jurkat;  ,  ) or resistant cells (Jurkat-R; -----,  ) were
stimulated with (A) mitomycin C or (B) agonistic anti-CD95 antibodies
at the indicated concentrations for 24 hours. Cells in the absence of
apoptotic stimuli were treated with the diluent of the respective
highest drug concentration. Induction of apoptosis was assessed by
propidium iodide staining of hypodiploid apoptotic nuclei and
subsequent flow cytometry.
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| Fig 2.
Induction of apoptosis mediated by daunorubicin,
doxorubicin, and etoposide is independent of CD95 receptor signaling.
Jurkat (, ) or Jurkat-R cells (-----, ) were stimulated with
(A) daunorubicin, (B) doxorubicin, and (C) etoposide at the indicated
concentrations. Assessment of hypodiploid apoptotic nuclei was
performed as described in Fig 1.
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To further exclude a possible participation of CD95 in drug-mediated
cell death, we preincubated cells with the neutralizing receptor decoy
construct GST-CD95, which prevents CD95L from receptor binding.
Induction of cell death was assessed in a flow cytometer by
annexin-V-FITC staining. As shown in Fig
3A, all antitumor agents induced apoptosis
in both Jurkat as well as in Jurkat-R cells, whereas
anti-CD95-triggered apoptosis occurred only in Jurkat cells. GST-CD95
inhibited anti-CD95-triggered cell death, whereas apoptosis elicited by
the anticancer drugs was virtually unaffected (Fig 3B). GST alone,
which was used as a negative control, had no effect on anti-CD95 nor on
drug-mediated cell death (Fig 3C). Similar data were obtained by
fluorescence-activated cell sorter analysis using propidium iodide
staining of dead cells and apoptotic nuclei (data not shown). Thus,
these results suggest that signaling through CD95 is not a prerequisite
for anticancer drug-mediated apoptosis.
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| Fig 3.
Inhibition of CD95 signaling does not affect antitumor
drug-mediated cell death. Jurkat ( ) or Jurkat-R ( ) cells
(3 × 104 per well) were preincubated with (A) medium,
(B) GST-CD95 (100 µg/mL), or (C) GST (100 µg/mL) for 1 hour, and
then treated with either anti-CD95 (20 ng/mL), daunorubicin (Dauno; 5 µg/mL), doxorubicin (Doxo; 1 µg/mL), etoposide (Etopo; 25 µg/mL),
or mitomycin C (Mito; 25 µg/mL). Apoptosis was assessed after 24 hours by measuring phosphatidylserine exposure by
annexin-V-FITC-staining and flow cytometry.
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Caspases are involved in induction of apoptosis by mitomycin C,
etoposide, daunorubicin, and doxorubicin.
Because caspases are involved in drug-mediated apoptosis, we
investigated whether caspases are also activated by anticancer drugs in
the absence of CD95/CD95L interaction. Therefore, we stimulated
CD95-sensitive and resistant cells under conditions analogous as in Fig
3 and used zVAD-fmk, a broad peptide inhibitor of caspases. Similarly,
as in the previous experiment, all cytotoxic agents induced cell death
in both Jurkat and Jurkat-R cells, whereas CD95 triggering only
resulted in apoptosis in Jurkat cells. Flow cytometric staining of
hypodiploid nuclei revealed that zVAD-fmk strongly attenuated anti-CD95
and drug-induced apoptosis (Fig 4A, upper
panel). Inhibition of caspases was also
able to prevent membrane damage and cell death as determined by
measuring the uptake of propidium iodide into cells (Fig 4A, lower
panel).
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| Fig 4.
Anticancer drugs mediate caspase-dependent apoptosis in
the absence of CD95 signaling. (A) CD95-susceptible Jurkat or
CD95-resistant Jurkat-R cells were pretreated with medium ( and  )
or 100 µmol/L zVAD-fmk () for 1 hour, and subsequently stimulated
with anti-CD95 (20 ng/mL), daunorubicin (Daun; 5 µg/mL), doxorubicin
(Doxo; 1 µg/mL), etoposide (Etop; 25 µg/mL), or mitomycin C (Mito;
25 µg/mL). After 24 hours, induction of apoptosis was measured by
propidium iodide staining of hypodiploid nuclei (upper panel) and cell
death was determined by the uptake of propidium iodide in dead cells
(lower panel). (B and C) stimulated were 1 × 106 Jurkat
or Jurkat-R cells for the indicated time with (B) daunorubicin (5 µg/mL) or with (C) different concentrations of etoposide (25 µg/mL
lanes 4,8; 12.5 µg/mL lanes 3,7; 6.25 µg/mL lanes 2,6 or diluent
control lanes 1,5) for 6 hours. Cellular proteins were resolved on a
8% to 15% sodium dodecyl sulfate-polyacrylamide gel electrophoresis
(SDS-PAGE) and caspase activity was detected by cleavage of the
caspase-specific substrate PARP by using immunoblot analysis. Filled
arrowheads ( ) indicate the uncleaved p116 and open arrowheads ( )
indicate the cleaved p89 form of PARP.
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We further investigated the involvement of caspases in drug-induced
cell death by measuring cleavage of a known caspase substrate in
immunoblot analyses. PARP, an enzyme involved in DNA repair, is cleaved
during apoptosis and has been shown to serve as a substrate for
caspase-3.35,36 Fig 4B shows that PARP, a 116-kD protein, was cleaved into the characteristic 89-kD fragment after 8 hours of
daunorubicin treatment in both CD95-sensitive and resistant Jurkat
cells (Fig 4B). Etoposide also induced PARP cleavage in a
dose-dependent manner in both cell lines (Fig 4C). Thus, these data
show that caspases are involved in drug-mediated apoptosis independently of CD95L/receptor interaction.
Caspase-8/FLICE is activated by anticancer drugs in the absence of
CD95L/receptor interaction.
During CD95-mediated apoptosis, the caspase cascade is initiated by the
recruitment and cleavage of caspase-8 at the DISC. Because anticancer
drugs obviously mediate activation of caspases independently of CD95,
we next investigated whether caspase-8 is activated during drug-induced
cell death. Caspase-8 is synthesized as two isoforms of about 55 kD
(caspase-8a and caspase-8b) which, after formation of intermediate
cleavage products of 43 and 41 kD, are processed to a p18 and p10
heterodimer.37,38 As assessed with an antibody directed
against the p18 subunit, CD95 ligation resulted in the cleavage of
procaspase-8 into its characteristic intermediate fragments and the
active p18 subunit (Fig 5A). Surprisingly, a similar cleavage pattern was obtained after treatment of Jurkat cells
with etoposide (Fig 5A), indicating that caspase-8 was also activated
by a receptor-independent apoptotic pathway.
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| Fig 5.
