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Department of Physiology, University of Connecticut Health Center, Farmington, CT 06030
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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increases cPLA2 phosphorylation, which is inhibited by SB
203580, but not PD 98059, suggesting a role for p38 MAP kinase. LPS
increases cPLA2 phosphorylation and arachidonic acid
release. However, neither of these actions is inhibited by either PD
98059 or SB 203580. PMA increases cPLA2 phosphorylation.
This action is inhibited by PD 98059 but not SB 203580. Finally, FMLP
increases cPLA2 phosphorylation and arachidonic acid
release. Interestingly, while the FMLP-induced phosphorylation of
cPLA2 is not affected by the inhibitors of the p38 MAP
kinase or erk cascades, both inhibitors significantly decrease
arachidonic acid release stimulated by FMLP. SB 203580 or PD 98059 has
no inhibitory effects on the activity of coenzyme A-independent
transacylase. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Until recently, the mitogen-activated protein (MAP) kinases (MAPK), also referred to as erk1 and erk2, were the only cloned and well-characterized mammalian MAPKs. Recently, two other MAPK subtypes, the c-Jun N-terminal kinase (JNK) stress-activated protein kinase (SAPK) and p38/reactivating kinase (mammalian equivalent of high osmolality glycerol response-1 (HOG-1) in yeast) were discovered 17, 18, 19, 20, 21 . Currently, there are three parallel kinase cascades involved in agonist-induced signal transduction: the erk1/2, the JNK/SAPK, and the p38. These three kinases are themselves activated by phosphorylation on threonine and tyrosine residues, and the upstream kinases that can phosphorylate these enzymes are MAP/erk kinase (MEK) (threonine glutamic acid tyrosine kinase), SAP/erk kinase (SEK) (threonine proline tyrosine kinase), and reactivating kinase kinase (TGY kinase), respectively 17, 18, 19 . In proliferating cells, substrates for the erk cascade include ribosomal S6 kinase (p90rsk), and transcription factors, such as c-jun and c-fos 17, 18, 19, 20, 21 . The substrates for the JNK/SAPK and p38 cascades in proliferating cells are just beginning to be identified. The substrates and the roles of all three MAPK subtypes in differentiated cells such as human neutrophils are unknown.
Recently, we and others demonstrated the presence of a novel 38-kDa
protein that is tyrosine phosphorylated and activated in human
neutrophils and terminally differentiated cells upon stimulation with
various agonists 12, 22, 23 . This 38-kDa protein was identified as
the mammalian homologue of HOG-1 in yeast, the p38 MAPK. The
present studies were undertaken to determine the roles of various
kinases in the phosphorylation of cPLA2 and the release of
arachidonic acid in human neutrophils stimulated by FMLP, PAF, LPS,
PMA, and TNF-. This was done using three recently developed
compounds; SB 203580, a highly specific inhibitor of the p38 MAPK
activity, which does not affect erk2 or Jun kinase subtypes; PD 98059,
a specific inhibitor of the erk cascade; and lastly, GO 6850
(bisindolylmaleimide), a highly specific inhibitor of protein kinase C
(PKC) 24, 25, 26 .
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Blood was obtained from human volunteers, and neutrophils were isolated using a Ficoll/Hypaque gradient. Contaminating RBC were lysed by hypotonic shock 27, 28 . The neutrophils were resuspended in modified HBSS containing 0.1% BSA and 10 mM HEPES, pH 7.35.
Fractionation of cells into soluble and particulate fractions
Fractionation of cells into soluble and particulate fractions was conducted as described previously 29, 30 . Briefly, 3 ml of cells (1 x 107 cells/ml) were treated with inhibitor or diluent for 30 min followed by the addition of buffer or agonist. Following reaction termination, the cells were resuspended at 108 cells/ml in iced buffered sucrose solution (10 mM HEPES, pH 7.5, 100 mM sucrose, 1 mM EGTA, 0.5 mM EDTA, 50 µg/ml leupeptin, 2 mM sodium orthovanadate, 5 mM sodium pyrophosphate, and 100 µM PMSF). Cells were disrupted by sonication, then centrifuged at 100,000 x g for 30 min at 4°C. The cytosolic fraction protein was measured, adjusted, and used in subsequent assays.
