Department of Pediatrics, University of Chicago, Chicago, Illinois 60637-1470
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
We have demonstrated that platelet-derived
growth factor (PDGF) stimulates p38 mitogen-activated protein (MAP)
kinase activation in bovine tracheal myocytes, suggesting that p38 is
involved in growth regulation. We therefore examined whether p38
regulates expression of cyclin D1, a G1 cyclin required for
cell cycle traversal. The chemical p38 inhibitors SB-202190 and
SB-203580 each increased basal and PDGF-induced cyclin D1 promoter
activity and protein abundance. Overexpression of a dominant negative
allele of MAP kinase kinase-3 (MKK3), an upstream activator of p38,
had similar effects. Conversely, active MKK3 and MKK6, both of which
increase p38
activity, each decreased transcription from the cyclin
D1 promoter. Together, these data demonstrate that p38 negatively regulates cyclin D1 expression. We tested whether p38 regulates cyclin
D1 expression via inhibition of extracellular signal-regulated kinase
(ERK) activation. Chemical inhibitors of p38 induced modest ERK
phosphorylation and activation. However, dominant negative MKK3 was
insufficient to activate ERK, and active MKK3 and MKK6 did not
attenuate platelet-derived growth factor-mediated ERK activation. These
data are consistent with the notion that p38
negatively regulates
cyclin D1 expression via an ERK-independent pathway.
bovine; extracellular signal-regulated kinase; platelet-derived growth factor; signal transduction; mitogen-activated protein kinase
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
THERE IS AMPLE EVIDENCE supporting the notion that excess airway smooth muscle mass contributes to the pathogenesis of airflow obstruction in asthma (6, 10-12, 19, 22, 49, 52, 55). Accordingly, we have examined the signaling pathways responsible for cell cycle traversal in primary bovine tracheal myocytes. Inhibition of mitogen-activated protein (MAP) kinase/extracellular signal-regulated kinase (ERK) kinase (MEK) reduced platelet-derived growth factor (PDGF)-induced DNA synthesis in a concentration-dependent manner, suggesting that catalytic activation of MEK-1 and ERK is required for DNA synthesis in these cells (28). In addition, we found that activation of MEK-1 is required and sufficient to activate transcription from the promoter of cyclin D1 (47), a critical regulator of G1 progression in these cells (66).
Growth factor treatment of airway smooth muscle cells also induces activation of the two stress-activated MAP kinases, p38 and c-Jun amino-terminal kinase (JNK) (41, 45, 50), consistent with the notion that these intermediates, like ERKs, play a role in the growth regulation. The precise role of p38 and JNK in airway smooth muscle cell cycle traversal has not been established, however.
The p38 MAP kinase family consists of four isoforms. p38 was
originally identified in lipopolysaccharide-stimulated mouse macrophages and was found to have substantial homology to the yeast
high-osmolarity glycerol kinase (9, 18, 36, 38). Since
this original identification, three additional isoforms,
,
, and
, have been cloned (23, 24, 53, 60). p38
, -
, and
-
are somewhat ubiquitously expressed, whereas p38-
is primarily
restricted to skeletal muscle (60). p38
and -
are
inhibitable by pyridinylimidazole compounds such as SB-202190 and
SB-203580, whereas p38
and -
are not (14, 24, 31). Three MAP kinase kinases (MKKs) for p38 have been identified. MKK6
strongly activates all p38 isoforms, and MKK3 preferentially activates
p38
and -
(14, 21, 23, 37, 46, 57). MKK4 appears to
phosphorylate and activate both JNK1 and p38
(9, 36).
Few studies in any cell system have addressed the potential role of p38 in the regulation of cell growth. Overexpression of p38 inhibits serum-induced NIH/3T3 cell cycle progression (39). Mutations of the MKK4 gene have been found in human pancreatic, lung, breast, testicular, and colorectal cancer cell lines (54, 56). Expression of an active MEK kinase (MEKK)-3 inhibited cell cycle progression and reversed Ras-induced transformation in NIH/3T3 cells (13). This effect was blocked by coexpression of an inactive MKK6, suggesting that cell cycle arrest was mediated by p38.
The mechanism by which signaling through the p38 pathway may negatively regulate growth has not been well studied. p38 attenuates transcription from the cyclin D1 promoter in CCL-39 hamster lung fibroblasts (33) but is necessary for optimal induction of cyclin D1 promoter activity in the human breast cancer MCF-7 cell line (34).
Recent evidence suggests that p38 exerts its effects, in part, via the
negative regulation of ERK. The triggering of stress-activated kinases,
concomitant with the inhibition of the ERK pathway, has been observed
in a number of cell systems undergoing apoptosis (4,
65). In rat-1 fibroblasts, stimulation of
1A-adrenoreceptors by phenylephrine stimulated p38
activation while inhibiting ERK phosphorylation (3).
Treatment with the chemical p38 inhibitors SB-202190 or SB-203580 has
been demonstrated to increase ERK activation in mouse keratinocytes
(48), HepG2 cells (51), and, in a preliminary report, canine tracheal myocytes (25). These data suggest
that p38 may negatively regulate the ERK pathway, although the
signaling intermediates linking p38 and ERK have not been established.
We tested the hypothesis that in primary bovine tracheal myocytes, p38 negatively regulates transcription from the cyclin D1 promoter. Our results suggest that p38 negatively regulates cyclin D1 expression in an ERK-independent manner.