Chemotherapeutic agents induce processing of caspase-8 in
the absence of CD95 signaling. (A) Jurkat cells
(1 × 106) were treated with different concentrations
of either etoposide (50 µg/mL lane 4; 5 µg/mL lane 3; 0.5 µg/mL
lane 2; diluent control lane 1) for 6 hours or anti-CD95 (1,000 ng/mL
lane 8; 50 ng/mL lane 7; 2.5 ng/mL lane 6; diluent control lane 5) for
3 hours. Cellular proteins were separated by SDS-PAGE and processing of
procaspase-8 was detected by immunoblotting with anti-caspase-8. Open
arrowheads () indicate the two different isoforms of procaspase-8
(caspase-8/a and caspase-8/b) that are cleaved into the intermediate
forms p43 and p41 () and finally processed to the active p18 subunit
( ). Ig light chain of stimulatory anti-CD95 antibody is indicated
with an asterisk. (B) Jurkat or Jurkat-R cells
(1 × 106) were treated for the indicated time with
anti-CD95 (1 µg/mL), doxorubicin (Doxo; 1 µg/mL), or (C) with
daunorubicin (Dauno; 5 µg/mL), etoposide (Etopo; 20 µg/mL), or
mitomycin C (Mito; 25 µg/mL). Cellular proteins were immunoblotted
with anti-caspase-8 as described in (A). Only a section of the
immunoblots indicating the cleaved intermediate forms (p43 and p41) of
caspase-8a and caspase-8b is shown.
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A time course experiment revealed that after CD95 ligation of sensitive
Jurkat cells caspase-8 was readily processed into the p43/41
intermediate forms within 1 hour, whereas no processing was observed in
CD95-resistant Jurkat-R cells (Fig 5B). Doxorubicin and daunorubicin
induced caspase-8 cleavage in a similar kinetic after 8 to 12 hours in
both Jurkat and Jurkat-R cells (Figs 5B and C). In contrast, etoposide
and mitomycin C-induced caspase-8 activation occurred slightly faster.
After treatment with etoposide significant caspase-8 activation was
observed within 5 hours, whereas mitomycin C induced full processing
after 8 hours (Fig 5C). In addition, a dose-response experiment showed
that all anticancer drugs induced proteolytic activation of caspase-8
at concentrations that are achievable in the plasma of patients during
therapy (Fig 6).
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| Fig 6.
Dose-dependent cleavage of caspase-8 by mitomycin C,
daunorubicin, and doxorubicin. Jurkat and Jurkat-R cells
(1 × 106) were stimulated with mitomycin C (25 µg/mL lanes 4,8; 12.5 µg/mL lanes 3,7; 6.25 µg/mL lanes 2,6;
diluent control lanes 1,5; for 6 hours), daunorubicin (5 µg/mL lanes
4,8; 2.5 µg/mL lanes 3,7; 1.25 µg/mL lanes 2,6; diluent control
lanes 1,5; for 10 hours) or doxorubicin (2 µg/mL lanes 4,8; 1 µg/mL
lanes 3,7; 0.5 µg/mL lanes 2,6; diluent control lanes 1,5; for 10 hours). Whole cell lysates were immunoblotted with anti-caspase-8 as
described in Fig 5A. Filled arrowheads () indicate the
cleaved intermediate forms of caspase-8a and caspase-8b.
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Inhibition of CD95L/receptor interaction does not affect
drug-mediated caspase-8 activation.
We further analyzed the effect of the neutralizing GST-CD95 fusion
protein on caspase-8 activation after drug treatment. Whereas GST-CD95
potently blocked cleavage of caspase-8 in response to anti-CD95 (Fig
7A) or recombinant CD95L (data not shown),
it had virtually no effect on the processing of caspase-8 accomplished by etoposide or mitomycin C in CD95-sensitive and resistant Jurkat T-cells. GST, which was used as a control, did not affect the processing of caspase-8. In contrast to GST-CD95, the caspase inhibitor
zVAD-fmk blocked not only anti-CD95, but also etoposide and mitomycin
C-induced cleavage of caspase-8. Essentially, the same results were
obtained in leukemic CEM cells, in which both anti-CD95 and anticancer
drugs induced PARP cleavage and the processing of caspase-8 (Fig 7B).
Similar to Jurkat cells, only CD95 but not drug-mediated caspase
activation was inhibited by the neutralizing CD95 decoy construct.
Thus, these data clearly show that mechanisms of cell death and caspase
activation in response to anticancer drugs are not dependent on a
functional interaction of CD95L with its cognate receptor.
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| Fig 7.
Inhibition of CD95 signaling with GST-CD95 does not
inhibit antitumor drug-mediated cleavage of caspase-8 and PARP. (A)
Jurkat, Jurkat-R cells (1 × 106) or (B) CEM cells
were pretreated with medium, zVAD-fmk (100 µmol/L), GST-CD95 (100 µg/mL), or GST (100 µg/mL) for 1 hour, and then stimulated with
anti-CD95 (20 ng/mL; 3 hours), etoposide (Etopo; 25 µg/mL; 6 hours)
and mitomycin C (Mito; 25 µg/mL; 6 hours) or medium (6 hours).
Cellular proteins were immunoblotted with (A and B) anti-caspase-8 or
(B) anti-PARP as described in Fig 5A. The immunoblots indicate the
cleaved intermediate fragments () of caspase-8a and caspase-8b. The
anti-PARP immunoblot shows the uncleaved p116 and the cleaved p89 form
of PARP ().
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Drug-induced activation of caspase-8 and apoptosis is not dependent
on death receptor signaling.
Because caspase-8 is the most proximal caspase involved in
receptor-mediated apoptosis, the previous data implicated other mechanisms responsible for caspase-8 processing. Caspase-8 is not only
recruited to the DISC of CD95, but also to the receptors for tumor
necrosis factor (TNF) and TNF-related apoptosis-inducing ligand
(TRAIL). To further exclude a potential role of death receptors in
drug-mediated activation of caspase-8, we analyzed the effect of
anticancer drugs in HeLa cells, which stably expressed a GFP-tagged dominant-negative mutant of the adapter protein FADD lacking the essential DED region.33 It has been shown that FADD
transduces apoptotic signals triggered by CD95, TNF-R1 and
the TRAIL receptors.22,33,39-43 In accordance, HeLa-FADD-DN
cells are resistant to anti-CD95 and TRAIL and have an intrinsic
resistance to TNF-mediated apoptosis.33 Microscopic
analysis revealed that stimulation with anti-CD95 did not induce any
apoptotic alterations in HeLa-FADD-DN cells, thus confirming the
protective effect of the dominant-negative FADD-mutant (Fig
8A). In contrast, treatment with
staurosporine and mitomycin C caused the appearance of typical
apoptotic features such as cell condensation and membrane blebbing (Fig
8A). The parallel assessment of DNA fragmentation revealed that all
cytotoxic drugs induced apoptosis in both HeLa-FADD-DN and vector
control cells, whereas anti-CD95 induced apoptosis only in HeLa cells transfected with the vector alone (Fig 8B). In accordance,
staurosporine and mitomycin-C activated caspase-8 in both HeLa-vector
and HeLa-FADD-DN cells, whereas anti-CD95 triggered activation of
caspase-8 only in HeLa-vector cells (Fig 8C). These data indicate that
apoptosis by anticancer drugs does not require death receptor signaling and that caspase-8 can be activated in the absence of a FADD-containing receptor signaling complex.
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| Fig 8.
