1 Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, Alabama 35294; 2 Department of Physiology and the Cardiovascular Institute, Loyola University Chicago, Stritch School of Medicine, Maywood, Illinois 60154; and 3 Department of Pharmacology, Columbia University, New York, New York 10032
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Abnormal vascular smooth muscle cell (VSMC) growth plays a key role in the pathogenesis of hypertension and atherosclerosis. Angiotensin II (ANG II) elicits a hypertrophic growth response characterized by an increase in protein synthesis without cell proliferation. The present study investigated the role of the nonreceptor tyrosine kinase PYK2 in the regulation of ANG II-induced signaling pathways that mediate VSMC growth. Using coimmunoprecipitation analysis, the role of PYK2 as an upstream regulator of both extracellular signal-related kinase (ERK) 1/2 mitogen-activated protein kinase and phosphatidylinositol 3-kinase (PI 3-kinase) pathways was examined in cultured rat aortic VSMC. ANG II (100 nM) promoted the formation of a complex between PYK2 and the ERK1/2 regulators Shc and Grb2. ANG II caused a rapid and Ca2+-dependent tyrosine phosphorylation of the adapter molecule p130Cas, which coimmunoprecipitated both PYK2 and PI 3-kinase in ANG II-treated VSMC. Complex formation between PI 3-kinase and p130Cas and PYK2 was associated with a rapid phosphorylation of the ribosomal p70S6 kinase in a Ca2+- and tyrosine kinase-dependent manner. These data suggest that PYK2 is an important regulator of multiple signaling pathways involved in ANG II-induced VSMC growth.
p130Cas, extracellular signal-regulated kinase, mitogen-activated protein kinase, phosphatidylinositol 3-kinase, tyrosine kinase
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
ANGIOTENSIN II (ANG II) regulates a variety of physiological responses including salt and water balance, blood pressure, and vascular tone. In addition to its vasoconstrictor effects, ANG II is also a potent growth factor for vascular tissue (19). These effects of ANG II have been linked to the development of various diseases associated with altered vascular smooth muscle cell (VSMC) growth such as hypertension, restenosis, and atherosclerosis (9, 20, 21).
ANG II induces hypertrophic growth in cultured VSMC as well as in intact aorta (reviewed in Ref. 19). In cultured VSMC, the hypertrophic growth response is defined as an increase in cell size and protein content without change in cell number and DNA replication (3, 7). These growth-promoting effects of ANG II include changes in gene expression, protein synthesis and turnover, and protein assembly into stress fibers.
A variety of intracellular signaling cascades are involved in the control of ANG II-induced VSMC growth, including the extracellular recognition kinases (ERK1/2) family of mitogen-activated protein (MAP) kinases (28) and phosphatidylinositol 3-kinase (PI 3-kinase) (25). The mechanisms by which ERK1/2 mediate ANG II induced-protein synthesis have not been fully identified but are thought to occur at the level of gene expression and/or the initiation of protein translation. The activation of PI 3-kinase and its downstream targets, Akt and the ribosomal p70S6 kinase, is critical for protein synthesis in many cell types, including VSMC (25). For example, p70S6 kinase is thought to be the major in vivo mediator of ribosomal S6 protein phosphorylation, a necessary step in ANG II-mediated protein synthesis in VSMC (8). In other cell types, both ERK1/2 and PI 3-kinase have been shown to regulate the phosphorylation of PHAS-1/eIF4E complex, a key regulator of translation initiation (16).
The precise molecular mechanisms that link AT1 receptor activation to the activation of the ERK1/2 and the PI 3-kinase signaling pathways have not been fully established. In other cell types, the proline-rich nonreceptor tyrosine kinase 2 (PYK2) has been shown to link G protein-coupled receptors to upstream regulators of ERK1/2 MAP kinases, such as Grb2, Shc, and the nucleotide exchange factor Sos (5). We have demonstrated that PYK2 is activated by ANG II in VSMC in a Ca2+- and protein kinase C (PKC)-dependent manner, resulting in its interaction with Src (23). More recent studies from Eguchi et. al. (6) suggest that interaction between Src and PYK2 leads to the recruitment of Grb2, a signaling intermediate in the ERK1/2 pathway. Thus PYK2 may be a key upstream regulator of ERK1/2 activation in VSMC.