![]() |
METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Materials.
Anti-human -smooth muscle actin, peroxidase-linked goat
anti-rabbit IgG, protein A Sepharose beads, and myelin basic protein were purchased from Sigma (St. Louis, MO). SB-203580 and SB-202190 were
from Calbiochem (La Jolla, CA). PDGF was obtained from Upstate Biotechnology (Lake Placid, NY). PD-98059 and recombinant activating transcription factor (ATF)-2 were obtained from New England
Biolabs (Beverly, MA). [
-32P]ATP and an enhanced
chemiluminescence kit were purchased from DuPont/NEN Research Products
(Wilmington, DE). LipofectAMINE and Opti-MEM were obtained from Life
Technologies (Gaithersburg, MD). An antibody against dually
phosphorylated ERK and a luciferase assay kit were purchased from
Promega (Madison, WI). Peroxidase-linked rat anti-mouse
light chain
IgG was obtained from Zymed Laboratories (South San Francisco, CA). A
polyclonal antibody against cyclin D1 was purchased from Santa Cruz
Biotechnology (Santa Cruz, CA). For in vitro phosphorylation assays, a
monoclonal antibody against hemagglutinin (HA; 12CA5) was obtained from
Babco (Berkeley, CA).
Cell culture.
Bovine tracheal smooth muscle cells were isolated as described
previously (1). Myocytes from passage 5 or less
were studied. Confluent cultures exhibited the typical
"hill-and-valley" appearance and showed specific immunostaining for
-smooth muscle actin. Cells were cultured in DMEM containing 10%
fetal bovine serum (FBS), 1% nonessential amino acids, 100 U/ml of
penicillin, and 100 µg/ml of streptomycin. A7r5 and COS cells were
obtained from the American Type Culture Collection (Manassas, VA).
Determination of cyclin D1 promoter transcriptional activity.
Cells were seeded into 60-mm dishes at 50-80% confluence and
incubated in 10% FBS-DMEM overnight. After being rinsed, the cells
were incubated with a liposome solution consisting of serum- and
antibiotic-free medium, plasmid DNA (total of 1.8 µg/plate), and
LipofectAMINE (12 µl/plate). Cells were transiently cotransfected with plasmids encoding the human cyclin D1 promoter subcloned into a
luciferase reporter and either the relevant expression vector or
control vector (42). After 5 h, the liposome solution was replaced with 10% FBS-DMEM. The next day, cells were serum starved
in DMEM. After 24 h of serum starvation, selected cultures were
treated with chemical p38 inhibitors and/or PDGF (30 ng/ml). Finally,
16 h after PDGF treatment, cells were harvested for analysis of
luciferase activity with lysis buffer provided in the Promega luciferase assay system. Luciferase activity was measured at room temperature with a luminometer (Turner Designs, Sunnyvale, CA). Luciferase content was assessed by measuring the light emitted during
the initial 30 s of the reaction, and the values are expressed in
arbitrary light units. The background activity from cell extracts was
typically >0.02 U compared with signals on the order of
102 to 103 U. Cyclin D1 promoter
transcriptional activation was normalized for transfection efficiency
by cotransfecting cells with a cDNA encoding -galactosidase (30 ng/plate).
-Galactosidase activity was assessed by colorimetric
assay with o-nitrophenyl-
-D-galactoside as a
substrate (43).
Preparation of cell extracts for immunoblotting.
Cells were cultured in six-well plates and serum starved for 48 h
before treatment. Selected cultures were treated with 3 µM SB-202190
or SB-203580 1 h before growth factor treatment or with 30 ng/ml
of PDGF (for 10 min or 16 h for ERK or cyclin D1 immunoblots,
respectively). Cells were washed in PBS (150 mM NaCl and 0.1 M
phosphate, pH 7.5) and extracted in a lysis buffer containing 50 mM
Tris, pH 7.5, 40 mM -glycerophosphate, 100 mM NaCl, 2 mM EDTA, 50 mM
NaF, 200 µM Na3VO4, 200 µM
phenylmethylsulfonyl fluoride, and 1% Triton X-100. Insoluble
materials were removed by centrifugation (13,000 rpm for 10 min at
4°C), and the supernatant was transferred to fresh microcentrifuge tubes.
Western analysis of whole cell extracts. Extracts (10 µg) were resolved on a 10% SDS-polyacrylamide gel and transferred to nitrocellulose by semidry transfer (Hoefer, San Francisco, CA). After incubation with the appropriate polyclonal antibody, signals were amplified and visualized with anti-rabbit IgG and enhanced chemiluminescence.
Measurement of ERK and p38 activation.
Cells were transiently cotransfected with cDNAs encoding HA-tagged ERK2
or p38 and the expression vector of interest. Forty-eight hours
after transfection, the cells were serum starved in DMEM. The next day,
selected cultures were treated with PDGF (30 ng/ml for 10 min) or
anisomycin (50 µg/ml for 30 min). Activation of ERK2 or p38
was
then assessed by immunoprecipitation of the epitope tag followed by an
in vitro phosphorylation assay with major basic protein (MBP)
and ATF-2 as substrates, respectively, as described (28,
41). Treated cells were washed twice with PBS and incubated in a
lysis buffer consisting of 50 mM Tris · HCl, pH 7.5, 1% Triton X-100, 40 mM
-glycerophosphate, 100 mM NaCl, 50 mM NaF, 2 mM EDTA,
200 µM Na3VO4, and 0.2 mM
phenylmethylsulfonyl fluoride (30 min at 4°C). Insoluble materials
were removed by centrifugation (13,000 rpm for 10 min at 4°C). Cell
lysates were then incubated for 3 h with 30 µl of protein A
Sepharose beads precoupled with the 12CA5 anti-HA antibody.