Drug-induced apoptosis and activation of caspase-8 is
independent of FADD signaling. (A and B) HeLa cells
(4 × 104) stably expressing a GFP-tagged
dominant-negative mutant of FADD (GFPFADD-DN)
or the GFP-tagged vector alone (GFPvector) were treated
with anti-CD95 (1 µg/mL), staurosporine (2.5 µmol/L), etoposide (25 µg/mL), mitomycin C (25 µg/mL) or left untreated. (A) After 24 hours, HeLa-FADD-DN cells were analyzed for the induction of apoptosis
under a fluorescent microscope. (B) Subsequently, HeLa-FADD-DN and
HeLa-vector cells were lysed in a hypotonic buffer, and the number of
hypodiploid apoptotic nuclei was determined in a flow cytometer. Mean
values of ± SD from triplicate experiments are shown. (C)
HeLa-FADD-DN (1.5 × 106) or HeLa-vector cells were
incubated with medium, anti-CD95 (1 µg/mL), staurosporine (2.5 µmol/L) or mitomycin C (25 µg/mL) for the indicated time. Cellular
proteins were immunoblotted with anti-caspase-8 as described in Fig 5A.
The intermediate cleavage forms (p43 and p41) of caspase-8a and
caspase-8b are shown.
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Inhibition of protein synthesis by cycloheximide activates caspase-8.
Because the family of death receptors is steadily growing, it could not
be excluded that the activation of caspase-8 by antitumor drugs is
mediated by yet unknown death receptors that transduce their signals
independently of FADD. Therefore, we asked whether de novo synthesis of
other death ligands, such as TNF or TRAIL, is involved in drug-induced
caspase-8 activation. To this end, cells were treated with
cycloheximide, an inhibitor of protein synthesis. In some cell types,
the sole inhibition of translation by cycloheximide induces apoptosis.
This apoptosis is independent on CD95 signaling and occurred to a
similar extent in both Jurkat and Jurkat-R cells (Fig
9A). Interestingly, treatment with
cycloheximide also induced processing of caspase-8 in the absence of
any additional stimuli in CD95 sensitive and resistant cells (Fig 9B).
These data finally exclude that inducible expression of CD95L or other death ligands is an indispensable prerequisite for the activation of
caspase-8 and subsequent apoptosis.
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| Fig 9.
Inhibition of protein synthesis by cycloheximide induces
apoptosis and processing of caspase-8. (A) Jurkat
(3 × 104) () or Jurkat-R cells () were treated
with the indicated concentrations of cycloheximide for 24 hours.
Assessment of apoptotic nuclei was accomplished by flow cytometry of
propidium iodide-stained hypodiploid nuclei. (B) Jurkat or Jurkat-R
cells (1 × 106) were incubated with cycloheximide
(lanes 4,8: 10 µg/mL; lanes 3,7: 2 µg/mL; lanes 2,6: 0.4 µg/mL;
lanes 1,5: diluent control) for 5 hours. Total cell lysates were
immunoblotted with anti-caspase-8 antibody as described in Fig 5A.
Filled arrowheads () indicate the cleaved intermediate forms of 43 kD and 41 kD of caspase-8a and caspase-8b.
|
|
| |
DISCUSSION |
In the present study, we investigated the mechanism of cell death
induced by a number of anticancer agents. We show that daunorubicin, doxorubicin, mitomycin C as well as etoposide induce caspase-dependent apoptosis in drug-sensitive target cells. This suggests that
drug-induced cytotoxicity is mediated by mechanisms that take place
downstream of primary intracellular target action. These data are in
line with previous results by us and others showing that effector
caspases, such as caspase-3, -6, and -7 are critically involved in
anticancer drug-mediated cell death.9-12
For the first time, it is shown that antineoplastic agents used in
conventional chemotherapy are also able to induce activation of
caspase-8. Previously, the role of caspase-8 appeared to be restricted
to apoptosis mediated by death receptors, such as CD95, TNF-R1, and the
TRAIL receptors. In the death receptor pathway, caspase-8 is a crucial
component of the DISC, in which it is recruited through its DEDs that
interact with receptor-associated FADD.22-24,44 Because
caspase-8, with the exception of caspase-10, is the only DED-containing
protease, it is assumed to act as a proximal initiator caspase, which
subsequently processes downstream effector caspases.
The activation of caspase-8 by anticancer drugs, as shown in this
study, could be explained by the notion that drug-induced cytotoxicity
might be mediated by a death receptor-dependent mechanism. Accordingly,
it was recently proposed that anticancer drugs induce the upregulation
of CD95L by ceramide-mediated mechanism, which then induces subsequent
CD95-dependent apoptosis in drug-sensitive target
cells.25-27,45 However, our present data convincingly show that this pathway may not be a principle and necessary mechanism of
anticancer drug action. This assumption is supported by a number of
independent evidences.
First, we show that anticancer drugs induce apoptosis in both
CD95-sensitive and resistant Jurkat cells with a similar kinetic and
dose dependency. Second, although both anticancer drug- and CD95-induced apoptosis was prevented by a broad-range caspase inhibitor, only CD95-mediated cell death was inhibitable by a neutralizing CD95 receptor decoy construct in leukemic Jurkat and CEM
cells. Third, the occurrence of drug-induced apoptosis in HeLa cells
stably transfected with a dominant-negative FADD mutant excludes the
possibility that other FADD-dependent death receptors are involved in
anticancer drug-mediated cell death. Finally, we show that caspase-8
activation and subsequent apoptosis can also be induced by
cycloheximide, an inhibitor of protein synthesis. Although this latter
finding does not exclude that the release of a preformed death ligand
may be involved, it suggests that inducible synthesis of CD95L or other
ligands is not a prerequisite for drug-induced caspase-8 activation and apoptosis.
The mechanism of caspase-8 processing and activation independently of
CD95 is intriguing. Besides the death receptor/FADD pathway, it has
become recently clear that a second, either independent or
interconnected pathway exists, which is essentially controlled by the
release of mitochondrial components and inhibited by antiapoptotic members of the Bcl-2 family.46-51 An early event in this
process is the redistribution of cytochrome c into the cytosol, which can be triggered by a variety of apoptotic conditions such as death
receptors, ceramide, and anticancer drugs.48,49,51-53 In the cytosol, cytochrome c interacts, together with dATP, with apoptotic
protease-activating factor-1 (Apaf-1), the mammalian homologue of the
Caenorhabditis elegans cell death regulator
Ced-4.54 Binding of these components then exposes an
interaction motif in Apaf-1, which serves as a so-called caspase
recruitment domain (CARD) by binding to caspases that have a similar
CARD motif at their N-terminus. A CARD motif has been identified in
caspase-1, -2, and -9.55 Caspase-8 and the structurally
related caspase-10 also contain a long prodomain that may exert a
similar regulatory function. In this respect, it has been found that
caspase-8 can interact with Ced-4 in cell-free systems.56
Therefore, it is possible that caspase-8 is either directly recruited
to Apaf-1 or activated by an upstream caspase of this cascade. It will
be interesting to investigate whether caspase-8 is activated by direct interaction with Apaf-1 or further downstream by another protease such
as caspase-9.
