Previous Article | Next Article 
Molecular and Cellular Biology, February 2001, p. 854-864, Vol. 21, No. 3
0270-7306/01/$04.00+0 DOI: 10.1128/MCB.21.3.854-864.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Downregulation of Bim, a Proapoptotic Relative of
Bcl-2, Is a Pivotal Step in Cytokine-Initiated Survival Signaling in
Murine Hematopoietic Progenitors
Tetsuharu
Shinjyo,1
Ryoko
Kuribara,1,2
Takeshi
Inukai,3
Hajime
Hosoi,4
Taisei
Kinoshita,5
Atsushi
Miyajima,5
Peter J.
Houghton,6
A. Thomas
Look,7
Keiya
Ozawa,1,2 and
Toshiya
Inaba1,8,*
Departments of Molecular Biology1
and Hematology,2 Jichi Medical School,
Tochigi 329-0498, Department of Pediatrics, Yamanashi Medical
University, Yamanashi 409-3898,3
Department of Pediatrics, Kyoto Prefectural Medical School,
Kyoto 606,4 Institute of Molecular
and Cellular Bioscience, University of Tokyo, Tokyo
174,5 and Department of
Molecular Oncology, Research Institute for Radation Biology and
Medicine, Hiroshima University, Hiroshima
734-8553,8 Japan; Department of
Molecular Pharmacology, St. Jude Children's Research Hospital,
Memphis Tennessee 381056; and Department
of Pediatric Oncology, Dana Farber Cancer Institute, Boston,
Massachusetts 021157
Received 17 July 2000/Returned for modification 20 September
2000/Accepted 2 November 2000
 |
ABSTRACT |
Two distinct signaling pathways regulate the survival of
interleukin-3 (IL-3)-dependent hematopoietic progenitors. One
originates from the membrane-proximal portion of the cytoplasmic domain
of the IL-3 receptor (
c chain), which is shared by IL-3 and
granulocyte-macrophage colony-stimulating factor and is involved in the
regulation of Bcl-xL through activation of STAT5. The other
pathway emanates from the distal region of the c chain and overlaps
with downstream signals from constitutively active Ras proteins.
Although the latter pathway is indispensable for cell survival, its
downstream targets remain largely undefined. Here we show that the
expression of Bim, a member of the BH3-only subfamily of cell death
activators, is downregulated by IL-3 signaling through either of two
major Ras pathways: Raf/mitogen-activated protein kinase and the
phosphatidylinositol 3-kinase/mammalian target of rapamycin.
Akt/phosphokinase B does not appear to play a significant role in this
regulatory cascade. Bim downregulation has important implications for
cell survival, since enforced expression of this death activator at
levels equivalent to those induced by cytokine withdrawal led to
apoptosis even in the presence of IL-3. We conclude that Bim is a
pivotal molecule in cytokine regulation of hematopoietic cell survival.
| |
INTRODUCTION |
Homeostasis, responses to emergency
situations, and the transition from emergency to normal status in the
hematopoietic system are all regulated by cytokines, through control of
cell division, differentiation, and survival. Cytokine regulation of
apoptosis not only plays a critical role in normal hematopoiesis but
also can contribute to leukemogenesis, when one or more control
elements fail to function properly. In recent attempts to clarify the
transduction pathways through which cytokines control cell survival, we
found that such signaling can involve multiple independent pathways (20, 31).
By contrast, cell death decisions are implemented through an
evolutionarily conserved mechanism (or general apoptosis program) in
which members of the Bcl-2 superfamily play the central roles (reviewed
in references 1 and 6). The anti- or proapoptotic family
members regulate the translocation of cytochrome c from mitochondria to the cytosol, an event that ultimately activates the
caspase cascade, while the BH3-only subfamily of cell death activators
inhibit the function of the antiapoptotic Bcl-2 family members by
binding to them. In the nematode Caenorhabditis elegans, Egl-1, the sole BH3-only death activator in this organism, functions at
the most upstream point of this system to inhibit the function of
Ced-9, again the sole antiapoptotic member of the Bcl-2 family in
C. elegans (8). In mammals, six proteins have
been isolated as members of the BH3-only subfamily and five have been
identified as antiapoptotic and three as proapoptotic members of the
Bcl-2 family. Redundancy in each category of the Bcl-2 superfamily
could be explained, at least partially, by the tissue- and/or
stimulus-specific response of each family member. For instance, the
antiapoptotic Bcl-w protein is required for the support of Sertoli
cells in the testes but not of other cells, as demonstrated in studies with Bcl-w-deficient mice (39), while Bid, a member of
BH3-only subfamily, is involved in Fas-mediated apoptosis in
hepatocytes (48). Thus, several members of the Bcl-2
superfamily might play key roles in the response of hematopoietic
progenitors to cytokine withdrawal.
Previous studies have demonstrated the importance of Ras pathways, not
only in oncogenesis when constitutively activated by oncogenic Ras
mutants (reviewed in reference 26) but also in the
regulation of apoptosis in hematopoietic progenitors (reviewed in
reference 33). The membrane-distal region of the
cytoplasmic domain of the common (c) chain is required for both
the survival and activation of Ras in interleukin-3 (IL-3)-dependent
hematopoietic cells (28). Recently, both
Raf/mitogen-activated protein kinase (MAPK) and
rapamycin/wortmannin-sensitive (most probably phosphatidylinositol 3-kinase [PI3-K]-dependent) pathways downstream of active Ras were
found to prevent apoptosis in cytokine-deprived hematopoietic cells
(30). These findings suggest that certain Bcl-2 family members under the control of Ras pathways may be pivotal factors in the
cytokine-mediated regulation of apoptosis in hematopoietic progenitors.
We and others reported previously that the expression levels of
Bcl-xL are downregulated in cytokine-deprived murine
IL-3-dependent cells and that enforced expression of Bcl-xL
in these cells markedly delays apoptosis due to IL-3 deprivation
(31, 32). We also demonstrated that signals originating
from the proximal portion of the c chain, but not from Ras pathways,
are required for the induction of Bcl-xL expression
(31), in agreement with recent published results
indicating that STAT5 is involved in the transcriptional regulation of
Bcl-xL (15, 42, 43). Thus, although
Bcl-xL is considered to be an important target of
cytokine-dependent survival pathways, it is probably not the Bcl-2
family member predicted to interact with Ras pathways.
Another candidate is Bad, a BH3-only subfamily member that is
inactivated by the phosphorylation of two serine residues through Akt
and other kinases downstream of Ras pathways (12, 13, 50).
However, the contribution of Akt/Bad pathways to the apoptosis regulation system in hematopoietic cells has been controversial. Some
investigators contend that Akt reverses apoptosis caused by cytokine
withdrawal but does so much less efficiently than activated Ras does
(2, 44), while others rule out the involvement of Akt/Bad
pathways in the survival of hematopoietic progenitors (3, 16,
41).
A final candidate is Bim/Bod. This BH3-only death activator was
isolated independently by two groups that exploited its ability to bind
Bcl-2 or Mcl1 (18, 34). Alternative splicing gives rise to
three variants, BimEL, BimL, and BimS, each of which contains the BH3
domain and functions as a death inducer. In certain cell types, BimEL
and BimL, but not BimS, bind to an Mr = 8,000 dynein light chain, LC8 (also known as PIN or Dlc-1) (11,
25, 27), which mediates the function of the former two isoforms
(38). The biological relevance of Bim to the control of
hematopoiesis was indicated in a recent study of Bim-deficient mice, in
which the homeostasis of the lymphoid system was disrupted without
resulting in apparent adverse effects on other organs (4).
Here we identify Bim downregulation through either of two major Ras
pathways (Raf/MAPK or PI3-K/mTOR) as a pivotal step in the maintenance
of hematopoietic cell survival under the control of cytokines.
| |
MATERIALS AND METHODS |
Cell culture and cell growth assay.
Murine IL-3-dependent
(Baf-3, FL5.12, and 32D) cells were cultured in RPMI 1640 medium
containing 10% fetal calf serum, 20 mM HEPES, 50 µM
2-mercaptoethanol, and 0.5% 10T1/2 conditioned medium as a source of
murine IL-3. In some experiments, recombinant mouse IL-3 (Wako Pure
Chemical, Osaka, Japan) was used at the concentrations indicated in the
figure legends. To deplete IL-3, we washed the cells twice with
IL-3-free growth medium. Viable-cell counts were determined by trypan
blue dye exclusion in triplicate assays. BOSC23 cells, an ecotropic
retrovirus packaging cell line, were purchased from the American Type
Culture Collection and cultured in Dulbecco's modified Eagle's medium
containing 10% fetal calf serum.
Enforced expression of genes of interest in Baf-3 cells.
Stable transfectants of truncated forms of the human
granulocyta-macrophage colony-stimulating factor (GM-CSF) receptor and dexamethasone (Dex)-inducible Ras mutants were established in Baf-3
cells, as described previously (30, 40). Zn-inducible cells were generated by previously published methods (24).
Transfected gene products were induced by the addition of
ZnSO4 in culture medium at 100 µM unless otherwise
specified in the figure legends. The BimELns mutant, which expresses
BimEL alone, was made by replacing a nucleotide in the splicing donor
site for BimL mRNA without amino acid replacement. Transfectants were
maintained in medium containing either 1 mg of G418 per ml or 200 µg
of hygromycin per ml. For retrovirus-mediated gene expression, we
constructed a control CD8-expressing vector plasmid (pMX/IRES-CD8) from
the pMX retroviral vector (a gift of T. Kitamura) (36) by
inserting an IRES-CD8 cassette in which the mouse CD8 cDNA was fused in frame to the internal ribosomal entry site (IRES) sequence. Particular genes were expressed by inserting their cDNAs immediately after the 5'
long terminal repeat sequence. A dominant negative mutant of Akt
(MAA-phosphokinase B [PKB]) was made by including the K179M, T308A,
and S473A substitutions as described by Wang et al. (46). Retrovirus was made by the method of Onishi et al. (36)
using BOSC23 cells. Retroviral infection of Baf-3 cells and the
selection of CD8-positive cells with a CD8 monoclonal antibody and MACS separation columns (Miltenyi Biotec) were performed by a method described previously (31). The selection procedure was
repeated until more than 95% cells were positive for CD8 by flow cytometry.