Immunoprecipitates were washed three times with lysis buffer and twice
with kinase buffer containing 20 mM HEPES (pH 7.4), 10 mM
MgCl2, 1 mM dithiothreitol, 200 µM Na3VO4, and 10 mM p-nitrophenyl
phosphate. Immune complexes were resuspended in a final volume of 30 µl of kinase buffer and incubated (20 min at 30°C) with 5 µCi of
[
-32P]ATP and either 0.25 mg/ml of MBP or 0.2 mg/ml of
ATF-2. Reactions were terminated by adding Laemmli buffer and boiling.
Samples were resolved on a 10% SDS gel, and the proteins were
transferred to a nitrocellulose membrane by semidry transfer. After
Ponceau staining, the membrane was exposed to film and substrate
phosphorylation assessed by optical scanning (Jandel Scientific, San
Rafael, CA).
Statistical analysis. When applicable, significance was assessed by one-way analysis of variance (ANOVA). Differences identified by ANOVA were pinpointed by the Student-Newman-Keuls multiple range test.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Chemical inhibition of p38 negatively regulates cyclin D1
transcription and protein abundance.
Inhibition of p38 has been demonstrated to enhance cyclin D1
promoter activity and protein abundance in Chinese hamster lung fibroblasts (33). To determine the effect of p38
inhibition on cyclin D1 expression in bovine tracheal myocytes, cells
were transiently transfected with a cDNA encoding the full-length
cyclin D1 promoter subcloned into a luciferase reporter gene
(2). SB-202190 or SB-203580 (3 µM each) each increased
basal and PDGF-induced transcription from the cyclin D1 promoter (Fig.
1A). Furthermore, SB-203580
increased basal and PDGF-induced cyclin D1 protein abundance (Fig.
1B).
|
MKK3 and MKK6 regulate transcription from the cyclin D1 promoter
and cyclin D1 protein abundance.
MKK3 and MKK6 phosphorylate and activate p38 (14, 21, 23,
37, 46, 57). To determine whether MKK3 regulates cyclin D1 expression, we transiently transfected bovine tracheal myocytes with
cDNAs encoding mutant alleles of MKK3 and the luciferase-tagged cyclin
D1 promoter. Similar to the chemical p38 inhibitors, overexpression of
a dominant negative form of MKK3 (MKK3-AL) increased transcription from
the cyclin D1 promoter (Fig.
2A). Overexpression of a
constitutively active MKK3 [MKK3(glu)] reduced basal and PDGF-induced
cyclin D1 promoter activity, and overexpression of active MKK6
[MKK6(glu)] had similar effects (Fig. 2B).
|
|
Chemical p38 inhibitors inhibit p38 and activate ERK in primary bovine tracheal myocytes. In certain cell systems, chemical inhibition of p38 (with SB-202190 or SB-203580) increases ERK activity (3, 25, 51). As in other cell types (2, 33, 62), it has been demonstrated in bovine tracheal myocytes that ERK activation is an upstream activator of transcription from the cyclin D1 promoter (47). Thus inhibition of p38 could increase cyclin D1 accumulation via the activation of ERK. We therefore investigated the potential interactions between the p38 and ERK signaling pathways in bovine tracheal smooth muscle cells.
To test the effects of chemical inhibitors on p38 activity, cells were transiently transfected with a HA-tagged p38
|
|
MKK3 and MKK6 regulate p38 but not ERK in primary bovine tracheal
myocytes.
To examine the effects of mutant forms of MKK3 and MKK6 on p38
activity, bovine tracheal myocytes were transiently cotransfected with cDNAs encoding a HA-tagged p38 (64) and mutant
alleles of either MKK3 [MKK3-AL and MKK3(glu)] or MKK6 [MKK6(glu)].
p38
activity was determined by immunoprecipitation with an anti-HA antibody followed by an in vitro phosphorylation assay with ATF-2 as a substrate. Overexpression of MKK3-AL reduced p38
activation (Fig. 6, A
and B), whereas overexpression of MKK3(glu) and MKK6(glu) each induced significant activation of p38
(Fig. 6, C and
D).
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
ERK activation has been shown to be required for DNA synthesis in
a wide variety of cell systems including airway smooth muscle (28, 35, 40, 59, 63). On the other hand, activation of p38
MAP kinase has been associated with growth inhibition (33) and apoptosis (4, 5, 7, 15, 26, 30, 32, 58, 61,
65). We have demonstrated that PDGF induces activation of both
ERK and p38 in bovine tracheal myocytes (41) and that ERK activation is required for PDGF-induced cyclin D1 expression and
DNA synthesis in these cells (28, 41, 47). If p38
negatively regulated cyclin D1 expression, then PDGF-induced activation
of p38 might limit ERK-mediated cell growth. Because excess airway smooth muscle proliferation has been noted in patients with fatal asthma, the concurrent regulation of growth-stimulatory and
growth-inhibitory pathways by growth factors might constitute an
important physiological mechanism limiting airflow obstruction in these patients.