Immunoblotting
Immunoblotting was performed as described previously 28, 29, 30 . Equivalent amounts of protein (100 µg) were loaded onto SDS-PAGE (8% for cPLA2 detection or 12% for p38 MAPK and erk2 detection) gels. After electrophoresis, proteins were transferred from the gel to polyvinylidene difluoride membranes in transfer buffer (20 mM Tris base, 150 mM glycine, 20% methanol, pH 8.9). Blots were washed in Tris-buffered saline/Tween 20, incubated with the desired Ab, and then probed with the appropriate horseradish peroxidase-conjugated Ab. The enhanced chemiluminescence method was used for detection.
In vitro phosphorylation
The p38 MAPK activity assay was performed using heat shock protein (hsp) 27 as a substrate in an in vitro kinase assay. In vitro phosphorylation of hsp 27 was conducted as described previously 22, 30, 31 . Briefly, 15 µl of soluble fraction containing 2 mg/ml protein prepared from control or stimulated cells was added to 15 µl of assay buffer comprised of 100 mM Tris, pH 7.4, 20 mM MgCi2, 40 mM ATP, 16 mM okadaic acid, 33 mg/ml hsp 27, and 0.24 mCi/ml [32P]ATP. Reactions were conducted for 10 min at 25°C and were stopped by adding 30 µl of 2x SDS-PAGE sample buffer. The samples were boiled for 5 min and electrophoresed on 12% polyacrylamide gels. Phosphorylation of hsp 27 protein was measured under conditions where it was linear with respect to the time of incubation and enzyme dilution.
Arachidonic acid release
Labeling the cells with [3H]arachidonic acid and the release of radiolabeled arachidonic acid were conducted as described previously 32 with slight modifications. Briefly, the cells (1 x 108/ml) were incubated with 2.5 µCi/ml [3H]arachidonic acid (which had been dried under N2 and resuspended in HBSS containing 0.1 mg/ml fatty acid-free BSA) for 45 min at 37°C. The inhibitors SB 203580 (15 µM), PD 98059 (15 µM), or DMSO were added to the radiolabeled cells for 30 min. Cells were then washed three times, resuspended at 5 x 106/ml, and incubated at 37°C for varying times with the appropriate stimulus. The cells were then vortexed, centrifuged, and an aliquot of the supernatant was counted.
Protocol for SB 203580, PD 98059, and GO 6850 treatment and agonist stimulation of human neutrophils
Neutrophils were treated either with diluent, 15 µM SB 203580,
15 µM PD 98059, or 5 µM GO 6850 for 30 min before stimulation by
500 nM PAF (1 min); 100 nM FMLP (2 min); 10 ng/ml TNF- (5 min); 100
ng/ml LPS in combination with 1% serum (30 min); 3–4 nM PMA (5 min);
or 1 mM okadaic acid (1 h). For arachidonic acid release assays, the
concentrations of PAF and FMLP were increased to 1 µM and 500 nM,
respectively, to cause a more substantial effect.
Coenzyme A-independent transacylase (CoA-IT) activity
CoA-IT activity was measured in microsomes isolated from human neutrophils. Microsomes were prepared as previously described by Winkler et al. 33 with minor modifications. Isolated neutrophils from whole human blood were washed with a buffered solution containing 250 mM sucrose, 1 mM EGTA, 1 mM MgCl2, 10 mM TrisHCl, pH 7.4, and sonicated with a small probe three times for 5 s each. The disrupted cells were centrifuged at 18,000 x g for 20 min. Microsomal fraction were pelleted from the supernatant by centrifugation at 170,000 x g for 40 min. The microsomes were washed once with PBS (138 mM NaCl, 10 mM Na2HPO4, 2.7 mM KCl) containing 1 mM EGTA, pH 7.4, pelleted by centrifugation, and resuspended in PBS and stored at -70°C until assayed. CoA-IT activity in microsome preperation was measured as described by Winkler et al. 33 . Briefly, microsomes were diluted in PBS containing 1 mM EGTA, and reaction was started by the addition of 0.1 µCi [[3H]]1-alkyl-2-lyso-GPC/tube and 1 µM final unlabeled 1-alkyl-2-lyso-glycerol-3 phosphorylcholine (GPC) and run for 10 min at 37°C. Lipids were extracted according to the method of Bligh and Dyer 34 , and aliquots of the chloroform phase were separated by TLC. The [3H]1-alkyl-2-lyso-GPC and [3H]1-alkyl-2-acyl-GPC bands were scraped and quantified by liquid scintillation spectroscopy.