Immunoprecipitation and immunoblot analysis.
For immunoblot
analysis, cells were solubilized in Nonidet P-40 (NP-40) lysis buffer
(150 mM NaCl, 1.0% NP-40, 50 mM Tris [pH 8.0]) containing protease
inhibitor mixture (Complete; Roche Molecular Biochemicals); total
cellular proteins were separated by sodium dodecyl
sulfate-polyacrylanide gel electrophoresis (SDS-PAGE). For detecting
the phosphorylation of MAPK, a phosphatase inhibitor mixture (50 mM
sodium fluoride, 10 mM sodium pyrophosphate, 2 mM sodium orthovanadate)
was added to the lysis buffer. Cell lysates extracted from
106 living cells were applied per lane unless otherwise
specified in the figure legends. After their wet electrotransfer onto
polyvinylidene difluoride membranes, the proteins were detected with
appropriate antibodies following standard procedures. The blots were
then stained with primary antibodies followed by horseradish
peroxidase-conjugated anti-rabbit or anti-mouse immunoglobulin
secondary antibodies and subjected to chemiluminescence detection as
specified by the manufacturer (Amersham). Metabolic labeling and
immunoprecipitation were performed by previously described standard
methods (49).
Phosphatase treatment of Bim.
Bim was immunoprecipitated
from 107 Baf-3 cells overexpressing BimEL and BimL
proteins. The protein A-Sepharose-coated beads-Bim complexes were
resuspended in 50 µl of the reaction buffer (100 mM NaCl, 50 mM Tris,
10 mM MgCl2, 1 mM dithiothreitol [pH 7.9] at 25°C)
containing protease inhibitor mixture. After 5 U of calf intestinal
alkaline phosphatase (New England Biolabs, Beverly, Mass.) were added,
the samples were incubated at 37°C for 30 min. Some samples contained
the phosphatase inhibitor mixture described above. The protein
A-Sepharose-coated beads were pelleted by centrifugation, washed three
times with NP-40 lysis buffer, resuspended in gel-loading buffer, and
examined by immunoblot analysis.
Immunocomplex kinase assay.
Akt was immunoprecipitated with
Akt antibody (New England Biolabs) from 107 parental Baf-3
cells or cells expressing a dominant negative form of Akt. The protein
A-Sepharose-coated beads-Akt complexes were resuspended in 50 µl of
reaction buffer (200 mM HEPES-NaOH, 100 mM MgCl2, 100 mM
MnCl2 [pH 7.4]) containing 100 µg of histone H2B (Roche
Molecular Biochemicals) per ml, 5 µM ribosomal ATP, and 10 µCi of
[
-32P]ATP. After incubation of the mixture for 30 min
at room temperature, gel-loading buffer was added to the beads, which
were then boiled for 5 min. Samples were separated on SDS-15%
polyacrylamide gels and viewed by autoradiography.
Analysis of mRNA expression.
Total cellular RNA was isolated
with the RNeasy kit as specified by the manufacturer (Qiagen, Hildes,
Germany). The RNase protection assay was performed with a commercial
kit (Ambion, Austin, Tex.) as specified by the, manufacturer. Briefly,
RNA samples (10 µg each) were hybridized with an
32S-UTP
S-labeled RNA probe, which protects a 265-bp
fragment in mouse BimEL and BimL mRNA and a 210-bp fragment in BimS
mRNA. After single-stranded RNA was digested by RNase A, samples were separated on a 5% acrylamide-8 M urea gel and viewed by autoradiography.
Reagents.
Bim polyclonal antibodies were raised against
glutathione S-transferase fusion proteins containing amino
acids 9 to 53 of mouse BimL as previously described (22).
mTOR and AU-1 monoclonal antibodies were described previously
(17). Bcl-x and Bcl-2 polyclonal antibodies were purchased
from Transduction Laboratories (Lexington, Ky.), while Akt and
phosphorylated (Ser473) Akt-specific antibodies were purchased from New
England Biolabs. Etoposide, wortmannin, and rapamycin were purchased
from Sigma (St. Louis, Mo).
| |
RESULTS |
Bim is induced in cytokine-deprived murine IL-3-dependent
cells.
We first analyzed the expression of Bcl-2 superfamily
members in Baf-3 murine IL-3-dependent pro-B-lymphoid cells in the
presence or absence of IL-3. As previously reported by us and others
(29, 31, 32), the expression levels of Bcl-xL
were downregulated in IL-3-starved Baf-3 cells (Fig.
1A, top panel). In addition, when cells
were cultured in IL-3-containing medium, Bim mRNA was barely detectable
by the RNase protection assay but its expression was rapidly induced by
IL-3 starvation (Fig. 1B, top panel). By immunoblot analysis, we
detected a small amount of BimEL and BimL but no BimS in cells cultured
in the presence of IL-3, while increased amounts of all three Bim
proteins were observed within 24 h in cells cultured in the absence of
IL-3 (Fig. 1C, top panel). BimEL proteins expressed in cells cultured
in IL-3-containing medium seemed to migrate with several additional
bands, indicating that BimEL could be phosphorylated by IL-3-signaling
pathways. Bim was similarly induced by IL-3 withdrawal in FL5.12
pro-B-lymphoid and 32D myeloid cells (data not shown).
View larger version (41K):
[in this window]
[in a new window]
|
FIG. 1.
Expression of Bcl-xL and Bim in Baf-3 cells
undergoing apoptosis induced by various death stimuli. (A to C) Baf-3
cells were cultured in IL-3-free medium (top panel), treated with 50 Gy
of ionizing radiation (IR) and cultured with IL-3 (middle panel), or
cultured in medium containing IL-3 and etoposide (VP16) at 3 µg/ml
(bottom panel) for the indicated times. The expression of
Bcl-xL protein (A), Bim mRNA (B), or Bim protein (C) was
detected by specific antibodies for each protein (A and C) or by an
RNase protection assay with a cDNA probe designed to detect mouse BimEL
and BimL mRNA (B). (D) Percent viability, determined by trypan blue dye
exclusion of Baf-3 cells treated with the various cell death inducers
at the time of sample collection.
|
|
To determine whether the simultaneous upregulation of Bim and
downregulation of Bcl-x
L is specific for cells undergoing
apoptosis
due to IL-3 withdrawal or is a common response of Baf-3 cells
to various apoptotic stimuli, we treated Baf-3 cells in the presence
of
IL-3 with 50 Gy of ionizing radiation or etoposide at 3 µg/ml,
each
of which induces apoptosis in roughly the same time as IL-3
withdrawal
does (
5) (Fig.
1D). This experiment showed that
there was
little change in protein expression levels of Bcl-x
L (Fig.
1A, middle and bottom panels). Although Bim expression was
somewhat
induced by these death stimuli, the induction levels
were not so
prominent as that induced by IL-3 starvation (Fig.
1B and C, middle and
bottom panels), indicating that upregulation
of Bim accompanied by
downregulation of Bcl-x
L appears specific
for cells
undergoing apoptosis induced by cytokine
withdrawal.
Enforced expression of Bim induces apoptosis in Baf-3 cells.
To test whether Bim protein contributes to apoptosis induced by IL-3
starvation, we established Baf-3 cells that expressed each Bim protein
after Zn induction. Because the coding region of BimEL cDNA contains
both the alternative splicing donor and acceptor sites for BimL mRNA,
cells transduced with native BimEL cDNA expressed roughly equal amounts
of BimEL and BimL proteins (see Fig. 3A, lane 3). Thus, before
determining the biological activity of each Bim protein, we mutated the
splice donor site in BimEL cDNA to generate BimEL alone (see Materials
and Methods). As shown in Fig. 2A, we
obtained clones that expressed BimEL, BimL, and BimS protein upon
induction with Zn at 100 µM. Cells strongly expressing any form of
Bim protein rapidly underwent apoptosis, even when the culture medium
contained an excess dose of IL-3 (Fig. 2B). To determine the
antiapoptotic effects of Bim proteins expressed at levels equivalent to
those in IL-3-starved Baf-3 cells, we induced the BimEL protein with
lower concentrations of Zn, 40 or 60 µM (Fig. 2C). Cells cultured
under these conditions still underwent apoptosis in the presence of
IL-3 (Fig. 2D); similar results were obtained with clones expressing
BimL (data not shown). Taken together, these data indicate that
expression levels of Bim associated with IL-3 withdrawal are high
enough to reverse the antiapoptotic effects of IL-3.
View larger version (34K):
[in this window]
[in a new window]
|
FIG. 2.
Induction of apoptosis by each Bim isoform. (A)
Immunoblot analysis of Baf-3 cells engeneered to express BimEL (lanes 1 and 2), BimL (lanes 3 and 4), or BimS (lanes 5 and 6) in the absence
(lanes 1, 3, and 5) or presence (lanes 2, 4, and 6) of zinc (100 µM).
Samples were collected 4 h after the addition of Zn. (B) Percent
viability of cells expressing BimEL, BimL, or BimS after the addition
of Zn (100 µM) in the presence of IL-3 (5 ng/ml). (C) Baf-3 cells
inducibly expressing BimEL, pMT(BimELns clone 10), were cultured in
IL-3-containing medium (5 ng/ml) with various Zn concentrations for
4 h (lanes 1 to 4). For comparison of expression levels, cell
extracts from parental Baf-3 cells cultured in the absence of IL-3 for
the indicated times (lanes 5 to 7) were blotted on the same membrane.
Bim proteins were detected by immunoblot analysis. (D) Percent
viability of pMT(BimELns clone 10) cells cultured in IL-3-containing
medium (5 ng/ml) with various concentration of Zn.
|
|
BimEL and BimL are phosphorylated by IL-3 signaling.