In this report, we show that inhibition of p38, either by chemical inhibitors or by overexpression of a dominant negative allele of MKK3, increases cyclin D1 promoter transcription and protein abundance in cultured primary airway smooth muscle cells. These data imply a basal level of p38 activation in serum-starved cells (Figs. 4 and 6). Furthermore, selective activation of p38 by overexpression of either MKK3 or MKK6 attenuated both basal and PDGF-induced transcription from the cyclin D1 promoter. These data suggest that p38 negatively regulates cyclin D1 expression in airway smooth muscle cells. Our data confirm previous experiments in CCL-39 fibroblasts in which overexpression of MKK3 decreased basal and serum-induced cyclin D1 expression, whereas treatment with a chemical p38 inhibitor or a dominant negative form of MKK3 increased transcription from the cyclin D1 promoter (33). Furthermore, our data are consistent with recent studies suggesting that p38 negatively regulates cyclin G1 expression. In PC12 cells, hypoxia increased p38 activation while decreasing cyclin D1 levels (8). In mouse embryonic kidney cells, chemical inhibition of p38 allowed Rac1 to induce anchorage-dependent cyclin A transcription (44).
In PC12 (65) and HeLa cells (4), p38
activation occurred concomitantly with inhibition of the ERK pathway,
suggesting that p38 negatively regulates ERK. Also, treatment with the
pyridinyl imidazole compounds SB-203580 and SB-202190 has been noted to increase ERK activation in mouse keratinocytes (48),
canine tracheal myocytes (25), and HepG2 cells
(51), suggesting that inhibition of p38 activates the ERK
pathway (51). We therefore examined the regulation of ERK
activity by p38 in bovine tracheal myocytes. We found that treatment of
bovine myocytes with the chemical p38 inhibitors SB-202190 and
SB-203580 increased ERK activation. However, overexpression of a
dominant negative form of MKK3, which inhibited anisomycin-induced
p38 activation, did not increase basal or PDGF-stimulated ERK
activity. Furthermore, when we overexpressed active forms of MKK3 and
MKK6, both of which increase p38
activation, there was no reduction
of either basal or PDGF-induced activation of ERK. Taken together,
these data suggest that the selective inhibition or activation of
p38
by upstream activators does not regulate ERK activation.
Furthermore, they suggest that the pyridinyl imidazole compounds
SB-202190 and SB-203580 may activate ERK via a MEK-dependent,
p38
-independent pathway.
It has recently been demonstrated that SB-203580 inhibits Raf-1 in vitro (17). However, in vivo, inhibition of Raf-1 by SB-203580 is counterbalanced by a novel feedback loop, leading to Raf-1 activation (16, 27). Our study, in which ERK activation was induced by both SB-202190 and SB-203580 but not a dominant negative inhibitor of MKK3, supports the notion that pyridinyl imidazole inhibitors may have p38-independent effects. Interestingly, neither chemical inhibitor increased ERK activation in COS cells (Page and Hershenson, unpublished results), indicating that the level of compensatory Raf-1 activation may vary with cell type. Our laboratory has previously shown (29) that in bovine tracheal myocytes, an alternative MEK activator besides Raf-1 is responsible for MEK phosphorylation. Thus the finding that pyridinyl imidazole compounds activate ERK in our system suggests that these reagents may inhibit other MAP kinase kinase kinases besides Raf.
Other possible explanations for the differential effects of chemical
p38 inhibitors and the dominant negative MKK3 need to be considered.
The pyridinyl imidazole compounds selectively inhibit p38 and -
,
and therefore it is conceivable that the activation of ERK by these
compounds relates to inhibition of p38
(31). Because
p38
is activated by MKK6 but not by MKK3, experiments that employed
a dominant negative mutant of MKK6 would address this possibility.
However, the two dominant negative MKK6 mutants we tested, although
they attenuated anisomycin-induced activation of p38, also
significantly increased basal p38 activity. Nevertheless, our finding
that a constitutively active MKK6 mutant that activates all isoforms of
p38 (14, 23, 24, 60) had no effect on ERK activation
suggests that p38
does not regulate ERK in our system.
It is important to note potential limitations of our approach, which employed the overexpression of constitutively active and dominant negative signaling intermediates. First, it is possible that overexpression of a constitutively active protein could induce a supraphysiological outcome. For example, it is conceivable that although selective activation of p38 by MKK3/6 inhibits cyclin D1 expression in our system, p38 activation alone may not be sufficient for this effect under normal conditions and may require the activation of other negative regulatory pathways. Second, dominant negative proteins may bind downstream signaling intermediates, thereby blocking the activity of other upstream activators that bind at the same site. However, due to the difficulty of selectively activating p38, we saw no alternative but to use these reagents. For example, chemical activators of p38 such as anisomycin, hyperosmolarity, and UV irradiation tend to cause multiple stress-related effects that would likely influence our results. Nevertheless, chemical inhibitors of p38 increased cyclin D1 promoter activation and protein abundance in our system, confirming the enhancing effects of the dominant negative MKK3 on transcription from the cyclin D1 promoter. Finally, we did not examine the effects of p38 on the transcription or protein abundance of other G1 cyclins, cyclin-dependent kinases, or cyclin-dependent kinase inhibitors. Thus while the negative regulation of cyclin D1 by p38 would tend to attenuate cell cycle traversal, the net effect of p38 activation on airway cell DNA synthesis and proliferation remains to be determined.