Much less is known about the upstream signaling pathways that link
AT1 receptor activation to the PI 3-kinase pathway. In other cell types, regulation of PI 3-kinase involves interactions of
the Src homology 2 (SH2) domains of the regulatory p85 -subunit with
tyrosine phosphorylated proteins (14). One
potential regulator of PI 3-kinase in VSMC is the adapter molecule
p130Cas (a Crk-associated substrate). p130Cas, initially identified as
a major tyrosine phosphorylated protein in v-Crk and v-Src transformed
cells (10), also contains proline-rich sequences that may
allow it to act as a docking protein for the SH3 and SH2 domains of PI
3-kinase. In addition, p130Cas is tyrosine phosphorylated in response
to ANG II in VSMC in a PKC- and Ca2+-dependent manner
(26). Because PYK2 activation by ANG II requires PKC and
Ca2+, we hypothesized that this kinase could also mediate
p130Cas phosphorylation, and thus PI 3-kinase activation.
The present study investigates the signaling pathways in VSMC that link the AT1 receptor activation to ERK1/2 and PI 3-kinase signaling. We have identified Shc as an additional component of a signaling complex formed among PYK2, Src, and Grb2 in VSMC. We have shown, for the first time, that PYK2 forms a complex with PI 3-kinase in response to ANG II. In addition, our data indicate that PYK2 is involved in p130Cas phosphorylation and its subsequent association with PI 3-kinase in VSMC. Finally, we have shown that the activation of p70S6 kinase is dependent on PI 3-kinase, a tyrosine kinase, and Ca2+.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Cell culture. VSMC from thoracic aortas of 10- to 12-wk-old male Sprague-Dawley rats were isolated by enzymatic digestion as described previously (17). VSMC (passages 3-7) were maintained in culture at 37°C in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% calf serum, 100 U/ml penicillin, 10 µg/ml streptomycin, and 2 mM glutamine. VSMC were grown to ~75% confluency and then growth arrested for 48 h in serum-free DMEM supplemented with 0.1% BSA.
Immunoblot analysis. Cell lysates were prepared as described previously (17). Equal amounts of protein (30 µg) were resolved by 10% SDS-PAGE and transferred to nitrocellulose. Immunoblot analysis was performed using phosphorylation state-specific antibodies against ERK1/2 MAP kinases (1:5,000; Promega), PYK2 (pY881, 1:1,000; Biosource), or p70S6 kinase (1:1,000; New England Biolabs). Bands were visualized by enhanced chemiluminescence (ECL; Amersham) and quantified using NIH Image software.
Immunoprecipitation. Cell lysates were prepared for immunoprecipitation as described (23). Equal amounts of protein (600 µg) were immunoprecipitated with anti-pTyr, anti-Shc polyclonal antibodies, or anti-PYK2 monoclonal antibodies (Transduction Labs) overnight at 4°C. Immune complexes were collected by incubation with protein A-Sepharose or protein G-agarose beads for 2 h at 4°C. For immunoprecipitations using glutathione-S-transferase (GST)-fusion proteins conjugated to agarose beads, cell lysates were precleared with protein G for 30 min. Immunoprecipitates were separated by SDS-PAGE, and proteins were detected by immunoblotting as described above using anti-PYK2 (1:1,000), anti-p130Cas (1:1,000), anti-pTyr, or anti-Grb2 (1:2,000, Transduction Labs) monoclonal antibodies.
Data analysis. Blots shown are representative of at least n = 3 experiments. One-way repeated-measures analysis of variance (ANOVA) followed by Bonferroni's t-test were used for comparisons among multiple groups. Differences among means were considered significant at P < 0.05. Data were analyzed using InStat statistical software (GraphPad).