The
additional bands of BimEL observed in the immunoblot analysis of
lysates from Baf-3 cells cultured in IL-3-containing medium (Fig. 1C,
lane 1) raised the possibility that Bim is phosphorylated through IL-3
signaling. Overexposure of immunoblots revealed that not only BimEL,
but also BimL, corresponded to a slowly migrating band in lysates from
cells growing in IL-3-containing medium but not those cultured in
IL-3-free medium (Fig. 3A, lanes 1 and
2). Extracts from cells overexpressing BimEL and BimL proteins from the
pMT-BimEL vector showed a similar pattern of Bim protein expression (lanes 3 and 4). To test whether these additional bands are
phosphorylated Bim proteins, we treated immunoprecipitated Bim proteins
with calf intestine alkaline phosphatase (CIAP). All Bim proteins were detected by Bim antibody but not by preimmune serum in
immunoprecipitates from cells overexpressing BimEL and BimL (Fig. 3B,
lanes 1 and 2). The more slowly migrating bands of BimEL and BimL were
eliminated, apparently by a shift to the faster-migrating bands due to
treatment with CIAP lacking phosphatase inhibitor (lane 3), but were
retained by treatment with CIAP containing phosphatase inhibitor (lane 4), indicating that BimEL and BimL are phosphorylated through IL-3
signaling. To confirm these results, we labeled Baf-3 cells overexpressing BimEL with [32P]orthophosphate for
2 h, with or without IL-3, and immunoprecipitated Bim protein. The
results in Fig. 3C clearly show that BimEL protein was
hyperphosphorylated in the presence of IL-3.
View larger version (21K):
[in this window]
[in a new window]
|
FIG. 3.
Bim phosphorylation. (A) Immunoblot analysis using the
Bim antibody with lysates from parental Baf-3 cells (pt) (lanes 1 and
2) or cells overexpressing BimEL and BimL (lanes 3 and 4) cultured in
the presence (lanes 1 and 3) or absence (lanes 2 and 4) of IL-3. To
demonstrate an extra band comigrating with BimL, we added five times
more cell extract to lane 1 than to the other lanes. (B) Lysates from
Baf-3 cells overexpressing BimEL and BimL proteins were
immunoprecipitated (IP) with preimmune serum (lane 1) or Bim antibody
(lanes 2 to 4). After treatment with CIAP in the absence (lane 3) or
presence (lane 4) of phosphatase inhibitor, precipitants were separated
with SDS-PAGE and Bim proteins were then detected with Bim antibody.
(C) Baf-3 cells overexpressing BimEL protein were cultured in medium
containing [32P]orthophosphate in the presence (lanes 1 and 2) or absence (lane 3) of IL-3 for 4 h. Immunoprecipitates
obtained by preimmune serum (lane 1) or Bim antibody (lanes 2 and 3)
were separated by SDS-PAGE. (D) Lysates from Baf-3 cells overexpressing
BimEL protein were immunoprecipitated with preimmune serum (lane 2) or
antibody against Bim (lane 3), Bcl-2 (lane 4), or Bcl-x (lane 5). After
immunoprecipitation and SDS-PAGE followed by blotting, Bim proteins
were detected with Bim antibody. Lane 1 is a positive control to detect
BimEL protein in whole-cell lysates.
|
|
Because phosphorylated Bad cannot interact with members of the
antiapoptotic Bcl-2 family (
50), we tested whether
phosphorylated
Bim proteins can bind to Bcl-x
L and Bcl-2.
Extracts of Baf-3 cells
inducibly expressing BimEL were
immunoprecipitated with preimmune
serum (Fig.
3D, lane 2), anti-Bim
(lane 3), anti-Bcl-2 (lane 4),
and anti-Bcl-x (lane 5) antibodies, and
the proteins bound to
protein A-Sepharose-coated beads were separated
and blotted. Immunodetection
by Bim antiserum revealed that both the
hyperphosphorylated and
hypophosphorylated forms of BimEL are
coimmunoprecipitated with
Bcl-2 and Bcl-x (lanes 4 and 5). Similar
results were obtained
with extracts from BimL-expressing cells (data
not shown). Thus,
unlike the case with Bad, phosphorylation of Bim does
not affect
the ability of the protein to bind to members of the
antiapoptotic
Bcl-2
family.
Either the Ras/Raf/MAPK or the Ras/PI3-K pathway is sufficient to
downregulate Bim expression.
To identify the signaling protein(s)
through which IL-3 regulates Bim expression, we first used Baf-3 cells
expressing the human c chain truncated at amino acid 544 (544
cells). Stimulation of the receptor with human GM-CSF activates
signaling molecules near the membrane-proximal region of the c
chain, such as JAK2/STAT5, but does not affect components of the Ras
signaling pathways (40).
As expected, the
544 cells grew well in IL-3-containing medium but
underwent rapid apoptosis in the absence of the cytokine.
When cultured
with human GM-CSF, the cells proliferated for approximately
72 h
essentially the same rate as did cells in IL-3-containing
medium with
no apparent loss of viability; thereafter, they underwent
cell cycle
arrest followed by rapid apoptosis, as in earlier studies
(
28,
31) (Fig.
4A). Immunoblot analysis
showed that the expression
levels of Bim remained unaltered for about
12 h, subsequently
reaching levels equivalent to those in
IL-3-starved Baf-3 cells
(Fig.
4B, upper panel). By contrast, as we
previously reported
(
31), Bcl-x
L protein
levels were maintained for more than 5
days before the cells underwent
apoptosis (Fig.
4B, bottom panel).
These results indicate that signals
originating from the membrane-distal
region of the
c chain are
required for the regulation of Bim
expression and hematopoietic cell
survival.
View larger version (45K):
[in this window]
[in a new window]
|
FIG. 4.
Identification of upstream pathways regulating Bim
expression. (A) IL-3 was washed from the culture medium of 544
cells, and the cells were cultured in the presence of human GM-CSF
(hGM-CSF) or murine IL-3 (mIL3) or in the absence of cytokines. The
number of viable cells was determined by trypan blue dye exclusion. (B)
Immunoblot analysis using lysates from 544 cells cultured in human
GM-CSF (lanes 1 to 14) or in the absence of cytokines (lanes 15 and 16)
for the indicated times. Bim (top panel) and Bcl-xL (bottom
panel) proteins were detected by specific antibodies to each protein.
(C) Phosphorylation of p42/MAPK. Parental Baf-3 cells (lanes 1 and 2)
or cells expressing Ras(S17N) induced by the addition of
10 7 M Dex (lanes 3 and 4) were cultured in IL-3-free
medium for 2 h (lanes 1 and 3), after which IL-3 was added at 20 ng/ml for 5 min (lanes 2 and 4). p42/MAPK was detected by immunoblot
analysis using a specific monoclonal antibody. Cell extracts from
105 cells were loaded per lane. (D) Parental Baf-3 cells
(lane 1) or cells containing Dex-inducible Ras(S17N) in the presence or
absence of Dex (lanes 2 and 3) were cultured in IL-3-containing medium.
The phosphorylated form of Akt was detected with antibody specifically
recognizing phosphorylated Akt at Ser473. (E) Parental Baf-3 cells and
cells expressing Ras(S17N) were cultured in the presence or absence of
wortmannin (WMN). The number of viable cells (left panel) and the
percent viability determined by trypan blue dye exclusion (right panel)
are shown. The results are from a representative experiment; similar
results were obtained in three other independent experiments. (F)
Immunoblot analysis with Bim antibodies. Parental Baf-3 cells treated
with wortmannin at 0.5 µM (top panel) or cells expressing Ras(S17N)
induced by addition of 107 M Dex in the absence (middle
panel) or presence (bottom panel) of wortmannin were cultured in
IL-3-containing medium (5 ng/ml). Cell extracts were prepared after the
indicated times.
|
|
To determine the downstream pathways of the distal
c chain that
regulate Bim expression, we used Baf-3 cells conditionally
expressing a
dominant negative Ras protein, Ras(S17N), and the
PI3-K inhibitor
wortmannin. Induction of Ras(S17N) in Baf-3 cells
by Dex abrogated the
activation of MAPK by IL-3 stimulation (Fig.
4C) but did not affect the
activation of PI3-K, as judged by the
phosphorylation of Akt (Fig.
4D).
In agreement with earlier reports
(
35,
45), expression of
Ras(S17N) in Baf-3 cells impeded cell
growth but did not induce
apoptosis (Fig.
4E). Baf-3 cells cultured
in wortmannin at 0.5 µM
also proliferated more slowly, but only
a limited number of cells
underwent apoptosis. However, when Baf-3
cells expressing Ras(S17N)
were cultured in medium containing
wortmannin, they virtually all
underwent apoptosis, confirming
the notion that at least one of the
Ras-activated pathways is
required for cell survival (
28).
Bim expression was downregulated in both Baf-3 cells expressing
Ras(S17N) and those cultured with wortmannin (Fig.
4F, top
and middle
panels). When cells expressing Ras(S17N) were cultured
in wortmannin,
Bim proteins were induced after 48 to 72 h (bottom
panel), a time
comparable to that when Bim protein was induced
in
544 cells
cultured in human GM-CSF (Fig.
4B). These results
indicate that
activation of either the Ras/MAPK or Ras/PI3-K pathway
is sufficient to
suppress Bim
expression.
Involvement of mTOR in cytokine-regulated cell survival.
To
further dissect the Ras/PI3-K signaling pathways, we used Baf-3 cells
conditionally expressing constitutively active Ras mutants induced by
Dex. We and others previously reported (28, 31) that the
Ras(G12V) mutant, when expressed in Baf-3 cells, activates both the
Raf/MAPK and PI3-K pathways, while the Ras(G12V/V45E) mutant activates
the PI3-K but not the Raf/MAPK pathway, as judged by the
phosphorylation of p42/MAPK, Akt, or p70/S6 kinase (Table 1). Baf-3 cells expressing either Ras
mutant survive for prolonged times in IL-3-free medium, while the
antiapoptotic effect of Ras(G12V/V45E) but not that of Ras(G12V) is
reversed by either the PI3-K inhibitor wortmannin or the mTOR inhibitor
rapamycin (Table 1; Fig. 5D)
To test whether Bim expression is involved in the wortmannin- or
rapamycin-sensitive pathways, we performed immunoblot analysis
using
cell lysates from Baf-3 cells that expressed either Ras(G12V)
or
Ras(G12V/V45E), with or without wortmannin or rapamycin, in
IL-3-free
medium. As expected, Bim was downregulated in cells
expressing
Ras(G12V) whether or not wortmannin was present (Fig.
5A). Downregulation of Bim was also
observed in Baf-3 cells expressing
Ras(G12V/V45E) (Fig.
