|
|
|
|||||||
|
|||||||||
(Received for publication, November 8, 1996, and in revised form, December 23, 1996)
From the Biology Department, Syracuse University,
Syracuse, New York 13244
ABSTRACT It is well established that mitogens inhibit
differentiation of skeletal muscle cells, but the insulin-like growth
factors (IGFs), acting through a single receptor, stimulate both
proliferation and differentiation of myoblasts. Although the IGF-I
mitogenic signaling pathway has been extensively studied in other cell
types, little is known about the signaling pathway leading to
differentiation in skeletal muscle. By using specific inhibitors of the
IGF signal transduction pathway, we have begun to define the signaling
intermediates mediating the two responses to IGFs. We found that
PD098059, an inhibitor of mitogen-activated protein (MAP) kinase kinase
activation, inhibited IGF-stimulated proliferation of L6A1 myoblasts
and the events associated with it, such as phosphorylation of the MAP kinases and elevation of c-fos mRNA and cyclin D protein.
Surprisingly, PD098059 caused a dramatic enhancement of
differentiation, evident both at a morphological (fusion of myoblasts
into myotubes) and biochemical level (elevation of myogenin and p21
cyclin-dependent kinase inhibitor expression, as well as
creatine kinase activity). In sharp contrast, LY294002, an inhibitor of
phosphatidylinositol 3-kinase, and rapamycin, an inhibitor of the
activation of p70 S6 kinase (p70S6k), completely abolished
IGF stimulation of L6A1 differentiation. We found that
p70S6k activity increased substantially during
differentiation, and this increase was further enhanced by PD098059.
Our results demonstrate that the MAP kinase pathway plays a primary
role in the mitogenic response and is inhibitory to the myogenic
response in L6A1 myoblasts, while activation of the
phosphatidylinositol 3-kinase/p70S6k pathway is essential
for IGF-stimulated differentiation. Thus, it appears that signaling
from the IGF-I receptor utilizes two distinct pathways leading either
to proliferation or differentiation.
INTRODUCTION It is widely believed that most mitogens stimulate skeletal muscle
cell proliferation but inhibit differentiation. However, the
insulin-like growth factors (IGF-I and
IGF-II)1 are unique among growth factors in
that they stimulate both proliferation and differentiation
of muscle cells in culture. Recently, it has been shown that IGF
actions occur through two phases (reviewed in Ref. 1). This involves an
initial mitogenic response in which intracellular mediators of
proliferation such as cyclin D mRNA and c-fos mRNA
are up-regulated and mediators of the myogenic response such as
elevation of myogenin mRNA are suppressed (2), followed by a strong
myogenic response characterized by expression of the muscle-specific
transcription factor myogenin and fusion of myoblasts into
myotubes.
Although IGF binding has been shown for both IGF receptors, the type I
IGF-I receptor and the type II IGF-II receptor, most biological actions
of IGF-I and IGF-II on L6A1 muscle cells appear to be mediated by the
type I IGF-I receptor (3). It is not obvious how stimulation of two
mutually exclusive processes such as proliferation and differentiation
can be mediated by the same receptor. A likely possibility is that
there is a step at which signal transduction pathways leading to
proliferation or differentiation diverge. Although many of the
signaling intermediates for the mitogenic actions of IGFs have been
described for other cell types, little is known about signal
transduction pathways leading to muscle differentiation.
The results of studies on signaling by the insulin and IGF-I receptors
in other cell types have revealed two primary pathways by which these
signals might be transmitted. Binding of IGF-I to the type I receptor
induces a conformational change resulting in receptor
autophosphorylation, followed by tyrosine phosphorylation of several
cellular substrates. Two primary substrates of the activated IGF
receptor are IRS-1 and Shc (4, 5). Phosphorylated tyrosines on IRS-1
serve as docking sites for multiple proteins containing SH-2 domains.
Among those proteins shown to bind to IRS-1 are the p85 regulatory
subunit of PI 3-kinase (6), SH-PTP2 (Syp) (7), the adaptor protein Nck
(8), and Grb-2 (9). This latter protein associates with mSos, which
activates Ras in turn activating the Raf-1/MAP kinase pathway (10).
Phosphorylated Shc also associates with Grb-2 and activates this
pathway (5, 11). Although both IRS-I and Shc can link the activated
IGF-I receptor with Grb-2/Sos, Sasaoka et al. (12) have
recently shown that the Shc·Grb-2/Sos complex is more important for
Ras activation.
A second pathway involving PI 3-kinases is also observed in
insulin/IGF-I receptor signal transduction (4, 13). Association of
IRS-1 with p85/p110 PI 3-kinase results in its activation, leading to
downstream phosphorylation and activation of the serine/threonine kinase p70S6k (14). This in turn leads to phosphorylation
of the ribosomal S6 protein and PHAS-I, resulting in an increased
translation of mRNAs containing polypyrimidine tracts (15). In
addition, Weng et al. (16) have shown that constitutively
active PI 3-kinase indirectly activates p70S6k, possibly
through the kinase Akt (17). However, p70S6k may also be
activated through a PI 3-kinase-independent pathway, since mutation of
the PDGF receptor has been shown to abolish PI 3-kinase activation in
transfected 293 cells, while p70S6k activation remains
unaffected (18). In addition, Lenormand et al. (19) have
recently shown that expression of an estradiol-regulated form of Raf-1
( In order to evaluate the relative importance of the MAP kinase and PI
3-kinase pathways in skeletal muscle proliferation and differentiation,
we have determined the effect of several inhibitors of these mediators
on L6A1 myoblasts. PD098059 is a noncompetitive inhibitor of MEK that
functions by blocking the activation of MEK by Raf-1 (20). Its
inhibitory effect on the MAP kinase pathway has been shown in several
cell types including mouse Swiss 3T3 fibroblasts and PC-12 cells (21)
and has been shown to block insulin-stimulated MAP kinase activation in
3T3-L1 adipocytes and L6 myotubes (22) but has no effect on a number of
other protein kinases (21). LY294002 is a PI 3-kinase inhibitor (23) that has recently been reported to block differentiation in L6E9 myoblasts (24). Rapamycin is an immunosuppressant drug known to block
progression of the cell cycle at the G1 phase. Rapamycin blocks activation of the serine/threonine kinase p70S6k
(25), resulting in the dephosphorylation of S6 ribosomal protein or
PHAS-I, leading to a decrease in translation of specific,
rapamycin-sensitive transcripts (15). Through the use of these
inhibitors we have separated the pathways involved in skeletal muscle
proliferation and differentiation and have shown that they diverge at a
very early point. Our studies demonstrate that the early stimulation of
cell proliferation involves the Ras/Raf-1/MAP kinase pathway, and the
later stimulation of differentiation utilizes the PI
3-kinase/p70S6k pathway.
EXPERIMENTAL PROCEDURES The IGF-I analog, R3 IGF-I, was a gift from Paul
Walton and John Ballard (GroPep Pty Ltd, Adelaide, Australia).
Rapamycin was purchased from ICN Pharmaceuticals, Inc. (Costa Mesa,
CA), and LY294002 was purchased from Calbiochem. The MEK inhibitor, PD098059, was a gift from Alan Saltiel (Parke-Davis Pharmaceuticals, Ann Arbor, MI). The c-fos cDNA probe was obtained from
ATCC (Rockville, MD), and the myogenin cDNA probe (E26) was a gift
from Eric N. Olson (University of Texas Southwestern Medical Center,
Dallas, TX). The anti-cyclin D antibody, p70S6k assay kit,
and recombinant protein A-agarose were from Upstate Biotechnology, Inc.
(Lake Placid, NY). Anti-phospho-MAPK antibody was purchased from New
England Biolabs, Inc. (Beverly, MA). Hybridoma cells secreting the
anti-myogenin antibody were a gift from W. R. Wright (University of
Texas, Austin, TX). Goat anti-rabbit and anti-mouse horseradish
peroxidase-conjugated IgG, the BCA protein assay reagents, and the
SuperSignal CL-HRP substrate system for enhanced chemiluminescence
(ECL) were purchased from Pierce. Tissue culture medium components were
purchased from Life Technologies, Inc. All other chemicals were from
Sigma.
L6A1 myoblasts were grown in Dulbecco's
modified Eagle's medium (DMEM) containing 10% horse serum, 1% chick
embryo extract, and 1% antibiotic, antimycotic solution (penicillin,
streptomycin, and amphotericin B). Cultures were plated at 1.2 × 105 cells/35-mm dish for cell number and creatine kinase
(CK) determination and at 1.2 × 106 cells/100-mm dish
for RNA extraction or immunoprecipitation of proteins. After incubation
overnight, the cultures were washed with DMEM before the addition of
IGFs or inhibitors in DMEM containing 0.5 mg/ml bovine serum albumin.
Stock solutions of inhibitors were dissolved in Me2SO at
the following concentrations and stored at To
quantitate cell proliferation, cells were trypsinized and counted in a
model ZBI Coulter Counter 1 day after the addition of growth factors
and inhibitors. Differentiation was quantitated 3 days after the
addition of growth factors and inhibitors by measuring the elevation in
CK activity using the NAD-coupled microtiter assay (27); CK levels were
normalized to DNA content as described previously (28).
