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Department of Internal Medicine and Interdisciplinary Program in Immunology, University of Iowa College of Medicine, Iowa City, IA 52242
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The mitogen-activated protein kinase (MAPK) family of enzymes has been implicated in the transduction of a wide variety of extracellular signals, both mitogenic and apoptotic (7, 8, 9, 10). A recent study of the PC12 cell line showed that c-Jun NH2-terminal kinase (JNK) and p38 MAPK enzymes are activated following nerve growth factor withdrawal leading to apoptosis (10). In this same study, activation of the MAPK enzyme extracellular signal-regulated kinase (ERK) was found to be important for cell survival. More recent work using an in vitro transfection system in MCF7 and HeLa cells, however, has demonstrated that TNF receptor-1 (TNFR1)-induced apoptosis does not involve JNK activation, while another study demonstrated that TNFR-mediated apoptosis requires JNK activity in Jurkat T cells (11, 12). The role of JNK in CD95-mediated apoptosis in T cells is controversial as well. Some studies support the importance of this signaling pathway, while others do not (11, 13, 14, 15, 16).
The role of MAPK enzymes in mediating B cell apoptosis is uncertain as well. One study demonstrated a correlation between ERK activity and IgM receptor-mediated cell death in WEHI 231 cells (17). This study showed that CD40 engagement blocks IgM receptor-mediated killing while inducing activation of JNK and p38 MAPK. The authors proposed that ERK activation correlates with cell death and that full activation of all three kinases correlates with cell survival. Another recent study, however, has demonstrated that activation of JNK and p38 correlates with cell death in the human B lymphoma line, B104 (18).
In the present study, we undertook both biochemical and genetic approaches to better understand the roles of ERK2 and JNK in surface IgM-mediated apoptosis. Our data support a correlation between ERK2 activation and apoptosis in WEHI 231 B cells. In contrast, we found that activation of JNK does not appear to play a significant role in IgM-mediated cell death. Furthermore, although CD40 is a potent stimulator of JNK, this signaling pathway may not be required for CD40 to rescue WEHI 231 cells from Ag receptor-stimulated apoptosis.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Hamster anti-mouse CD40 mAb (HM40-3) and hamster anti-mouse CD95 mAb (Jo2) were purchased from PharMingen (San Diego, CA). Goat anti-mouse µ-chain-specific IgM was obtained from Sigma (St. Louis, MO). Mouse anti-ERK1/2 mAb from Zymed (South San Francisco, CA) and rabbit anti-ERK2 Ab (C-14) from Santa Cruz Biotechnology (Santa Cruz, CA) were used. Horseradish peroxidase-conjugated goat anti-mouse Ab was purchased from Bio-Rad (Hercules, CA).
NAC, PMA, glutathione-agarose, and myelin basic protein (MBP) were
purchased from Sigma. GammaBind Plus Sepharose from Pharmacia
(Piscataway, NJ) and D-sorbitol from Fisher Scientific
(Pittsburgh, PA) were used. Propidium iodide (PI), RNase A, and T1 were
obtained from Boehringer Mannheim (Indianapolis, IN).
[
-32P]ATP (10 mCi/ml) and enhanced chemiluminescence
reagents were obtained from Amersham (Arlington Heights, IL). The
Galacto-Light Plus reporter gene assay system was purchased from Tropix
(Bedford, MA). pGEX/GST c-Jun1–79 and pEF/MKP-1 plasmids
were gifts from Dr. Jeffrey Pessin (Iowa City, IA). CMV
ß-galactosidase (CMV-ß-gal) plasmid was provided by Dr. Grant
MacGregor (Atlanta, GA). Mouse rIL-4 was obtained from the National
Cancer Institute (Bethesda, MD).
