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(Received for publication, July
13, 1995; and in revised form, December 14, 1995) From the
In many cell types, position in the cell cycle appears to play a
role in determining susceptibility to apoptosis (programmed cell
death), and expression of various cyclins and activation of
cyclin-dependent kinases (CDKs) have been shown to correlate with the
onset of apoptosis in a number of experimental systems. To assess the
role of CDK-mediated cell cycle events in apoptosis, we have expressed CDK dominant negative mutants in human HeLa cells. Dominant
negative mutants of CDC2, CDK2, and CDK3 each suppressed apoptosis induced by both staurosporine and tumor
necrosis factor
A number of studies suggest that apoptosis is linked with cell
cycle events(1) . Within the organism, apoptosis is found
primarily in proliferating tissues (2, 3) and is
associated with induction of proliferation-associated
genes(4, 5, 6) . Quiescent fibroblasts are
resistant to cytotoxic lymphocyte-induced apoptosis(7) , and
resting peripheral blood T cells are resistant to activation-induced
death by ligation of the T cell receptor(8) . Blocking
proliferation with c-myc antisense oligonucleotides blocks
activation-induced death of T cells(9) , and E1A mutants that are defective in induction of DNA synthesis also fail
to induce apoptosis in susceptible cells(10) . Cells undergoing
apoptosis frequently seem to do so from late in G Cell
proliferation is regulated by cell cycle-specific synthesis of cyclins
and activation of cyclin-dependent kinases (CDKs). (
Figure 4:
Expression of wild type CDKs and cyclin A
overcomes the inhibitory effect of CDK-dns on TNF-
Figure 1:
Inhibition of TNF-
Figure 2:
Quantitation of apoptosis in transfected
cells. 300-600 cells prepared as described in Fig. 1were
counted by phase contrast or fluorescence microscopy and scored as
normal or apoptotic on the basis of morphology. The data for
TNF-
Figure 3:
Association of cdk3 with cyclin
A. HeLa cells were transfected with pCMVcdk2, pCMVcdk3, HA
epitope-tagged pCMVcdk2-dn-HA, pCMVcdk3-dn-HA, pCMVcdk5-dn-HA, or
pCMVcdk3-dn-HA in the presence of a 10-fold excess of pCMVcdk3. 48 h
following transfection, cell extracts were immunoprecipitated with
cyclin A antibody(19) . Western blots of SDS-polyacrylamide gel
electrophoresis-resolved immunoprecipitates were probed with anti-HA
antibody and detected with [
Figure 5:
Overexpression of wild type CDKs and
cyclin A increases apoptosis in the presence of BCL-2. A HeLa
line (clone HB14)(19) , which stably overexpresses human BCL-2 under control of the SV40 enhancer and
promoter(24) , was transfected with the plasmids indicated and
treated with cycloheximide and TNF-
We have shown that three different dominant negative CDK mutants are capable of blocking apoptosis in response to two
general inducers of apoptosis, TNF- These data are consistent with other published
results suggesting that events downstream of cyclin A-dependent kinase
activation are important for apoptosis. For example, cyclin A is
transcribed in response to two inducers of apoptosis, c-myc and E1A(47, 48, 49, 50) .
Elevated cyclin A expression from an inducible promoter can induce
apoptosis in serum-starved fibroblasts(48) , and cyclin
A-dependent kinases are specifically activated when apoptosis is
induced in T cell lines by human immunodeficiency virus tat(51) and in HeLa cells by a variety of physiological and
pharmacological agents(19) . In target cells killed by granzyme
B, CDC2 activity was increased(52) , including activity
associated with cyclin A. ( Results from
experiments performed with dominant negative CDK mutants must be
interpreted cautiously. Such mutants may be expected to exert
pleiotropic effects due to their interference with the cell cycle.
Although we have shown that overexpression of cyclin A or its wild type
catalytic partners can drive apoptosis to high levels in BCL-2
Volume 271,
Number 17,
Issue of April 26, 1996 pp. 10205-10209
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
FOOTNOTES
ACKNOWLEDGEMENTS
REFERENCES
, whereas a dominant negative mutant of CDK5 was without effect. Like CDC2 and CDK2, CDK3 was shown to form a
complex with cyclin A in vivo. CDK5 did not bind cyclin A to
any detectable extent. Overexpression of wild type CDC2, CDK2, CDK3, or cyclin A (but not cyclin B) markedly
elevated the incidence of apoptosis in BCL-2
cells, which otherwise fail to respond to these agents. These
results help identify cell cycle events that are also important for
efficient apoptosis.