Total RNA was isolated from cultures
using the RNeasy total RNA kit (Qiagen, Chatsworth, CA) according to
the manufacturer's instructions. Ten-µg samples were analyzed on
Northern blots. Consistency of RNA loading was monitored by
visualization of ethidium bromide-stained ribosomal RNA bands.
32P-Labeled probe for c-fos mRNA was
prepared by random priming of the 1-kilobase pair PstI
restriction fragment, for myogenin the 1.1-kilobase pair
BamHI/EcoRI restriction fragment, and for p21 the
420-base pair EcoRI restriction fragment. Blots were
hybridized at 42 °C and washed as described previously (29). The
autoradiographs and photographs were scanned with a Microtek Scanmaker
II at 600 dots per inch using Adobe Photoshop software.
Total cell lysates were
prepared by washing the cell monolayers (100-mm dish) twice with
ice-cold PBS followed by lysis in 0.5 ml of boiling 1 × Laemmli
sample buffer containing 100 mM dithiothreitol. Proteins
(10 µg) were separated by 10% SDS-polyacrylamide gel electrophoresis
following the procedures of Laemmli (30) and transferred onto
Immobilon-nitrocellulose (Millipore Corp.) in transfer buffer
containing 48 mM Tris base, 39 mM glycine, 0.037% SDS, and 20% methanol at 0.8 mA/cm2 for 1 h
using a SemiPhor Transfer Unit (Hoefer). The blots were blocked for
1 h at room temperature in blocking buffer containing 5% nonfat
dry milk in PBST (10 mM phosphate buffer, pH 7.5, 150 mM NaCl, 0.1% Tween 20) and incubated overnight at 4 °C
with antibodies against cyclin D1 (1 µg/ml) or phospho-MAPK (1:1000)
in 5% milk, PBST, or myogenin (1:50) in 1% milk, 10% goat serum,
PBST. After washing six times in PBST, the membranes were incubated for
1 h at room temperature with horseradish peroxidase-conjugated
goat anti-rabbit IgG diluted 1:3000 in 5% milk, PBST (except myogenin, which was incubated with horseradish peroxidase-conjugated goat anti-mouse IgG, 1:2000 dilution). The blots were washed six times in
PBST at room temperature, and immunolabeling was detected by ECL
according to the manufacturer's directions.
The cell monolayers in 100-mm dishes
were washed twice with ice-cold PBS, and the cells were lysed by
incubating the cultures for 20 min in 1 ml of cold modified
radioimmunoprecipitation assay buffer (50 mM Tris-HCl, pH
7.4, 1% Nonidet P-40, 0.25% sodium deoxycholate, 150 mM
NaCl, 1 mM EGTA, 1 mM PMSF, 1 µg/ml
aprotinin, 1 µg/ml leupeptin, 1 µg/ml pepstatin, 1 mM
Na3VO4, 1 mM NaF). The lysates were
centrifuged for 10 min in a microcentrifuge to remove any insoluble
material. The protein content of the supernatants was determined by the
BCA method. p70S6k activities were assayed by following a
modification of the manufacturer's protocols. After cell lysis in
radioimmunoprecipitation assay buffer, 200 µg of protein were
incubated with 1 µg of anti-p70S6k antibody for 1 h
at 4 °C. The immunocomplex was captured with 50 µl of protein
A-agarose (50% slurry) for 1 h at 4 °C. The beads were washed
two times with an equal volume of cold PBS followed by one wash with
assay dilution buffer. The beads were resuspended in 20 µl of assay
dilution buffer, 10 µl of substrate, 10 µl of inhibitor mixture,
and 10 µl of [ RESULTS The actions of IGF-I are modified by six
IGF-binding proteins (IGFBPs), three of which are secreted by L6A1
muscle cells and have both inhibitory and stimulatory effects on
proliferation and differentiation (Refs.
31-33).2 In order to minimize the actions
of these IGFBPs in this study, we used an analog of IGF-I with a low
affinity for IGFBPs, R3 IGF-I. We have previously reported that several
similar IGF-I analogs with reduced affinity for IGFBPs were not only
more potent than IGF-I in stimulating L6A1 myoblast proliferation and
differentiation, but caused much more extensive myotube formation (31).
The effects of adding increasing concentrations of R3 IGF-I on L6A1
cell proliferation and differentiation are shown in Fig.
1. Stimulation of proliferation after 1 day reached a
maximum at approximately 30-100 ng/ml R3 IGF-I. Maximum stimulation of
differentiation measured after 3 days was attained with 1-3 ng/ml of
R3 IGF-I, while exposure to higher concentrations resulted in
progressively lower CK levels accompanied by decreased fusion of
myoblasts into myotubes as well as increased cell division; this
biphasic response was also seen at higher concentrations of unmodified
IGF-I (34). Therefore, in subsequent studies, R3 IGF-I was used at low
concentrations of 1-3 ng/ml in experiments monitoring inhibitor
effects on differentiation and at higher concentrations of 10-30 ng/ml
when we studied effects on proliferation.
In
order to determine which signaling pathways play a role in mediating R3
IGF-I-stimulated proliferation and differentiation, we used three
compounds that have been reported to inhibit specific signaling
molecules in other cells. PD098059 specifically inhibits the activation
of MEK by Raf-1, thus suppressing MAP kinase activation (20). LY294002
is a specific inhibitor of PI 3-kinase (23), and rapamycin treatment
inhibits p70S6k (25). When these inhibitors were added in
the presence of R3 IGF-I, we observed striking differences in the
response of L6A1 myoblast cultures. Fig. 2 summarizes
the concentration dependences of the effects of each of the three
inhibitors on cell proliferation and differentiation when added in the
presence of 3 ng/ml R3 IGF-I. As shown in Fig. 2, increasing
concentrations of PD098059 gave progressive decreases in the mitogenic
response to R3-IGF-I, inhibiting proliferation approximately 90% at
concentrations of 30 µM, while the same levels of
inhibitor showed a dramatic 3-fold enhancement of R3 IGF-I-stimulated
differentiation quantitated as CK in units/mg of DNA. In sharp
contrast, incubation with increasing concentrations of either LY294002
or rapamycin progressively inhibited the myogenic response to R3 IGF-I
but had much less effect on the mitogenic response. At concentrations
of 10 µM and 1 ng/ml, LY294002 and rapamycin,
respectively, completely abolished R3 IGF-I-stimulated differentiation
but inhibited cell proliferation only 30-40%.
There were equally striking changes in cell morphology following
treatment with these inhibitors. Upon the removal of growth medium,
cells maintained in serum-free DMEM remained relatively quiescent, with
little, if any cell division or fusion into myotubes (Fig.
3A). After the addition of R3 IGF-I to L6A1
cultures, the myoblasts first underwent a round of cell division before
fusing into myotubes; after 2 days the cultures consisted of both
unfused myoblasts and differentiated myotubes (Fig. 3B).
However, when PD098059 was added in the presence of R3 IGF-I, there was
less cell division, and the myoblasts began to differentiate sooner, forming an extensive network of myotubes; after 2 days in culture, few
if any unfused myoblasts could be detected (Fig. 3C). In
sharp contrast, differentiation was completely inhibited in cultures treated with either LY294002 or rapamycin in the presence of R3 IGF-I
(Fig. 3, D and E). Myoblasts treated with these
two inhibitors continued to proliferate, as demonstrated by the
increased cell density when compared with those in DMEM, but they did
not fuse into myotubes. These results (Figs. 2 and 3) suggest that
signaling by the MAP kinase pathway is required for R3 IGF-I-stimulated L6A1 proliferation, but not for differentiation, and that inhibition of
this pathway by PD098059 permits, and even enhances, R3
IGF-I-stimulated L6A1 myoblast differentiation. In contrast, a
functional PI 3-kinase/p70S6k pathway is an absolute
requirement for IGF-stimulated differentiation.
As shown in Figs. 2 and
3, the inhibitors used here have dramatically opposite effects on
responses of myoblasts to R3 IGF-I. This raises the question of which
effect predominates when cells are incubated with combinations of these
inhibitors. As illustrated in Fig. 4B, the
results are unequivocal; the inhibitory effects of LY294002 and
rapamycin overcome both the stimulation of myogenesis by R3 IGF-I and
its enhancement by PD098059. Differentiation of myoblasts in response
to R3 IGF-I is completely blocked by these inhibitors, both in the
absence and presence of PD098059; morphological differentiation
(i.e. fusion) of the cultures closely parallels the
quantitation by CK levels presented in Fig. 4B. Thus, it
appears that the PI 3-kinase/p70S6k pathway is essential
for myogenesis even when the mitogenic response through MAP kinase is
blocked.
The results presented in Fig. 4A indicate a major effect of
the MAP kinase pathway and a smaller contribution of the PI
3-kinase/p70S6k pathway to the mitogenic response; there
was much greater inhibition of proliferation by PD098059 than by either
of the other inhibitors, but when added in combination with PD098059,
there was an even lower level of cell proliferation; indeed, there was
some loss of cells below the level found in DMEM controls.