There are also other evidences supporting the idea that anticancer
drug-induced and CD95-induced apoptosis involve distinct proximal
effector pathways. It has been observed that Bcl-2 and Bcl-XL, which exert their antiapoptotic function at
mitochondria, could potently inhibit anticancer drug-induced cell
death, whereas CD95-mediated or TNF-R1-mediated apoptosis was not
affected.11,57,58 In addition, recent gene targeting showed
that FADD-deficient fibroblasts became resistant to apoptosis mediated
by several death receptors, although these cells remained sensitive to
anticancer drug treatment.59 In line with this, we found
that expression of a dominant/negative FADD construct in HeLa cells
interfered with CD95-drug induced but not anticancer-drug induced
apoptosis. This further suggests that a FADD-independent pathway of
drug-induced apoptosis is not restricted to Jurkat T cells.
Overall, understanding the mechanisms of anticancer drug-induced
apoptosis is of principal importance for developing effective strategies in tumor therapy. It has been suggested that CD95L is
responsible for maintaining the immunoprivilege of some
organs.60,61 Although the role of CD95 in immunoprivilege
is controversially discussed,62-64 this would mean that
drug-triggered CD95L expression would be consequently associated with a
loss of immunosurveillance induced by drug-resistant tumor cells
bearing CD95L. However, the present data clearly show that apoptosis
mediated by anticancer drugs does not necessarily require a functional
CD95 system. Although anticancer drugs and CD95 ligation converge at a
common downstream pathway, both pathways can act independently to
induce apoptosis in sensitive target cells.
| |
ACKNOWLEDGMENT |
We thank Dr E. Gulbins for kindly providing us with the GST-CD95
construct and Dr P. Scheurich and Dr H. Wajant for HeLa cells stably
transfected with GFP-FADD-DN.
| |
FOOTNOTES |
Submitted June 18, 1998; accepted December 28, 1998.
Supported in part by grants from the Deutsche Forschungsgemeinschaft
(SFB 364/A7) and the European Union Biomed-2 program. S.W. acknowledges
a fellowship from the Bundesministerium für Bildung und Forschung.
The publication costs of this
article were defrayed in part by
page charge payment. This article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. section
1734 solely to indicate this fact.
Address reprint requests to Klaus Schulze-Osthoff, PhD,
Department of Internal Medicine I, Eberhard-Karls-University,
Otfried-Müller-Str. 10, D-72076 Tübingen, Germany; e-mail:
schulze-osthoff{at}uni-tuebingen.de.
| |
REFERENCES |
1.
Barry MA, Behnke CA, Eastman A:
Activation of programmed cell death (apoptosis) by cisplatin, other anticancer drugs, toxin and hyperthermia.
Biochem Pharmacol
40:2353, 1990[Medline]
[Order article via Infotrieve]
2.
Hannun YA:
Apoptosis and the dilemma of cancer chemotherapy.
Blood
89:1845, 1997[Free Full Text]
3.
Fisher DE:
Apoptosis in cancer therapy: Crossing the threshold.
Cell
78:539, 1994[Medline]
[Order article via Infotrieve]
4.
Miyashita T, Reed JC:
Bcl-2 oncoprotein blocks chemotherapy-induced apoptosis in a human leukemia cell line.
Blood
81:151, 1993[Abstract/Free Full Text]
5.
Lotem J, Sachs L:
Regulation by bcl-2, c-myc, and p53 of susceptibility to induction of apoptosis by heat shock and cancer chemotherapy compounds in differentiation-competent and a defective myeloid leukemic cells.
Cell Growth Differ
4:41, 1993[Abstract]
6.
Yang E, Korsmeyer SJ:
Molecular thanatopsis: A discourse on the BCL2 family and cell death.
Blood
88:386, 1996[Free Full Text]
7.
Lowe SW, Ruley HE, Jacks T, Housman DE:
p53-dependent apoptosis modulates the cytotoxicity of anticancer agents.
Cell
74:957, 1993[Medline]
[Order article via Infotrieve]
8.
Strasser A, Harris AW, Jacks T, Cory S:
DNA damage can induce apoptosis in proliferating lymphoid cells via p53-independent mechanisms inhibitable by Bcl-2.
Cell
79:329, 1994[Medline]
[Order article via Infotrieve]
9.
Zhu H, Fearnhead HO, Cohen GM:
An ICE-like protease is a common mediator of apoptosis induced by diverse stimuli in human monocytic THP.1 cells.
FEBS Lett
374:303, 1995[Medline]
[Order article via Infotrieve]
10.
Chen Z, Naito M, Mashima T, Tsuruo T:
Activation of actin-cleavable interleukin 1 -converting enzyme (ICE) family protease CPP-32 during chemotherapeutic agent-induced apoptosis in ovarian carcinoma cells.
Cancer Res
56:5224, 1996[Abstract/Free Full Text]
11.
Datta R, Banach D, Kojima H, Talanian RV, Alnemri ES, Wong WW, Kufe DW:
Activation of the CPP32 protease in apoptosis induced by 1--D-arabinofuranosylcytosine and other DNA-damaging agents.
Blood
88:1936, 1996[Abstract/Free Full Text]
12.
Los M, Herr I, Friesen C, Fulda S, Schulze-Osthoff K, Debatin KM:
Cross-resistance of CD95- and drug-induced apoptosis as a consequence of deficient activation of caspases (ICE/Ced-3 proteases).
Blood
90:3118, 1997[Abstract/Free Full Text]
13.
Cohen GM:
Caspases: The executioners of apoptosis.
Biochem J
326:1, 1997
14.
Nicholson DN, Thornberry NA:
Caspases: Killer proteases.
Trends Biochem Sci
22:299, 1997[Medline]
[Order article via Infotrieve]
15.
Cryns V, Yuan J:
Proteases to die for.
Genes Dev
12:1551, 1998[Free Full Text]
16.
Stroh C, Schulze-Osthoff K:
Death by a thousand cuts: an ever increasing list of caspase substrates.
Cell Death Differ
5:997, 1998[Medline]
[Order article via Infotrieve]
17.
Enari M, Sakahiera H, Yokoyama H, Okawa K, Iwamatsu A, Nagata S:
A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD.
Nature
391:43, 1998[Medline]
[Order article via Infotrieve]
18.
Kuida K, Zheng TS, Na S, Kuan C, Yang D, Karasuyama H, Rakic P, Flavell RA:
Decreased apoptosis in the brain and premature lethality in CPP32-deficient mice.
Nature
384:368, 1996[Medline]
[Order article via Infotrieve]
19.
Krammer PH, Dhein J, Walczak H, Behrmann I, Mariani S, Matiba B, Fath M, Daniel PT, Knipping E, Westendorp MO, Stricker K, Baumler C, Hellbrandt S, Germer M, Peter ME, Debatin KM:
The role of APO-1-mediated apoptosis in the immune system.
Immunol Rev
142:175, 1994[Medline]
[Order article via Infotrieve]
20.
Schulze-Osthoff K, Ferrari D, Los M, Wesselborg S, Peter ME:
Apoptosis signaling by death receptors.