Materials
Electrophoresis reagents and m.w. markers were obtained from
Bio-Rad Laboratories (Melville, NY); polyvinylidene fluoride protein
transfer membrane (Immobilon-P) was obtained from Millipore Corporation
(Bedford, MA); enhanced chemiluminescence and other Western blotting
reagents were obtained from Pierce (Rockford, IL); and
[
-32P]ATP, 6000 Ci/mmol, and
[3H]arachidonic acid were purchased from Dupont NEN
(Boston, MA). TNF- the inhibitor SB 203580 and GO 6850 were
purchased from Calbiochem (San Diego, CA). The recently developed
anti-phospho p38 Ab (this Ab does not cross-react with other MAPK
subtypes) and anti-phospho MAPK (which recognizes the
phosphorylated forms of erk1 and erk2) were obtained from Santa Cruz
Biotechnology (Santa Cruz, CA). Anti-cPLA2 Ab was
generously provided by the Immunology Department of the Genetics
Institute (Cambridge MA). PMA and FMLP were purchased from Sigma (St.
Louis, Mo). PD 98059 was purchased from New England Biolabs (Beverly,
MA). The reagent [3H]1-alkyl-2-lyso-GPC (58 Ci/mol) was
purchased from New England Nuclear Life Science Products (Boston MA),
and 1-alkyl-2-lyso-GPC was obtained from Biomol Research Laboratory
(Plymouth, PA).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The effects of SB 203580, the p38 MAPK inhibitor, and PD
98059, the MEK inhibitor, on the agonist-stimulated tyrosine
phosphorylation of the p38 MAPK and erk2 in human neutrophils were
examined. In these experiments, the cells were incubated with diluent,
SB 203580, or PD 98059 and then stimulated with FMLP, PAF, TNF-, or
LPS. After reaction termination, cells were sonicated and cytosolic
fractions subjected to SDS-PAGE. Immunoblot analysis was performed
using anti-phospho-MAPK or anti-phospho-p38 kinase. The results
are summarized in Fig. 1
. These data show
several points. First, PAF and FMLP stimulate the tyrosine
phosphorylation of both erk2 and p38 MAPK. Second, as previously
reported 13, 15, 22 , TNF- and LPS increase the tyrosine
phosphorylation only of p38 MAPK. Third, PD 98059 inhibits the
stimulated phosphorylation of erk2 but has no effect on the
phosphorylation of p38 MAPK. Fourth, SB 203580 does not affect the
phosphorylation of either erk2 or p38 MAPK. The inhibitory action of SB
203580 is on the enzymatic activity of the p38 MAPK so its
phosphorylation is not affected. On the other hand, the inhibitory
effect of PD 98059 on the erk2 cascade is through mek1 inhibition and,
therefore, erk2 tyrosine phosphorylation. Also note that SB 203580
alone causes a slight increase in p38 MAPK tyrosine phosphorylation.
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An important in vivo substrate for p38 MAPK is MAPK/activated
protein kinase-2 (MAPKAPK-2). The activated MAPKAPK-2 increases the
phosphorylation of the small m.w. hsp 27. Accordingly, the activity of
p38 MAPK was determined using the small m.w. hsp 27 as substrate. In
these experiments, designed to show the efficacy of SB 203580, human
neutrophils in suspension were incubated with diluent or SB 203580 and
then stimulated with TNF-, LPS, PAF, or FMLP. Lysates were prepared
and used in an in vivo kinase assay using hsp 27 as substrate as
described in Materials and Methods. The results summarized
in Fig. 2 clearly show that all four
stimuli induce an increase in the phosphorylation of hsp 27, and the
stimuli-induced increases are abolished by the inhibitor SB 203580. To
verify that the mek1 inhibitor PD 98059 has no effect on the p38 MAPK,
we also tested it on cells stimulated by FMLP and PAF. It was found
that PD 98059 has no effect on the FMLP- or PAF-stimulated increase in
the in vitro phosphorylation of hsp 27.