or early
in S phase whether the stimulus is ligation of antigen
receptors(11, 12) , growth factor
deprivation(5, 13, 14) , or restoration of
wild type p53 function(15, 16) . Arrest in G
or early G suppresses apoptosis in response to a wide
range of
agents(9, 14, 15, 16, 17, 18, 19, 20) ,
whereas arrest late in G or in S phase can accelerate or
potentiate apoptosis (15, 16, 18, 19) . These data imply
the existence of molecules present in late G and S phase
whose activities facilitate the execution of apoptosis.)Response to growth factors and passage through G is mediated by D-type cyclins, whereas control of entry into S
phase (commencement of DNA replication) is regulated by protein kinases
associated with cyclin E and cyclin A (see reviews in (21) and (22) ). Cyclin D- and cyclin E-associated kinases phosphorylate
pRB, the protein product of the retinoblastoma susceptibility gene,
allowing E2F-dependent transcription of genes whose products are
required for DNA replication (e.g. DNA polymerase ),
whereas cyclin A-associated kinases participate in the phosphorylation
of substrates associated with formation of the replication fork (e.g. replication factor A(21, 22) ). We have
shown previously that arrest of HeLa cells in late G or
early S phase greatly potentiates the apoptosis-inducing ability of a
variety of agents(19) . Induction of apoptosis was uniformly
associated with activation of cyclin A-dependent kinases but not
activity associated with cyclins E or B(19) . These results
suggested that cyclin A might act as a cell cycle-dependent facilitator
of apoptosis. To define at a molecular level cell cycle events that
modify the ability to undergo apoptosis, wild type and dominant
negative mutants of CDKs (23) were transiently expressed in
HeLa cells prior to exposure to apoptosis-inducing agents. We report
that dominant negative mutants of CDC2, CDK2, and CDK3 suppressed apoptosis in response to both staurosporine
and TNF-. In addition, overexpression of the wild type form of
these kinases or cyclin A circumvented the anti-apoptosis activity of
the oncogene BCL-2.
Cell Lines and Plasmids
Human HeLa cells derived
from a cervical carcinoma were grown in monolayer culture as described
previously(19) . A HeLa line (clone HB14), which stably
overexpresses human BCL-2 under control of the SV40 enhancer
and promoter (24) , was maintained in G418 as described
previously(19) . Wild type CDK-expressing plasmids under
control of the CMV promoter (pCMVcdc2, pCMVcdk2, pCMVcdk3, and
pCMVcdk5) and dominant negative CDK-expressing plasmids (pCMVcdc2-dn,
pCMVcdk2-dn, pCMVcdk3-dn, and pCMVcdk5-dn) with or without the
hemagglutinin epitope (HA) tag were a gift from Drs. Sander van den
Heuvel and Ed Harlow (23) (Massachusetts General Hospital). The lacZ-expressing plasmid pCMV
was a gift of Dr. Arthur Lee
(Harvard School of Public Health), and the cyclin A-expressing plasmid
pCMVcycA was from Dr. Phil Hinds (Harvard Medical School)(25) .
A plasmid expressing human cyclin B containing a myc-derived
epitope tag was a gift of Dr. Frank McKeon (Harvard Medical
School)(26) .Transfection and Analysis of Transfected Cells
2
10
HeLa cells grown on glass coverslips were
co-transfected with 0.4 µg pCMV (lacZ) and 2 µg
of either pCMVcdc2-dn, pCMVcdk2-dn, pCMVcdk3-dn, pCMVcdk5-dn, or
pCMVcdk3-dn-HA using LipofectACE (Life Technologies, Inc.) according to
the manufacturer's instructions. For the experiments shown in Fig. 4, 10 µg of wild type plasmid was included along with 1
µg of pCMV and 5 µg of each CDK-dn construct. 48 h after
transfection, cells were induced to undergo apoptosis by the addition
of 100 ng/ml TNF- (Research Bioproducts) and 30 µg/ml
cycloheximide for 3 h or by the addition of 500 ng/ml staurosporine and
30 µg/ml cycloheximide for 5 h. The cells were fixed and stained
for -galactosidase activity using 5-bromo-4-chloro-3-indolyl
-D-galactopyranoside (X-gal) or for indirect
immunofluorescence using an anti--galactosidase antibody (Organon
Teknika/Cappel) at 1:1000 or an anti-HA antibody (Babco) at 1:1000, all
exactly as described(27) . Immunofluorescence of myc-tagged cyclin B expression (26) and fluorescent
DNA staining with Hoechst 33258 (19) were carried out as
described. For quantitation, 300-600 cells were counted by phase
contrast or fluorescence microscopy and scored as normal or apoptotic
on the basis of nuclear and cytoplasmic morphology. For
autoradiography, cells were labeled for 3 h with 10 µCi/ml
[
H]thymidine prior to fixation and X-gal
staining. Incorporation was visualized with Ilford Nuclear Emulsion K-5
and Kodak D19 developer with an exposure time of 8 days. 600-1200 cells
were examined from each treatment group.