Eur J Biochem
254:439, 1998[Medline]
[Order article via Infotrieve]
21.
Nagata S:
Apoptosis by death factor.
Cell
88:355, 1997[Medline]
[Order article via Infotrieve]
22.
Boldin MP, Goncharov TM, Goltsev YV, Wallach D:
Involvement of MACH, a novel MORT1/FADD-interacting protease, in Fas/APO-1- and TNF receptor-induced cell death.
Cell
85:803, 1996[Medline]
[Order article via Infotrieve]
23.
Kischkel FC, Hellbardt S, Behrmann I, Germer M, Pawlita M, Krammer PH, Peter ME:
Cytotoxicity-dependent APO-1 (Fas/CD95)-associated proteins form a death-inducing signaling complex (DISC) with the receptor.
EMBO J
14:5579, 1995[Medline]
[Order article via Infotrieve]
24.
Muzio M, Chinnaiyan AM, Kischkel FC, O'Rourke K, Shevchenko A, Ni J, Scaffidi C, Bretz JD, Zhang M, Gentz R, Mann M, Krammer PH, Peter ME, Dixit VM:
FLICE, a novel FADD-homologous ICE/CED-3-like protease, is recruited to the CD95 (Fas/APO-1) death-inducing signaling complex.
Cell
85:817, 1996[Medline]
[Order article via Infotrieve]
25.
Friesen C, Herr I, Krammer PH, Debatin KM:
Involvement of the CD95 (APO-1/FAS) receptor/ligand system in drug-induced apoptosis in leukemia cells.
Nat Med
2:574, 1996[Medline]
[Order article via Infotrieve]
26.
Müller M, Strand S, Hug H, Heinemann E-M, Walczak H, Hofmann WJ, Stremmel W, Krammer PH, Galle PR:
Drug-induced apoptosis in hepatoma cells is mediated by the CD95 (Apo-1/Fas) receptor/ligand system and involves activation of wild-type p53.
J Clin Invest
99:403, 1997[Medline]
[Order article via Infotrieve]
27.
Fulda S, Sieverts H, Friesen C, Herr I, Debatin KM:
The CD95 (APO-1/Fas) system mediates drug-induced apoptosis in neuroblastoma cells.
Cancer Res
57:3823, 1997[Abstract/Free Full Text]
28.
Eischen CM, Kottke TJ, Martins LM, Basi GS, Tung JS, Earnshaw WC, Leibson PJ, Kaufmann SH:
Comparison of apoptosis in wild-type and Fas-resistant cells: Chemotherapy-induced apoptosis is not dependent on Fas/Fas ligand interactions.
Blood
90:935, 1997[Abstract/Free Full Text]
29.
Gamen S, Anel A, Lasierra P, Alava MA, Martinez Lorenzo MJ, Pineiro A, Naval J:
Doxorubicin-induced apoptosis in human T-cell leukemia is mediated by caspase-3 activation in a Fas-independent way.
FEBS Lett
417:360, 1997[Medline]
[Order article via Infotrieve]
30.
Villunger A, Egle A, Kos M, Hartmann BL, Geley S, Kofler R, Greil R:
Drug-induced apoptosis is associated with enhanced Fas (Apo-1/CD95) ligand expression but occurs independently of Fas (Apo-1/CD95) signaling in human T-acute lymphatic leukemia cells.
Cancer Res
57:3331, 1997[Abstract/Free Full Text]
31.
Fulda S, Friesen C, Los M, Scaffidi C, Mier W, Benedict M, Nunez G, Krammer PH, Peter ME, Debatin KM:
Betulinic acid triggers CD95 (APO-1/Fas)- and p53-independent apoptosis via activation of caspases in neuroectodermal tumors.
Cancer Res
57:4956, 1997[Abstract/Free Full Text]
32.
Tolomeo M, Dusonchet L, Meli M, Grimaudo S, D'Alessandro N, Papoff G, Ruberti G, Rausa L:
The CD95/CD95 ligand system is not the major effector in anticancer drug-mediated apoptosis.
Cell Death Differ
5:735, 1998[Medline]
[Order article via Infotrieve]
33.
Wajant H, Johannes FJ, Haas E, Siemienski K, Schwenzer R, Schubert G, Weiss T, Grell M, Scheurich P:
Dominant-negative FADD inhibits TNFR60-, Fas/Apo1- and TRAIL-R/Apo2-mediated cell death but not gene induction.
Curr Biol
8:113, 1998[Medline]
[Order article via Infotrieve]
34.
Nicoletti I, Migliorati G, Pagliacci MC, Grignani F, Riccardi C:
A rapid and simple method for measuring thymocyte apoptosis by propidium iodide staining and flow cytometry.
J Immunol Methods
139:271, 1991[Medline]
[Order article via Infotrieve]
35.
Kaufmann SH, Desnoyers S, Ottaviano Y, Davidson NE, Poirier GG:
Specific proteolytic cleavage of poly(ADP-ribose) polymerase: An early marker of chemotherapy-induced apoptosis.
Cancer Res
53:3976, 1993[Abstract/Free Full Text]
36.
Tewari M, Quan LT, O'Rourke K, Desnoyers S, Zeng Z, Beidler DR, Poirier GG, Salvesen GS, Dixit VM:
Yama/CPP32, a mammalian homolog of CED-3, is a CrmA-inhibitable protease that cleaves the death substrate poly(ADP-ribose) polymerase.
Cell
81:801, 1995[Medline]
[Order article via Infotrieve]
37.
Medema JP, Scaffidi C, Kischkel FC, Shevchenko A, Mann M, Krammer PH, Peter ME:
FLICE is activated by association with the CD95 death-inducing signaling complex (DISC).
EMBO J
16:2794, 1997[Medline]
[Order article via Infotrieve]
38.
Scaffidi C, Medema JP, Krammer PH, Peter ME:
FLICE is predominantly expressed as two functionally active isoforms, caspase-8/a and caspase-8/b.
J Biol Chem
272:26953, 1997[Abstract/Free Full Text]
39.
Chinnaiyan AM, O'Rourke K, Yu GL, Lyons RH, Garg M, Duan DR, Xing L, Gentz R, Ni J, Dixit VM:
Signal transduction by DR3, a death domain-containing receptor related to TNFR-1 and CD95.
Science
274:990, 1996[Abstract/Free Full Text]
40.
Pan G, O'Rourke K, Chinnaiyan AM, Gentz R, Ebner R, Ni J, Dixit VM:
The receptor for the cytotoxic ligand TRAIL.
Science
276:111, 1997[Abstract/Free Full Text]
41.
Walczak H, Degli Esposti MA, Johnson RS, Smolak PJ, Waugh JY, Boiani N, Timour MS, Gerhart MJ, Schooley KA, Smith CA, Goodwin RG, Rauch CT:
TRAIL-R2: A novel apoptosis-mediating receptor for TRAIL.
EMBO J
16:5386, 1997[Medline]
[Order article via Infotrieve]
42.
Chaudhary PM, Eby M, Jasmin A, Bookwalter A, Murray J, Hood L:
Death receptor 5, a new member of the TNFR family, and DR4 induce FADD-dependent apoptosis and activate the NF- B pathway.