5B, top panel);
however, in contrast to Ras(G12V),
this effect was reversed by
wortmannin or rapamycin (top and lower
middle panels), suggesting that
rapamycin-sensitive pathways (mTOR/S6-K)
play a role in the
antiapoptotic pathways downstream of PI3-K.
View larger version (46K):
[in this window]
[in a new window]
|
FIG. 5.
mTOR participates in the regulation of Bim by PI3-K
pathways. (A and B) Immunoblot analysis with Bim antibodies. Cells were
cultured in IL-3-free medium for the indicated times. Baf-3 cells
containing Dex-inducible Ras(G12V) were cultured with 107
M Dex in the absence (upper panel) or presence (lower panel) of
wortmannin (WMN) (0.5 µM) (A). Baf-3 cells containing Dex-inducible
Ras(G12V/V45E) were cultured with 107 M Dex (top panel),
in the presence of wortmannin (upper middle panel) or rapamycin (Rapa)
(10 ng/ml; lower middle panel); Baf-3 cells containing Dex-inducible
Ras(G12V/V45E) were engeneered to express an AU1-tagged
rapamycin-resistant form of mTOR (AU1-mTOR-rr) (see Materials and
Methods) and were cultured with rapamycin (bottom panel) (B). (C)
Immunoblot analysis of AU1-tagged mTOR-rr and an endogenous mTOR
protein in cell extracts from Baf-3 cells containing Dex-inducible
Ras(G12V/V45E) (V45E; lanes 1 and 3) or expressing AU1-mTOR-rr (rr,
lanes 2 and 4), using monoclonal antibodies specific for AU1 (lanes 1 and 2) and mTOR (lanes 3 and 4). (D) Percent viability of cells
expressing Ras(G12V/V45E), with or without mTOR-rr (rr), in the
presence or absence of rapamycin in IL-3-free medium. Also shown is
that of cells expressing Ras(G12V/V45E) and MAA-PKB in IL-3-free
medium.
|
|
To confirm that mTOR is involved in the regulation of cytokine-mediated
cell survival, we used a mutant form of mTOR that
is resistant to
rapamycin (Ser
2035 
Ile; designated mTOR-rr)
(
7). Retrovirus containing the cDNA
of mTOR-rr, tagged
with the AU1 sequence at its 5' end and murine
CD8 (lyt-2) as a marker,
was transfected to Baf-3 cells inducibly
expressing Ras(G12V/V45E) (see
Materials and Methods). CD8-expressing
cells, selected by magnetic
beads, were demonstrated to express
mTOR-rr by immunoblot analysis with
a monoclonal antibody specific
for the AU1 peptide (Fig.
5C, lane 2),
which comigrated with endogenously
expressed mTOR protein (lane 3).
These cells were then cultured
in IL-3-free, Dex-containing medium,
with or without rapamycin.
As shown in Fig.
5D, cells expressing both
Ras(G12V/V45E) and
mTOR-rr survived in the presence as well as the
absence of rapamycin.
Moreover, Bim was downregulated in these cells
(Fig.
5B, bottom
panel), indicating that mTOR contributes to cell
survival by downregulating
Bim
expression.
Lack of Akt involvement in cytokine-mediated regulation of
apoptosis.
The apparent reversal of the antiapoptotic effects of
Ras(G12V/V45E) by rapamycin raised questions about the contribution of
Akt to downstream regulation of apoptosis through Ras pathways. Indeed,
the phosphorylation levels of Akt in dying cells
[Ras(G12V/V45E)-expressing cells cultured in IL-3-free medium with
rapamycin] (Fig. 6A, lane 6) were
equivalent to those in healthy cells (i.e., those without rapamycin or
parental Baf-3 cells cultured in IL-3-containing medium) (lanes 1 and
5). To directly assess the regulatory role of Akt in Baf-3 cells, we
established cells that expressed myristylated Akt (myr-Akt) after Zn
induction. Although myr-Akt was readily induced with Zn, migrating
slightly slower than endogenous Akt (Fig. 6B, lane 2), its expression
was quickly downregulated when IL-3 was removed. Substitution of a
retroviral long terminal repeat for the metallothionein promoter failed
to stimulate constant expression of myr-Akt in IL-3-starved Baf-3 cells
(data not shown), suggesting that this myristylated protein is
downregulated by posttranscriptional mechanisms. Because rapid
downregulation of myr-Akt was also observed in 32D and FL5.12 cells
(data not shown), we could not determine the antiapoptotic effect of
myr-Akt.
View larger version (20K):
[in this window]
[in a new window]
|
FIG. 6.
(A) Phosphorylation of Akt in Baf-3 cells. Parental (Pt)
Baf-3 cells were cultured in the presence (lane 1) or absence (lane 2)
of IL-3 for 12 h. Baf-3 cells containing Dexinducible
Ras(G12V/V45E) were cultured in IL-3-containing medium without (lane 3)
or with (lane 4) 107 M Dex for 16 h. The Dex-treated
cells were then cultured in IL-3-free, Dex-containing medium for
12 h without wortmannin (WMN) or rapamycin (Rap) (lane 5), with 10 ng/ml rapamycin (lane 6), or with 100 nM wortmannin (lane 7). Akt
phosphorylated at Ser473 (top panel) was detected using specific
antibodies. (B) Expression of total Akt protein in IL-3-starved Baf-3
cells as detected by immunoblot analysis with antibody recognizing both
phosphorylated and nonphosphorylated Akt. Baf-3 cells engineered to
express myr-Akt upon addition of Zn were cultured in IL-3-containing
medium in the absence (lane 1) or presence (lane 2) of 100 µM Zn for
16 h. The cells were then transferred into IL-3-free medium
containing Zn for the indicated times. (C) Akt kinase assay. Parental
cells (Pt) (lanes 1 and 2) and cells expressing a dominant-negative
form of Akt (dn- kt) (lanes 3 and 4) were starved of IL-3 for 2 h (lanes 1 and 3) and then cultured in the presence of IL-3 (20 ng/ml)
for 5 min (lanes 2 and 4). The kinase activity of Akt was determined by
histone H2B phosphorylation in immunoprecipitates using Akt antibody.
(D) Baf-3 cells containing Dex-inducible Ras(G12V/V45E) were
retrovirally engineered to express a dominant negative form of Akt
(MAA-PKB) (dnkt) and then were cultured in IL-3-free, Dex-containing
medium for the indicated times.
|
|
We then used a dominant negative form of Akt (MAA-PKB; see Materials
and Methods) (
46) to investigate the possible role
of Akt
kinase in apoptosis regulation by Ras/PI3-K pathways. Retrovirus
containing the cDNAs of MAA-PKB and CD8 was used to infect Baf-3
cells
inducibly expressing Ras(G12V/V45E). Although selected cells
lacked Akt
kinase activity (Fig.
6C), there were no discernible
differences in
survival between cells expressing Ras(G12V/V45E)
alone and those
expressing MAA-PKB in addition to the Ras mutant
in IL-3-free medium
(Fig.
5D). Moreover, Bim expression was suppressed
in these cells (Fig.
6D), suggesting that Akt is not required
for suppression of this BH3
family member. These results, taken
together, show that Akt does not
appear to contribute to the cytokine-mediated
survival of murine
IL-3-dependent hematopoietic cell
lines.
| |
DISCUSSION |
In earlier studies, we identified two distinct signaling pathways
that appear to regulate cell survival. One originates from the
membrane-proximal portion of the c chain and is involved in the
rapid and stable regulation of Bcl-xL, while the other emanates from the distal region of the c chain and overlaps with the
downstream pathways of constitutively active Ras proteins (31). The first pathway has been described by others
(37), and recently STAT5 was shown to play a pivotal role
in upregulating Bcl-xL through signals from the proximal
domain of the c chain (15, 42, 43). However, such
induction of Bcl-xL is not sufficient to protect cells from
apoptosis (28) (see Fig. 4B, where human GM-CSF was
used to stimulate 544 cells). Because Ras is activated through
signals from the distal c chain (40) and because an oncogenic Ras mutant [Ras(G12V)] rescued 544 cells from apoptosis (28), it appears likely that in addition to
Bcl-xL one or more genes downstream of Ras(G12V) are
required for cell survival. The findings presented here implicate Bim,
a member of the BH3-only subfamily of cell death activators, as a key
target in the regulation of apoptosis by cytokines, acting through Ras
pathways (summarized in Fig. 7).
Importantly Bim expression was downregulated not only through
constitutively active Ras proteins but also through the physiologically
relevant Ras/Raf or Ras/PI3-K pathway (Fig. 4E).
View larger version (22K):
[in this window]
[in a new window]
|
FIG. 7.
Proposed pathways for the cytokine regulation of Baf-3
cell survival. Signals arising from the membrane-proximal portion of
the c chain contribute to the rapid and stable expression of
Bcl-xL through activation of STAT5 but downregulate Bim
only transiently. By contrast, signals from the membrane-distal domain
of the c chain activate the P13-K/mTOR and Raf/MAPK pathways through
Ras stimulation. Either pathway can downregulate Bim expression. When
suppressed, Bim cannot inhibit the antiapoptotic functions of Bcl-2 and
Bcl-xL and thus contributes to the cytokine-mediated
survival of Baf-3 cells.
|
|
The biological significance of Bim expression in hematopoietic cells
was demonstrated by several experiments. First, in the presence of
cytokine, mRNA and protein expression of Bim was very low in the murine
IL-3-dependent cells used in this study, which included both lymphoid
(Baf-3 and FL5.12) and myeloid (32D) cells. Second, the induction of
Bim protein in Baf-3 cells uniformly correlated with cell fate (Fig. 4
to 6). Third, enforced expression of Bim at levels equivalent to those
in IL-3-deprived Baf-3 cells induced apoptosis in the presence of IL-3
(Fig. 2C and D), suggesting that Bim might be able to induce cell death
by itself. The expression level-dependent apoptosis was also noted in
studies with Bim-deficient mice: that is, lymphocytes isolated from
Bim/ mice were much more resistant to apoptosis than
were those from wild-type mice, while cells from Bim+/
mice showed intermediate resistance (4).