Cell culture
The mouse B lymphoma cell line WEHI 231 was maintained at 37°C in 5% CO2-95% O2 in RPMI medium supplemented with 10% heat-inactivated FBS, penicillin (1000 U/ml), streptomycin (1000 U/ml), glutamine (20 mM), and 2-ME (5 x 10-5 M). When cells were treated with NAC, they were preincubated at a density of 1 x 106 cells/ml for at least 2 h before stimulation. Experiments were then performed in the continuous presence of NAC.
Nuclei staining and analysis of hypodiploid nuclei
Cells (1 x 105/ml) were left untreated (control) or were stimulated with anti-IgM (1 µg/ml), anti-CD95 (0.2 µg/ml), anti-CD40 (0.1 µg/ml), or rIL-4 (50 U/ml). Anti-IgM was also applied in combination with anti-CD40, anti-CD95, or rIL-4. After 24 or 48 h of incubation, cells were harvested and washed once with HBSS containing 5 mM EDTA. Following fixation with 70% ethanol for 30 min at room temperature, cells were washed once with HBSS, then treated with RNase (100 µg/ml RNase A plus 200 U/ml RNase T1). After a 20-min incubation at room temperature, cell nuclei were stained with PI (1%), and hypodiploid nuclei were determined by FACS analysis using a FACScan and LYSIS II software (Becton Dickinson, Mountain View, CA).
ERK2 activation assay
ERK2 activation following receptor ligation was determined
either by electrophoretic mobility shift resulting from the
phosphorylation of ERK2 or by an in vitro ERK2 enzymatic assay using
MBP as a substrate (19). Cells (5 x 106) were
incubated in medium alone or in medium containing anti-IgM (1
µg/ml), anti-CD95 (2 µg/ml), anti-CD40 (1 µg/ml), rIL-4
(500 U/ml), or PMA (50 nM) at 37°C for various periods of time. Cells
were lysed in 1% Nonidet P-40 lysis buffer containing 150 mM NaCl, 10
mM Tris (pH 7.4), protease inhibitors (50 µg/ml aprotinin, 10 µg/ml
leupeptin, 40 µg/ml pepstatin A, and 1 mM PMSF), and phosphatase
inhibitors (400 µM sodium vanadate, 10 mM sodium fluoride, and 10 mM
sodium pyrophosphate). For the ERK2 mobility shift assay, lysates from
1 x 106 cells were mixed with 2x Laemmli’s sample
buffer, boiled, and subjected to SDS-12% PAGE. After transferring
proteins to a nitrocellulose membrane, Western blot was performed by
blocking the membrane with 5% nonfat dried milk and incubating with
anti-ERK1/2 mAb, followed by horseradish peroxidase-conjugated
secondary Ab; detection was conducted with enhanced chemiluminescence
reagents. Quantitation of phosphorylated ERK2 was performed by scanning
the slower migrating band, followed by analyzing the intensity with
National Institutes of Health Image/Gel Plotting software. For ERK2
enzymatic activity, lysates from 5 x 106 cells were
immunoprecipitated for 2 h with rabbit anti-ERK2 Ab conjugated
to GammaBind Plus Sepharose. The kinase activities of the
immunoprecipitates were assessed using [-32P]ATP and
MBP as a substrate and were visualized by SDS-15% PAGE and
autoradiography as previously described (20).