Immunity
7:821, 1997[Medline]
[Order article via Infotrieve]
43.
Schneider P, Thome M, Burs K, Bodmer JL, Hofman K, Kataoka T, Holler N, Tschopp J:
TRAIL receptors 1 (DR4) and 2 (DR5) signal FADD-dependent apoptosis and activate NF-B.
Immunity
7:831, 1997[Medline]
[Order article via Infotrieve]
44.
Pan G, Ni J, Wei YF, Yu G, Gentz R, Dixit VM:
An antagonist decoy receptor and a death domain-containing receptor for TRAIL.
Science
277:815, 1997[Abstract/Free Full Text]
45.
Herr I, Wilhelm D, Bohler T, Angel P, Debatin KM:
Activation of CD95 (APO-1/Fas) signaling by ceramide mediates cancer therapy-induced apoptosis.
EMBO J
16:6200, 1998[Medline]
[Order article via Infotrieve]
46.
Strasser A, Harris AW, Huang DC, Krammer PH, Cory S:
Bcl-2 and Fas/APO-1 regulate distinct pathways to lymphocyte apoptosis.
EMBO J
14:6136, 1995[Medline]
[Order article via Infotrieve]
47.
Smith KG, Strasser A, Vaux DL:
CrmA expression in T lymphocytes of transgenic mice inhibits CD95 (Fas/APO-1)-transduced apoptosis, but does not cause lymphadenopathy or autoimmune disease.
EMBO J
15:5167, 1996[Medline]
[Order article via Infotrieve]
48.
Kluck RM, Bossy Wetzel E, Green DR, Newmeyer DD:
The release of cytochrome c from mitochondria: a primary site for Bcl-2 regulation of apoptosis.
Science
275:1132, 1997[Abstract/Free Full Text]
49.
Yang J, Liu X, Bhalla K, Kim CN, Ibrado AM, Cai J, Peng TI, Jones DP, Wang X:
Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked.
Science
275:1129, 1997[Abstract/Free Full Text]
50.
Newton K, Harris AW, Bath ML, Smith KGC, Strasser A:
A dominant interfering mutant of FADD/MORT1 enhances deletion of autoreactive thymocytes and inhibits proliferation of mature T lymphocytes.
EMBO J
17:706, 1998[Medline]
[Order article via Infotrieve]
51.
Scaffidi C, Fulda S, Srinivasan A, Friesen C, Li F, Tomaselli KJ, Debatin K-M, Krammer P, Peter ME:
Two CD95 (Apo-1/Fas) signaling pathways.
EMBO J
17:1675, 1998[Medline]
[Order article via Infotrieve]
52.
Adachi S, Cross AR, Babior BM, Gottlieb RA:
Bcl-2 and the outer mitochondrial membrane in the inactivation of cytochrome c during Fas-mediated apoptosis.
J Biol Chem
272:21878, 1997[Abstract/Free Full Text]
53.
Kharbanda S, Pandey P, Schofield L, Israels S, Roncinske R, Yoshida K, Bharti A, Yuan Z-M, Saxena S, Weichselbaum R, Nalin C, Kufe D:
Role of Bcl-xL as an inhibitor of cytosolic cytochrome C accumulation in DNA damage-induced apoptosis.
Proc Natl Acad Sci USA
94:6939, 1997[Abstract/Free Full Text]
54.
Zou H, Henzel WJ, Liu X, Lutschg A, Wang X:
Apaf-1, a human protein, homologous to C. elegans Ced-4, participates in cytochrome c-dependent activation of caspase-3.
Cell
90:405, 1997[Medline]
[Order article via Infotrieve]
55.
Hofmann K, Bucher P, Tschopp J:
The CARD domain: a new apoptotic signalling motif.
Trends Biochem Sci
22:155, 1997[Medline]
[Order article via Infotrieve]
56.
Chinnaiyan AM, O'Rourke K, Lane BR, Dixit VM:
Interaction of CED-4 with CED-3 and CED-9: A molecular framework for cell death.
Science
275:1122, 1997[Abstract/Free Full Text]
57.
Moreno MB, Memon SA, Zacharchuk CM:
Apoptosis signaling pathways in normal T cells: Differential activity of Bcl-2 and IL-1-converting enzyme family protease inhibitors on glucocorticoid- and Fas-mediated cytotoxicity.
J Immunol
157:3845, 1996[Abstract]
58.
Erhardt P, Cooper GM:
Activation of the CPP32 apoptotic protease by distinct signaling pathways with differential sensitivity to Bcl-xL.
J Biol Chem
271:17601, 1996[Abstract/Free Full Text]
59.
Yeh WC, Pompa JL, McCurrach ME, Shu HB, Elia AJ, Shahinian A, Ng M, Wakeham A, Khoo W, Mitchell K, El-Deiry WS, Lowe SW, Goeddel DV, Mak TW:
FADD: Essential for embryo development and signaling from some, but not all, inducers of apoptosis.
Science
279:1954, 1998[Abstract/Free Full Text]
60.
Bellgrau D, Gold D, Selawry H, Moore J, Franzusoff A, Duke RC:
A role for CD95 ligand in preventing graft rejection.
Nature
377:630, 1995[Medline]
[Order article via Infotrieve]
61.
Griffith TS, Brunner T, Fletcher SM, Green DR, Ferguson TA:
Fas ligand-induced apoptosis as a mechanism of immune privilege.
Science
270:1189, 1995[Abstract/Free Full Text]
62.
Seino K, Kayagaki N, Okumura K, Yagita H:
Antitumor effect of locally produced CD95 ligand.
Nat Med
3:165, 1997[Medline]
[Order article via Infotrieve]
63.
Allison J, Georgiou HM, Strasser A, Vaux DL:
Transgenic expression of CD95 ligand on islet -cells induces a granulocytic infiltration but does not confer immune privilege upon islet allografts.
Proc Natl Acad Sci USA
94:3943, 1997[Abstract/Free Full Text]
64.
Kang SM, Schneider DB, Lin Z, Hanahan D, Dichek DA, Stock OG, Baekkeskov S:
Fas ligand expression in islets of Langerhans does not confer immune privilege and instead targets them for rapid destruction.
Nat Med
3:738, 1997[Medline]
[Order article via Infotrieve]
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|
S. Bergeron, M. Beauchemin, and R. Bertrand
Camptothecin- and etoposide-induced apoptosis in human leukemia cells is independent of cell death receptor-3 and -4 aggregation but accelerates tumor necrosis factor-related apoptosis-inducing ligand-mediated cell death
Mol. Cancer Ther.,
December 1, 2004;
3(12):
1659 - 1669.
[Abstract]
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D. Chandra, G. Choy, X. Deng, B. Bhatia, P. Daniel, and D. G. Tang
Association of Active Caspase 8 with the Mitochondrial Membrane during Apoptosis: Potential Roles in Cleaving BAP31 and Caspase 3 and Mediating Mitochondrion-Endoplasmic Reticulum Cross Talk in Etoposide-Induced Cell Death
Mol. Cell. Biol.,
August 1, 2004;
24(15):
6592 - 6607.