There are at least two other mechanisms by which Bim could be regulated
by cytokines. One is phosphorylation. Like expression level-dependent
regulation, signals from the distal portion of the c chain are
required for stable phosphorylation of the Bim protein. Signals from
the proximal c chain can also phosphorylate Bim protein, but only
for a limited time (Fig. 4B). The function of Bad, another member of
the BH3-only family of death activators, is regulated through the
phosphorylation of two serine residues, which causes the loss of its
capacity to bind to antiapoptotic Bcl-2 family members, such as Bcl-2
or Bcl-xL (12, 13, 50). Unlike Bad, however,
hyperphosphorylated forms of Bim also bind to Bcl-2 or
Bcl-xL as avidly as hypophosphorylated forms do (Fig. 3D).
We cannot exclude the possibility that the phosphorylation of Bim
affects its function through mechanisms other than the regulation of
binding to members of the antiapoptotic Bcl-2 family. Even so, it is
likely to be a relatively minor mechanism, at least in Baf-3 cells,
because enforced expression of BimEL and BimL proteins in Baf-3 cells
induced rapid apoptosis in the presence of IL-3, even though both Bim
proteins were hyperphosphorylated by IL-3 signaling (Fig. 2 and 3C).
Regulation of the subcellular localization of Bim protein by cytokines
is another mechanism. Puthalakath et al. reported (38) that Bim protein forms a complex with an Mr
8,000 dynein light chain, LC8 (11, 25, 27), which
functions as a component of the dynein motor complex. In the presence
of IL-3, the Bim-LC8 complex binds to the intermediate chain of the
dynein motor complex on the microtubules, physically separated from
antiapoptotic Bcl-2 family members on the surface of mitochondria. IL-3
withdrawal dissociates the Bim-LC8 complex from the dynein intermediate
chain by undetermined mechanisms, enabling Bim to bind to antiapoptotic Bcl-2 family members and inhibit their function. Expression levels of
LC8 in Baf-3 or 32D cells were very low; indeed, the LC8 mRNA in these
cells was not detectable by Northern blot analysis using a unique
probe, despite the fact that high-level expression was readily detected
in tissues and organs with the same probe (our unpublished data). In
addition, as discussed above, enforced (but not excessive) expression
of Bim induces apoptosis in Baf-3 cells, even in cultures containing
excess concentrations of IL-3 (Fig. 2C and D). All evidence considered,
we prefer Bim expression levels to phosphorylation or subcellular
localization as the means by which cytokines regulate apoptosis in
these particular cell systems we used. Further investigation is needed
to substantiate this interpretation with native hematopoietic progenitors.
The biological significance of PI3-K and the lack of significance of
Raf/MAPK pathways in cell survival were established by using nerve
growth factor to stimulate rat pheochromocytoma PC-12 cells
(47). Akt, a downstream kinase of PI3-K, was then shown to
be involved in the PI3-K-mediated survival of cerebellum granular cells
(14). In these cells, a dominant negative form of Akt blocks the prosurvival effects of insulin-like growth factor I, while a
constitutively active form of Akt promotes cell survival without
neurotropic factors. Moreover, rapamycin does not affect cell survival,
indicating that Akt (not mTOR) plays the major role in the regulation
of apoptosis in neuronal cells. By contrast, both Raf/MAPK and PI3-K
can protect hematopoietic progenitors from apoptosis (28,
30), while the role of Akt in this system has been
controversial. Although several reports indicate a marginal capacity of
Akt to reverse apoptosis due to cytokine withdrawal (2,
44), accumulating evidence refutes a contribution of this
protein to cell survival (3, 16, 41). Our data not only
support these previous negative findings but also suggest that mTOR is
a key downstream mediator of hematopoietic progenitor cell survival, a
role supported by recent observations on mTOR in human rhabdomyosarcoma
cells (17). In light of the above observations, it is not
surprising that Bad phosphorylation, which occurs downstream of Akt,
does not play an important role in the cytokine-mediated survival of
hematopoietic cells (16).
Aberrant control of apoptosis in hematopoietic progenitors may
predispose the cells to leukemic conversion. Several chimeric gene
products formed by nonrandom chromosomal translocations, such as the
BCR-ABL tyrosine kinase implicated in chronic myelogenous leukemia
(CML) and the E2A-HLF transcription factor in adolescent acute
lymphoblastic leukemia (ALL) (19, 21) can block apoptosis due to cytokine withdrawal when expressed in Baf-3 cells (9, 10,
23). In the present study, Bim expression was not affected by
the E2A-HLF chimera in Baf-3 cells (data not shown); however, it was
suppressed in Baf-3 cells expressing BCR-ABL kinase and in
BCR-ABL-positive human leukemia cell lines established from ALL or CML
patients (our unpublished results). Indeed, the accumulation of mature
hematopoietic cells, a prominent feature of CML, is also seen in
Bim-deficient mice (4), suggesting that Bim downregulation might be involved in the pathogenesis of CML. Whether Bim contributes to leukemogenesis induced by BCR-ABL or oncogenic Ras mutants is
currently under investigation in our laboratory.
| |
ACKNOWLEDGMENTS |
We are indebted to T. Kitamura for providing the pMX retrovirus
vector and to John Gilbert for editorial review.
This research was supported by Grants-in-Aid from the Ministry of
Education, Science and Culture of Japan, by the Uehara Memorial Foundation and the Japan Leukemia Research Fund.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Department of
Molecular Oncology, Research Institute for Radiation Biology and
Medicine, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima
734-8553, Japan. Phone: 81-82-257-5834. Fax: 81-82-256-7103.
| |
REFERENCES |
| 1.
|
Adams, J. M., and S. Cory.
1998.
The Bcl-2 protein family: arbiters of cell survival.
Science
281:1322-1326[Abstract/Free Full Text].
|
| 2.
|
Ahmed, N. N.,
H. L. Grimes,
A. Bellacosa,
T. O. Chan, and P. N. Tsichlis.
1997.
Transduction of interleukin-2 antiapoptotic and proliferative signals via Akt protein kinase.
Proc. Natl. Acad. Sci. USA
94:3627-3632[Abstract/Free Full Text].
|
| 3.
|
Bao, H.,
S. M. Jacobs-Helber,
A. E. Lawson,
K. Penta,
A. Wickrema, and S. T. Sawyer.
1999.
Protein kinase B (c-Akt), phosphatidylinositol 3-kinase, and STAT5 are activated by erythropoietin (EPO) in HCD57 erythroid cells but are constitutively active in an EPO-independent, apoptosis-resistant subclone (HCD57-SREI cells).
Blood
93:3757-3773[Abstract/Free Full Text].
|
| 4.
|
Bouillet, P.,
D. Metcalf,
D. C. S. Huang,
D. M. Tarlinton,
T. W. H. Kay,
F. Koentgen,
J. M. Adams, and A. Strasser.
1999.
Proapoptotic Bcl-2 relative Bim required for certain apoptotic responses, leukocyte homeostasis, and to preclude autoimmunity.
Science
286:1735-1738[Abstract/Free Full Text].
|
| 5.
|
Britos-Bray, M.,
M. Ramirez,
W. Cao,
X. Wang,
P. P. Liu,
C. I. Civin, and A. D. Friedman.
1998.
CBF-SMMHC, expressed in M4eo acute myeloid leukemia, reduces p53 induction and slows apoptosis in hematopoietic cells exposed to DNA-damaging agents.
Blood
92:4344-4352[Abstract/Free Full Text].
|
| 6.
|
Chao, D. T., and S. J. Korsmeyer.
1998.
BCL-2 family: regulators of cell death.
Annu. Rev. Immunol.
16:395-419[CrossRef][Medline].
|
| 7.
|
Chen, J,
X. F. Zheng,
E. J. Brown, and S. L. Schreiber.
1995.
Identification of an 11-kDa FKBP12-rapamycin-binding domain within the 289-kDa FKBP12-rapamycin-associated protein and characterization of a critical serine residue.
Proc. Natl. Acad. Sci. USA
92:4947-4951[Abstract/Free Full Text].
|
| 8.
|
Conradt, B., and H. R. Horvitz.
1998.
The C. elegans protein EGL-1 is required for programmed cell death and interacts with the Bcl-2-like protein CED-9.
Cell
93:519-529[CrossRef][Medline].
|
| 9.
|
Cortez, D.,
L. Kadlec, and A. M. Pendergast.
1995.
Structural and signaling requirements for BCR-ABL-mediated transformation and inhibition of apoptosis.
Mol. Cell. Biol.
15:5531-5541[Abstract].
|
| 10.
|
Cortez, D.,
G. Stoica,
J. H. Pierce, and A. M. Pendergast.
1996.
The BCR-ABL tyrosine kinase inhibits apoptosis by activating a Ras-dependent signaling pathway.
Oncogene
13:2589-2594[Medline].
|
| 11.
|
Crépieux, P.,
H. Kwon,
N. Leclerc,
W. Spencer,
S. Richard,
R. Lin, and J. Hiscott.
1997.
I B physically interacts with a cytoskeleton-associated protein through its signal response domain.
Mol. Cell. Biol.
17:7375-7385[Abstract].
|
| 12.
|
Datta, S. R.,
H. Dudek,
X. Tao,
S. Masters,
H. Fu,
Y. Gotoh, and M. E. Greenberg.
1997.
Akt phosphorylation of BAD couples survival signals to the cell-intrinsic death machinery.
Cell
91:231-241[CrossRef][Medline].
|
| 13.
|
del Peso, L.,
M. Gonzalez-Garcia,
C. Page,
R. Herrera, and G. Nunez.
1997.
Interleukin-3-induced phosphorylation of BAD through the protein kinase Akt.
Science
278:687-689[Abstract/Free Full Text].
|
| 14.
|
Dudek, H.,
S. R. Datta,
T. F. Franke,
M. J. Birnbaum,
R. Yao,
G. M. Cooper,
R. A. Segal,
D. R. Kaplan, and M. E. Greenberg.
1997.
Regulation of neuronal survival by the serine-threonine protein kinase Akt.
Science
275:661-665[Abstract/Free Full Text].
|
| 15.
|
Dumon, S.,
S. C. Santos,
F. Debierre-Grockiego,
V. Gouilleux-Gruart,
L. Cocault,
C. Boucheron,
P. Mollat,
S. Gisselbrecht, and F. Gouilleux.
1999.