-induced
apoptosis. Cells were transfected with the combination of plasmids
indicated and treated and analyzed as described in the legend to Fig. 1. Transfections contained wild type and dominant negative
plasmids at a 2:1 ratio, respectively (see ``Experimental
Procedures''). Transfected cells were identified by
-galactosidase activity, and apoptotic cells were identified by
phase microscopy. The percentage of inhibition of apoptosis was
calculated as [1 - (% of apoptosis in the presence of
CDK-dn/% of apoptosis with lacZ alone)] 100. The
data are the the means ± S.E. for six (cdc2-dn, cdk2-dn, and
cyclin A) and four (cdk3-dn) independent
determinations.
- and
staurosporine-induced apoptosis by dominant negative CDK mutants. a and b, HeLa cells co-transfected with lacZ and
an excess of either pCMVcdk2-dn (a) or pCMVcdk5-dn (b). 48 h after transfection, cells were induced to undergo
apoptosis by addition of 100 ng/ml TNF- and 30 µg/ml
cycloheximide for 3 h, at which time they were fixed and stained with
X-gal for -galactosidase activity. Note the characteristic
rounded, shriveled appearance of apoptotic cells. Arrowheads identify cells displaying -galactosidase activity. c and d, HeLa cells were co-transfected with lacZ
and pCMVcdk2-dn and induced to undergo apoptosis 48 h later by
treatment with 500 ng/ml staurosporine and 30 µg/ml cycloheximide
for 5 h and then fixed and stained for indirect immunofluorescence
using an anti--galactosidase antibody. c, fluorescent DNA
staining with Hoechst 33258; d, anti--galactosidase
immunofluorescence staining of the same cells. Note that the
characteristic apoptotic nuclei are present only in cells not
expressing the transfection marker (arrowheads). e and f, HeLa cells were transfected with pCMVcdk3-dn-HA.
Cells were treated with staurosporine and cycloheximide and stained
with an anti-HA antibody. e, DNA staining with Hoechst 33258; f, anti-HA immunofluorescence staining of the same cells. Note
that the sole apoptotic cell (arrowhead) is the only cell not
expressing cdk3-dn-HA.
Immunoprecipitations and Western Analysis
2
10
HeLa cells were transfected by lipofection with
2 µg of pCMVcdk2, pCMVcdk3, HA epitope-tagged pCMVcdk2-dn-HA,
pCMVcdk3-dn-HA, pCMVcdk5-dn-HA, or pCMVcdk3-dn-HA in the presence of 20
µg of pCMVcdk3. 48 h following transfection, cell extracts were
immunoprecipitated with cyclin A antibody as described
previously(19) . Western blots of SDS-polyacrylamide gel
electrophoresis-resolved immunoprecipitates (5 10 cell equivalents per lane) were probed with anti-HA antibodies
(Babco), which were detected with [
I]protein A
(1 µCi/ml), all according to previously described
methods(19, 28) . Exposure time was 48 h on Kodak XAR
film with an intensifying screen.
Inhibition of Apoptosis by CDK Dominant Negative
Mutants
Apoptosis was induced in HeLa cells by the addition of
either TNF- and cycloheximide or staurosporine and cycloheximide.
These treatments were selected because they rapidly induce apoptosis in
numerous cell types, both primary and
transformed(29, 30, 31) , and are thus
believed to activate elements of a death program common to all
cells(31, 32) . To assess the role of CDKs in
apoptosis induced by these agents, CDK-dns were transfected into cells
48 h prior to induction of apoptosis. When expressed under control of a
strong promoter, the catalytically inactive CDK-dns compete with wild
type CDKs for cyclin binding and thus act as competitive inhibitors of
CDK activity(23) . Following induction of apoptosis, HeLa cells
co-transfected with CDK-dns and a lacZ marker plasmid were
fixed and stained for -galactosidase activity with a chromogenic
substrate (Fig. 1, a and b) or for
-galactosidase expression by indirect immunofluorescence (Fig. 1, c and d), thereby allowing
microscopic determination of the apoptosis frequency among lacZ cells. Alternatively, expression of
HA-tagged CDK mutants (CDK-dn-HA) permitted direct detection of
CDK-dn-transfected cells by indirect immunofluorescence with a
hemagglutinin-specific monoclonal antibody (Fig. 1, e and f). Note that the presence of transfection markers
for cdk2-dn and cdk3-dn correlates with the absence of apoptosis in the
representative fields shown. All three methods for detecting
transfected cells yielded equivalent results, and the combined data are
shown quantitatively in Fig. 2. Cdc2-dn, cdk2-dn, and cdk3-dn
were capable of inhibiting apoptosis induced by either TNF- or
staurosporine with cdc2-dn being the least effective. Although CDC2 can
bind to cyclin A at G/S (33, 34) and
although there is evidence for CDC2 activity being required
for S phase entry in some cell
types(35, 36, 37) , Cdc2-dn has its
predominant effect during G
(23) , which may account
for its reduced effectiveness here. Simultaneous transfection with
pairs of CDK-dns or with all three inhibitory CDK-dns did not lead to
inhibition greater than that seen with either cdk2-dn or cdk3-dn alone.