Materials.
PI 3-kinase p85 -subunit NH2- and COOH-SH2 domain
GST-fusion proteins conjugated to agarose beads and polyclonal PI
3-kinase antibodies were from Upstate Biotechnology. Anti-p130Cas,
anti-PYK2, and anti-phosphotyrosine antibodies were from Transduction
Laboratories. Anti-phosphoPYK2 was from Biosource. A monoclonal anti-PI
3-kinase antibody was from Panvera. Anti-phospho ERK1/2 were from
Promega. Anti-p70S6 kinase antibodies were from New England
Biolabs. 1,2-Bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA-AM) was from Alexis Laboratories. LY-294002 and genistein were purchased from Calbiochem.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Ca2+-dependent p130Cas
phosphorylation and PYK2-p130Cas complex formation in response to ANG
II.
In other cell types, PYK2 directly interacts with the docking
protein p130Cas (15). Once tyrosine is phosphorylated,
p130Cas can bind to SH2 domain-containing proteins. Therefore, we
speculated that p130Cas might link PYK2 to PI 3-kinase in VSMC because
the regulatory p85 -subunit of PI 3-kinase contains several SH2
domains. In Fig. 1A, we
determined whether ANG II induces p130Cas phosphorylation. VSMC were
treated with 100 nM ANG II for 0-60 min. VSMC were then lysed and
immunoprecipitated with a polyclonal anti-pTyr antibody or monoclonal
anti-p130Cas antibodies. Phosphorylated p130Cas in the
immunoprecipitates was measured by immunoblot analysis with a
monoclonal anti-p130Cas or anti-pTyr antibody. Little basal p130Cas
phosphorylation occurred in the absence of ANG II. Upon treatment with
ANG II, a rapid and sustained increase in p130Cas tyrosine
phosphorylation was observed as early as 1 min, was maximal at 2-5
min, and returned toward control levels at 30 min (Fig. 1A).
Pretreatment with BAPTA-AM to chelate intracellular Ca2+
inhibited ANG II-dependent p130Cas phosphorylation at all time points
(Fig. 1B).
|
ANG II stimulates a Ca2+-dependent
interaction among PYK2, p130Cas, and PI 3-kinase in VSMC.
We next determined whether phosphorylated p130Cas could bind to the
p85 regulatory subunit of PI 3-kinase. GST-fusion proteins containing the NH2- or COOH-terminal SH2 domains of PI
3-kinase p85
-subunit were immobilized on agarose beads and
incubated with lysates from control and ANG II-treated cells. After the
beads were washed, affinity-purified proteins were subjected to
SDS-PAGE and immunoblotted using anti-p130Cas or anti-PYK2 antibodies.
Under basal conditions, little p130Cas and PYK2 were bound to both SH2
domains of PI 3-kinase p85
. ANG II stimulation caused a significant
increase in the amount of p130Cas and PYK2 bound to the SH2 domains
(Figs. 2A and
3A). The time course for this
interaction was consistent with that for ANG II-mediated p130Cas
tyrosine phosphorylation. Pretreatment with 50 µM BAPTA-AM prevented
the association of the PI 3-kinase p85
-subunit SH2 domains with
p130Cas and PYK2 (Figs. 2B and 3B). To confirm
results obtained with the use of the SH2 domains of PI 3-kinase, we
also used coimmunoprecipitation analysis to demonstrate that ANG II
stimulates an endogenous complex formation between PI 3-kinase and
p130Cas and PYK2 (Figs. 2C and 3C). To confirm
equal lane loading, blots shown in Figs. 2C and
3C were stripped and reprobed with anti-PI 3-kinase and
anti-PYK2 antibodies.
|
|
Effects of PI 3-kinase inhibition on PYK2 and p130Cas activation.