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It is generally agreed that erk2 increases the phosphorylation of
cPLA2 on serine 505 in an in vitro assay 7 . However, it
is not known if the phosphorylation of cPLA2 in intact
cells by physiological stimuli is mediated by erk2. To examine this
point, we measured the phosphorylation of cPLA2 in control
and PD 98059-treated human neutrophils in suspension stimulated by PAF
and FMLP. Both of these stimuli increase the tyrosine phosphorylation
of erk2, and this phosphorylation is inhibited by PD 98059 (see Fig. 1
). In these experiments, the cells were treated with PD 98059 and then
stimulated with PAF or FMLP. After reaction termination, cells were
sonicated and cytosolic fractions subjected to SDS-PAGE. Immunoblot
analysis was performed using anti-cPLA2. The
phosphorylation of cPLA2 is detected by the retarded
mobility of the phosphorylated enzyme. The results summarized in Fig. 3 clearly show that the PAF-induced, but
not FMLP-induced, phosphorylation of cPLA2 is inhibited by
PD 98059. As expected, PD 98059 had no effect on the phosphorylation of
cPLA2 in human neutrophils in suspension stimulated by
TNF- or LPS (data not shown).
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, and
FMLP
Recently, we and others 8, 9, 10, 11, 12 have presented evidence that
indicates that the phosphorylation of cPLA2 can be achieved
independently of erk2. To address this point further, the
phosphorylation of cPLA2 in control and SB 203580-treated
human neutrophils upon stimulation with LPS, PAF, FMLP, or TNF- was
examined. In these experiments, human neutrophils in suspension were
incubated with diluent or SB 203580, and cells were then stimulated
with FMLP, LPS, TNF-, or PAF. Cytosolic fractions were prepared,
electrophoresed, transferred, and the resulting immunoblot probed with
anti-cPLA2 Ab. The data summarized in Fig. 4 clearly show that only the
TNF--induced (Fig. 4A) phosphorylation of
cPLA2 is inhibited in cells pretreated with SB 203580.
Also, the FMLP-stimulated phosphorylation was not affected in cells
pretreated with both SB 203580 and PD 98059 added together (Fig. 4B).
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As shown in the previous sections, the FMLP-induced
phosphorylation of cPLA2 is not inhibited by PD 98059 or SB
203580. This strongly suggests that FMLP can phosphorylate
cPLA2 independently of the activation of erk2 and p38 MAPK.
The chemotactic tripeptide FMLP is known to activate the PKC system and
calcium-activated kinases. To examine the possibility that the PKC
system may phosphorylate cPLA2 directly, we measured the
effects of the PKC inhibitor, GO 6850, on the phosphorylation of
cPLA2 in human neutrophils stimulated by FMLP and PMA, an
activator of the PKC system. The results summarized in Fig. 5 show three main points. First,
stimulation of human neutrophils with PMA induces the phosphorylation
of cPLA2, and this phosphorylation is greatly diminished by
the PKC inhibitor GO 6850 (Fig. 5A). Second, while the PKC
inhibitor decreases the phosphorylation of cPLA2 in cells
stimulated with PMA, it has no effect on the actions of FMLP (Fig. 5A). Third, stimulation of human neutrophils with PMA
increases the tyrosine phosphorylation of erk1 and erk2 (Fig. 5B) and p38 MAPK (Fig. 5C). However, while GO
6850 inhibits the PMA-induced tyrosine phosphorylation of both p38 and
the erk kinases, it has no effect on the tyrosine phosphorylation of
these kinases induced by FMLP. Note that in Fig. 5B we were
able to detect clearly the phosphorylated form of erk1 (p44), which is
stimulated by both PMA and FMLP. Like the erk2 phosphorylation, only
PMA-stimulated phosphorylation is inhibited by GO 6850.
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clearly show that the PMA-induced
phosphorylation of cPLA2 is inhibited by PD 98059 (Fig. 6A). Also note that PMA-induced phosphorylation of
cPLA2 is not affected by SB 203580 (Fig. 6B). On
the other hand, SB 203580 in combination with PD 98059 inhibits this
phosphorylation (data not shown). When examining the action of PD 98059
on PMA-induced effects, it is extremely important to control for
concentrations of PMA. Because of the nature of the inhibitory action
of PD 98059 on the tyrosine phosphorylation of erk1 and 2, it is
possible to overcome this inhibition by using relatively high
concentrations of PMA (data not shown).