We usually measure proliferative responses of L6A1
myoblasts to mitogenic stimuli by counting the actual number of cells
in the cultures 1 day after the specific treatment. In addition, we
also determined the effects of these inhibitors on several intracellular mediators closely associated with early aspects of the
proliferative response in other cell systems. Of these, c-fos mRNA is an established early marker for cell
proliferation and has been shown to be induced during IGF-I-stimulated
mitogenesis in L6 myoblasts (35, 36). We investigated the effects of
PD098059, LY294002, and rapamycin on c-fos mRNA levels
after 30 min as shown in Fig. 5A. In the
presence of R3 IGF-I, c-fos mRNA levels were enhanced
about 3-fold above control levels. As expected, this increase in
c-fos mRNA was significantly reduced in the presence of
the MEK inhibitor, PD098059. In contrast, LY294002 and rapamycin, at
concentrations that abolished myogenic differentiation, had little or
no effect on c-fos mRNA levels compared with R3 IGF-I treatment alone. Both combinations of inhibitors resulted in decreased c-fos mRNA levels similar to those seen for PD098059
plus R3 IGF-I alone.
Effects of inhibitors on specific mediators
of the early proliferative response to R3 IGF-I. L6A1 myoblast
cultures were established overnight in growth medium at 1.2 × 106 cells/100-mm dish, washed once with DMEM, and then
treated with R3 IGF-I (30 ng/ml for c-fos mRNA and 10 ng/ml for phospho-MAPK and cyclin D determinations) in the absence or
presence of PD098059 (30 µM), LY294002 (10 µM), or rapamycin (1 ng/ml). The following analyses were
made in parallel cultures. A, c-fos mRNA.
Total RNA was prepared 30 min after the addition of R3 IGF-I with
or without inhibitors, 10 µg was analyzed on Northern blots (middle part), and
c-fos mRNA abundance was quantitated by densitometry (upper part); equal loading was verified with ethidium
bromide-stained ribosomal 18 S RNA (lower part).
B, phospho-MAPK. Cell lysates were prepared 1 h after
the addition of R3 IGF-I with or without inhibitors, and 10 µg of
protein was separated by 10% SDS-polyacrylamide gel electrophoresis
and immunoblotted with anti-phospho-MAPK. The Western blot for
phospho-MAPK is shown in the lower part, and the
densitometric quantitation of ERK 1 (bars on
left) and ERK 2 (bars on right) is
shown in the upper part. C, cyclin D. Cell
lysates were prepared 4 h after the addition of R3 IGF-I with or
without inhibitors, and 10 µg of protein was separated by 10%
SDS-polyacrylamide gel electrophoresis and immunoblotted with an
antibody to cyclin D. The Western blot is shown in the lower
part, and the densitometric quantitation is shown in the upper part.
The primary substrates of MEK are the p42/p44MAPK isoforms,
ERK 2 and ERK 1, respectively (37). PD098059 prevents the activation of
MEK, thereby inhibiting phosphorylation and activation of MAP kinases.
It has been postulated that the MAP kinases are required for activation
of the G1 cyclin-dependent complexes (38).
Therefore, levels of phosphorylated MAP kinase were measured in cells
incubated with PD098059, LY294002, and rapamycin, both individually and in combination. As shown in Fig. 5B, PD098059 had the
expected effect of inhibiting R3 IGF-I-induced phosphorylation of both ERK 1 and 2 within 1 h. LY294002 and rapamycin, on the other hand, had little or no effect. When LY294002 and rapamycin were added in
combination with PD098059, the inhibition was equivalent to PD098059 on
its own. This confirms that the action of PD098059 is upstream of MAP
kinase and that LY294002 and rapamycin have no effect on the MAP kinase
pathway or, therefore, on c-fos transcription.
The inhibitory effects of PD098059 on the levels of cyclin D1, a
protein essential to cell cycle progression, were similar to, although
not as dramatic as those seen for phosphorylated MAP kinase. However,
PD098059 in the presence of either LY294002 or rapamycin was almost
completely inhibitory (Fig. 5C). This suggests that although
proliferation occurs primarily through the MAP kinase pathway,
inhibiting this pathway does not block all aspects of the proliferative
response, suggesting that the PI 3-kinase/p70S6k pathway
may also be involved. However, since R3 IGF-I and LY294002 or rapamycin
in the absence of PD098059 had no effect on cyclin D1 levels, the
proliferative role of the PI 3-kinase/p70S6k pathway is
most probably minor.
In a similar study on early mediators of
differentiation, we measured levels of myogenin and p21 mRNA 18 and
42 h after the R3 IGF-I addition in the absence or presence of the
inhibitors (Fig. 6). Myogenin is a
basic-helix-loop-helix transcription factor of the MyoD family that is
required for terminal differentiation and myotube formation (reviewed
in Ref. 1). Induction of the mRNA for the
cyclin-dependent kinase inhibitor, p21, is an early event
in myogenic differentiation marking the exit of myoblasts from the cell
cycle (39).
Effects of inhibitors on R3 IGF-I stimulation
of myogenin and the cyclin-dependent kinase inhibitor,
p21. Cultures were prepared and treated as described under Fig. 5, except that R3 IGF-I was added at 3 ng/ml. The following analyses were made in
parallel cultures. A, myogenin mRNA. Total RNA was
prepared 18 and 42 h after the addition of R3 IGF-I with or
without inhibitors, 10 µg was analyzed on Northern blots
(middle part), and myogenin mRNA abundance was
quantitated by densitometry (upper part); equal loading was
verified with ethidium bromide-stained ribosomal 18 S RNA (lower
part). B, myogenin protein. Total cell lysates were prepared, and 10 µg of total protein was separated by 10%
SDS-polyacrylamide gel electrophoresis and immunoblotted with
anti-myogenin. The Western blot is shown in the lower part,
and densitometric quantitation is shown in the upper part.
C, p21 mRNA. RNA preparation and Northern analyses were
performed as described in A for myogenin.
At early times of incubation (18 h), R3 IGF-I-treated cells exhibited
very little increase in myogenin mRNA compared with DMEM control
cells (Fig. 6A). In the presence of R3 IGF-I and PD098059,
myogenin mRNA levels were elevated more than 3-fold compared with
R3 IGF-I treatment alone, indicating an early enhancement of the
myogenic response. LY294002 and rapamycin-treated cells showed a
significant reduction in myogenin mRNA at that time. The
combination of PD098059 with rapamycin and LY294002 reduced the
enhancing effect of PD098059 on myogenin mRNA levels, although they
were still slightly elevated above control and R3 IGF-I levels.
By 42 h, myogenin mRNA in R3 IGF-I-treated cells was elevated
almost 4-fold above DMEM-treated control cells. This is consistent with
observations that myogenin mRNA levels rise rather late after the
IGF-I addition (2, 29). Cells treated with R3 IGF-I and PD098059 had
myogenin levels similar to those in R3 IGF-I-treated cells;
i.e. by this late time, myogenin levels in R3 IGF-I-treated cells reached those levels seen in cells treated with both R3 IGF-I and
PD098059. LY294002 and rapamycin with R3 IGF-I alone or in combination
with PD098059 showed patterns similar to those seen at 18 h.
At 18 h, myogenin protein levels correlated well with mRNA
values showing enhanced myogenin protein in the presence of R3 IGF-I
and PD098059 (Fig. 6B). The effect of the inhibitors on myogenin protein levels was less pronounced by 48 h, although R3
IGF-I-treated cells showed enhanced levels of myogenin compared with
DMEM-treated cells. This suggests that translation of myogenin mRNA
may be affected differently than transcription of the myogenin gene in
the presence of these inhibitors.
Levels of p21 mRNA were also elevated in the presence of R3 IGF-I
and R3 IGF-I plus PD098059 at 18 and 42 h (Fig. 6C).
However, the inhibitors LY294002 and rapamycin, alone or in combination with PD098059, completely blocked p21 mRNA expression compared with
DMEM-treated cells, again suggesting the importance of the PI
3-kinase/p70S6k pathway in differentiation. These results
reflect the well established coordination of cell cycle exit with the
onset of myogenic differentiation.
Results
with the inhibitors LY294002 and rapamycin strongly indicated a major
role for the PI 3-kinase/p70S6k pathway in the stimulation
of myogenesis by the IGFs, suggesting that p70S6k activity
should be elevated during differentiation. To test this possibility, we
measured the enzymatic activity of p70S6k in lysates from
cells treated with R3 IGF-I as shown in Fig. 7. R3 IGF-I
increased p70S6k activity above control levels over an
extended period of time (48 h), showing an initial increase over
control values (Fig. 7, inset), followed by a much larger
increase at 48 h. The later activity was substantially elevated in
the presence of PD098059, just as was differentiation of the cells.
Treatment with rapamycin and LY294002 reduced p70S6k
activity to the level found in DMEM controls, demonstrating that inhibition of p70S6k activity parallels the effects of
these inhibitors on myogenesis. These results suggest that the early
increase in p70S6k activity may contribute to the
relatively rapid mitogenic response, while the larger subsequent
increase is necessary for the stimulation of L6A1 myoblast
differentiation.
DISCUSSION The results presented here represent the first steps toward
understanding the signaling pathways involved in two major responses of
myoblasts to the IGFs. When we (40) first reported that the known
mitogens, IGFs, stimulate differentiation as well as proliferation of
skeletal muscle cells in culture, there was some reluctance to accept
this observation because of the long held (41) and well supported
(42-45) view that mitogens block myogenesis by forcing myoblasts in
the G1 stage of the cell cycle to reenter S phase rather
than to fuse to form postmitotic myotubes. However, the stimulation of
myogenic differentiation by the IGFs has been demonstrated independently in a number of laboratories (reviewed in Ref. 1), and the
effect is now well established although not fully explained.