In conclusion, our data strongly suggest that in bovine tracheal
myocytes, p38 negatively regulates transcription from the cyclin D1
promoter via an ERK-independent pathway. Furthermore, chemical
inhibitors of p38 may activate ERK in a p38
-independent manner.
![]() |
ACKNOWLEDGEMENTS |
---|
We thank Drs. Richard Pestell, Michael Karin, Roger Davis,
and James Woodgett for kindly providing the cDNAs encoding
1745CD1LUC, p38
, MKK3, and MKK6, respectively. We also thank Dr.
Marsha Rosner for thoughtful critique of the manuscript.
![]() |
FOOTNOTES |
---|
These studies were supported by National Heart, Lung, and Blood Institute Grants HL-54685, HL-56399, and HL-63314.
Address for reprint requests and other correspondence: M. B. Hershenson, Univ. of Chicago Children's Hospital, 5841 S. Maryland Ave., MC 4064, Chicago, IL 60637-1470 (E-mail: mhershen{at}midway.uchicago.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 14 June 2000; accepted in final form 17 November 2000.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Abe, MK,
Chao TS,
Solway J,
Rosner MR,
and
Hershenson MB.
Hydrogen peroxide stimulates mitogen-activated protein kinase in bovine tracheal myocytes: implications for human airway disease.
Am J Respir Cell Mol Biol
11:
577-585,
1994[Abstract].
2.
Albanese, C,
Johnson J,
Watanabe G,
Eklund N,
Vu D,
Arnold A,
and
Pestell RG.
Transforming p21ras mutants and c-Ets-2 activate the cyclin D1 promoter through distinguishable regions.
J Biol Chem
270:
23589-23597,
1995
3.
Alexandrov, A,
Keffel S,
Goepel M,
and
Michel MC.
Stimulation of 1A-adrenoreceptors in rat-1 cells inhibits extracellular signal-regulated kinase by activated p38 mitogen-activated protein kinase.
Mol Pharmacol
54:
755-760,
1998
4.
Berra, E,
Diaz-Meco MT,
and
Moscat J.
The activation of p38 and apoptosis by the inhibition of ERK is antagonized by the phosphoinositide 3-kinase/Akt pathway.
J Biol Chem
273:
10792-10797,
1998
5.
Brenner, B,
Koppenhoefer U,
Weinstock C,
Linderkamp O,
Lang F,
and
Gulbins E.
Fas- or ceramide-induced apoptosis is mediated by a Rac1-regulated activation of Jun N-terminal kinase/p38 kinases and GADD153.
J Biol Chem
272:
22173-22181,
1997
6.
Carroll, N,
Elliot J,
Morton A,
and
James A.
The structure of large and small airways in nonfatal and fatal asthma.
Am Rev Respir Dis
147:
405-410,
1993[ISI][Medline].
7.
Chen, YR,
Wang X,
Templeton D,
Davis RJ,
and
Tan TH.
The role of c-Jun N-terminal kinase (JNK) in apoptosis induced by ultraviolet C and gamma radiation. Duration of JNK activation may determine cell death and proliferation.
J Biol Chem
271:
31929-31936,
1996
8.
Conrad, PW,
Rust RT,
Han J,
Millhorn DE,
and
Beitner-Johnson D.
Selective activation of p38 alpha and p38 gamma by hypoxia. Role in regulation of cyclin D1 by hypoxia in PC12 cells.
J Biol Chem
274:
23570-23576,
1999
9.
Derijard, B,
Raingeaud J,
Barrett T,
Wu IH,
Han J,
Ulevitch RJ,
and
Davis RJ.
Independent human MAP-kinase signal transduction pathways defined by MEK and MKK isoforms.
Science
267:
682-685,
1995[ISI][Medline].
10.
Dunnill, MS,
Massarella GR,
and
Anderson JA.
A comparison of the quantitative anatomy of the bronchi in normal subjects, in status asthmaticus, in chronic bronchitis and in emphysema.
Thorax
24:
176-179,
1969[ISI][Medline].
11.
Ebina, M,
Takahashi T,
Chiba T,
and
Motomiya M.
Cellular hypertrophy and hyperplasia of airway smooth muscles underlying bronchial asthma. A 3-D morphometric study.
Am Rev Respir Dis
148:
720-726,
1993[ISI][Medline].
12.
Ebina, M,
Yaegashi H,
Chiba R,
Takahashi T,
Motomiya M,
and
Tanemura M.
Hyperreactive site in the airway tree of asthmatic patients revealed by thickening of bronchial muscles. A morphometric study.
Am Rev Respir Dis
141:
1327-1332,
1990[ISI][Medline].
13.
Ellinger-Ziegelbauer, H,
Kelly K,
and
Siebenlist U.
Cell cycle arrest and reversion of Ras-induced transformation by a conditionally activated form of mitogen-activated protein kinase kinase kinase 3.
Mol Cell Biol
19:
3857-3868,
1999
14.
Enslen, H,
Raingeaud J,
and
Davis RJ.
Selective activation of p38 mitogen-activated protein (MAP) kinase isoforms by the MAP kinase kinases MKK3 and MKK6.
J Biol Chem
273:
1741-1748,
1998
15.
Frasch, SC,
Nick JA,
Fadok VA,
Bratton DL,
Worthen GS,
and
Henson PM.
p38 mitogen-activated protein kinase-dependent and -independent intracellular signal transduction pathways leading to apoptosis in human neutrophils.