To further strengthen the idea that both PYK2 and p130Cas lie upstream
of PI 3-kinase activation, we pretreated VSMC with the PI 3-kinase
inhibitors LY-294002 and wortmannin. ANG II-induced phosphorylation of
either PYK2 or p130Cas was not affected by these inhibitors (Fig.
4, A and B).
|
ANG II activates p70S6 kinase, a downstream target of
PI 3-kinase, in a Ca2+- and tyrosine
kinase-dependent manner.
To determine whether ANG II activates p70S6 kinase in a PI
3-kinase- and tyrosine kinase-dependent manner, we pretreated
growth-arrested VSMC with LY-294002, wortmannin, and genistein.
Activation was assessed with the use of anti-p70S6 kinase
antibodies that recognize the phosphorylated, active form of the kinase
(pThr-421/Ser-424 and pThr-401). ANG II-activation of p70S6
kinase was completely blocked by each inhibitor, suggesting that both
tyrosine kinase(s) and PI 3-kinase are required for activation (Fig.
5A). p70S6 kinase
phosphorylation was also completely inhibited by pretreatment with 50 µM BAPTA-AM (Fig. 5B). Drug treatment had no effect on p70S6 kinase expression as detected by Western blotting
with anti-total p70S6 kinase antibodies (Fig.
5C). The apparent higher molecular weight of the bands in
lanes 3-5 was due to a reduced mobility of the fully
phosphorylated form of p70S6 kinase. In fact, this band
shift detected in the total p70S6 kinase blot provides an
independent confirmation of the data in Fig. 5B.
|
PYK2 regulates upstream activators of ERK1/2.
We next identified some of the tyrosine kinases involved in the
regulation of the ERK1/2 signaling pathway. We previously showed that
ANG II causes a rapid activation of PYK2 that is associated with the
formation of a PYK2-Src complex (23). Because Src-induced tyrosine phosphorylation of the adapter molecule Shc leads to the
formation of a Shc-Grb2 complex (30), we next determined whether PYK2 is associated with these signaling intermediates. To assay
for PYK2-Shc-Grb2 complex formation, VSMC were treated with ANG II for
0-20 min and then lysed and immunoprecipitated with an anti-Shc
antibody. The PYK2-Shc complex formation was determined by
immunoblotting with a monoclonal anti-PYK2 antibody. PYK2-Shc complexes
were detected as early as 1 min and were maintained at 30 min (Fig.
6A). The blots were then
stripped and reprobed with anti-Grb2 antibodies and with anti-Shc
antibodies to confirm equal lane loading. The association of Shc with
Grb2 is also increased in ANG II-treated VSMC (Fig. 6A).
These results, along with our previous data (23), indicate
that treatment with ANG II stimulates the formation of a signaling
complex that consists of at least PYK2, Shc, Src, and Grb2. These
results were confirmed by immunoprecipitation with an anti-PYK2
antibody and blotting for Grb2 or Src (data not shown). The PYK2-Grb2
complex formation was associated with an increase in phosphorylation of
Y881 of PYK2 (Fig. 6B), which is necessary for PYK-Grb2
interaction in other cell types (4).
|
PI 3-kinase inhibition has no effect on ANG II-induced ERK1/2
activation.
Previous reports suggest that PI 3-kinase may regulate cell growth
through the activation of ERK1/2 MAP kinases (11, 22). In
these experiments, we pretreated VSMC with the specific PI 3-kinase
inhibitor LY-294002 (1-20 µM). PI 3-kinase inhibition had no
effect on ERK1/2 activation as measured by Western blotting with
anti-active ERK1/2 antibodies (Fig. 7).