In contrast, a dominant negative mutant of CDK5(38) ,
a CDK of limited tissue distribution (39) that binds
to D-type cyclins in vitro and is activated in vivo by p35, a protein with little sequence homology to known
vertebrate cyclins(40, 41) , had no effect on
apoptosis.
-induced apoptosis are the means ± S.E. for seven
independent determinations; those for staurosporine-induced apoptosis
are for three independent determinations. sham, cells taken
through the transfection procedure in the absence of added
DNA.
Cdk3 Associates with Cyclin A in Vivo
A cyclin
partner for CDK3, which, like CDK2, is required for the G/S
phase transition, has not been described previously. However, as shown
in Fig. 3, cdk3-dn-HA, like cdk2-dn-HA, was co-precipitated with
antibodies to cyclin A from extracts of cells overexpressing these
constructs. Cdk5-dn-HA failed to associate with cyclin A.
Co-transfection of cdk3-dn-HA with a 10-fold excess of wild type CDK3 lacking the epitope tag blocked the association of cyclin
A with cdk3-dn-HA, indicating that wild type CDK3 can compete with
cdk3-dn-HA for cyclin A binding. Control cyclin A immunoprecipitations
from cells overexpressing either wild type CDC2 or wild type CDK2 displayed no nonspecific HA-reactive species. These data
show that CDK3, when overexpressed, will bind cyclin A in vivo and imply that at least part of its activity in this model system
may be cyclin A-dependent. Although the subcellular localization of
CDK3 is unknown, CDC2, CDK2, and cyclin A reside in both the nucleus
and cytoplasm of HeLa cells(19, 34, 42) .
Because apoptotic events within these compartments can occur
independently(43, 44) , it is noteworthy that
inhibition of CDKs blocked both the cytoplasmic (cell rounding and
surface blebbing; Fig. 1a) and nuclear events
(chromatin condensation; Fig. 1, c and e)
associated with apoptosis. We note that complete inhibition of
apoptosis is not seen with any individual CDK-dn or with any
combination of CDK-dns. This may be due to the elevated levels of
cyclins that are found in human papillomavirus-positive cells (45) (making competition for positive regulatory factors by
CDK-dns more difficult in HeLa cells), to variations in the level of
CDK-dn protein expressed transiently in each cell, or to additional
apoptotic pathways that are independent of CDK activity.
I]protein A.
HA-immunoreactive species are indicated by the arrowhead.