J Biol Chem
273:
8389-8397,
1998
16.
Hall-Jackson, CA,
Eyers PA,
Cohen P,
Goedert M,
Boyle FT,
Hewitt N,
Plant H,
and
Hedge P.
Paradoxical activation of Raf by a novel Raf inhibitor.
Chem Biol
6:
559-568,
1999[ISI][Medline].
17.
Hall-Jackson, CA,
Goedert M,
Hedge P,
and
Cohen P.
Effect of SB-203580 on the activity of c-Raf in vitro and in vivo.
Oncogene
18:
2047-2054,
1999[ISI][Medline].
18.
Han, J,
Lee JD,
Tobias PS,
and
Ulevitch RJ.
Endotoxin induces rapid protein tyrosine phosphorylation in 70Z/3 cells expressing CD14.
J Biol Chem
268:
25009-25014,
1993
19.
Heard, BE,
and
Hossain S.
Hyperplasia of bronchial smooth muscle in asthma.
J Pathol
10:
319-332,
1973.
20.
Hershenson, MB,
Chao TS,
Abe MK,
Gomes I,
Kelleher MD,
Solway J,
and
Rosner MR.
Histamine antagonizes serotonin and growth factor-induced mitogen-activated protein kinase activation in bovine tracheal smooth muscle cells.
J Biol Chem
270:
19908-19913,
1995
21.
Holland, PM,
Suzanne M,
Campbell JS,
Noselli S,
and
Cooper JA.
MKK7 is a stress-activated mitogen-activated protein kinase kinase functionally related to hemipterous.
J Biol Chem
272:
24994-24998,
1997
22.
James, AL,
Pare PD,
and
Hogg JC.
The mechanics of airway narrowing in asthma.
Am Rev Respir Dis
139:
242-246,
1989[ISI][Medline].
23.
Jiang, Y,
Chen C,
Li Z,
Guo W,
Gegner JA,
Lin S,
and
Han J.
Characterization of the structure and function of a new mitogen-activated protein kinase (p38).
J Biol Chem
271:
17920-17926,
1996
24.
Jiang, Y,
Gram H,
Zhao M,
New L,
Gu J,
Feng L,
Di Padova F,
Ulevitch RJ,
and
Han J.
Characterization of the structure and function of the fourth member of p38 group mitogen-activated protein kinases, p38delta.
J Biol Chem
272:
30122-30128,
1997
25.
Jones, CA,
and
Brown JK.
p38 inhibition increases ERK1/ERK2 kinase activity and DNA synthesis in airway smooth muscle cells: evidence for cross-talk between mitogen-activated protein (MAP) kinase pathways (Abstract).
Am J Respir Crit Care Med
159:
A722,
1999[ISI].
26.
Juo, P,
Kuo CJ,
Reynolds SE,
Konz RF,
Raingeaud J,
Davis RJ,
Biemann HP,
and
Blenis J.
Fas activation of the p38 mitogen-activated protein kinase signaling pathway requires ICE/CED-3 family proteases.
Mol Cell Biol
17:
24-35,
1997[Abstract].
27.
Kalmes, A,
Deou J,
Clowes AW,
and
Daum G.
Raf-1 is activated by the p38 mitogen-activated protein kinase inhibitor, SB-203580.
FEBS Lett
444:
71-74,
1999[ISI][Medline].
28.
Karpova, AY,
Abe MK,
Li J,
Liu P,
Rhee JM,
Kuo WL,
and
Hershenson MB.
MEK1 is required for PDGF-induced ERK activation and DNA synthesis in tracheal myocytes.
Am J Physiol Lung Cell Mol Physiol
272:
L558-L565,
1997
29.
Kartha, S,
Naureckas ET,
Li J,
and
Hershenson MB.
Partial characterization of a novel mitogen-activated protein kinase/extracellular signal-regulated kinase activator in airway smooth-muscle cells.
Am J Respir Cell Mol Biol
20:
1041-1048,
1999
30.
Kawasaki, H,
Morooka T,
Shimohama S,
Kimura J,
Hirano T,
Gotoh Y,
and
Nishida E.
Activation and involvement of p38 mitogen-activated protein kinase in glutamate-induced apoptosis in rat cerebellar granule cells.
J Biol Chem
272:
18518-18521,
1997
31.
Kumar, S,
McDonnell PC,
Gum RJ,
Hand AT,
Lee JC,
and
Young PR.
Novel homologues of CSBP/p38 MAP kinase: activation, substrate specificity and sensitivity to inhibition by pyridinyl imidazoles.
Biochem Biophys Res Commun
235:
533-538,
1997[ISI][Medline].
32.
Kummer, JL,
Rao PK,
and
Heidenreich KA.
Apoptosis induced by withdrawal of trophic factors is mediated by p38 mitogen-activated protein kinase.
J Biol Chem
272:
20490-20494,
1997
33.
Lavoie, J,
L'Allemain G,
Brunet A,
Muller R,
and
Pouyssegur J.
Cyclin D1 expression is regulated positively by p42/p44MAPK and negatively by p38/HOGMAPK pathway.
J Biol Chem
271:
20608-20616,
1996
34.