IL-3 dependent regulation of Bcl-xL gene expression by STAT5 in a bone marrow derived cell line.
Oncogene
18:4191-4199[CrossRef][Medline].
|
| 16.
|
Hinton, H. J., and M. J. Welham.
1999.
Cytokine-induced protein kinase B activation and Bad phosphorylation do not correlate with cell survival of hemopoietic cells.
J. Immunol.
162:7002-7009[Abstract/Free Full Text].
|
| 17.
|
Hosoi, H.,
M. B. Dilling,
T. Shikata,
L. N. Liu,
L. Shu,
R. A. Ashmun,
G. S. Germain,
R. T. Abraham, and P. J. Houghton.
1999.
Rapamycin causes poorly reversible inhibition of mTOR and induces p53-independent apoptosis in human rhabdomyosarcoma cells.
Cancer Res.
59:886-894[Abstract/Free Full Text].
|
| 18.
|
Hsu, S. Y.,
P. Lin, and A. J. Hsueh.
1998.
BOD (Bcl-2-related ovarian death gene) is an ovarian BH3 domain-containing proapoptotic Bcl-2 protein capable of dimerization with diverse antiapoptotic Bcl-2 members.
Mol. Endocrinol.
12:1432-1440[Abstract/Free Full Text].
|
| 19.
|
Hunger, S. P.,
K. Ohyashiki,
K. Toyama, and M. L. Cleary.
1992.
HLF, a novel hepatic bZIP protein, shows altered DNA-binding properties following fusion to E2A in t(17; 19) acute lymphoblastic leukemia.
Genes Dev.
6:1608-1620[Abstract/Free Full Text].
|
| 20.
|
Ikushima, S.,
T. Inukai,
T. Inaba,
S. D. Nimer,
J. L. Cleveland, and A. T. Look.
1997.
Pivotal role for the NFIL3/E4BP4 transcription factor in IL-3-mediated survival of pro-B lymphocytes.
Proc. Natl. Acad. Sci. USA
94:2609-2614[Abstract/Free Full Text].
|
| 21.
|
Inaba, T.,
W. M. Roberts,
L. H. Shapiro,
K. W. Jolly,
S. C. Raimondi,
S. D. Smith, and A. T. Look.
1992.
Fusion of the leucine zipper gene HLF to the E2A gene in human acute B-lineage leukemia.
Science
257:531-534[Abstract/Free Full Text].
|
| 22.
|
Inaba, T.,
L. H. Shapiro,
T. Funabiki,
A. E. Sinclair,
B. G. Jones,
R. A. Ashmun, and A. T. Look.
1994.
DNA-binding specificity and trans-activating potential of the leukemia-associated E2A-HLF fusion protein.
Mol. Cell. Biol.
14:3403-3413[Abstract/Free Full Text].
|
| 23.
|
Inaba, T.,
T. Inukai,
T. Yoshihara,
H. Seyschab,
R. A. Ashmun,
C. E. Canman,
S. J. Laken,
M. B. Kastan, and A. T. Look.
1996.
Reversal of apoptosis by the leukemia-associated E2A-HLF chimeric transcription factor.
Nature
382:541-544[CrossRef][Medline].
|
| 24.
|
Inukai, T.,
T. Inaba,
S. Ikushima, and A. T. Look.
1998.
The AD1 and AD2 transactivation domains of E2A are essential for the antiapoptotic activity of the E2A-HLF chimeric oncoprotein.
Mol. Cell. Biol.
18:6035-6043[Abstract/Free Full Text].
|
| 25.
|
Jaffrey, S. R., and S. H. Snyder.
1996.
PIN: an associated protein inhibitor of neuronal nitric oxide synthase.
Science
274:774-777[Abstract/Free Full Text].
|
| 26.
|
Joneson, T., and D. Bar Sagi.
1997.
Ras effectors and their role in mitogenesis and oncogenesis.
J. Mol. Med.
75:587-593[CrossRef][Medline].
|
| 27.
|
King, S. M.,
E. Barbarese,
J. F. Dillman III,
R. S. Patel-King,
H. Carson, and K. K. Pfister.
1996.
Brain cytoplasmic and flagellar outer arm dyneins share a highly conserved Mr 8,000 light chain.
J. Biol. Chem.
271:19358-19366[Abstract/Free Full Text].
|
| 28.
|
Kinoshita, T.,
T. Yokota,
K. Arai, and A. Miyajima.
1995.
Suppression of apoptotic death in hematopoietic cells by signaling through the IL-3/GM-CSF receptors
EMBO J.
14:266-275[Medline].
|
| 29.
|
Kinoshita, T.,
T. Yokota,
K. Arai, and A. Miyajima.
1995.
Regulation of Bcl-2 expression by oncogenic Ras protein in hematopoietic cells.
Oncogene
10:2207-2212[Medline].
|
| 30.
|
Kinoshita, T.,
M. Shirouzu,
A. Kamiya,
K. Hashimoto,
S. Yokoyama, and A. Miyajima.
1997.
Raf/MAPK and rapamycin-sensitive pathways mediate the anti-apoptotic function of p21Ras in IL-3-dependent hematopoietic cells.
Oncogene
15:619-627[CrossRef][Medline].
|
| 31.
|
Kuribara, R.,
T. Kinoshita,
A. Miyajima,
T. Shinjyo,
T. Yoshihara,
T. Inukai,
K. Ozawa,
A. T. Look, and T. Inaba.
1999.
Two distinct interleukin-3-mediated signal pathways, Ras-NFIL3(E4BP4) and Bcl-xL, regulate the survival of murine pro-B lymphocytes.
Mol. Cell. Biol.
19:2754-2762[Abstract/Free Full Text].
|
| 32.
|
Leverrier, Y.,
J. Thomas,
G. R. Perkins,
M. Mangeney,
M. K. L. Collins, and J. Marvel.
1997.
In bone marrow derived Baf-3 cells, inhibition of apoptosis by IL-3 is mediated by two independent pathways.
Oncogene
14:425-430[CrossRef][Medline].
|
| 33.
|
Miyajima, A.,
Y. Ito, and T. Kinoshita.
1999.
Cytokine signaling for proliferation, survival, and death in hematopoietic cells.
Int. J. Hematol.
69:137-146[Medline].
|
| 34.
|
O'Connor, L.,
A. Strasser,
L. A. O'Reilly,
G. Hausmann,
J. M. Adams,
S. Cory, and D. C. Huang.
1998.
Bim: a novel member of the Bcl-2 family that promotes apoptosis.
EMBO J.
17:384-395[CrossRef][Medline].
|
| 35.
|
Okuda, K.,
T. J. Ernst, and J. D. Griffin.
1994.
Inhibition of p21ras activation blocks proliferation but not differentiation of interleukin-3-dependent myeloid cells.
J. Biol. Chem.
269:24602-24607[Abstract/Free Full Text].
|
| 36.
|
Onishi, M.,
S. Kinoshita,
Y. Morikawa,
A. Shibuya,
J. Phillips,
L. L. Lanier,
D. M. Gorman,
G. P. Nolan,
A. Miyajima, and T. Kitamura.
1996.
Application of retrovirus-mediated expression cloning.
Exp. Hematol.
24:324-329[Medline].
|
| 37.
|
Packham, G.,
E. L. White,
C. M. Eischen,
H. Yang,
E. Parganas,
J. N. Ihle,
D. A. Grillot,
G. P. Zambetti,
G. Nunez, and J. L. Cleveland.
1998.
Selective regulation of Bcl-XL by a Jak kinase-dependent pathway is bypassed in murine hematopoietic malignancies.
Genes Dev.
12:2475-2487[Abstract/Free Full Text].
|
| 38.
|
Puthalakath, H.,
D. C. Huang,
L. A. O'Reilly,
S. M. King, and A. Strasser.
1999.
The proapoptotic activity of the Bcl-2 family member Bim is regulated by interaction with the dynein motor complex.
Mol. Cell
3:287-296[CrossRef][Medline].
|
| 39.
|
Ross, A. J.,
K. G. Waymire,
J. E. Moss,
A. F. Parlow,
M. K. Skinner,
L. D. Russell, and G. R. MacGregor.
1998.
Testicular degeneration in Bclw-deficient mice.
Nat. Genet.
18:251-256[CrossRef][Medline].
|
| 40.
|
Sato, N.,
K. Sakamaki,
N. Terada,
K. Arai, and A. Miyajima.
1993.
Signal transduction by the high-affinity GM-CSF receptor: two distinct cytoplasmic regions of the common subunit responsible for different signaling.
EMBO J.
12:4181-4189[Medline].
|
| 41.
|
Scheid, M. P., and V. Duronio.
1998.
Dissociation of cytokine-induced phosphorylation of Bad and activation of PKB/akt: involvement of MEK upstream of Bad phosphorylation.
Proc. Natl. Acad. Sci. USA
95:7439-7444[Abstract/Free Full Text].
|
| 42.
|
Silva, M.,
A. Benito,
C. Sanz,
F. Prosper,
D. Ekhterae,
G. Nunez, and J. L. Fernandez-Luna.
1999.
Erythropoietin can induce the expression of bcl-x(L) through Stat5 in erythropoietin-dependent progenitor cell lines.
J. Biol. Chem.
274:22165-22169[Abstract/Free Full Text].
|
| 43.
|
Socolovsky, M.,
A. E. Fallon,
S. Wang,
C. Brugnara, and H. F. Lodish.
1999.
Fetal anemia and apoptosis of red cell progenitors in Stat5a-/-5b-/- mice: a direct role for Stat5 in Bcl-X(L) induction.
Cell
98:181-191[CrossRef][Medline].
|
| 44.
|
Songyang, Z.,
D. Baltimore,
L. C. Cantley,
D. R. Kaplan, and T. F. Franke.
1997.
Interleukin 3-dependent survival by the Akt protein kinase.
Proc. Natl. Acad. Sci. USA
94:11345-11350[Abstract/Free Full Text].
|
| 45.
|
Terada, K.,
Y. Kaziro, and T. Satoh.
1995.
Ras is not required for the interleukin 3-induced proliferation of a mouse pro-B cell line, Baf-3.