[Abstract]
[Full Text]
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S. Yang and F. G. Haluska
Treatment of Melanoma with 5-Fluorouracil or Dacarbazine In Vitro Sensitizes Cells to Antigen-Specific CTL Lysis through Perforin/Granzyme- and Fas-Mediated Pathways
J. Immunol.,
April 1, 2004;
172(7):
4599 - 4608.
[Abstract]
[Full Text]
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P. Pandiyan, D. Gartner, O. Soezeri, A. Radbruch, K. Schulze-Osthoff, and M. C. Brunner-Weinzierl
CD152 (CTLA-4) Determines the Unequal Resistance of Th1 and Th2 Cells against Activation-induced Cell Death by a Mechanism Requiring PI3 Kinase Function
J. Exp. Med.,
March 15, 2004;
199(6):
831 - 842.
[Abstract]
[Full Text]
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X. D. Zhang, S. K. Gillespie, and P. Hersey
Staurosporine induces apoptosis of melanoma by both caspase-dependent and -independent apoptotic pathways
Mol. Cancer Ther.,
February 1, 2004;
3(2):
187 - 197.
[Abstract]
[Full Text]
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C. Pingoud-Meier, D. Lang, A. J. Janss, L. B. Rorke, P. C. Phillips, T. Shalaby, and M. A. Grotzer
Loss of Caspase-8 Protein Expression Correlates with Unfavorable Survival Outcome in Childhood Medulloblastoma
Clin. Cancer Res.,
December 15, 2003;
9(17):
6401 - 6409.
[Abstract]
[Full Text]
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J. Geller, I. Petak, K. S. Szucs, K. Nagy, D. M. Tillman, and J. A. Houghton
Interferon-{gamma}-Induced Sensitization of Colon Carcinomas to ZD9331 Targets Caspases, Downstream of Fas, Independent of Mitochondrial Signaling and the Inhibitor of Apoptosis Survivin
Clin. Cancer Res.,
December 15, 2003;
9(17):
6504 - 6515.
[Abstract]
[Full Text]
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S. F. Schlosser, M. Schuler, C. P. Berg, K. Lauber, K. Schulze-Osthoff, F. W. Schmahl, and S. Wesselborg
Ribavirin and Alpha Interferon Enhance Death Receptor-Mediated Apoptosis and Caspase Activation in Human Hepatoma Cells
Antimicrob. Agents Chemother.,
June 1, 2003;
47(6):
1912 - 1921.
[Abstract]
[Full Text]
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F. J. Lopez-Hernandez, M. A. Ortiz, Y. Bayon, and F. J. Piedrafita
Z-FA-fmk Inhibits Effector Caspases but not Initiator Caspases 8 and 10, and Demonstrates That Novel Anticancer Retinoid-related Molecules Induce Apoptosis via the Intrinsic Pathway
Mol. Cancer Ther.,
March 1, 2003;
2(3):
255 - 263.
[Abstract]
[Full Text]
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J.-W. Hyun, Y.-C. Jung, H.-S. Kim, E.-Y. Choi, J.-E. Kim, B.-H. Yoon, S.-H. Yoon, Y.-S. Lee, J. Choi, H.-J. You, et al.
8-Hydroxydeoxyguanosine Causes Death of Human Leukemia Cells Deficient in 8-Oxoguanine Glycosylase 1 Activity by Inducing Apoptosis
Mol. Cancer Res.,
February 1, 2003;
1(4):
290 - 299.
[Abstract]
[Full Text]
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M.-T. Park, J. A Kang, J.-A Choi, C.-M. Kang, T.-H. Kim, S. Bae, S. Kang, S. Kim, W.-I. Choi, C.-K. Cho, et al.
Phytosphingosine Induces Apoptotic Cell Death via Caspase 8 Activation and Bax Translocation in Human Cancer Cells
Clin. Cancer Res.,
February 1, 2003;
9(2):
878 - 885.
[Abstract]
[Full Text]
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J. Nitobe, S. Yamaguchi, M. Okuyama, N. Nozaki, M. Sata, T. Miyamoto, Y. Takeishi, I. Kubota, and H. Tomoike
Reactive oxygen species regulate FLICE inhibitory protein (FLIP) and susceptibility to Fas-mediated apoptosis in cardiac myocytes
Cardiovasc Res,
January 1, 2003;
57(1):
119 - 128.
[Abstract]
[Full Text]
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B. Neumeister, M. Faigle, K. Lauber, H. Northoff, and S. Wesselborg
Legionella pneumophila induces apoptosis via the mitochondrial death pathway
Microbiology,
November 1, 2002;
148(11):
3639 - 3650.
[Abstract]
[Full Text]
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J. Pifer, D. Robison, and P. E. Funk
The Avian Chb6 Alloantigen Triggers Apoptosis in a Mammalian Cell Line
J. Immunol.,
August 1, 2002;
169(3):
1372 - 1378.
[Abstract]
[Full Text]
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M. Beil, J. Leser, M. P. Lutz, A. Gukovskaya, T. Seufferlein, G. Lynch, S. J. Pandol, and G. Adler
Caspase 8-mediated cleavage of plectin precedes F-actin breakdown in acinar cells during pancreatitis
Am J Physiol Gastrointest Liver Physiol,
March 1, 2002;
282(3):
G450 - G460.
[Abstract]
[Full Text]
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E. Papouli, M. Defais, and F. Larminat
Overexpression of Metallothionein-II Sensitizes Rodent Cells to Apoptosis Induced by DNA Cross-linking Agent through Inhibition of NF-kappa B Activation
J. Biol. Chem.,
February 8, 2002;
277(7):
4764 - 4769.
[Abstract]
[Full Text]
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D. R. Jones, R. M. Broad, L. D. Comeau, S. J. Parsons, and M. W. Mayo
Inhibition of nuclear factor {kappa}B chemosensitizes non-small cell lung cancer through cytochrome c release and caspase activation
J. Thorac. Cardiovasc. Surg.,
February 1, 2002;
123(2):
310 - 317.
[Abstract]
[Full Text]
[PDF]
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A. D. Schimmer, D. W. Hedley, L. Z. Penn, and M. D. Minden
Receptor- and mitochondrial-mediated apoptosis in acute leukemia: a translational view
Blood,
December 15, 2001;
98(13):
3541 - 3553.
[Full Text]
[PDF]
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A. de Thonel, A. Bettaieb, C. Jean, G. Laurent, and A. Quillet-Mary
Role of protein kinase C zeta isoform in Fas resistance of immature myeloid KG1a leukemic cells
Blood,
December 15, 2001;
98(13):
3770 - 3777.
[Abstract]
[Full Text]
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K. Stahnke, S. Fulda, C. Friesen, G. Strau{beta}, and K.-M. Debatin
Activation of apoptosis pathways in peripheral blood lymphocytes by in vivo chemotherapy
Blood,
November 15, 2001;
98(10):
3066 - 3073.