It has been difficult to understand how the same agent, IGF-I (or
IGF-II or insulin at high concentrations), acting through a single
receptor (3), can stimulate these two responses, which are generally
considered to be mutually antagonistic. One suggested explanation for
our observations was that IGFs acted only to maintain the cells in a
viable state, thus allowing differentiation rather than
actively stimulating the process. In earlier studies, we demonstrated that IGFs did not stimulate differentiation solely by
preventing cell death by showing that the IGFs caused significant differentiation in medium containing horse serum at levels as high as
5% (34). Another possibility, that the greater fusion rate might
result from the higher cell density in cultures stimulated to
proliferate by IGF-I, was eliminated by experiments (46) that showed
that the stimulation of myogenesis by IGFs was readily detectable in
cells incubated under conditions in which DNA synthesis was blocked or
proliferating cells were killed by cytosine arabinoside. However, it
has recently been shown that IGF-II does, in fact, act as an autocrine
survival factor for differentiating myoblasts (47). Although our
earlier results show that this does not account for the stimulation of
differentiation, we had previously shown that IGF-II was produced by
differentiating muscle cells and that the rate of differentiation of
different muscle cell lines correlated with the amount of IGF-II
secreted by the cell (48). Additional direct evidence implicating
IGF-II as an autocrine myogenic factor was provided by Stewart et
al. (49) and Quinn et al. (50), who showed that
overexpression of IGF-II or the type I IGF-I receptor in C2 cells
resulted in greatly accelerated differentiation. Thus, it appears that
although the IGFs may act as survival factors and stimulate
proliferation, the role of IGFs as positive regulators of myogenesis is
firmly established.
A beginning of an explanation of the two responses (proliferation and
differentiation) to IGFs was provided by the finding that there is a
temporal separation in the responses of myoblasts to IGFs; cells first
proliferate and then differentiate (2, 28). But this does not explain
how myoblasts can respond in the two different ways or what signals
mediate those responses.
The results presented here suggest that the signaling pathways leading
to proliferation or differentiation diverge at a very early step,
possibly as early as the interaction of the IGF-I receptor or of IRS-1
with intracellular signaling pathways. As in many other cell types, the
stimulation of proliferation in L6A1 skeletal muscle cells by IGF
appears to be mediated by the Ras/Raf-1/MAP kinase pathway. Several
years ago it was shown that overexpression of ras oncogenes
in myoblasts inhibited muscle differentiation (51, 52). Since Ras
activation is a critical component of the signaling pathway leading to
MAP kinase activation and proliferation, it is reasonable to predict
that the stimulation of the MAP kinase pathway would inhibit
differentiation and that inhibition of the MAP kinase pathway would
stimulate differentiation. We have now shown that the addition of
PD098059, the specific inhibitor of MEK activation by Raf-1, not only
blocked cell proliferation, but resulted in a dramatic increase in
myogenesis above that obtained with R3 IGF-I alone (Figs. 2 and 3).
This was evident not only by morphologic changes, but also by changes
in specific mediators of proliferation and differentiation. Levels of
mRNA for the transcription factor, c-fos, an early
marker of proliferation, and of cyclin D protein were suppressed by
PD098059 (Fig. 5, A and C). In contrast, the
increase in myogenin mRNA and protein was significantly enhanced in
the presence of PD098059, particularly during early stages of
differentiation (18 h) (Fig. 6, A and B) when
myogenin levels are not normally elevated in IGF-I-treated myoblasts
(2, 29). This suggests the possibility that proliferation is the
primary response of muscle cells to IGFs and that differentiation
occurs when there is an interruption of the MAP kinase pathway. What external stimuli bring about this inhibition in vivo is
unclear. Our results are similar to those of Lazar et al.
(22) who, using PD098059 to block MEK activation by insulin,
demonstrated that the MAP kinase pathway is not required for many of
the metabolic actions of insulin, such as glucose uptake, lipogenesis,
and glycogen synthesis. However, in their studies, PD098059 treatment
did not result in additional stimulation of any of the metabolic
actions as we have observed in the case of IGF-stimulated L6A1
myogenesis.
In apparent disagreement with our finding that activation of the MAP
kinase pathway is not required for L6A1 myogenesis, Hashimoto et
al. (53) concluded that activation of MAP kinase played a positive
role in the expression of myogenin and subsequent differentiation of
C2C12 myoblasts. Their conclusion was based on their observations that
genistein, a tyrosine kinase inhibitor, inhibited phosphorylation of
MAP kinase, myogenin expression, and fusion of myoblasts into myotubes.
Since genistein is a general inhibitor that can affect the
phosphorylation of many signaling intermediates, it is possible that
the inhibitory effects on myogenesis could result from effects on
mediators other than MAP kinase. When we added genistein to L6A1
myoblasts at concentrations lower than the 50 µM used by Hashimoto et al., we found that it inhibited proliferation
as well as differentiation (data not shown), and concluded that this was not a useful inhibitor for our studies on the separate
responses.
The importance of the MAP kinase pathway in proliferation has been
shown in several cell types for many different mitogens (37). Lavoie
et al. (38) have reported that activation of the
p42/p44MAPK isoforms, which leads to activation of various
transcription factors, also results in positive regulation of cyclin D1
expression. Growth factor-dependent accumulation of cyclin
D1 has been shown to be necessary in order for cells to pass the
G1 restriction point of the cell cycle (54).
As shown in Fig. 5, A and B, neither LY294002 nor
rapamycin was inhibitory to ERK 1 and 2 phosphorylation or
c-fos expression, and neither further enhanced the
inhibition of ERK 1 and ERK 2 phosphorylation by PD098059. However, the
decrease in cyclin D1 levels at 4 h by PD098059 (Fig.
5C), although significant, was further enhanced by either
LY294002 or rapamycin, although neither had any effect individually.
Cyclin D1 has been postulated to be the "nuclear sensor" of
extracellular signals (38), and our results indicate that cyclin D1
levels may be regulated by both the MAP kinase and the PI
3-kinase/p70S6k pathways. This suggests that although the
MAP kinase pathway is essential for proliferation, the
p70S6k pathway also plays a role. This is further
substantiated by our finding (Fig. 7) that there was an initial
increase of p70S6k activity 30 min after R3 IGF-I
stimulation, a time when proliferative signals such as c-fos
mRNA are up-regulated. IGF-I, at 80 ng/ml, which gives a greater
mitogenic response than R3 IGF-I, caused an even greater early increase
in p70S6k activity (data not shown), supporting the role of
p70S6k in proliferation. At the high levels required for
complete suppression of myogenesis, both LY294002 and rapamycin exhibit
some (40%) inhibition of the proliferative response to R3 IGF-I.
However, it appears that the PI 3-kinase/p70S6k pathway
plays a much more critical role in the myogenic response to IGFs.
Inhibition of PI 3-kinase or p70S6k by LY294002 or
rapamycin, respectively, resulted in little or no myotube formation
(Fig. 3) accompanied by sharply decreased levels of myogenin and p21
mRNA (Fig. 6). In addition, p70S6k activity increased
with time as R3 IGF-I-stimulated differentiation proceeded (Fig. 7).
This elevated activity was further enhanced by PD098059, suggesting
that inhibition of the MAP kinase pathway stimulates differentiation.
The increase in p70S6k activity was completely abolished in
the presence of LY294002 and rapamycin (Fig. 7), supporting our
interpretation that myogenic signaling proceeds from PI 3-kinase to
p70S6k. Our results are in direct contrast with those
reported by Jayaraman and Marks (55), who concluded that rapamycin
treatment of BC3H1 myoblasts induced differentiation. BC3H1 myoblasts
are not typical of most skeletal muscle cells, since they do not fuse
to form myotubes, and biochemical differentiation can be reversed. In the cited study, much higher levels of rapamycin were used, and elevation of Our results also suggest that changes in the expression of biochemical
markers alone do not indicate differentiation. Myogenin mRNA and
protein levels were elevated at 42 h in cells treated with R3
IGF-I and the combination of PD098059 and LY294002 or rapamycin (Fig.
6). However, under these conditions, no differentiation was seen to
occur (Fig. 4B). Therefore, elevated myogenin levels alone
are not sufficient to induce differentiation; LY294002 and rapamycin
may be inhibiting an additional component of the differentiation pathway, possibly the myogenic factor MEF-2 (56).
As this manuscript was being completed, Pilch's group (57) published
the results of a study of effects of FGF and IGF on the proliferative
response of C2C12 myoblasts. They found complete inhibition of MAP
kinase activation but only partial inhibition of
[3H]thymidine incorporation by PD098059 and concluded
that a MAP kinase-independent pathway makes a substantial contribution
to the mitogenic response to IGF-I. To the extent that they overlap, our results are in good overall agreement with theirs; we too find that
PI 3-kinase activation plays some role in the mitogenic response, and
the myogenic response is suppressed by LY294002, which was not used by
Milasincic et al. (57).
Our findings are consistent with the recent report of Kaliman et
al. (24), who also observed that LY294002 and wortmannin inhibited
the capacity of L6E9 myoblasts to form myotubes when the cultures were
placed in low serum differentiation medium and concluded that PI
3-kinase activation was required for differentiation. The effects of
IGFs on their system were not considered, although our previous
observations of autocrine expression of IGFs by myoblasts in low serum
medium (48) indicate that IGFs may have been acting in those cells as
well.