Cyclin A or Wild Type CDK Expression Reverses the Effect
of CDK-dns
Although cdk5-dn provides a useful comparison for the
effects of the other CDK-dns, the best control is to counter the effect
of a mutant CDK by co-expression of the wild type CDK or its cyclin partner(23) . Thus, wild type CDKs
and cyclin A were co-transfected with the CDK-dns to see if inhibition
of apoptosis could be reversed. As shown in Fig. 4,
co-expression of either cyclin A or the wild type CDKs was able to
overcome the inhibitory effect on apoptosis of each CDK-dn. Among the
wild type CDKs, CDC2 was least effective, whereas CDK2 and CDK3 were similarly effective at blocking inhibition
by all three CDK-dns. Inhibition was blocked most effectively by cyclin
A co-transfection. Indeed, in some cases cyclin A was able to drive
cells to apoptosis frequencies higher than those seen in control cells
transfected with lacZ alone. These results suggest redundant
functions of CDC2, CDK2, and CDK3 in
apoptosis. The ability of any one dominant negative mutant to inhibit
apoptosis is likely due to competition between the mutant and the three
wild type kinases for the same CDK activating component(s).Cyclin A or CDK Overexpression Can Promote Apoptosis in a
BCL-2
Although the above experiments
indicate that inhibition of certain CDKs suppresses apoptosis, it is
possible that this suppression is accomplished through an indirect
mechanism. For example, Cdk2-dn and Cdk3-dn mutants are known to
prevent entry of cells into S phase(23) , and a biochemical
pathway required for apoptosis may be active only during this phase of
the cell cycle. If, however, overexpression of cyclin A or wild type CDKs in asynchronously growing cell populations increases
apoptosis in response to agents such as TNF- Background, a closer
relationship between these proteins and apoptosis may be implied,
particularly if increases occur in the absence of gross cell cycle
perturbation. To address this issue, we overexpressed wild type CDKs and cyclin A in asynchronously growing HB14 cells, a HeLa
cell line that stably overexpresses the BCL-2 oncogene and is
resistant to staurosporine- and TNF--induced
apoptosis(19) . When HB14 cells were transiently transfected
with either wild type CDC2, CDK2, CDK3, or
cyclin A, the levels of TNF--induced apoptosis increased from
approximately 2-9% in control transfections to as high as 70%, a
level approaching that seen in the BCL-2-deficient parental
HeLa cells (Fig. 5). The proportion of cells in S phase was not
significantly altered in cells expressing cyclin A (32%) when compared
with nonexpressing cells in the same dish (34%), as detemined by
autoradiography of cells labeled with
[H]thymidine and stained for -galactosidase
activity, indicating that the increase in apoptosis was not due to an
increase in the number of cells in S phase. In contrast to cyclin A,
expression of cyclin B in HB14 cells had no effect on apoptosis.
Consistent with the earlier CDK-dn experiments, CDC2 was less
effective than CDK2, CDK3, and cyclin A at enhancing
apoptosis in the presence of BCL-2. Similar results were
obtained when apoptosis was induced by staurosporine or
6-dimethylaminopurine and when a second independent BCL-2 clone was used (data not shown). Only
modest increases in apoptosis were obtained in HeLa cells, indicating
that the level of expression of these genes is not rate-limiting for
apoptosis in the parental cells. Minimal toxicity was seen when CDKs or cyclin A were overexpressed in the absence of
TNF-, in agreement with the findings of others for CDC2 and CDK2(23, 26, 46) .
as described in the legend to Fig. 1. Transfected cells were identified by -galactosidase
activity, and apoptotic cells were identified by phase microscopy (lacZ, cdc2, cdk2, cdk3, and cyclin A). Cyclin B-transfected
cells were visualized by indirect immunofluorescence staining using an
antibody to the myc-epitope tag, with apoptotic cells
identified by Hoechst staining. NT, not transfected (cells
processed in parallel but without LipofectACE treatment). The data are
the means ± S.E. for two (cdk3, cyclin A, and cyclin B) or three (NT, lacZ, cdc2, and cdk2) independent
determinations.
and staurosporine. The
inhibitory effect could be overcome by co-expression of cyclin A or by
the wild type form of any of the other three wild type CDKs.
This functional redundancy suggests that the dominant negative mutants
exert their effects via competition for a common activating factor or
factors, a conclusion further supported by the finding that
co-transfection of two or more CDK-dns was no more effective than
single CDK-dns for suppressing apoptosis. It is possible that this
shared activating factor is cyclin A, because all three mutants can
bind this cyclin.)Use of a temperature-sensitive CDC2 mutant significantly reduced apoptosis in this
system(52) , and transfection of target cells with wee1, a tyrosine kinase that phosphorylates and inactivates
CDKs, also inhibited apoptosis(53) . cells in the absence of gross cell
cycle perturbations, these data do not demonstrate a direct involvement
of cyclins or CDKs in apoptosis. The biochemical pathways that emanate
from activation of any cyclin-CDK complex are most certainly complex,
wide-ranging, and intricately involved in many different facets of cell
proliferation. However, these findings do identify cell cycle
components whose activity, when modulated, can promote or impede
apoptosis. The existence of such components supports the notion that
specific cell cycle events must be completed before apoptosis can occur
in an efficient manner and suggests that cell cycle regulatory proteins
might be useful therapeutic targets for manipulating apoptosis.
),
tumor necrosis factor ; X-gal, 5-bromo-4-chloro-3-indolyl
-D-galactopyranoside; HA, hemagglutinin epitope.)
We are grateful to S. van den Heuvel and E. Harlow of
the Massachusetts General Hospital for their generous gifts of CDK-dn,
epitope-tagged, and wild type plasmid constructs; to P. Hinds of
Harvard Medical School for the pCMVcycA plasmid; to F. McKeon and R.