Transient expression of MKP-1 and ß-gal reporter gene assay
Cells were washed twice with PBS and resuspended at 10 x 106 cells in 400 µl of cytomix (21). The indicated amount of vector (pEF), MKP-1 (pEF/MKP-1), or CMV-ß-gal plasmid DNA was added, and electroporation was performed with a Bio-Rad gene pulser at 960 µFd and 270 mV. Transfected cells were cultured in growth medium for 24 h, harvested, and then divided into two aliquots. One aliquot was used to determine the amount of cell-associated ß-gal activity, and the other was used to measure the percentage of cells with hypodiploid nuclei. For the ß-gal reporter gene assay, viable cells (5 x 105/ml) were incubated in medium alone (control) or stimulated with anti-IgM or a combination of anti-IgM plus anti-CD95 for various periods of time. Cell-associated ß-gal activity following each stimulation was determined using a Galacto-Light Plus reporter gene assay system, and luminescence was quantitated with a Monolight 2010 luminometer (San Diego, CA). Cell-associated ß-gal activity in stimulated samples was compared with that in unstimulated cells. The percentage of cell-associated ß-gal activity in stimulated samples was calculated as (cell-associated ß-gal activity in stimulated cells/cell-associated ß-gal activity in unstimulated cells) x 100. We then plotted % ß-gal activity lost (100 - % cell-associated ß-gal activity) against the percentage of cells with hypodiploid nuclei in samples incubated under the same conditions, and linear regression analysis was performed. A similar assay has been described previously (22).
c-Jun kinase assay
JNK activity was assayed in vitro using affinity-purified GST
c-Jun1–79 fusion protein (13). Briefly, cells (5 x
106) were incubated in medium alone or were treated with
anti-IgM (1 µg/ml), anti-CD95 (2 µg/ml), anti-CD40 (1
µg/ml), rIL-4 (500 U/ml), or D-sorbitol (0.6 M) at 37°C
for various times. Cellular lysates were incubated with GST
c-Jun1–79 fusion protein (1 µg) for 4 h at 4°C.
The fusion protein complex was then washed and incubated in 30 µl of
kinase reaction mixture (20 mM HEPES (pH 8.0), 20 mM MgCl2,
20 mM ß-glycerophosphate, 100 µM sodium vanadate, 2 mM DTT, 20 µM
ATP, and 0.3 µCi [-32P]ATP) for 30 min at 25°C.
Reactions were stopped by adding HEPES binding buffer (20 mM HEPES (pH
8.0), 2.5 mM MgCl2, 0.1 mM EDTA, 50 mM NaCl, and 0.05%
Triton X-100). The complexes were washed gently, mixed with 30 µl of
1x Laemmli’s sample buffer, boiled, and then resolved by SDS-10%
PAGE. The gel was stained with Coomassie brilliant blue and dried, and
phosphorylated fusion protein was detected by autoradiography.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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We first examined the functional consequence of ligation of
several surface receptors on WEHI 231 cells. Cells were incubated in
medium alone (negative control) or were treated with Ab against IgM,
CD95, or CD40 or with rIL-4. Anti-IgM was also used in combination with
anti-CD95, anti-CD40, or rIL-4. After 24 or 48 h of
incubation, cells were fixed, and their nuclei were stained with PI.
Hypodiploid nuclei, as an indicator of apoptosis, were then assessed by
FACS analysis. As shown in Figure 1
and
as others have reported for this cell line, anti-IgM causes
significant apoptosis (30–55% at 48 h). Costimulation with
anti-CD95 enhances surface IgM-mediated killing (65 to 85% at
48 h), whereas costimulation with anti-CD40 or rIL-4
interferes markedly with IgM-mediated apoptosis. Stimulation with
anti-CD95, anti-CD40, or rIL-4 alone has no effect on WEHI 231
cell survival.
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We next examined ERK2 activity in cells stimulated as described in
Figure 1
. We assayed ERK2 phosphorylation by measuring a shift in
mobility by SDS-PAGE and quantitated the intensity of the slower
migrating band, which indicates the level of phosphorylated, active
ERK2. In Figure 2A the
intensities of the bands of phosphorylated ERK2 following each
stimulation are compared. As shown, we found that ERK2 is significantly
phosphorylated at 5 min following anti-IgM treatment and is
phosphorylated further at 10 min. The increase in ERK2 phosphorylation
lasts for at least 30 min. Cotreatment with anti-IgM plus
anti-CD95 enhances ERK2 phosphorylation at 5 min over that seen
with anti-IgM alone. This enhancement of ERK2 phosphorylation
persists for the 30-min course of the experiment. In contrast,
cotreatment of anti-IgM with anti-CD40 or rIL-4 decreases ERK2
phosphorylation at all time points studied. Anti-CD40, rIL-4, or
anti-CD95 treatment alone has no effect on ERK2 phosphorylation. A
representative experiment demonstrating the raw data from which the
graph in Figure 2A is derived is shown in Figure 2B.