Lee, RJ,
Albanese C,
Stenger RJ,
Watanabe G,
Inghirami G,
Haines GK,
Webster M,
Muller WJ,
Brugge JS,
Davis RJ,
and
Pestell RG.
pp60(v-src) induction of cyclin D1 requires collaborative interactions between the extracellular signal-regulated kinase, p38 and Jun kinase pathways. A role for cAMP response element-binding protein and activating transcription factor-2 in pp60(v-src) signaling in breast cancer cells.
J Biol Chem
274:
7341-7350,
1999
35.
Lew, DB,
Dempsey BK,
Zhao Y,
Muthalif M,
Fatima S,
and
Malik KU.
Beta-hexosaminidase-induced activation of p44/42 mitogen-activated protein kinase is dependent on p21Ras and protein kinase C and mediates bovine airway smooth-muscle proliferation.
Am J Respir Cell Mol Biol
21:
111-118,
1999
36.
Lin, A,
Minden A,
Martinetto H,
Claret FX,
Lange-Carter C,
Mercurio F,
Johnson GL,
and
Karin M.
Identification of a dual specificity kinase that activates the Jun kinases and p38-Mpk2.
Science
268:
286-290,
1995[ISI][Medline].
37.
Lu, X,
Nemoto S,
and
Lin A.
Identification of c-Jun NH2-terminal protein kinase (JNK)-activating kinase 2 as an activator of JNK but not p38.
J Biol Chem
272:
24751-42475,
1997
38.
Minden, A,
Lin A,
McMahon M,
Lange-Carter C,
Derijard B,
Davis RJ,
Johnson GL,
and
Karin M.
Differential activation of ERK and JNK mitogen-activated protein kinases by Raf-1 and MEKK.
Science
266:
1719-1723,
1994[ISI][Medline].
39.
Molnar, A,
Theodoras AM,
Zon LI,
and
Kyriakis JM.
Cdc42Hs, but not Rac1, inhibits serum-stimulated cell cycle progression at G1/S through a mechanism requiring p38/RK.
J Biol Chem
272:
13229-13235,
1997
40.
Orsini, MJ,
Krymskaya VP,
Eszterhas AJ,
Benovic JL,
Panettieri RA,
and
Penn RB.
MAPK superfamily activation in human airway smooth muscle: mitogenesis requires prolonged p42/p44 activation.
Am J Physiol Lung Cell Mol Physiol
277:
L479-L488,
1999
41.
Page, K,
Li J,
and
Hershenson MB.
Platelet-derived growth factor stimulation of mitogen-activated protein kinases and cyclin D1 promoter activity in cultured airway smooth muscle cells: role of Ras.
Am J Respir Cell Mol Biol
20:
1294-1302,
1999
42.
Page, K,
Li J,
Hodge JA,
Liu PT,
Vanden Hoek TL,
Becker LB,
Pestell RG,
Rosner MR,
and
Hershenson MB.
Characterization of a Rac1 signaling pathway to cyclin D1 expression in airway smooth muscle cells.
J Biol Chem
274:
22065-22071,
1999
43.
Parmacek, MS,
Vora AJ,
Shen T,
Barr E,
Jung F,
and
Leiden JM.
Identification and characterization of a cardiac-specific transcriptional regulatory element in the slow cardiac tropinin C gene.
Mol Cell Biol
12:
1967-1976,
1992[Abstract].
44.
Philips, A,
Roux P,
Coulon V,
Bellanger JM,
Vie A,
Vignais ML,
and
Blanchard JM.
Differential effect of rac and cdc42 on p38 kinase activity and cell cycle progression of nonadherent primary mouse fibroblasts.
J Biol Chem
275:
5911-5917,
2000
45.
Pyne, NJ,
and
Pyne S.
Platelet-derived growth factor activates a mammalian Ste20 coupled mitogen-activated protein kinase in airway smooth muscle.
Cell Signal
9:
311-317,
1997[ISI][Medline].
46.
Raingeaud, J,
Whitmarsh AJ,
Barrett T,
Derijard B,
and
Davis RJ.
MKK3- and MKK6-regulated gene expression is mediated by the p38 mitogen-activated protein kinase signal transduction pathway.
Mol Cell Biol
16:
1247-1255,
1996[Abstract].
47.
Ramakrishnan, M,
Musa NL,
Li J,
Liu P,
Pestell RG,
and
Hershenson MB.
Catalytic activation of extracellular signal-regulated kinases induces cyclin D1 expression in airway smooth muscle.
Am J Respir Cell Mol Biol
18:
736-740,
1998
48.
Rosenberger, SF,
Gupta A,
and
Bowden GT.
Inhibition of p38 MAP kinase increases okadaic acid mediated AP-1 expression and DNA binding but has no effect on TRE dependent transcription.
Oncogene
18:
3626-3632,
1999[ISI][Medline].
49.
Saetta, M,
Di Stefano A,
Rosina C,
Thiene G,
and
Fabbri LM.
Quantitative structural analysis of peripheral airways and arteries in sudden fatal asthma.
Am Rev Respir Dis
143:
138-143,
1991[ISI][Medline].
50.
Shapiro, PS,
Evans JN,
Davis RJ,
and
Posada JA.
The seven-transmembrane-spanning receptors for endothelin and thrombin cause proliferation of airway smooth muscle cells and activation of the extracellular regulated kinase and c-Jun NH2-terminal kinase groups of mitogen-activated protein kinases.
J Biol Chem
271:
5750-5754,
1996
51.