Heald of Harvard Medical School for the myc-tagged cyclin B
expression construct; to Sun W. Tam (currently at Mitotix, Inc.) for
carrying out the cyclin B transfections; and to Arthur (Mu En) Lee of
Harvard School of Public Health for the pCMV plasmid and for
valuable comments on the manuscript.
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
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H. Yuan, Y.-M. Xie, and I. S. Y. Chen Depletion of Wee-1 Kinase Is Necessary for both Human Immunodeficiency Virus Type 1 Vpr- and Gamma Irradiation-Induced Apoptosis J. Virol., February 1, 2003; 77(3): 2063 - 2070. [Abstract] [Full Text] [PDF]
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C. V. Finkielstein, L. G. Chen, and J. L. Maller A Role for G1/S Cyclin-dependent Protein Kinases in the Apoptotic Response to Ionizing Radiation J. Biol. Chem., October 4, 2002; 277(41): 38476 - 38485. [Abstract] [Full Text] [PDF]
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 D. Sampath, Z. Shi, and W. Plunkett Inhibition of Cyclin-Dependent Kinase 2 by the Chk1-Cdc25A Pathway during the S-Phase Checkpoint Activated by Fludarabine: Dysregulation by 7-Hydroxystaurosporine Mol. Pharmacol., September 1, 2002; 62(3): 680 - 688. [Abstract] [Full Text] [PDF]
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T. Sandal, C. Stapnes, H. Kleivdal, L. Hedin, and S. O. Doskeland A Novel, Extraneuronal Role for Cyclin-dependent Protein Kinase 5 (CDK5). MODULATION OF cAMP-INDUCED APOPTOSIS IN RAT LEUKEMIA CELLS J. Biol. Chem., May 31, 2002; 277(23): 20783 - 20793. [Abstract] [Full Text] [PDF]
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 S. Mazumder, B. Gong, Q. Chen, J. A. Drazba, J. C. Buchsbaum, and A. Almasan Proteolytic Cleavage of Cyclin E Leads to Inactivation of Associated Kinase Activity and Amplification of Apoptosis in Hematopoietic Cells Mol. Cell. Biol., April 1, 2002; 22(7): 2398 - 2409. [Abstract] [Full Text] [PDF]
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 S. C. Coleman, Z. A. Stewart, T. A. Day, J. L. Netterville, B. B. Burkey, and J. A. Pietenpol Analysis of Cell-Cycle Checkpoint Pathways in Head and Neck Cancer Cell Lines: Implications for Therapeutic Strategies Arch Otolaryngol Head Neck Surg, February 1, 2002; 128(2): 167 - 176. [Abstract] [Full Text] [PDF]
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K. Ye, J. Zhou, J. W. Landen, E. M. Bradbury, and H. C. Joshi Sustained Activation of p34cdc2 Is Required for Noscapine-induced Apoptosis J. Biol. Chem., December 7, 2001; 276(50): 46697 - 46700. [Abstract] [Full Text] [PDF]
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S. Leonce, V. Perez, S. Lambel, D. Peyroulan, F. Tillequin, S. Michel, M. Koch, B. Pfeiffer, G. Atassi, J. A Hickman, et al. Induction of Cyclin E and Inhibition of DNA Synthesis by the Novel Acronycine Derivative S23906-1 Precede the Irreversible Arrest of Tumor Cells in S Phase Leading to Apoptosis Mol. Pharmacol., December 1, 2001; 60(6): 1383 - 1391. [Abstract] [Full Text] [PDF]
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D. Chattopadhyay, M. K. Ghosh, A. Mal, and M. L. Harter Inactivation of p21 by E1A Leads to the Induction of Apoptosis in DNA-Damaged Cells J. Virol., October 15, 2001; 75(20): 9844 - 9856. [Abstract] [Full Text] [PDF]
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S. Adachi, A. J. Obaya, Z. Han, N. Ramos-Desimone, J. H. Wyche, and J. M. Sedivy c-Myc Is Necessary for DNA Damage-Induced Apoptosis in the G2 Phase of the Cell Cycle Mol. Cell. Biol., August 1, 2001; 21(15): 4929 - 4937. [Abstract] [Full Text] [PDF]
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 S. Adachi, H. Ito, M. Tamamori-Adachi, Y. Ono, T. Nozato, S. Abe, M.-a. Ikeda, F. Marumo, and M. Hiroe Cyclin A/cdk2 Activation Is Involved in Hypoxia-Induced Apoptosis in Cardiomyocytes Circ. Res., March 2, 2001; 88(4): 408 - 414. [Abstract] [Full Text] [PDF]