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C ERK2-mediated phosphorylation of MBP after ligation of
the various cell surface receptors correlates with ERK2 phosphorylation
seen by the shift in mobility (Fig. 2B). PMA-induced ERK2
activation served as a positive control in these experiments. These
data demonstrate that surface IgM ligation leads to the phosphorylation
and activation of ERK2. Additionally, we found that ERK2
phosphorylation and activation are enhanced by costimulation with
anti-CD95 and are reduced by costimulation with anti-CD40 and
rIL-4. These data, when combined with those in Figure 1, indicate that
ERK2 activation correlates with anti-IgM-induced apoptosis in WEHI
231 B cells. These data also suggest the possibility that the
modulatory effects of CD40L and rIL-4 on surface IgM-mediated apoptosis
may be mediated at least in part through effects on IgM-mediated ERK2
activation. Inhibition of ERK2 activity by overexpression of MKP-1 interferes with anti-IgM- or anti-IgM- plus anti-CD95-mediated apoptosis in WEHI 231 B cells
Next, we investigated further whether ERK2 plays an active role in mediating apoptosis following surface IgM ligation. We reasoned that if ERK2 activation was important for this process, specific inactivation of ERK2 should decrease cell death following anti-IgM treatment. We therefore examined the effects of decreasing ERK2 activity on surface IgM-mediated apoptosis. To do this we overexpressed MKP-1, a molecule that dephosphorylates and subsequently reduces ERK2 activity (23).
We first studied the effects of MKP-1 overexpression on ERK2
phosphorylation. WEHI 231 cells were transfected transiently with
various amounts of vector (pEF) or MKP-1 (pEF/MKP-1) plasmid as
indicated in Figure 3. The transfected
cells were treated with anti-IgM or a combination of anti-IgM
plus anti-CD95. ERK2 phosphorylation following IgM engagement was
reduced considerably in cells transfected with MKP-1 in a
dose-dependent fashion at both 5 and 15 min after stimulation (Fig. 3A). Additionally, overexpression of MKP-1 effectively
interferes with ERK2 activation in cells treated with a combination of
anti-CD95 and anti-IgM (Fig. 3B).
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C, there is a positive
correlation between the amount of ß-gal lost in stimulated cells and
percentage of cells with hypodiploid nuclei, probably reflecting loss
of the integrity of the plasma membrane of cells undergoing apoptosis.
After validating the ß-gal loss assay as a measure of apoptosis, we
transfected WEHI 231 cells transiently with pEF or pEF/MKP-1 along with
the CMV-ß-gal plasmid. Twenty to twenty-four hours following
transfection, cells were treated with medium alone (control),
anti-IgM, or a combination of anti-IgM plus anti-CD95. The
amount of ß-gal lost from pEF or pEF/MKP-1 cotransfected cells was
compared 28 to 36 h after treatment. As shown in Figure 3D, ß-gal was lost following treatment of pEF
vector-transfected cells with anti-IgM alone (20%) and
anti-IgM plus anti-CD95 cotreatment (50%) compared with the
amount of ß-gal lost from cells incubated in medium. In contrast, in
MKP-1-transfected cells, treatment with anti-IgM alone resulted in
only a small amount of ß-gal loss. Additionally, cotreatment with
anti-IgM plus anti-CD95 was less effective in inducing ß-gal
loss in the MKP-1 transfectants than in cells transfected with the
vector alone. Although when expressed at high levels, MKP-1 may also
modulate the activity of other members of the MAPK family, these data
support our findings indicating that ERK2 plays an active role in
mediating IgM-induced apoptosis.