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The intracellular signaling components that link AT1
receptors to VSMC growth involve a complex network of protein-protein interactions and kinase cascades. Recent evidence suggests that ERK1/2
MAP kinase (2) and PI 3-kinase (25) play a
crucial role in ANG II-induced VSMC hypertrophy, but little is known
about the intracellular signaling intermediates that link
AT1 receptor to these pathways. Here, we have shown, for
the first time, that the Ca2+-sensitive, nonreceptor
tyrosine kinase PYK2 links the AT1 receptor to the PI
3-kinase signaling pathway. Using coimmunoprecipitation analysis and
the SH2 domains of PI 3-kinase in "pull-down" assays, we have
demonstrated that PYK2 activation is associated with the tyrosine
phosphorylation of the adapter molecule p130Cas and complex formation
with the p85 regulatory subunit of PI 3-kinase.
In other cell types, the adapter molecule p130Cas is an important mediator of both PYK2 (15) and focal adhesion kinase (FAK) (10) signaling. Because of its combination of proline-rich sequences, an SH3 domain, and multiple tyrosine phosphorylation sites, p130Cas serves as a docking protein for SH2 and SH3 domain-containing proteins (24). Moreover, p130Cas can constitutively interact with proline-rich sequences in PYK2 via its SH3 domains (1). The present data provide the first evidence that ANG II stimulates the association of PYK2 with p130Cas in VSMC. There was a modest PYK2-p130Cas interaction in control cells that was dramatically increased with ANG II activation and tyrosine phosphorylation of PYK2 (Fig. 1).
Further support for an interaction between PYK2 and p130Cas comes from our finding that p130Cas tyrosine phosphorylation in response to ANG II in VSMC is Ca2+ dependent. These results are confirmed by Sayeski et al. (26), who demonstrated PKC- and Ca2+-dependent p130Cas tyrosine phosphorylation by ANG II in cultured VSMC. However, Takahashi et al. (29) observed not only that the tyrosine phosphorylation of p130Cas and its association with c-Crk II was Ca2+ and PKC independent but also that there was a lack of PYK2-p130Cas interaction in response to ANG II. The reasons for these discrepancies are unclear but may reflect differences in the passages of VSMC used or in the differences in the detergents used in the lysis buffer.
To gain further insight into the functional consequences of
p130Cas-PYK2 interaction in VSMC, we examined their interaction with PI
3-kinase. Our results provide the first demonstration of an interaction
among PYK2, p130Cas, and PI 3-kinase. Using the NH2- and
COOH-terminal SH2 domains of the p85 -subunit of PI 3-kinase coupled
to agarose beads, we were able to detect binding to both p130Cas and
PYK2 in a Ca2+-dependent manner (Figs. 2 and 3). This
interaction seems to be associated with the activation of PI 3-kinase
as determined by the LY-294002- and BAPTA-sensitive phosphorylation of
the downstream effector p70S6 kinase (Fig. 5). Furthermore,
the ANG II-dependent activation of p70S6 kinase was also
prevented by tyrosine kinase inhibition. However, neither LY-294002 nor
wortmannin had any effect on the tyrosine phosphorylation of either
PYK2 or p130Cas (Fig. 4). These findings, along with the time course
for ANG II-induced phosphorylation of these molecules, are consistent
with our hypothesis that PYK2 lies upstream of the PI 3-kinase pathway;
however, studies with dominant negative constructs of PYK2 are
necessary to conclusively confirm these findings.