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 J. J. Smith, E. K. Evans, M. Murakami, M. B. Moyer, M. A. Moseley, G. V. Woude, and S. Kornbluth Wee1-regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts J. Cell Biol., December 18, 2000; 151(7): 1391 - 1400. [Abstract] [Full Text] [PDF]
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 M. T. Winkler, L. S. Schang, A. Doster, T. Holt, and C. Jones Analysis of cyclins in trigeminal ganglia of calves infected with bovine herpesvirus-1 J. Gen. Virol., December 1, 2000; 81(12): 2993 - 2998. [Abstract] [Full Text]
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M. Nishizawa, M. Kamata, R. Katsumata, and Y. Aida A Carboxy-Terminally Truncated Form of the Human Immunodeficiency Virus Type 1 Vpr Protein Induces Apoptosis via G1 Cell Cycle Arrest J. Virol., July 1, 2000; 74(13): 6058 - 6067. [Abstract] [Full Text]
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L. A. Porter, G. Singh, and J. M. Lee Abundance of cyclin B1 regulates gamma -radiation-induced apoptosis Blood, April 15, 2000; 95(8): 2645 - 2650. [Abstract] [Full Text] [PDF]
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 A. M. Senderowicz and E. A. Sausville Preclinical and Clinical Development of Cyclin-Dependent Kinase Modulators J Natl Cancer Inst, March 1, 2000; 92(5): 376 - 387. [Abstract] [Full Text] [PDF]
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 I. Petrache, L. E. Otterbein, J. Alam, G. W. Wiegand, and A. M. K. Choi Heme oxygenase-1 inhibits TNF-alpha -induced apoptosis in cultured fibroblasts Am J Physiol Lung Cell Mol Physiol, February 1, 2000; 278(2): L312 - L319. [Abstract] [Full Text] [PDF]
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L. Michea, D. R. Ferguson, E. M. Peters, P. M. Andrews, M. R. Kirby, and M. B. Burg Cell cycle delay and apoptosis are induced by high salt and urea in renal medullary cells Am J Physiol Renal Physiol, February 1, 2000; 278(2): F209 - F218. [Abstract] [Full Text] [PDF]
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K. J. Harvey, D. Lukovic, and D. S. Ucker Caspase-dependent Cdk Activity Is a Requisite Effector of Apoptotic Death Events J. Cell Biol., January 10, 2000; 148(1): 59 - 72. [Abstract] [Full Text] [PDF]
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 S. K. Banerjee, A. P. Weston, M. N. Zoubine, D. R. Campbell, and R. Cherian Expression of Cdc2 and Cyclin B1 in Helicobacter pylori-Associated Gastric MALT and MALT Lymphoma : Relationship to Cell Death, Proliferation, and Transformation Am. J. Pathol., January 1, 2000; 156(1): 217 - 225. [Abstract] [Full Text] [PDF]
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J. Ciacci-Zanella, M. Stone, G. Henderson, and C. Jones The Latency-Related Gene of Bovine Herpesvirus 1 Inhibits Programmed Cell Death J. Virol., December 1, 1999; 73(12): 9734 - 9740. [Abstract] [Full Text]
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H.-M. Koo, M. J. McWilliams, W. G. Alvord, and G. F. Vande Woude Ras Oncogene-Induced Sensitization to 1-{{beta}}-D-Arabinofuranosylcytosine Cancer Res., December 1, 1999; 59(24): 6057 - 6062. [Abstract] [Full Text] [PDF]
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K. S. Choi, Y. W. Eom, Y. Kang, M. J. Ha, H. Rhee, J.-W. Yoon, and S.-J. Kim Cdc2 and Cdk2 Kinase Activated by Transforming Growth Factor-beta 1 Trigger Apoptosis through the Phosphorylation of Retinoblastoma Protein in FaO Hepatoma Cells J. Biol. Chem., November 5, 1999; 274(45): 31775 - 31783. [Abstract] [Full Text] [PDF]
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Z. A. Stewart, D. Mays, and J. A. Pietenpol Defective G1-S Cell Cycle Checkpoint Function Sensitizes Cells to Microtubule Inhibitor-induced Apoptosis Cancer Res., August 1, 1999; 59(15): 3831 - 3837. [Abstract] [Full Text] [PDF]
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T.-H. Wang, D. M. Popp, H.-S. Wang, M. Saitoh, J. G. Mural, D. C. Henley, H. Ichijo, and J. Wimalasena Microtubule Dysfunction Induced by Paclitaxel Initiates Apoptosis through Both c-Jun N-terminal Kinase (JNK)-dependent and -Independent Pathways in Ovarian Cancer Cells J. Biol. Chem., March 19, 1999; 274(12): 8208 - 8216. [Abstract] [Full Text] [PDF]