JNK activity does not correlate with IgM-mediated apoptosis
We next examined JNK activation in WEHI 231 cells following
incubation of cells in medium alone or in medium containing
anti-IgM, anti-CD95, anti-CD40, and/or rIL-4. JNK enzyme
activity was determined by phosphorylation of a GST
c-Jun1–79 fusion protein as an in vitro substrate. As
shown in Figure 4, we found that a 5-min
incubation of cells with medium alone results in some degree of JNK
activation (slight phosphorylation of a GST c-Jun1–79
fusion protein), which diminishes shortly thereafter (data not shown;
see medium control incubated for 30 min in Fig. 5A). In our hands, treatment
of cells with anti-CD95 or anti-IgM alone or in combination
results in only a minimal increase in JNK activity above that observed
in the control (Fig. 4A). It should be noted that this
differs somewhat from a recent report by Purkerson and Parker (24), who
found an approximately 2-fold increase in JNK activation following
anti-IgM stimulation of WEHI 231 cells. In this study, however, the
investigators used 10-fold more stimulating Ab directed against surface
IgM than we used in our protocol.
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A). This activation
lasts for upward of 15 min following anti-CD40 treatment and is
significantly diminished by 30 min. Cotreatment with anti-IgM
enhances anti-CD40-induced JNK activation (Fig. 4A).
These data are in agreement with previous reports examining WEHI 231
and other cell lines, demonstrating that CD40 is a much more potent
activator of JNK than is anti-IgM (17, 24, 25, 26, 27).
The data in Figure 4B illustrate that rIL-4 does not
increase JNK activity significantly above control levels. Thus, it is
clear that JNK activation is not a major factor in the inhibition of
IgM-mediated apoptosis by rIL-4. However, the enhancement of JNK
activation by IgM plus CD40 suggests the possibility that CD40
engagement may rescue IgM-mediated apoptosis at least in part through
enhanced JNK activity. Therefore, we examined the role of anti-CD40
induced JNK activity in inhibiting surface IgM-induced apoptosis in
WEHI 231 cells.
NAC inhibition of JNK activity does not affect the anti-CD40 blockade of surface IgM-mediated apoptosis
Since anti-CD40 is a potent inducer of JNK activation in WEHI
231 cells, we addressed whether this signaling event is important in
the rescue of cells from anti-IgM-mediated apoptosis. One mechanism
of JNK activation has been linked to the production of reactive oxygen
intermediates by engagement of TNFR family members (28, 29, 30). We
hypothesized that CD40-induced JNK activation may similarly involve
reactive oxygen intermediates second messengers, since CD40 is a member
of the TNFR family. We approached this by asking if pretreatment of
cells with the reducing agent, NAC (which interferes with
anti-CD40-induced JNK activity (J.R.L. and G.A.K., manuscript in
preparation)), would block CD40-mediated inhibition of IgM-induced
apoptosis. In the experiment shown in Figure 5, we incubated WEHI 231
cells with NAC (10 mM) for 2 h before treatment with anti-IgM
and/or anti-CD40. Although at 48 h, NAC treatment results in
decreased anti-IgM-mediated cell death (data not shown) similar to
previous reports in thymocytes and WEHI 231 cells (31, 32, 33), surface
IgM-stimulated apoptosis at 24 h is still seen.