J. Biol. Chem.
270:27880-27886[Abstract/Free Full Text].
|
| 46.
|
Wang, Q.,
R. Somwar,
P. J. Bilan,
Z. Liu,
J. Jin,
J. R. Woodgett, and A. Klip.
1999.
Protein kinase B/Akt participates in GLUT4 translocation by insulin in L6 myoblasts.
Mol. Cell. Biol.
19:4008-4018[Abstract/Free Full Text].
|
| 47.
|
Yao, R., and G. M. Cooper.
1995.
Requirement for phosphatidylinositol-3 kinase in the prevention of apoptosis by nerve growth factor.
Science
267:2003-2006[Abstract/Free Full Text].
|
| 48.
|
Yin, X. M.,
K. Wang,
A. Gross,
Y. Zhao,
S. Zinkel,
B. Klocke,
K. A. Roth, and S. J. Korsmeyer.
1999.
Bid-deficient mice are resistant to Fas-induced hepatocellular apoptosis.
Nature
400:886-891[CrossRef][Medline].
|
| 49.
|
Yoshihara, T.,
T. Inaba,
L. H. Shapiro,
J. Kato, and A. T. Look.
1995.
E2A-HLF-mediated cell transformation requires both the trans-activation domains of E2A and the leucine zipper dimerization domain of HLF.
Mol. Cell. Biol.
15:3247-3255[Abstract].
|
| 50.
|
Zha, J.,
H. Harada,
E. Yang,
J. Jockel, and S. J. Korsmeyer.
1996.
Serine phosphorylation of death agonist Bad in response to survival factor results in binding to 14-3-3 not Bcl-xL.
Cell
87:619-628[CrossRef][Medline].
|
Molecular and Cellular Biology, February 2001, p. 854-864, Vol. 21, No. 3
0270-7306/01/$04.00+0 DOI: 10.1128/MCB.21.3.854-864.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
This article has been cited by other articles:
-
Sharma, S., Lichtenstein, A.
(2008). Dexamethasone-induced apoptotic mechanisms in myeloma cells investigated by analysis of mutant glucocorticoid receptors. Blood
112: 1338-1345
[Abstract]
[Full Text]
-
Zhang, L., Zambon, A. C., Vranizan, K., Pothula, K., Conklin, B. R., Insel, P. A.
(2008). Gene Expression Signatures of cAMP/Protein Kinase A (PKA)-promoted, Mitochondrial-dependent Apoptosis: COMPARATIVE ANALYSIS OF WILD-TYPE AND cAMP-DEATHLESS S49 LYMPHOMA CELLS. J. Biol. Chem.
283: 4304-4313
[Abstract]
[Full Text]
-
Woodland, R. T., Fox, C. J., Schmidt, M. R., Hammerman, P. S., Opferman, J. T., Korsmeyer, S. J., Hilbert, D. M., Thompson, C. B.
(2008). Multiple signaling pathways promote B lymphocyte stimulator dependent B-cell growth and survival. Blood
111: 750-760
[Abstract]
[Full Text]
-
Deng, J., Shimamura, T., Perera, S., Carlson, N. E., Cai, D., Shapiro, G. I., Wong, K.-K., Letai, A.
(2007). Proapoptotic BH3-Only BCL-2 Family Protein BIM Connects Death Signaling from Epidermal Growth Factor Receptor Inhibition to the Mitochondrion. Cancer Res.
67: 11867-11875
[Abstract]
[Full Text]
-
Pei, X.-Y., Dai, Y., Tenorio, S., Lu, J., Harada, H., Dent, P., Grant, S.
(2007). MEK1/2 inhibitors potentiate UCN-01 lethality in human multiple myeloma cells through a Bim-dependent mechanism. Blood
110: 2092-2101
[Abstract]
[Full Text]
-
Hacker, G., Suttner, K., Harada, H., Kirschnek, S.
(2006). TLR-dependent Bim phosphorylation in macrophages is mediated by ERK and is connected to proteasomal degradation of the protein. Int Immunol
18: 1749-1757
[Abstract]
[Full Text]
-
McCarty, M. F., Block, K. I.
(2006). Preadministration of High-Dose Salicylates, Suppressors of NF-{kappa}B Activation, May Increase the Chemosensitivity of Many Cancers: An Example of Proapoptotic Signal Modulation Therapy. Integr Cancer Ther
5: 252-268
[Abstract]
-
Yano, T., Ito, K., Fukamachi, H., Chi, X.-Z., Wee, H.-J., Inoue, K.-i., Ida, H., Bouillet, P., Strasser, A., Bae, S.-C., Ito, Y.
(2006). The RUNX3 Tumor Suppressor Upregulates Bim in Gastric Epithelial Cells Undergoing Transforming Growth Factor {beta}-Induced Apoptosis. Mol. Cell. Biol.
26: 4474-4488
[Abstract]
[Full Text]
-
Meller, R., Cameron, J. A., Torrey, D. J., Clayton, C. E., Ordonez, A. N., Henshall, D. C., Minami, M., Schindler, C. K., Saugstad, J. A., Simon, R. P.
(2006). Rapid Degradation of Bim by the Ubiquitin-Proteasome Pathway Mediates Short-term Ischemic Tolerance in Cultured Neurons. J. Biol. Chem.
281: 7429-7436
[Abstract]
[Full Text]
-
Haneline, L. S., White, H., Yang, F.-C., Chen, S., Orschell, C., Kapur, R., Ingram, D. A.
(2006). Genetic reduction of class IA PI-3 kinase activity alters fetal hematopoiesis and competitive repopulating ability of hematopoietic stem cells in vivo. Blood
107: 1375-1382
[Abstract]
[Full Text]
-
Qi, X.-J., Wildey, G. M., Howe, P. H.
(2006). Evidence That Ser87 of BimEL Is Phosphorylated by Akt and Regulates BimEL Apoptotic Function. J. Biol. Chem.
281: 813-823
[Abstract]
[Full Text]
-
Zhao, Y., Tan, J., Zhuang, L., Jiang, X., Liu, E. T., Yu, Q.
(2005). Inhibitors of histone deacetylases target the Rb-E2F1 pathway for apoptosis induction through activation of proapoptotic protein Bim. Proc. Natl. Acad. Sci. USA
102: 16090-16095
[Abstract]
[Full Text]
-
Aichberger, K. J., Mayerhofer, M., Krauth, M.-T., Vales, A., Kondo, R., Derdak, S., Pickl, W. F., Selzer, E., Deininger, M., Druker, B. J., Sillaber, C., Esterbauer, H., Valent, P.
(2005). Low-Level Expression of Proapoptotic Bcl-2-Interacting Mediator in Leukemic Cells in Patients with Chronic Myeloid Leukemia: Role of BCR/ABL, Characterization of Underlying Signaling Pathways, and Reexpression by Novel Pharmacologic Compounds. Cancer Res.
65: 9436-9444
[Abstract]
[Full Text]
-
Austin, M., Cook, S. J.
(2005). Increased Expression of Mcl-1 Is Required for Protection against Serum Starvation in Phosphatase and Tensin Homologue on Chromosome 10 Null Mouse Embryonic Fibroblasts, but Repression of Bim Is Favored in Human Glioblastomas. J. Biol. Chem.
280: 33280-33288
[Abstract]
[Full Text]
-
Clybouw, C., Mchichi, B., Mouhamad, S., Auffredou, M. T., Bourgeade, M. F., Sharma, S., Leca, G., Vazquez, A.
(2005). EBV Infection of Human B Lymphocytes Leads to Down-Regulation of Bim Expression: Relationship to Resistance to Apoptosis. J. Immunol.
175: 2968-2973
[Abstract]
[Full Text]
-
Moller, C., Alfredsson, J., Engstrom, M., Wootz, H., Xiang, Z., Lennartsson, J., Jonsson, J.-I., Nilsson, G.
(2005). Stem cell factor promotes mast cell survival via inactivation of FOXO3a-mediated transcriptional induction and MEK-regulated phosphorylation of the proapoptotic protein Bim. Blood
106: 1330-1336
[Abstract]
[Full Text]
-
Nyunoya, T., Monick, M. M., Powers, L. S., Yarovinsky, T. O., Hunninghake, G. W.
(2005). Macrophages Survive Hyperoxia via Prolonged ERK Activation Due to Phosphatase Down-regulation. J. Biol. Chem.
280: 26295-26302
[Abstract]
[Full Text]
-
Reginato, M. J., Mills, K. R., Becker, E. B. E., Lynch, D. K., Bonni, A., Muthuswamy, S. K., Brugge, J. S.
(2005). Bim Regulation of Lumen Formation in Cultured Mammary Epithelial Acini Is Targeted by Oncogenes. Mol. Cell. Biol.
25: 4591-4601
[Abstract]
[Full Text]
-
Luciano, F., Zhai, D., Zhu, X., Bailly-Maitre, B., Ricci, J.-E., Satterthwait, A. C., Reed, J. C.
(2005). Cytoprotective Peptide Humanin Binds and Inhibits Proapoptotic Bcl-2/Bax Family Protein BimEL. J. Biol. Chem.
280: 15825-15835
[Abstract]
[Full Text]
-
Abrams, M. T., Robertson, N. M., Yoon, K., Wickstrom, E.
(2004). Inhibition of Glucocorticoid-induced Apoptosis by Targeting the Major Splice Variants of BIM mRNA with Small Interfering RNA and Short Hairpin RNA. J. Biol. Chem.
279: 55809-55817
[Abstract]
[Full Text]
-
Harada, H., Quearry, B., Ruiz-Vela, A., Korsmeyer, S. J.
(2004). Survival factor-induced extracellular signal-regulated kinase phosphorylates BIM, inhibiting its association with BAX and proapoptotic activity. Proc. Natl. Acad. Sci. USA
101: 15313-15317
[Abstract]
[Full Text]
-
Becker, E. B. E., Howell, J., Kodama, Y., Barker, P. A., Bonni, A.
(2004). Characterization of the c-Jun N-Terminal Kinase-BimEL Signaling Pathway in Neuronal Apoptosis. J. Neurosci.
24: 8762-8770
[Abstract]
[Full Text]
-
Fukazawa, H., Noguchi, K., Masumi, A., Murakami, Y., Uehara, Y.