Singh, RP,
Dhawan P,
Golden C,
Kapoor GS,
and
Mehta KD.
One way cross-talk between p38MAPK and p42/44MAPK: inhibition of p38MAPK induces low density lipoprotein receptor expression through activation of the p42/44MAPK cascade.
J Biol Chem
274:
19593-19600,
1999
52.
Sobonya, RE.
Quantitative structural alterations in long-standing allergic asthma.
Am Rev Respir Dis
130:
289-292,
1984[ISI][Medline].
53.
Stein, B,
Yang MX,
Young DB,
Janknecht R,
Hunter T,
Murray BW,
and
Barbosa MS.
p38-2, a novel mitogen-activated protein kinase with distinct properties.
J Biol Chem
272:
19509-19517,
1997
54.
Su, GH,
Hilgers W,
Shekher MC,
Tang DJ,
Yeo CJ,
Hruban RH,
and
Kern SE.
Alterations in pancreatic, biliary, and breast carcinomas support MKK4 as a genetically targeted tumor suppressor gene.
Cancer Res
58:
2339-2342,
1998[Abstract].
55.
Takizawa, T,
and
Thurlbeck WM.
Muscle and mucous gland size in the major bronchi of patients with chronic bronchitis, asthma and asthmatic bronchitis.
Am Rev Respir Dis
104:
331-336,
1971[ISI][Medline].
56.
Teng, DH,
Perry WL,
Hogan JK,
Baumgard M,
Bell R,
Berry S,
Davis T,
Frank D,
Frye C,
Hattier T,
Hu R,
Jammulapati S,
Janecki T,
Leavit A,
Mitchell JT,
Pero R,
Sexton D,
Schroeder M,
Su PH,
Swedlund B,
Kyriakis JM,
Avruch J,
Bartel P,
Wong AK,
and
Tavtigian SV.
Human mitogen-activated protein kinase kinase 4 as a candidate tumor suppressor.
Cancer Res
57:
4177-4182,
1997[Abstract].
57.
Tournier, C,
Whitmarsh AJ,
Cavanagh J,
Barrett T,
and
Davis RJ.
Mitogen-activated protein kinase kinase 7 is an activator of the c-Jun NH2-terminal kinase.
Proc Natl Acad Sci USA
94:
7337-7342,
1997
58.
Toyoshima, F,
Moriguchi T,
and
Nishida E.
Fas induces cytoplasmic apoptotic responses and activation of the MKK7-JNK/SAPK and MKK6-p38 pathways independent of CPP32-like proteases.
J Cell Biol
139:
1005-1015,
1997
59.
Walker, TR,
Moore SM,
Lawson MF,
Panettieri RAJ,
and
Chilvers ER.
Platelet-derived growth factor-BB and thrombin activate phosphoinositide 3-kinase and protein kinase B: role in mediating airway smooth muscle proliferation.
Mol Pharmacol
54:
1007-1015,
1998
60.
Wang, XS,
Diener K,
Manthey CL,
Wang S,
Rosenzweig B,
Bray J,
Delaney J,
Cole CN,
Chan-Hui PY,
Mantlo N,
Lichenstein HS,
Zukowski M,
and
Yao Z.
Molecular cloning and characterization of a novel p38 mitogen-activated protein kinase.
J Biol Chem
272:
23668-23674,
1997
61.
Wang, Y,
Huang S,
Sah VP,
Ross J, Jr,
Brown JH,
Han J,
and
Chien KR.
Cardiac muscle cell hypertrophy and apoptosis induced by distinct members of the p38 mitogen-activated protein kinase family.
J Biol Chem
273:
2161-2168,
1998
62.
Watanabe, G,
Howe A,
Lee R,
Albanese C,
Shu I,
Karnezis A,
Zon L,
Kyriakis J,
Rundell K,
and
Pestell R.
Induction of cyclin D1 by simian virus 40 small T tumor antigen.
Proc Natl Acad Sci USA
93:
12861-12866,
1996
63.
Whelchel, A,
Evans J,
and
Posada J.
Inhibition of ERK activation attenuates endothelin-stimulated airway smooth muscle proliferation.
Am J Respir Cell Mol Biol
16:
589-596,
1997[Abstract].
64.
Xia, Y,
Wu Z,
Su B,
Murray B,
and
Karin M.
JNKK1 organizes a MAP kinase module through specific and sequential interactions with upstream and downstream components mediated by its amino-terminal extension.
Genes Dev
12:
3369-3381,
1998
65.
Xia, Z,
Dickens M,
Raingeaud J,
Davis RJ,
and
Greenberg ME.
Opposing effects of ERK and JNK-p38 MAP kinases on apoptosis.
Science
270:
1326-1331,
1995[Abstract].
66.
Xiong, W,
Pestell RG,
Watanabe G,
Li J,
Rosner MR,
and
Hershenson MB.
Cyclin D1 is required for S phase traversal in bovine tracheal myocytes.
Am J Physiol Lung Cell Mol Physiol
272:
L1205-L1210,
1997
67.
Zanke, BW,
Rubie EA,
Winnett E,
Chan J,
Randall S,
Parsons M,
Boudreau K,
McInnis M,
Yan M,
Templeton DJ,
and
Woodgett JR.
Mammalian mitogen-activated protein kinase pathways are regulated through formation of specific kinase-activator complexes.
J Biol Chem
271:
29876-29881,
1996