In addition to establishing a link between PYK2 and PI 3-kinase, the role of PYK2 in ANG II-dependent ERK1/2 MAP kinase activation was also investigated. Previous data had shown that PYK2 autophosphorylates on Y402 upon activation, creating a binding site for the SH2 domain of Src (5). We have shown that Src forms a complex with PYK2 in response to ANG II in VSMC (23). In other cell types, recruitment and activation of Src leads to tyrosine phosphorylation of PYK2 at Y881 to create a binding site for the SH2 domain of Grb2 (4). In addition, Src may also tyrosine phosphorylate the adapter molecule Shc, leading to its interaction with Grb2 (30). Grb2 could then recruit the guanine nucleotide exchange factor Sos, leading to Ras activation and, ultimately, ERK1/2 stimulation (2). In the present study, we have demonstrated that PYK2 forms a complex with both Grb2 and Shc in VSMC (Fig. 6A) and that this association is Ca2+ and tyrosine kinase dependent (Rocic P and Lucchesi PA, unpublished observations). We have concluded that the association of PYK2 with Src is necessary for Shc-Grb2 complex formation and ERK1/2 stimulation. This conclusion is supported by recent work from Berk's laboratory (12), which demonstrated a role of Src in mediating ANG II-induced ERK1/2 activation, and from the work of Schieffer et al. (27), which defined a role for both Src and p21 in ANG II-induced VSMC growth. Eguchi et al. (6) also reported that a Grb2-GST fusion protein could immunoprecipitate PYK2 in VSMC but did not show an interaction with either endogenous Grb2 or Shc.
It is not clear whether PYK2 can directly interact with Shc because we were not consistently able to coimmunoprecipitate Shc with an anti-PYK2 antibody (data not shown). It may be that PYK2 interaction with Shc is indirect, mediated by Src. For example, Blaukat et al. (4) showed that Src is required not only for PYK2 regulation of Shc-Grb2 complex formation but also for the direct interaction of PYK2 and Grb2 via phosphorylation of Y881 of PYK2. In our experiments, the association of Shc, PYK2, and Grb2 correlated with an increase in the tyrosine phosphorylation of Y881 (Fig. 6B).
There is some evidence that PI 3-kinase is involved in the activation
of the ERK1/2 MAP kinase pathway (11). This modulation is
proposed to involve the activation of Ras by the p110 isoform of PI
3-kinase (22). In this study, LY-294002 as high as 20 µM
failed to block ANG II-induced ERK1/2 activation at all time points
examined (Fig. 7). Moreover, we were consistently unable to detect the
p110
-isoform in VSMC cell lysates by immunoblot analysis (data not
shown). Thus it appears that there is no cross talk between these
pathways in response to ANG II in VSMC.
Our data suggest that PYK2 is an upstream regulator of two parallel
signaling pathways, the ERK1/2 and the PI 3-kinase pathways, and thus
represents a bifurcation point for the ANG II signal (Fig.
8). The formation of a PYK2-Grb2
and/or the Src-Shc-Grb2 complex leads to ERK1/2 regulation indirectly
via activation of Ras. PYK2-dependent tyrosine phosphorylation and
interaction with the adapter molecule p130Cas lead to their association
with the p85 -subunit of PI 3-kinase. We speculate that this complex
formation leads to the activation of PI 3-kinase and its downstream
targets Akt and p70S6 kinase. Our results do not rule out
the involvement of other upstream tyrosine kinases or signaling
molecules. For example, p125FAK is activated by ANG II in
VSMC (18) and has been shown to regulate Src and p130Cas
(13).
|
In summary, this study establishes a role for PYK2 in linking AT1 receptor activation to two distinct signaling pathways in VSMC. The precise role this kinase plays in mediating ANG II-induced VSMC growth remains to be elucidated. Future studies with dominant negative constructs of PYK2 are needed to determine whether this kinase regulates rate-limiting steps necessary for VSMC hypertrophy in response to AT1 receptor activation.
![]() |
ACKNOWLEDGEMENTS |
---|
This work was supported by National Heart, Lung, and Blood Institute Grant HL-56046, American Heart Association National Grant-in Aid 96006970 (P. A. Lucchesi), and the John and Marian Falk Trust for Medical Research (A. Sabri).
![]() |
FOOTNOTES |
---|
Address for reprint requests and other correspondence: P. A. Lucchesi, UAB Dept. of Physiology and Biophysics, 986 MCLM, 1530 3rd Ave. S, Birmingham, AL 35294-0005 (E-mail: lucchesi{at}physiology.uab.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 11 May 2000; accepted in final form 7 August 2000.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Astier, A,
Avraham H,
Manie SN,
Groopman J,
Canty T,
Avraham S,
and
Freedman AS.