NAC treatment effectively inhibits CD40-induced JNK activation (Fig. 5A). However, CD40 engagement still interferes with the
activation of ERK2 induced by surface IgM cross-linking in NAC-treated
cells (Fig. 5B). Furthermore, we found that anti-CD40
still rescues cells from IgM-mediated death following NAC treatment at
24 h (Fig. 6). These data also
suggest that JNK activation is not a major factor for CD40-mediated
interference of Ag receptor-induced cell death in WEHI 231 B cells.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Many recent studies have implicated members of the MAPK family as regulators of signals leading to apoptosis or cellular growth in various systems, including B cells. Interestingly, MAP kinases appear to be activated differentially depending upon the cell types and ligands studied, making it difficult to define the exact roles of these family members. For example, in some systems activation of JNK leads to cell survival, while in others stimulation of this enzyme correlates with potentiation of programmed cell death (10, 11, 12, 13, 14, 15, 16, 17, 18, 34, 35, 36). Similarly, there is controversy regarding the roles of both ERK and p38 as mediators of cell growth or apoptosis (10, 18). A number of laboratories have examined the role of MAPK family members in B cell signal transduction leading to both activation and cell death. Again, there is considerable disagreement in the literature, with some studies suggesting that surface IgM-induced apoptosis depends on activation of ERK, while other studies have shown that cell death depends on JNK stimulation (17, 18, 24, 25, 26).
In this study we focused on the potential roles of ERK2 and JNK as modulators of apoptotic signals following surface IgM engagement on WEHI 231 B cells in the presence and the absence of costimulation. Our results support previous reports that ERK2 activation is an important signal leading to IgM-mediated apoptosis in these cells, since modulation of ERK2 activity by costimulation via various other receptors correlates with the degree of cell survival. We also provide several lines of evidence to suggest that in contrast to ERK2, activation of JNK does not appear to play a crucial role in modulating surface IgM-mediated apoptosis in WEHI 231 cells. First, we found that although rIL-4 is a potent inhibitor of IgM-stimulated apoptosis, engagement of the IL-4R does not lead to significant activation of JNK. Similarly, although CD40 ligation results in both a rescue from apoptosis and JNK activation, we found that interference of CD40-mediated JNK stimulation with NAC does not inhibit the ability of CD40 to block anti-IgM-mediated WEHI 231 cell death. Interestingly, both CD40 and IL-4R engagement lead to inhibition of IgM-stimulated ERK2 activation. Together, these findings suggest that the rescue from apoptosis is more likely a function of interference with ERK2 activation than a function of JNK stimulation, although it is possible that low levels of residual JNK activity may be involved in protection from apoptosis.
We next took a genetic approach to address the causal role of ERK2 activation in surface IgM-induced apoptosis. We took advantage of the phosphatase, MKP-1, which dephosphorylates ERK2, leading to termination of this signaling pathway (23). Although in our transient transfection system, we were not able to completely interfere with surface IgM- and/or CD95-mediated ERK2 activation, we were able to substantially decrease this signaling event. The decrease in ERK2 activation correlates well with inhibition of apoptosis in the transfected cells, as shown by the ß-gal release assay. Although MKP-1 may affect other MAPK pathways, our findings are consistent with the correlation of ERK2 activation with Ag receptor-mediated apoptosis.
It should be noted that our results differ somewhat from those reported previously. Sutherland et al. (17) showed that costimulation of WEHI 231 cells with both anti-IgM and anti-CD40 resulted in ERK2 activation indistinguishable from that seen with anti-IgM alone. Additionally, this group as well as Purkerson and Parker (24) found that stimulation with anti-IgM alone resulted in a small degree of JNK activation. One possible explanation for the differences in our results is that we used a 10- to 20-fold lower dose of anti-IgM for our experiments. Furthermore, the focus of these other studies was on the biochemical consequences of receptor ligation and did not extend to evaluation of cell survival. Additionally, in contrast to our results, Graves et al. (18) found that surface IgM stimulation results in activation of JNK. One difference between their study and ours is that Graves et al. examined the B104 human B cell lymphoma line. A second difference between the studies is that Graves et al. examined much later time points than we did following receptor ligation.