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Because cPLA2 has been shown to mediate the production
of agonist-induced arachidonic acid release and phosphorylation of the
enzyme is necessary for its activation 1, 2, 3, 4, 5, 6, 7, 8 , the relationship between
inhibition of cPLA2 phosphorylation and arachidonic acid
release in human neutrophils was examined. To determine this,
neutrophils were treated with SB 203580 or PD 98059, stimulated with
either LPS, TNF-, PAF, FMLP, LPS plus PAF, LPS plus FMLP, TNF-
plus PAF, or TNF- plus FMLP, and arachidonic acid release was
determined as described in Materials and Methods (Table I). The data summarized in this table
reveal two interesting points. First, LPS, which does not induce a rise
in the intracellular concentration of free calcium, causes arachidonic
acid release. This is consistent with the recent observation that PMA
and okadaic acid induce arachidonic acid release without
increasing the intracellular concentration of free calcium 35 in
macrophages. The LPS-induced arachidonic acid release is only, if any,
slightly affected by PD 98059 or SB 203580. Second, and more
interestingly, while neither of these inhibitors affect the
phosphorylation of cPLA2 produced by FMLP, both inhibitors
greatly reduce the release of arachidonic acid stimulated by this
agonist.
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One possible explanation for the observed inhibition of
FMLP-induced arachidonic acid release by SB 203580 is that this agent
inhibits CoA-IT activity. This enzyme selectively remodels arachidonate
between different phospholipids 33 . Consistent with this possibility
is the recent finding that ß-lactams SB 212047 and other similar
compounds are irreversible inhibitors of this enzyme 33 . These
authors have reported that the inhibition of CoA-IT blocks the release
of arachidonic acid in stimulated neutrophils 33 . To examine this
possibility, we measured the effects of SB 203580 and PD 98059 on the
activity of CoA-IT in microsomes isolated from human neutrophils. The
results summarized in Fig. 7 clearly show
that these two inhibitors have no effect on the activity of CoA-IT.
Several concentrations (as high as 80 µM) were tested, and the
results were the same. Note that tosylamido-2-phenylethyl chloromethyl
ketone (TPCK) (1 mM), a known inhibitor of this enzyme, reduces
the activity of this enzyme greatly 33 . Also PMSF (1 mM), which is
known to inhibit this enzyme by 40%, produces similar inhibition 33 .
We also found that 1-alkyl-2-acyl-GPC exhibits a negative feedback on
CoA-IT (Fig. 7). This effect was concentration dependent (data not
shown).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The data presented here show several new and interesting interrelated
points concerning the kinases that phosphorylate cPLA2 in
human neutrophils, and probably other cell types. First, in intact
cells, cPLA2 can be phosphorylated by several kinases.
These include erk 1/2, p38 MAPK, and as yet unidentified kinases.
Second, the kinase involved in the phosphorylation of cPLA2
depends on the stimulus used. For example, erk1/2 and p38 MAPK mediate
the phosphorylation of cPLA2 in cells stimulated by PAF and
TNF-, respectively. On the other hand, other kinases can also
mediate the phosphorylation of cPLA2 in cells stimulated by
LPS or FMLP. Third, while PMA-induced phosphorylation of
cPLA2 is inhibited by the PKC inhibitor GO 6850 in intact
neutrophils, the effect of PKC on cPLA2 is mediated through
erk1/2. One possible kinase that may mediate the action of FMLP on
cPLA2 is the calcium calmodulin protein kinase(s). However,
utilization of a potent inhibitor of calcium calmodulin protein kinase
II, K-252a 37 , failed to inhibit the FMLP-induced phosphorylation of
cPLA2 in human neutrophils (data not shown).
Interestingly, while the FMLP-induced phosphorylation of
cPLA2 is not affected by inhibitors of the p38 MAPK cascade
or erk, the FMLP-stimulated arachidonic acid release is greatly reduced
by inhibitors of the p38 MAPK and erk cascades. This is not due to
possible action of these inhibitors on CoA-IT, because, unlike
ß-lactams SB 212047 33 , SB 203580 or PD 98059 does not inhibit the
activity of CoA-IT (Fig. 7). It is possible that these inhibitors
affect the translocation of cPLA2 to where its substrates
are located and/or reduce the rise in the intracellular concentration
of free calcium. In preliminary experiments, we found that the rise in
calcium is qualitatively similar in both control and SB 203580-treated
cells, which were stimulated with FMLP (data not shown). The
determination of the kinases that mediate the phosphorylation of
cPLA2 in human neutrophils stimulated by LPS and FMLP, the
site(s) that are phosphorylated by each stimulus, and the effects of
these inhibitors on the basal and stimulated distribution of
cPLA2 remain to be examined and are the subjects of future
studies.