Although other mitogens have been shown to stimulate PI 3-kinase
activity in non-muscle cells, we have found no reports of stimulation
of PI 3-kinase in muscle by FGF, PDGF, or EGF. Milasincic et
al. (57) have recently reported that in C2C12 cells, IGF-I and
insulin increase PI 3-kinase activity by approximately 2-fold, whereas
basic FGF, a potent inhibitor of myogenesis, was inconsistent in
stimulating PI 3-kinase activity. Also, Kudla et al. (58) have shown that PDGF-BB stimulation of MM14 cells overexpressing the
PDGF- We cannot rule out the possibility that these other mitogens may also
act through PI 3-kinase in skeletal muscle. It has already been shown
that different mitogens acting through the same signaling pathway can
generate separate responses. In PC12 cells, FGF or nerve growth factor
stimulation results in differentiation, whereas EGF stimulation leads
to increased cell proliferation (reviewed in Ref. 62). All of these
mitogens stimulate MAP kinase activity; however, the duration of the
MAP kinase signal is sustained in FGF/nerve growth factor stimulation
but is transient in EGF stimulation. In addition to signal duration,
other factors may potentially play a role in modulating signaling
pathways for different mitogenic stimuli.
The role of Raf-1 and its importance in linking Ras to the MAP kinase
pathway is well established. However, some studies suggest that Raf-1
may have actions other than activation of the Ras/MAP kinase pathway.
Alessi et al. (21) have reported that inhibition of MEK by
PD098059 in PDGF- and insulin-stimulated Swiss 3T3 cells or
IGF-I-stimulated L6 cells resulted in an increase in Raf-1 activity,
although the consequences of this elevated Raf-1 activity were not
investigated. They suggest that downstream components of the MAP kinase
pathway may act to suppress Raf-1 activity, possibly through
hyperphosphorylation. However, in hippocampal neuronal cells, elevated
levels of Raf-1 activity, but not MEK or ERK, were sufficient for
differentiation (63). Transfection with a Raf-1:estrogen receptor
fusion gene, It is striking that two such different responses of myoblasts to the
IGFs are mediated by quite distinct signaling pathways rather than
resulting from subtle differences in timing or concentrations of
signaling intermediates. Although many recent studies have examined IGF
signaling pathways, none have directly addressed the issue of how IGF-I
can stimulate both proliferation and differentiation in a single cell
type such as skeletal muscle. In this study we have now shown that
different signaling pathways are specifically associated with each
response; cell proliferation is mediated primarily by the Ras/Raf-1/MAP
kinase pathway, whereas stimulation of differentiation utilizes the PI
3-kinase/p70S6k pathway.
FOOTNOTES Acknowledgments We thank Paul Walton and John Ballard (GroPep
Pty Ltd, Adelaide, Australia) for a generous gift of R3 IGF-I. We also
thank Alan Saltiel (Parke-Davis Pharmaceuticals, Ann Arbor, MI) for the
MEK inhibitor, PD098059, Eric Olson (UT Southwestern Medical Center,
Dallas, TX) for the myogenin cDNA probe, and W. R. Wright (University of Texas, Austin, TX) for the anti-myogenin antibody. We
would especially like to thank Cathleen Jenney for superb technical assistance.
REFERENCES
Volume 272, Number 10,
Issue of March 7, 1997
pp. 6653-6662
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
§,,,
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
FOOTNOTES
Acknowledgments
REFERENCES
Raf-1:ER) results in p70S6k activation via a
MAPK-independent pathway. Therefore, it appears that multiple signaling
pathways lead to p70S6k activation.
20 °C: PD098059, 30 mM; LY294002, 10 mM; rapamycin, 1 mg/ml.
32P]ATP mixture (75 mM
MgCl2, 500 µM ATP, 10 µCi of
[32P]ATP). The reaction mixtures were incubated for 10 min at 30 °C and then centrifuged for 1 min. Aliquots (20 µl) of
the supernatants were spotted onto P81 phosphocellulose paper squares
and washed three times in 0.75% phosphoric acid, followed by one wash
in acetone. The squares were transferred to scintillation vials and counted in a Beckman LS 7500 scintillation counter.
Fig. 1.
Concentration dependence of the mitogenic and
myogenic responses of L6A1 myoblasts to R3 IGF-I. L6A1 myoblast
cultures were established overnight in growth medium at 1.2 × 105 cells/35-mm dish, washed once with DMEM, and then
treated with the indicated concentrations of R3 IGF-I. Effects on
proliferation (cell number) were determined after 1 day. Effects on
differentiation, expressed as CK/DNA, were determined after 3 days. The
results are expressed as means ± S.E. of triplicate
determinations.
[View Larger Version of this Image (21K GIF file)]
Fig. 2.
Concentration dependence of effects of
inhibitors on the mitogenic and myogenic responses to R3 IGF-I.
Cultures were prepared and analyzed as detailed in the legend to Fig. 1
and under "Experimental Procedures." R3 IGF-I was added at 3 ng/ml in the presence of the indicated concentrations of PD098059, LY294002, or rapamycin. Effects on proliferation (cell number) were measured after 1 day, and effects on differentiation (CK/DNA) were measured after 3 days. Results are expressed as percentage of the R3 IGF-I effect in triplicate determinations.
[View Larger Version of this Image (21K GIF file)]
Fig. 3.
Morphological effects of R3 IGF-I and
inhibitors on L6A1 myoblasts. L6A1 myoblast cultures were prepared
as described in the legend to Fig. 1. R3 IGF-I (3 ng/ml) was added in
the presence of 30 µM PD098059, 10 µM
LY294002, or 1 ng/ml rapamycin. After 2 days, the cultures were washed
with PBS, fixed with MeOH, and stained with Wright's and Giemsa
stains.
[View Larger Version of this Image (149K GIF file)]
Fig. 4.
Effects of combinations of inhibitors
on the stimulation of proliferation and differentiation by R3
IGF-I. L6A1 myoblast cultures were established as described in the
legend to Fig. 1. R3 IGF-I (3 ng/ml) was added in the presence of 30 µM PD098059, 10 µM LY294002, or 1 ng/ml
rapamycin, alone or in combinations as specified. Proliferation (cell
number) was measured after 1 day, and differentiation (CK/DNA) was
measured after 3 days. Results are expressed as means ± S.E. of
triplicate determinations.
[View Larger Version of this Image (31K GIF file)]
Fig. 5.
[View Larger Version of this Image (25K GIF file)]
Fig. 6.
[View Larger Version of this Image (27K GIF file)]
Fig. 7.
Assay of p70S6k after the
addition of R3 IGF-I and inhibitors. L6A1 myoblast cultures were
established overnight in growth medium at 1.2 × 106
cells/100-mm dish, washed once with DMEM, and then treated with R3
IGF-I (1 ng/ml) with or without PD098059 (10 µM),
LY294002 (10 µM), or rapamycin (1 ng/ml). At the
indicated times, p70S6k immunoprecipitates were prepared
and assayed for p70S6k activity as described under
"Experimental Procedures." The inset provides a clearer
indication of the effect of treatments after 30 min. The data obtained
with R3 IGF-I plus LY294002 or R3 IGF-I plus rapamycin were essentially
identical to those obtained with the DMEM controls, so separate points
are not shown for them.
[View Larger Version of this Image (30K GIF file)]
-actin expression was the only myogenic response that
was reported.
receptor resulted in autophosphorylation of the PDGF- receptor, activation of MAP kinase, and increased jun B and
c-fos mRNA. However, no effect on proliferation or
differentiation was observed. In other muscle cell types, PDGF has been
shown to repress myogenesis and promote proliferation (59-61).
However, the signaling mechanisms involved were not investigated.
Raf-1:ER, resulted in increased Raf-1 activity and
extended differentiation following stimulation with estradiol.
Prolonged activation of MAP kinases was not sufficient for
differentiation, suggesting an alternative pathway where Raf-1, but not
MAP kinases, functions in differentiation of neuronal cells. Yen
et al. (64) have reported that expression of an activated
form of Raf-1 accelerates terminal differentiation of promyelocytic
leukemia cells, and this is accompanied by down-regulation of
retinoblastoma gene product phosphorylation. However, Ming et
al. (18) have reported that neither Ras nor Raf-1 plays a role in
activation of p70S6k. Dominant negative mutants of Ras or
Raf-1 both failed to inhibit p70S6k activation by EGF in
human 293 cells but did inhibit MAP kinase activation. We have shown
that p70S6k activity is critical for differentiation in
L6A1 skeletal muscle cells, and we are currently exploring the specific
role that Raf-1 plays in this process.
The first three authors contributed equally to this work.
§
To whom correspondence should be addressed: Biology Department,
Syracuse University, 130 College Place, Syracuse, NY 13244-1220. Tel.:
315-443-9174; Fax: 315-443-2012; E-mail:
sacoolic{at}mailbox.syr.edu.