The related adhesion focal tyrosine kinase is tyrosine-phosphorylated after 1-integrin stimulation in B cells.
J Biol Chem
272:
228-232,
1997
2.
Berk, BC,
and
Corson MA.
Angiotensin II signal transduction in vascular smooth muscle.
Circ Res
80:
607-616,
1997
3.
Berk, BC,
Vekshtein V,
Gordon HM,
and
Tsuda T.
Angiotensin II-stimulated protein synthesis in cultured vascular smooth muscle cells.
Hypertension
13:
305-314,
1989[Abstract].
4.
Blaukat, A,
Ivankovic-Dikic I,
Grönroos E,
Dolfi F,
Tokiwa G,
Vuori K,
and
Dikic I.
Adaptor proteins Grb2 and Crk couple Pyk2 with activation of specific mitogen-activated protein kinase cascades.
J Biol Chem
274:
14893-14901,
1999
5.
Dikic, I,
Tokiwa G,
Lev S,
Courtneidge SA,
and
Schlessinger J.
A role for Pyk2 and Src in linking G-protein-coupled receptors with MAP kinase activation.
Nature
383:
547-550,
1996[ISI][Medline].
6.
Eguchi, S,
Iwasaki H,
Inagami T,
Numaguchi K,
Yamakawa T,
Motley ED,
Owada KM,
Marumo F,
and
Hirata Y.
Involvement of PYK2 in angiotensin II signaling of vascular smooth muscle cells.
Hypertension
33:
201-206,
1999
7.
Geisterfer, AA,
Peach MJ,
and
Owens GK.
Angiotensin II induces hypertrophy, not hyperplasia, of cultured rat aortic smooth muscle cells.
Circ Res
62:
749-756,
1988[Abstract].
8.
Giasson, E,
and
Meloche S.
Role of p70 S6 protein kinase in angiotensin II-induced protein synthesis in vascular smooth muscle cells.
J Biol Chem
270:
5225-5231,
1995
9.
Gibbons, GH,
Pratt RE,
and
Dzau VJ.
Vascular smooth muscle cell hypertrophy vs. hyperplasia Autocrine transforming growth factor-beta 1 expression determines growth response to angiotensin II.
J Clin Invest
90:
456-461,
1992[ISI][Medline].
10.
Harte, MT,
Hildebrand JD,
Burnham MR,
Bouton AH,
and
Parsons JT.
p130Cas, a substrate associated with v-Src and v-Crk, localizes to focal adhesions and binds to focal adhesion kinase.
J Biol Chem
271:
13649-13655,
1996
11.
Ikeda, M,
Kito H,
and
Sumpio B.
Phosphatidylinositol-3 kinase dependent MAP kinase activation via p21ras in endothelial cells exposed to cyclic strain.
Biochem Biophys Res Commun
257:
668-671,
1999[ISI][Medline].
12.
Ishida, M,
Ishida T,
Thomas SM,
and
Berk BC.
Activation of extracellular signal-regulated kinases (ERK1/2) by angiotensin II is dependent on c-Src in vascular smooth muscle cells.
Circ Res
82:
7-12,
1998
13.
Ishida, T,
Ishida M,
Suero J,
Takahashi M,
and
Berk BC.
Agonist-stimulated cytoskeletal reorganization and signal transduction at focal adhesions in vascular smooth muscle cells require c-Src.
J Clin Invest
103:
789-797,
1999
14.
Kapeller, R,
and
Cantley LC.
Phosphatidylinositol 3-kinase.
Bioessays
16:
565-576,
1994[ISI][Medline].
15.
Lakkakorpi, PT,
Nakamura I,
Nagy RM,
Parsons JT,
Rodan GA,
and
Duong LT.
Stable association of PYK2 and p130Cas in osteoclasts and their co-localization in the sealing zone.
J Biol Chem
274:
4900-4907,
1999
16.