[Abstract]
[Full Text]
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H. Bantel, B. Sinha, W. Domschke, G. Peters, K. Schulze-Osthoff, and R. U. Janicke
{alpha}-Toxin is a mediator of Staphylococcus aureus-induced cell death and activates caspases via the intrinsic death pathway independently of death receptor signaling
J. Cell Biol.,
November 12, 2001;
155(4):
637 - 648.
[Abstract]
[Full Text]
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D. T. Jones, K. Ganeshaguru, A. E. Virchis, N. I. Folarin, M. W. Lowdell, A. B. Mehta, H. G. Prentice, A. V. Hoffbrand, and R. G. Wickremasinghe
Caspase 8 activation independent of Fas (CD95/APO-1) signaling may mediate killing of B-chronic lymphocytic leukemia cells by cytotoxic drugs or gamma radiation
Blood,
November 1, 2001;
98(9):
2800 - 2807.
[Abstract]
[Full Text]
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J. Pan, G. Xu, and S.-C. J. Yeung
Cytochrome c Release Is Upstream to Activation of Caspase-9, Caspase-8, and Caspase-3 in the Enhanced Apoptosis of Anaplastic Thyroid Cancer Cells Induced by Manumycin and Paclitaxel
J. Clin. Endocrinol. Metab.,
October 1, 2001;
86(10):
4731 - 4740.
[Abstract]
[Full Text]
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D. Chatterjee, I. Schmitz, A. Krueger, K. Yeung, S. Kirchhoff, P. H. Krammer, M. E. Peter, J. H. Wyche, and P. Pantazis
Induction of Apoptosis in 9-Nitrocamptothecin-treated DU145 Human Prostate Carcinoma Cells Correlates with de Novo Synthesis of CD95 and CD95 Ligand and Down-Regulation of c-FLIPshort
Cancer Res.,
October 1, 2001;
61(19):
7148 - 7154.
[Abstract]
[Full Text]
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N. Schrantz, M.-F. Bourgeade, S. Mouhamad, G. Leca, S. Sharma, and A. Vazquez
p38-mediated Regulation of an Fas-associated Death Domain Protein-independent Pathway Leading to Caspase-8 Activation during TGFbeta -induced Apoptosis in Human Burkitt Lymphoma B Cells BL41
Mol. Biol. Cell,
October 1, 2001;
12(10):
3139 - 3151.
[Abstract]
[Full Text]
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A. Renz, W. E. Berdel, M. Kreuter, C. Belka, K. Schulze-Osthoff, and M. Los
Rapid extracellular release of cytochrome c is specific for apoptosis and marks cell death in vivo
Blood,
September 1, 2001;
98(5):
1542 - 1548.
[Abstract]
[Full Text]
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G. Laurent and J.-P. Jaffrezou
Signaling pathways activated by daunorubicin
Blood,
August 15, 2001;
98(4):
913 - 924.
[Abstract]
[Full Text]
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L. Besnault, N. Schrantz, M. T. Auffredou, G. Leca, M. F. Bourgeade, and A. Vazquez
B Cell Receptor Cross-Linking Triggers a Caspase-8- Dependent Apoptotic Pathway That Is Independent of the Death Effector Domain of Fas-Associated Death Domain Protein
J. Immunol.,
July 15, 2001;
167(2):
733 - 740.
[Abstract]
[Full Text]
[PDF]
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P Latham, E K Lund, J C Brown, and I T Johnson
Effects of cellular redox balance on induction of apoptosis by eicosapentaenoic acid in HT29 colorectal adenocarcinoma cells and rat colon in vivo
Gut,
July 1, 2001;
49(1):
97 - 105.
[Abstract]
[Full Text]
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D. Bellarosa, A. Ciucci, A. Bullo, F. Nardelli, S. Manzini, C. A. Maggi, and C. Goso
Apoptotic Events in a Human Ovarian Cancer Cell Line Exposed to Anthracyclines
J. Pharmacol. Exp. Ther.,
April 13, 2001;
296(2):
276 - 283.
[Abstract]
[Full Text]
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P. J. Frost, L. H. Butterfield, V. B. Dissette, J. S. Economou, and B. Bonavida
Immunosensitization of Melanoma Tumor Cells to Non-MHC Fas-Mediated Killing by MART-1-Specific CTL Cultures
J. Immunol.,
March 1, 2001;
166(5):
3564 - 3573.
[Abstract]
[Full Text]
[PDF]
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T. Wieder, F. Essmann, A. Prokop, K. Schmelz, K. Schulze-Osthoff, R. Beyaert, B. Dorken, and P. T. Daniel
Activation of caspase-8 in drug-induced apoptosis of B-lymphoid cells is independent of CD95/Fas receptor-ligand interaction and occurs downstream of caspase-3
Blood,
March 1, 2001;
97(5):
1378 - 1387.
[Abstract]
[Full Text]
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B.R Brehm, S.C Wolf, D Bertsch, M Klaussner, S Wesselborg, S Schuler, and K Schulze-Osthoff
Effects of nebivolol on proliferation and apoptosis of human coronary artery smooth muscle and endothelial cells
Cardiovasc Res,
February 1, 2001;
49(2):
430 - 439.
[Abstract]
[Full Text]
[PDF]
|
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C. G. Ferreira, S. W. Span, G. J. Peters, F. A. E. Kruyt, and G. Giaccone
Chemotherapy Triggers Apoptosis in a Caspase-8-dependent and Mitochondria-controlled Manner in the Non-Small Cell Lung Cancer Cell Line NCI-H460
Cancer Res.,
December 1, 2000;
60(24):
7133 - 7141.
[Abstract]
[Full Text]
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P. A. Svingen, J. E. Karp, S. Krajewski, P. W. Mesner Jr, S. D. Gore, P. J. Burke, J. C. Reed, Y. A. Lazebnik, and S. H. Kaufmann
Evaluation of Apaf-1 and procaspases-2, -3, -7, -8, and -9 as potential prognostic markers in acute leukemia
Blood,
December 1, 2000;
96(12):
3922 - 3931.
[Abstract]
[Full Text]
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C. Ruiz-Ruiz, C. Muñoz-Pinedo, and A. López-Rivas
Interferon-{{gamma}} Treatment Elevates Caspase-8 Expression and Sensitizes Human Breast Tumor Cells to a Death Receptor-induced Mitochondria-operated Apoptotic Program
Cancer Res.,
October 1, 2000;
60(20):
5673 - 5680.
[Abstract]
[Full Text]
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M. Schuler, E. Bossy-Wetzel, J. C. Goldstein, P. Fitzgerald, and D. R. Green
p53 Induces Apoptosis by Caspase Activation through Mitochondrial Cytochrome c Release
J. Biol. Chem.,
March 15, 2000;
275(10):
7337 - 7342.
[Abstract]
[Full Text]
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K. Lauber, H. A. E. Appel, S. F. Schlosser, M. Gregor, K. Schulze-Osthoff, and S. Wesselborg
The Adapter Protein Apoptotic Protease-activating Factor-1 (Apaf-1) Is Proteolytically Processed during Apoptosis
J. Biol. Chem.,
August 3, 2001;
276(32):
29772 - 29781.
[Abstract]
[Full Text]
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TOP
ABSTRACT
INTRODUCTION