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Ramadan I. Sha’afi, Department of Physiology, University of Connecticut Health Center, Farmington, CT 06030. E-mail address:
3 Abbreviations used in this paper: PLA2, phospholipase A2; cPLA2, cystolic PLA2; PAF, platelet-activating factor; MAP, mitogen-activated protein; MAPK, MAP kinase; erk, extracellular signal-regulating kinase; MEK, MAP/erk kinase; MAPKAP kinase-2, MAPK/activated protein kinase-2; hsp, heat shock protein; PKC, protein kinase C; GO 6850, bisindolylmaleimide; CoA-IT, coenzyme A-independent transacylase; JNK, c-Jun N-terminal kinase; HOG-1, high osmolality glycerol response-1; SAPK, stress-activated protein kinase; GPC, glycerol-3 phosphorylcholine.
Received for publication October 2, 1997. Accepted for publication November 2, 1998.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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. Biochem. Biophys. Res. Commun. 209:271.[Medline]
-induced phosphorylation and activation of cytosolic phospholipase A2 are abrogated by an inhibitor of the p38 MAP kinase cascade in human neutrophils. Biochem. J. 319:17.
on tyrosine phosphorylation and activation of mitogen-activated protein kinases in human neutrophils. Biochem. J. 307:39.
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T. Pomorski, T. F. Meyer, and M. Naumann Helicobacter pylori-induced Prostaglandin E2 Synthesis Involves Activation of Cytosolic Phospholipase A2 in Epithelial Cells J. Biol. Chem., January 5, 2001; 276(1): 804 - 810. [Abstract] [Full Text] [PDF]
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M. W. Anthonsen, A. Solhaug, and B. Johansen Functional Coupling between Secretory and Cytosolic Phospholipase A2 Modulates Tumor Necrosis Factor-alpha - and Interleukin-1beta -induced NF-kappa B Activation J. Biol. Chem., August 3, 2001; 276(32): 30527 - 30536. [Abstract] [Full Text] [PDF]
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G. Siegel, L. Sternfeld, A. Gonzalez, I. Schulz, and A. Schmid Arachidonic Acid Modulates the Spatiotemporal Characteristics of Agonist-evoked Ca2+ Waves in Mouse Pancreatic Acinar Cells J. Biol. Chem., May 11, 2001; 276(20): 16986 - 16991. [Abstract] [Full Text] [PDF]
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M.-H. Yoo, C.-H. Woo, H.-J. You, S.-H. Cho, B.-C. Kim, J.-E. Choi, J.-S. Chun, B. H. Jhun, T.-S. Kim, and J.-H. Kim Role of the Cytosolic Phospholipase A2-linked Cascade in Signaling by an Oncogenic, Constitutively Active Ha-Ras Isoform J. Biol. Chem., June 29, 2001; 276(27): 24645 - 24653. [Abstract] [Full Text] [PDF]
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C. S. T. Hii, N. Moghadammi, A. Dunbar, and A. Ferrante Activation of the Phosphatidylinositol 3-Kinase-Akt/Protein Kinase B Signaling Pathway in Arachidonic Acid-stimulated Human Myeloid and Endothelial Cells. INVOLVEMENT OF THE ErbB RECEPTOR FAMILY J. Biol. Chem., July 13, 2001; 276(29): 27246 - 27255. [Abstract] [Full Text] [PDF]
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M. W. Anthonsen, S. Andersen, A. Solhaug, and B. Johansen Atypical lambda /iota PKC Conveys 5-Lipoxygenase/Leukotriene B4-mediated Cross-talk between Phospholipase A2s Regulating NF-kappa B Activation in Response to Tumor Necrosis Factor-alpha and Interleukin-1beta J. Biol. Chem., September 14, 2001; 276(38): 35344 - 35351. [Abstract] [Full Text] [PDF]
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M. A. Gijon, D. M. Spencer, A. R. Siddiqi, J. V. Bonventre, and C. C. Leslie Cytosolic Phospholipase A2 Is Required for Macrophage Arachidonic Acid Release by Agonists That Do and Do Not Mobilize Calcium. NOVEL ROLE OF MITOGEN-ACTIVATED PROTEIN KINASE PATHWAYS IN CYTOSOLIC PHOSPHOLIPASE A2 REGULATION J. Biol. Chem., June 23, 2000; 275(26): 20146 - 20156. [Abstract] [Full Text] [PDF]
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