Lawrence, JCJ,
and
Abraham RT.
PHAS/4E-BPs as regulators of mRNA translation and cell proliferation.
Trends Biochem Sci
22:
345-349,
1997[ISI][Medline].
17.
Lucchesi, PA,
Bell JM,
Willis LS,
Byron KL,
Corson MA,
and
Berk BC.
Calcium-dependent MAP kinase activation in SHR defines a hypertensive signal transduction phenotype.
Circ Res
78:
962-979,
1996
18.
Okuda, M,
Kawahara Y,
Nakayama I,
Hoshijima M,
and
Yokoyama M.
Angiotensin II transduces its signal to focal adhesions via angiotensin II type 1 receptors in vascular smooth muscle cells.
FEBS Lett
368:
343-347,
1995[ISI][Medline].
19.
Owens, GK.
Control of hypertrophic versus hyperplastic growth of vascular smooth muscle cells.
Am J Physiol Heart Circ Physiol
257:
H1755-H1765,
1989
20.
Pratt, RE.
Regulation of vascular smooth-muscle cell growth by angiotensin II.
Blood Press Suppl
2:
6-9,
1996[Medline].
21.
Rosendorff, C.
Renin-angiotensin system and vascular hypertrophy.
J Am Coll Cardiol
28:
803-812,
1996[ISI][Medline].
22.
Rubio, I,
Rodriguez-Viciana P,
Downward J,
and
Wetzker R.
Interaction of Ras with phosphoinositide 3-kinase .
Biochem J
326:
891-895,
1997[ISI][Medline].
23.
Sabri, A,
Govindarajan G,
Griffin TM,
Byron KL,
Samarel AM,
and
Lucchesi PA.
Calcium and protein kinase C dependent activation of the tyrosine kinase PYK2 by angiotensin II in vascular smooth muscle.
Circ Res
83:
841-851,
1998
24.
Sakai, R,
Iwamatsu A,
Hirano N,
Ogawa S,
Tanaka T,
Mano H,
Yazaki Y,
and
Hirai H.
A novel signaling molecule, p130, forms stable complexes in vivo with v-Crk and v-Src in a tyrosine phosphorylation-dependent manner.
EMBO J
13:
3748-3756,
1994[Abstract].
25.
Saward, L,
and
Zahradka P.
Angiotensin II activates phosphatidylinositol 3-kinase in vascular smooth muscle cells.
Circ Res
81:
249-257,
1997
26.
Sayeski, PP,
Ali MS,
Harp JB,
Marrero MB,
and
Bernstein KE.
Phosphorylation of p130Cas by angiotensin II is dependent on c-Src intracellular Ca2+ and protein kinase C.
Circ Res
82:
1279-1288,
1998
27.
Schieffer, B,
Drexler H,
Ling BL,
and
Marrero MB.
G protein-coupled receptors control vascular smooth muscle cell proliferation via pp60 and p21.
Am J Physiol Cell Physiol
272:
C2019-C2030,
1997
28.
Servant, MJ,
Giasson E,
and
Meloche S.
Inhibition of growth-factor induced protein synthesis by a selective MEK inhibitor in aortic smooth muscle cells.
J Biol Chem
271:
16047-16052,
1996
29.
Takahashi, T,
Kawahara T,
Taniguchi T,
and
Yokoyama M.
Tyrosine phosphorylation and association of p130 Cas and c-Crk II by ANG II in vascular smooth muscle cells.
Am J Physiol Heart Circ Physiol
274:
H1059-H1065,
1998
30.
Yamaguchi, M,
Tanaka T,
Wakai M,
Kitanaka A,
Kamano H,
Kubota Y,
Ohnishi H,
Takahara J,
and
Irino S.
Antisense Src expression inhibits tyrosine phosphorylation of Shc and its association with Grb2 and Sos which leads to MAP kinase activation in U937 human leukemia cells.
Leukemia
11:
497-503,
1997[ISI][Medline].