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 A. Hakem, T. Sasaki, I. Kozieradzki, and J. M. Penninger The Cyclin-dependent Kinase Cdk2 Regulates Thymocyte Apoptosis J. Exp. Med., March 15, 1999; 189(6): 957 - 968. [Abstract] [Full Text] [PDF]
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C. D. Scatena, Z. A. Stewart, D. Mays, L. J. Tang, C. J. Keefer, S. D. Leach, and J. A. Pietenpol Mitotic Phosphorylation of Bcl-2 during Normal Cell Cycle Progression and Taxol-induced Growth Arrest J. Biol. Chem., November 13, 1998; 273(46): 30777 - 30784. [Abstract] [Full Text] [PDF]
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Y. Jiang, A. Hossain, M. T. Winkler, T. Holt, A. Doster, and C. Jones A Protein Encoded by the Latency-Related Gene of Bovine Herpesvirus 1 Is Expressed in Trigeminal Ganglionic Neurons of Latently Infected Cattle and Interacts with Cyclin-Dependent Kinase 2 during Productive Infection J. Virol., October 1, 1998; 72(10): 8133 - 8142. [Abstract] [Full Text] [PDF]
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B.-B. Zhou, H. Li, J. Yuan, and M. W. Kirschner Caspase-dependent activation of cyclin-dependent kinases during Fas-induced apoptosis in Jurkat cells PNAS, June 9, 1998; 95(12): 6785 - 6790. [Abstract] [Full Text] [PDF]
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 S. Illenberger, Q. Zheng-Fischhöfer, U. Preuss, K. Stamer, K. Baumann, B. Trinczek, J. Biernat, R. Godemann, E.-M. Mandelkow, and E. Mandelkow The Endogenous and Cell Cycle-dependent Phosphorylation of tau Protein in Living Cells: Implications for Alzheimer's Disease Mol. Biol. Cell, June 1, 1998; 9(6): 1495 - 1512. [Abstract] [Full Text]
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K. J. Harvey, J. F. Blomquist, and D. S. Ucker Commitment and Effector Phases of the Physiological Cell Death Pathway Elucidated with Respect to Bcl-2, Caspase, and Cyclin-Dependent Kinase Activities Mol. Cell. Biol., May 1, 1998; 18(5): 2912 - 2922. [Abstract] [Full Text]
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N. J. Donato and M. Perez Tumor Necrosis Factor-induced Apoptosis Stimulates p53 Accumulation and p21WAF1 Proteolysis in ME-180 Cells J. Biol. Chem., February 27, 1998; 273(9): 5067 - 5072. [Abstract] [Full Text] [PDF]
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B. W. Parker, G. Kaur, W. Nieves-Neira, M. Taimi, G. Kohlhagen, T. Shimizu, M. D. Losiewicz, Y. Pommier, E. A. Sausville, and A. M. Senderowicz Early Induction of Apoptosis in Hematopoietic Cell Lines After Exposure to Flavopiridol Blood, January 15, 1998; 91(2): 458 - 465. [Abstract] [Full Text] [PDF]
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D. S. Park, B. Levine, G. Ferrari, and L. A. Greene Cyclin Dependent Kinase Inhibitors and Dominant Negative Cyclin Dependent Kinase 4 and 6 Promote Survival of NGF-Deprived Sympathetic Neurons J. Neurosci., December 1, 1997; 17(23): 8975 - 8983. [Abstract] [Full Text] [PDF]
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R. Beyaert, V. J. Kidd, S. Cornelis, M. Van de Craen, G. Denecker, J. M. Lahti, R. Gururajan, P. Vandenabeele, and W. Fiers Cleavage of PITSLRE Kinases by ICE/CASP-1 and CPP32/CASP-3 during Apoptosis Induced by Tumor Necrosis Factor J. Biol. Chem., May 2, 1997; 272(18): 11694 - 11697. [Abstract] [Full Text] [PDF]
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D. S. Park, E. J. Morris, L. A. Greene, and H. M. Geller G1/S Cell Cycle Blockers and Inhibitors of Cyclin-Dependent Kinases Suppress Camptothecin-Induced Neuronal Apoptosis J. Neurosci., February 15, 1997; 17(4): 1256 - 1270. [Abstract] [Full Text] [PDF]
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D. Kornitzer, R. Sharf, and T. Kleinberger Adenovirus E4orf4 protein induces PP2A-dependent growth arrest in Saccharomyces cerevisiae and interacts with the anaphase-promoting complex/cyclosome J. Cell Biol., July 23, 2001; 154(2): 331 - 344. [Abstract] [Full Text] [PDF]
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