(2004). BimEL is an important determinant for induction of anoikis sensitivity by mitogen-activated protein/extracellular signal-regulated kinase kinase inhibitors. Molecular Cancer Therapeutics
3: 1281-1288
[Abstract]
[Full Text]
-
Andoniou, C. E., Andrews, D. M., Manzur, M., Ricciardi-Castagnoli, P., Degli-Esposti, M. A.
(2004). A novel checkpoint in the Bcl-2-regulated apoptotic pathway revealed by murine cytomegalovirus infection of dendritic cells. JCB
166: 827-837
[Abstract]
[Full Text]
-
Valentijn, A. J., Gilmore, A. P.
(2004). Translocation of Full-length Bid to Mitochondria during Anoikis. J. Biol. Chem.
279: 32848-32857
[Abstract]
[Full Text]
-
Kuribara, R., Honda, H., Matsui, H., Shinjyo, T., Inukai, T., Sugita, K., Nakazawa, S., Hirai, H., Ozawa, K., Inaba, T.
(2004). Roles of Bim in Apoptosis of Normal and Bcr-Abl-Expressing Hematopoietic Progenitors. Mol. Cell. Biol.
24: 6172-6183
[Abstract]
[Full Text]
-
Zhang, L., Insel, P. A.
(2004). The Pro-apoptotic Protein Bim Is a Convergence Point for cAMP/Protein Kinase A- and Glucocorticoid-promoted Apoptosis of Lymphoid Cells. J. Biol. Chem.
279: 20858-20865
[Abstract]
[Full Text]
-
del Carmen Rodriguez, M., Bernad, A., Aracil, M.
(2004). Interleukin-6 deficiency affects bone marrow stromal precursors, resulting in defective hematopoietic support. Blood
103: 3349-3354
[Abstract]
[Full Text]
-
Matsunaga, T., Inaba, T., Matsui, H., Okuya, M., Miyajima, A., Inukai, T., Funabiki, T., Endo, M., Look, A. T., Kurosawa, H.
(2004). Regulation of annexin II by cytokine-initiated signaling pathways and E2A-HLF oncoprotein. Blood
103: 3185-3191
[Abstract]
[Full Text]
-
Ley, R., Ewings, K. E., Hadfield, K., Howes, E., Balmanno, K., Cook, S. J.
(2004). Extracellular Signal-regulated Kinases 1/2 Are Serum-stimulated "BimEL Kinases" That Bind to the BH3-only Protein BimEL Causing Its Phosphorylation and Turnover. J. Biol. Chem.
279: 8837-8847
[Abstract]
[Full Text]
-
Sandalova, E., Wei, C.-H., Masucci, M. G., Levitsky, V.
(2004). Regulation of expression of Bcl-2 protein family member Bim by T cell receptor triggering. Proc. Natl. Acad. Sci. USA
101: 3011-3016
[Abstract]
[Full Text]
-
Mouhamad, S., Besnault, L., Auffredou, M. T., Leprince, C., Bourgeade, M. F., Leca, G., Vazquez, A.
(2004). B Cell Receptor-Mediated Apoptosis of Human Lymphocytes Is Associated with a New Regulatory Pathway of Bim Isoform Expression. J. Immunol.
172: 2084-2091
[Abstract]
[Full Text]
-
Pellegrini, M., Belz, G., Bouillet, P., Strasser, A.
(2003). Shutdown of an acute T cell immune response to viral infection is mediated by the proapoptotic Bcl-2 homology 3-only protein Bim. Proc. Natl. Acad. Sci. USA
100: 14175-14180
[Abstract]
[Full Text]
-
Minami, Y., Yamamoto, K., Kiyoi, H., Ueda, R., Saito, H., Naoe, T.
(2003). Different antiapoptotic pathways between wild-type and mutated FLT3: insights into therapeutic targets in leukemia. Blood
102: 2969-2975
[Abstract]
[Full Text]
-
White, E.
(2003). The pims and outs of survival signaling: role for the Pim-2 protein kinase in the suppression of apoptosis by cytokines. Genes Dev.
17: 1813-1816
[Full Text]
-
Wang, Z., Malone, M. H., He, H., McColl, K. S., Distelhorst, C. W.
(2003). Microarray Analysis Uncovers the Induction of the Proapoptotic BH3-only Protein Bim in Multiple Models of Glucocorticoid-induced Apoptosis. J. Biol. Chem.
278: 23861-23867
[Abstract]
[Full Text]
-
Ley, R., Balmanno, K., Hadfield, K., Weston, C., Cook, S. J.
(2003). Activation of the ERK1/2 Signaling Pathway Promotes Phosphorylation and Proteasome-dependent Degradation of the BH3-only Protein, Bim. J. Biol. Chem.
278: 18811-18816
[Abstract]
[Full Text]
-
Wildey, G. M., Patil, S., Howe, P. H.
(2003). Smad3 Potentiates Transforming Growth Factor beta (TGFbeta )-induced Apoptosis and Expression of the BH3-only Protein Bim in WEHI 231 B Lymphocytes. J. Biol. Chem.
278: 18069-18077
[Abstract]
[Full Text]
-
Tepera, S. B., McCrea, P. D., Rosen, J. M.
(2003). A {beta}-catenin survival signal is required for normal lobular development in the mammary gland. J. Cell Sci.
116: 1137-1149
[Abstract]
[Full Text]
-
Lei, K., Davis, R. J.
(2003). JNK phosphorylation of Bim-related members of the Bcl2 family induces Bax-dependent apoptosis. Proc. Natl. Acad. Sci. USA
100: 2432-2437
[Abstract]
[Full Text]
-
Komatsu, N., Watanabe, T., Uchida, M., Mori, M., Kirito, K., Kikuchi, S., Liu, Q., Tauchi, T., Miyazawa, K., Endo, H., Nagai, T., Ozawa, K.
(2003). A Member of Forkhead Transcription Factor FKHRL1 Is a Downstream Effector of STI571-induced Cell Cycle Arrest in BCR-ABL-expressing Cells. J. Biol. Chem.
278: 6411-6419
[Abstract]
[Full Text]
-
Biswas, S. C., Greene, L. A.
(2002). Nerve Growth Factor (NGF) Down-regulates the Bcl-2 Homology 3 (BH3) Domain-only Protein Bim and Suppresses Its Proapoptotic Activity by Phosphorylation. J. Biol. Chem.
277: 49511-49516
[Abstract]
[Full Text]
-
Yamaguchi, H., Wang, H.-G.
(2002). Bcl-XL Protects BimEL-induced Bax Conformational Change and Cytochrome c Release Independent of Interacting with Bax or BimEL. J. Biol. Chem.
277: 41604-41612
[Abstract]
[Full Text]
-
Stahl, M., Dijkers, P. F., Kops, G. J. P. L., Lens, S. M. A., Coffer, P. J., Burgering, B. M. T., Medema, R. H.
(2002). The Forkhead Transcription Factor FoxO Regulates Transcription of p27Kip1 and Bim in Response to IL-2. J. Immunol.
168: 5024-5031
[Abstract]
[Full Text]
-
Li, L., Liu, D., Hutt-Fletcher, L., Morgan, A., Masucci, M. G., Levitsky, V.
(2002). Epstein-Barr virus inhibits the development of dendritic cells by promoting apoptosis of their monocyte precursors in the presence of granulocyte macrophage-colony-stimulating factor and interleukin-4. Blood
99: 3725-3734
[Abstract]
[Full Text]
-
Kirito, K., Watanabe, T., Sawada, K.-i., Endo, H., Ozawa, K., Komatsu, N.
(2002). Thrombopoietin Regulates Bcl-xL Gene Expression through Stat5 and Phosphatidylinositol 3-Kinase Activation Pathways. J. Biol. Chem.
277: 8329-8337
[Abstract]
[Full Text]
-
Claessens, Y.-E., Bouscary, D., Dupont, J.-M., Picard, F., Melle, J., Gisselbrecht, S., Lacombe, C., Dreyfus, F., Mayeux, P., Fontenay-Roupie, M.
(2002). In vitro proliferation and differentiation of erythroid progenitors from patients with myelodysplastic syndromes: evidence for Fas-dependent apoptosis. Blood
99: 1594-1601
[Abstract]
[Full Text]
-
Dijkers, P. F., Birkenkamp, K. U., Lam, E. W.-F., Thomas, N. S. B., Lammers, J.-W. J., Koenderman, L., Coffer, P. J.
(2002). FKHR-L1 can act as a critical effector of cell death induced by cytokine withdrawal: protein kinase B-enhanced cell survival through maintenance of mitochondrial integrity. JCB
156: 531-542
[Abstract]
[Full Text]
-
Wang, X.
(2001). The expanding role of mitochondria in apoptosis. Genes Dev.
15: 2922-2933
[Full Text]
-
Castedo, M., Ferri, K. F., Blanco, J., Roumier, T., Larochette, N., Barretina, J., Amendola, A., Nardacci, R., Metivier, D., Este, J. A., Piacentini, M., Kroemer, G.
(2001). Human Immunodeficiency Virus 1 Envelope Glycoprotein Complex-Induced Apoptosis Involves Mammalian Target of Rapamycin/Fkbp12-Rapamycin-Associated Protein-Mediated P53 Phosphorylation. JEM
194: 1097-1110
[Abstract]
[Full Text]
-
Han, J.-w., Flemington, C., Houghton, A. B., Gu, Z., Zambetti, G. P., Lutz, R. J., Zhu, L., Chittenden, T.
(2001). Expression of bbc3, a pro-apoptotic BH3-only gene, is regulated by diverse cell death and survival signals. Proc. Natl. Acad. Sci. USA
98: 11318-11323
[Abstract]
[Full Text]
-
Dijkers, P. F., Birkenkamp, K. U., Lam, E. W.-F., Thomas, N. S. B., Lammers, J.-W. J., Koenderman, L., Coffer, P. J.
(2002). FKHR-L1 can act as a critical effector of cell death induced by cytokine withdrawal: protein kinase B-enhanced cell survival through maintenance of mitochondrial integrity. JCB
156: 531-542
[Abstract]
[Full Text]
Top
Abstract
Introduction