The activation of ERK and JNK appears to be a convergence point for
signal transduction events following engagement of multiple surface
receptors on WEHI 231 B cells. It is likely that substrates of these
kinases play important roles in determining whether membrane proximal
events will result in cellular activation or death (37, 38, 39, 40, 41, 42). In support
of this, recent studies demonstrated that surface IgM cross-linking on
WEHI 231 increases the activity of I
B-
, a specific NF-B/Rel
inhibitor (43, 44, 45, 46, 47, 48). One postulated means by which CD40 engagement
rescues these cells from apoptosis is by inducing NF-B/Rel family
members and maintaining c-myc RNA and protein levels
(43, 44, 45, 46, 47, 48). Several studies have also shown in WEHI 231 cells that
expression levels of Bcl-xL, an antiapoptotic member of the Bcl-2
family, are regulated differentially following either surface IgM or
CD40 stimulation (33, 49, 50, 51, 52, 53). In these experiments, surface IgM
cross-linking reduces the level of Bcl-xL, while CD40 treatment
up-regulates the expression of this protein. Although it is not yet
clear whether receptor-activated MAPK family enzymes play direct roles
in activation of transcription factors leading to regulation of
proteins involved in apoptosis, it is likely that ERK and JNK affect
signaling cascades that are important in cell fate decisions. Current
studies are underway to identify the downstream effector molecules of
MAPK family enzymes in B lymphocytes.
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Acknowledgments |
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Footnotes |
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2 Established Investigator with the American Heart Association.
3 Address correspondence and reprints requests to Dr. Gary A. Koretzky, 540 EMRB, Department of Internal Medicine, University of Iowa, Iowa City, IA 52242. E-mail address:
4 Abbreviations used in this paper: CD40L, CD40 ligand; MAPK, mitogen-activated protein kinase; JNK, c-Jun NH2-terminal kinase; ERK, extracellular signal-regulated kinase; TNFR1, TNF receptor-1; NAC, N-acetyl-L-cysteine; MBP, myelin basic protein; PI, propidium iodide; GST, glutathione-S-transferase; ß-gal, ß-galactosidase; MKP-1, mitogen-activated protein kinase phosphatase-1.
Received for publication January 22, 1998. Accepted for publication April 9, 1998.
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B activation prevents cell death. Cell 87:565.[Medline]
B/Rel induces apoptosis of murine B cells. EMBO J. 15:4682.[Medline]
B proteins. J. Immunol. 157:81.[Abstract]
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E. Takada, H. Toyota, J. Suzuki, and J. Mizuguchi Prevention of Anti-IgM-Induced Apoptosis Accompanying G1 Arrest in B Lymphoma Cells Overexpressing Dominant-Negative Mutant Form of c-Jun N-Terminal Kinase 1 J. Immunol., February 1, 2001; 166(3): 1641 - 1649. [Abstract] [Full Text] [PDF]
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C. K. Sen, S. Khanna, S. Roy, and L. Packer Molecular Basis of Vitamin E Action. TOCOTRIENOL POTENTLY INHIBITS GLUTAMATE-INDUCED pp60c-Src KINASE ACTIVATION AND DEATH OF HT4 NEURONAL CELLS J. Biol. Chem., April 21, 2000; 275(17): 13049 - 13055. [Abstract] [Full Text] [PDF]
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J. M. Swart, D. M. Bergeron, and T. C. Chiles Identification of a Membrane Ig-Induced p38 Mitogen-Activated Protein Kinase Module That Regulates cAMP Response Element Binding Protein Phosphorylation and Transcriptional Activation in CH31 B Cell Lymphomas J. Immunol., March 1, 2000; 164(5): 2311 - 2319. [Abstract] [Full Text] [PDF]
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M.-Y. Cao, F. Shinjo, S. Heinrichs, J.-W. Soh, J. Jongstra-Bilen, and J. Jongstra Inhibition of Anti-IgM-induced Translocation of Protein Kinase C beta I Inhibits ERK2 Activation and Increases Apoptosis J. Biol. Chem., June 29, 2001; 276(27): 24506 - 24510. [Abstract] [Full Text] [PDF]
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