1
The abbreviations used are: IGF, insulin-like
growth factor; DMEM, Dulbecco's modified Eagle's medium; ERK,
extracellular regulated kinase; FGF, fibroblast growth factor; Grb-2:
growth factor receptor binding protein-2; IRS, insulin receptor
substrate; LY, LY294002, an inhibitor of the activation of PI 3-kinase;
MAP, mitogen-activated protein; MAPK, MAP kinase; MEK, MAP kinase
kinase; p21, a cyclin-dependent kinase inhibitor;
p70S6k, p70 S6 kinase; PD, PD098059, an inhibitor of the
activation of MEK by Raf-1; PDGF, platelet-derived growth factor; PI,
phosphatidylinositol; R3 IGF-I, the R3 analog of IGF-I in which an
arginine has replaced glutamate in the third residue to suppress
binding to IGF-binding proteins; Rapa, rapamycin, an inhibitor of the
activation of p70S6k; CK, creatine kinase; PBS,
phosphate-buffered saline; IGFBP, IGF-binding protein; EGF, epidermal
growth factor.
2
D. Z. Ewton, S. A. Coolican, S. Mohan, S. D. Chernausek, and J. R. Florini, manuscript in preparation.
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  A. PHILIPPOU, E. PAPAGEORGIOU, G. BOGDANIS, A. HALAPAS, A. SOURLA, M. MARIDAKI, N. PISSIMISSIS, and M. KOUTSILIERIS Expression of IGF-1 Isoforms after Exercise-induced Muscle Damage in Humans: Characterization of the MGF E Peptide Actions In Vitro In Vivo, July 1, 2009; 23(4): 567 - 575. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J.-E. Choi, S.-s. Lee, D. A. Sunde, I. Huizar, K. L. Haugk, V. J. Thannickal, R. Vittal, S. R. Plymate, and L. M. Schnapp Insulin-like Growth Factor-I Receptor Blockade Improves Outcome in Mouse Model of Lung Injury Am. J. Respir. Crit. Care Med., February 1, 2009; 179(3): 212 - 219. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. I. Bower, X. Li, R. Taylor, and I. A. Johnston Switching to fast growth: the insulin-like growth factor (IGF) system in skeletal muscle of Atlantic salmon J. Exp. Biol., December 15, 2008; 211(24): 3859 - 3870. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Wang, C. Wang, F. Xiao, H. Wang, and Z. Wu JAK2/STAT2/STAT3 Are Required for Myogenic Differentiation J. Biol. Chem., December 5, 2008; 283(49): 34029 - 34036. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. R. McKay, C. E. O'Reilly, S. M. Phillips, M. A. Tarnopolsky, and G. Parise Co-expression of IGF-1 family members with myogenic regulatory factors following acute damaging muscle-lengthening contractions in humans J. Physiol., November 15, 2008; 586(22): 5549 - 5560. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. G. McDaneld and D. M. Spurlock Ankyrin repeat and suppressor of cytokine signaling (SOCS) box-containing protein (ASB) 15 alters differentiation of mouse C2C12 myoblasts and phosphorylation of mitogen-activated protein kinase and Akt J Anim Sci, November 1, 2008; 86(11): 2897 - 2902. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. W. Hammers, E. K. Merritt, W. Matheny, M. L. Adamo, T. J. Walters, J. S. Estep, and R. P. Farrar Functional deficits and insulin-like growth factor-I gene expression following tourniquet-induced injury of skeletal muscle in young and old rats J Appl Physiol, October 1, 2008; 105(4): 1274 - 1281. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Ren, P. Yin, and C. Duan IGFBP-5 regulates muscle cell differentiation by binding to IGF-II and switching on the IGF-II auto-regulation loop J. Cell Biol., September 8, 2008; 182(5): 979 - 991. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Koyama, Y. Nakaoka, Y. Fujio, H. Hirota, K. Nishida, S. Sugiyama, K. Okamoto, K. Yamauchi-Takihara, M. Yoshimura, S. Mochizuki, et al. Interaction of Scaffolding Adaptor Protein Gab1 with Tyrosine Phosphatase SHP2 Negatively Regulates IGF-I-dependent Myogenic Differentiation via the ERK1/2 Signaling Pathway J. Biol. Chem., August 29, 2008; 283(35): 24234 - 24244. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Yahiaoui, D. Gvozdic, G. Danialou, M. Mack, and B. J. Petrof CC family chemokines directly regulate myoblast responses to skeletal muscle injury J. Physiol., August 15, 2008; 586(16): 3991 - 4004. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Al-Shanti and C. E Stewart PD98059 enhances C2 myoblast differentiation through p38 MAPK activation: a novel role for PD98059 J. Endocrinol., July 1, 2008; 198(1): 243 - 252. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Ding, K. J. Choi, J. H. Kim, X. Han, Y. Piao, J.-H. Jeong, W. Choe, I. Kang, J. Ha, H. J. Forman, et al. Endogenous Hydrogen Peroxide Regulates Glutathione Redox via Nuclear Factor Erythroid 2-Related Factor 2 Downstream of Phosphatidylinositol 3-Kinase during Muscle Differentiation Am. J. Pathol., June 1, 2008; 172(6): 1529 - 1541. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Nedachi, A. Kadotani, M. Ariga, H. Katagiri, and M. Kanzaki Ambient glucose levels qualify the potency of insulin myogenic actions by regulating SIRT1 and FoxO3a in C2C12 myocytes Am J Physiol Endocrinol Metab, April 1, 2008; 294(4): E668 - E678. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. De Alvaro, I. Nieto-Vazquez, J. M. Rojas, and M. Lorenzo Nuclear Exclusion of Forkhead Box O and Elk1 and Activation of Nuclear Factor-{kappa}B Are Required for C2C12-RasV12C40 Myoblast Differentiation Endocrinology, February 1, 2008; 149(2): 793 - 801. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M.-S. Yoon and J. Chen PLD regulates myoblast differentiation through the mTOR-IGF2 pathway J. Cell Sci., February 1, 2008; 121(3): 282 - 289. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Mandl, D. Sarkes, V. Carricaburu, V. Jung, and L. Rameh Serum Withdrawal-Induced Accumulation of Phosphoinositide 3-Kinase Lipids in Differentiating 3T3-L6 Myoblasts: Distinct Roles for Ship2 and PTEN Mol. Cell. Biol., December 1, 2007; 27(23): 8098 - 8112. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. S. Quinn, B. G. Anderson, and S. R. Plymate Muscle-specific overexpression of the type 1 IGF receptor results in myoblast-independent muscle hypertrophy via PI3K, and not calcineurin, signaling Am J Physiol Endocrinol Metab, December 1, 2007; 293(6): E1538 - E1551. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Odemis, K. Boosmann, M. T. Dieterlen, and J. Engele The chemokine SDF1 controls multiple steps of myogenesis through atypical PKC{zeta} J. Cell Sci., November 15, 2007; 120(22): 4050 - 4059. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. De Santa, S. Albini, E. Mezzaroma, L. Baron, A. Felsani, and M. Caruso pRb-Dependent Cyclin D3 Protein Stabilization Is Required for Myogenic Differentiation Mol. Cell. Biol., October 15, 2007; 27(20): 7248 - 7265. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Sun, K. Ma, H. Wang, F. Xiao, Y. Gao, W. Zhang, K. Wang, X. Gao, N. Ip, and Z. Wu JAK1 STAT1 STAT3, a key pathway promoting proliferation and preventing premature differentiation of myoblasts J. Cell Biol., October 8, 2007; 179(1): 129 - 138. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. J. Lim, K. J. Choi, Y. Ding, J. H. Kim, B. S. Kim, Y. H. Kim, J. Lee, W. Choe, I. Kang, J. Ha, et al. RhoA/Rho Kinase Blocks Muscle Differentiation via Serine Phosphorylation of Insulin Receptor Substrate-1 and -2 Mol. Endocrinol., September 1, 2007; 21(9): 2282 - 2293. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Pelosi, F. Marampon, B. M. Zani, S. Prudente, E. Perlas, V. Caputo, L. Cianetti, V. Berno, S. Narumiya, S. W. Kang, et al. ROCK2 and Its Alternatively Spliced Isoform ROCK2m Positively Control the Maturation of the Myogenic Program Mol. Cell. Biol., September 1, 2007; 27(17): 6163 - 6176. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Dogra, S. L. Hall, N. Wedhas, T. A. Linkhart, and A. Kumar Fibroblast Growth Factor Inducible 14 (Fn14) Is Required for the Expression of Myogenic Regulatory Factors and Differentiation of Myoblasts into Myotubes: EVIDENCE FOR TWEAK-INDEPENDENT FUNCTIONS OF Fn14 DURING MYOGENESIS J. Biol. Chem., May 18, 2007; 282(20): 15000 - 15010. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S.-E. Chen, B. Jin, and Y.-P. Li TNF-{alpha} regulates myogenesis and muscle regeneration by activating p38 MAPK Am J Physiol Cell Physiol, May 1, 2007; 292(5): C1660 - C1671. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Ciarmatori, D. Kiepe, A. Haarmann, U. Huegel, and B. Tonshoff Signaling mechanisms leading to regulation of proliferation and differentiation of the mesenchymal chondrogenic cell line RCJ3.1C5.18 in response to IGF-I J. Mol. Endocrinol., April 1, 2007; 38(4): 493 - 508. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Faenza, G. Ramazzotti, A. Bavelloni, R. Fiume, G. C. Gaboardi, M. Y. Follo, R. S. Gilmour, A. M. Martelli, K. Ravid, and L. Cocco Inositide-Dependent Phospholipase C Signaling Mimics Insulin in Skeletal Muscle Differentiation by Affecting Specific Regions of the Cyclin D3 Promoter Endocrinology, March 1, 2007; 148(3): 1108 - 1117. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Jacquemin, G. S. Butler-Browne, D. Furling, and V. Mouly IL-13 mediates the recruitment of reserve cells for fusion during IGF-1-induced hypertrophy of human myotubes J. Cell Sci., February 15, 2007; 120(4): 670 - 681. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Zhan, B. Jin, S.-E. Chen, J. M. Reecy, and Y.-P. Li TACE release of TNF-{alpha} mediates mechanotransduction-induced activation of p38 MAPK and myogenesis J. Cell Sci., February 15, 2007; 120(4): 692 - 701. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. A. Lovett, I. Gonzalez, D. A. M. Salih, L. J. Cobb, G. Tripathi, R. A. Cosgrove, A. Murrell, P. J. Kilshaw, and J. M. Pell Convergence of Igf2 expression and adhesion signalling via RhoA and p38 MAPK enhances myogenic differentiation J. Cell Sci., December 1, 2006; 119(23): 4828 - 4840. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Kim, A. L. Clark, A. Kiss, J. W. Hahn, R. Wesselschmidt, C. J. Coscia, and M. M. Belcheva {micro}- and {kappa}-Opioids Induce the Differentiation of Embryonic Stem Cells to Neural Progenitors J. Biol. Chem., November 3, 2006; 281(44): 33749 - 33760. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Kuninger, A. Wright, and P. Rotwein Muscle cell survival mediated by the transcriptional coactivators p300 and PCAF displays different requirements for acetyltransferase activity Am J Physiol Cell Physiol, October 1, 2006; 291(4): C699 - C709. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. H. Parker, R. L. S. Perry, M. C. Fauteux, C. A. Berkes, and M. A. Rudnicki MyoD Synergizes with the E-Protein HEB{beta} To Induce Myogenic Differentiation Mol. Cell. Biol., August 1, 2006; 26(15): 5771 - 5783. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. R. Barton Viral expression of insulin-like growth factor-I isoforms promotes different responses in skeletal muscle J Appl Physiol, June 1, 2006; 100(6): 1778 - 1784. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Kwintkiewicz, R. Z. Spaczynski, N. Foyouzi, T. Pehlivan, and A. J. Duleba Insulin and Oxidative Stress Modulate Proliferation of Rat Ovarian Theca-Interstitial Cells Through Diverse Signal Transduction Pathways Biol Reprod, June 1, 2006; 74(6): 1034 - 1040. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Dogra, H. Changotra, S. Mohan, and A. Kumar Tumor Necrosis Factor-like Weak Inducer of Apoptosis Inhibits Skeletal Myogenesis through Sustained Activation of Nuclear Factor-{kappa}B and Degradation of MyoD Protein J. Biol. Chem., April 14, 2006; 281(15): 10327 - 10336. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. S. Kraft, C. M. R. LeMoine, C. N. Lyons, D. Michaud, C. R. Mueller, and C. D. Moyes Control of mitochondrial biogenesis during myogenesis Am J Physiol Cell Physiol, April 1, 2006; 290(4): C1119 - C1127. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Riuzzi, G. Sorci, and R. Donato The Amphoterin (HMGB1)/Receptor for Advanced Glycation End Products (RAGE) Pair Modulates Myoblast Proliferation, Apoptosis, Adhesiveness, Migration, and Invasiveness: FUNCTIONAL INACTIVATION OF RAGE IN L6 MYOBLASTS RESULTS IN TUMOR FORMATION IN VIVO J. Biol. Chem., March 24, 2006; 281(12): 8242 - 8253. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Um and H. F. Lodish Antiapoptotic Effects of Erythropoietin in Differentiated Neuroblastoma SH-SY5Y Cells Require Activation of Both the STAT5 and AKT Signaling Pathways J. Biol. Chem., March 3, 2006; 281(9): 5648 - 5656. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. L. J. van der Velden, R. C. J. Langen, M. C. J. M. Kelders, E. F. M. Wouters, Y. M. W. Janssen-Heininger, and A. M. W. J. Schols Inhibition of glycogen synthase kinase-3{beta} activity is sufficient to stimulate myogenic differentiation Am J Physiol Cell Physiol, February 1, 2006; 290(2): C453 - C462. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S.-E. Chen, E. Gerken, Y. Zhang, M. Zhan, R. K. Mohan, A. S. Li, M. B. Reid, and Y.-P. Li Role of TNF-{alpha} signaling in regeneration of cardiotoxin-injured muscle Am J Physiol Cell Physiol, November 1, 2005; 289(5): C1179 - C1187. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. J. Fahey, J. M. Brameld, T. Parr, and P. J. Buttery Ontogeny of factors associated with proliferation and differentiation of muscle in the ovine fetus J Anim Sci, October 1, 2005; 83(10): 2330 - 2338. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I.-H. Park and J. Chen Mammalian Target of Rapamycin (mTOR) Signaling Is Required for a Late-stage Fusion Process during Skeletal Myotube Maturation J. Biol. Chem., September 9, 2005; 280(36): 32009 - 32017. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. de Alvaro, N. Martinez, J. M. Rojas, and M. Lorenzo Sprouty-2 Overexpression in C2C12 Cells Confers Myogenic Differentiation Properties in the Presence of FGF2 Mol. Biol. Cell, September 1, 2005; 16(9): 4454 - 4461. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C.-H. Fang, B.-G. Li, J. H. James, J.-K. King, A. R. Evenson, G. D. Warden, and P.-O. Hasselgren Protein Breakdown in Muscle from Burned Rats Is Blocked by Insulin-Like Growth Factor I and Glycogen Synthase Kinase-3{beta} Inhibitors Endocrinology, July 1, 2005; 146(7): 3141 - 3149. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Kiepe, S. Ciarmatori, A. Hoeflich, E. Wolf, and B. Tonshoff Insulin-Like Growth Factor (IGF)-I Stimulates Cell Proliferation and Induces IGF Binding Protein (IGFBP)-3 and IGFBP-5 Gene Expression in Cultured Growth Plate Chondrocytes via Distinct Signaling Pathways Endocrinology, July 1, 2005; 146(7): 3096 - 3104. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M Fernandez, F Sanchez-Franco, N Palacios, I Sanchez, and L Cacicedo IGF-I and vasoactive intestinal peptide (VIP) regulate cAMP-response element-binding protein (CREB)-dependent transcription via the mitogen-activated protein kinase (MAPK) pathway in pituitary cells: requirement of Rap1 J. Mol. Endocrinol., June 1, 2005; 34(3): 699 - 712. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. C. Jones, K. J. Tyner, L. Nibarger, H. M. Stanley, D. D.W. Cornelison, Y. V. Fedorov, and B. B. Olwin The p38{alpha}/{beta} MAPK functions as a molecular switch to activate the quiescent satellite cell J. Cell Biol., April 11, 2005; 169(1): 105 - 116. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. E. Spangenburg SOCS-3 Induces Myoblast Differentiation J. Biol. Chem., March 18, 2005; 280(11): 10749 - 10758. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. S. Bickel, J. Slade, E. Mahoney, F. Haddad, G. A. Dudley, and G. R. Adams Time course of molecular responses of human skeletal muscle to acute bouts of resistance exercise J Appl Physiol, February 1, 2005; 98(2): 482 - 488. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y.-C. Huang, R. G. Dennis, L. Larkin, and K. Baar Rapid formation of functional muscle in vitro using fibrin gels J Appl Physiol, February 1, 2005; 98(2): 706 - 713. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. G. Miller, J. D. Aplin, and M. Westwood Adenovirally Mediated Expression of Insulin-Like Growth Factors Enhances the Function of First Trimester Placental Fibroblasts J. Clin. Endocrinol. Metab., January 1, 2005; 90(1): 379 - 385. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Tiffin, S. Adi, D. Stokoe, N.-Y. Wu, and S. M. Rosenthal Akt Phosphorylation Is Not Sufficient for Insulin-Like Growth Factor-Stimulated Myogenin Expression but Must Be Accompanied by Down-Regulation of Mitogen-Activated Protein Kinase/Extracellular Signal-Regulated Kinase Phosphorylation Endocrinology, November 1, 2004; 145(11): 4991 - 4996. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Natalicchio, L. Laviola, C. De Tullio, L. A. Renna, C. Montrone, S. Perrini, G. Valenti, G. Procino, M. Svelto, and F. Giorgino Role of the p66Shc Isoform in Insulin-like Growth Factor I Receptor Signaling through MEK/Erk and Regulation of Actin Cytoskeleton in Rat Myoblasts J. Biol. Chem., October 15, 2004; 279(42): 43900 - 43909. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Rochat, A. Fernandez, M. Vandromme, J.-P. Moles, T. Bouschet, G. Carnac, and N. J. C. Lamb Insulin and Wnt1 Pathways Cooperate to Induce Reserve Cell Activation in Differentiation and Myotube Hypertrophy Mol. Biol. Cell, October 1, 2004; 15(10): 4544 - 4555. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Hayashi, K. Shibata, T. Morita, K. Iwasaki, M. Watanabe, and K. Sobue Insulin Receptor Substrate-1/SHP-2 Interaction, a Phenotype-dependent Switching Machinery of Insulin-like Growth Factor-I Signaling in Vascular Smooth Muscle Cells J. Biol. Chem., September 24, 2004; 279(39): 40807 - 40818. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. J. Halayko, S. Kartha, G. L. Stelmack, J. McConville, J. Tam, B. Camoretti-Mercado, S. M. Forsythe, M. B. Hershenson, and J. Solway Phophatidylinositol-3 Kinase/Mammalian Target of Rapamycin/p70S6K Regulates Contractile Protein Accumulation in Airway Myocyte Differentiation Am. J. Respir. Cell Mol. Biol., September 1, 2004; 31(3): 266 - 275. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Huang, L. Shu, J. Easton, F. C. Harwood, G. S. Germain, H. Ichijo, and P. J. Houghton Inhibition of Mammalian Target of Rapamycin Activates Apoptosis Signal-regulating Kinase 1 Signaling by Suppressing Protein Phosphatase 5 Activity J. Biol. Chem., August 27, 2004; 279(35): 36490 - 36496. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 V. Sartorelli and M. Fulco Molecular and Cellular Determinants of Skeletal Muscle Atrophy and Hypertrophy Sci. Signal., August 3, 2004; 2004(244): re11 - re11. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. I. Kontaridis, S. Eminaga, M. Fornaro, C. I. Zito, R. Sordella, J. Settleman, and A. M. Bennett SHP-2 Positively Regulates Myogenesis by Coupling to the Rho GTPase Signaling Pathway Mol. Cell. Biol., June 15, 2004; 24(12): 5340 - 5352. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Sorci, F. Riuzzi, C. Arcuri, I. Giambanco, and R. Donato Amphoterin Stimulates Myogenesis and Counteracts the Antimyogenic Factors Basic Fibroblast Growth Factor and S100B via RAGE Binding Mol. Cell. Biol., June 1, 2004; 24(11): 4880 - 4894. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Gonzalez, G. Tripathi, E. J. Carter, L. J. Cobb, D. A. M. Salih, F. A. Lovett, C. Holding, and J. M Pell Akt2, a Novel Functional Link between p38 Mitogen-Activated Protein Kinase and Phosphatidylinositol 3-Kinase Pathways in Myogenesis Mol. Cell. Biol., May 1, 2004; 24(9): 3607 - 3622. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. J. Cobb, D. A. M. Salih, I. Gonzalez, G. Tripathi, E. J. Carter, F. Lovett, C. Holding, and J. M. Pell Partitioning of IGFBP-5 actions in myogenesis: IGF-independent anti-apoptotic function J. Cell Sci., May 1, 2004; 117(9): 1737 - 1746. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. E. Spangenburg, D. K. Bowles, and F. W. Booth Insulin-Like Growth Factor-Induced Transcriptional Activity of the Skeletal {alpha}-Actin Gene Is Regulated by Signaling Mechanisms Linked to Voltage-Gated Calcium Channels during Myoblast Differentiation Endocrinology, April 1, 2004; 145(4): 2054 - 2063. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I.-J. Kim, K. M. Drahushuk, W.-Y. Kim, E. A. Gonsiorek, P. Lein, D. A. Andres, and D. Higgins Extracellular Signal-Regulated Kinases Regulate Dendritic Growth in Rat Sympathetic Neurons J. Neurosci., March 31, 2004; 24(13): 3304 - 3312. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. F. Kuemmerle, H. Zhou, and J. G. Bowers IGF-I stimulates human intestinal smooth muscle cell growth by regulation of G1 phase cell cycle proteins Am J Physiol Gastrointest Liver Physiol, March 1, 2004; 286(3): G412 - G419. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Safi, M. Vandromme, S. Caussanel, L. Valdacci, D. Baas, M. Vidal, G. Brun, L. Schaeffer, and E. Goillot Role for the Pleckstrin Homology Domain-Containing Protein CKIP-1 in Phosphatidylinositol 3-Kinase-Regulated Muscle Differentiation Mol. Cell. Biol., February 1, 2004; 24(3): 1245 - 1255. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. M. Wilson, J. Tureckova, and P. Rotwein Permissive Roles of Phosphatidyl Inositol 3-Kinase and Akt in Skeletal Myocyte Maturation Mol. Biol. Cell, February 1, 2004; 15(2): 497 - 505. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
X. Wang, S. R. Thomson, J. D. Starkey, J. L. Page, A. D. Ealy, and S. E. Johnson Transforming Growth Factor {beta}1 Is Up-regulated by Activated Raf in Skeletal Myoblasts but Does Not Contribute to the Differentiation-defective Phenotype J. Biol. Chem., January 23, 2004; 279(4): 2528 - 2534. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Haddad and G. R. Adams Inhibition of MAP/ERK kinase prevents IGF-I-induced hypertrophy in rat muscles J Appl Physiol, January 1, 2004; 96(1): 203 - 210. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Erbay, I.-H. Park, P. D. Nuzzi, C. J. Schoenherr, and J. Chen IGF-II transcription in skeletal myogenesis is controlled by mTOR and nutrients J. Cell Biol., December 8, 2003; 163(5): 931 - 936. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. P. Kirk, J. M. Oldham, F. Jeanplong, and J. J. Bass Insulin-like Growth Factor-II Delays Early but Enhances Late Regeneration of Skeletal Muscle J. Histochem. Cytochem., December 1, 2003; 51(12): 1611 - 1620. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. C. Sundgren, G. D. Giraud, J. M. Schultz, M. R. Lasarev, P. J. S. Stork, and K. L. Thornburg Extracellular signal-regulated kinase and phosphoinositol-3 kinase mediate IGF-1 induced proliferation of fetal sheep cardiomyocytes Am J Physiol Regulatory Integrative Comp Physiol, December 1, 2003; 285(6): R1481 - R1489. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Kataoka, I. Matsumura, S. Ezoe, S. Nakata, E. Takigawa, Y. Sato, A. Kawasaki, T. Yokota, K. Nakajima, A. Felsani, et al. Reciprocal Inhibition between MyoD and STAT3 in the Regulation of Growth and Differentiation of Myoblasts J. Biol. Chem., November 7, 2003; 278(45): 44178 - 44187. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I Sermet-Gaudelus, J C Souberbielle, I Azhar, J C Ruiz, P Magnine, V Colomb, C Le Bihan, D Folio, and G Lenoir Insulin-like growth factor I correlates with lean body mass in cystic fibrosis patients Arch. Dis. Child., November 1, 2003; 88(11): 956 - 961. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. L. Hribal, J. Nakae, T. Kitamura, J. R. Shutter, and D. Accili Regulation of insulin-like growth factor-dependent myoblast differentiation by Foxo forkhead transcription factors J. Cell Biol., August 18, 2003; 162(4): 535 - 541. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. R. Broussard, R. H. MCCusker, J. E. Novakofski, K. Strle, W. Hong Shen, R. W. Johnson, G. G. Freund, R. Dantzer, and K. W. Kelley Cytokine-Hormone Interactions: Tumor Necrosis Factor {alpha} Impairs Biologic Activity and Downstream Activation Signals of the Insulin-Like Growth Factor I Receptor in Myoblasts Endocrinology, July 1, 2003; 144(7): 2988 - 2996. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. Ostrovsky and E. Bengal The Mitogen-activated Protein Kinase Cascade Promotes Myoblast Cell Survival by Stabilizing the Cyclin-dependent Kinase Inhibitor, p21WAF1 Protein J. Biol. Chem., May 30, 2003; 278(23): 21221 - 21231. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. M. Cox, M. Du, M. Marback, E. C. C. Yang, J. Chan, K. W. M. Siu, and J. C. McDermott Phosphorylation Motifs Regulating the Stability and Function of Myocyte Enhancer Factor 2A J. Biol. Chem., April 18, 2003; 278(17): 15297 - 15303. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. De Arcangelis, D. Coletti, M. Conti, M. Lagarde, M. Molinaro, S. Adamo, G. Nemoz, and F. Naro IGF-I-induced Differentiation of L6 Myogenic Cells Requires the Activity of cAMP-Phosphodiesterase Mol. Biol. Cell, April 1, 2003; 14(4): 1392 - 1404. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. F. Kuemmerle IGF-I elicits growth of human intestinal smooth muscle cells by activation of PI3K, PDK-1, and p70S6 kinase Am J Physiol Gastrointest Liver Physiol, March 1, 2003; 284(3): G411 - G422. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. M. Bamman, V. J. Hill, G. R. Adams, F. Haddad, C. J. Wetzstein, B. A. Gower, A. Ahmed, and G. R. Hunter Gender Differences in Resistance-Training-Induced Myofiber Hypertrophy Among Older Adults J. Gerontol. A Biol. Sci. Med. Sci., February 1, 2003; 58(2): B108 - 116. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. E. Spangenburg, T. Abraha, T. E. Childs, J. S. Pattison, and F. W. Booth Skeletal muscle IGF-binding protein-3 and -5 expressions are age, muscle, and load dependent Am J Physiol Endocrinol Metab, February 1, 2003; 284(2): E340 - E350. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Cellier, M. Mage, J. Duchene, C. Pecher, R. Couture, J.-L. Bascands, and J.-P. Girolami Bradykinin reduces growth factor-induced glomerular ERK1/2 phosphorylation Am J Physiol Renal Physiol, February 1, 2003; 284(2): F282 - F292. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Chaum and H. Yang Transgenic Expression of IGF-1 Modifies the Proliferative Potential of Human Retinal Pigment Epithelial Cells Invest. Ophthalmol. Vis. Sci., December 1, 2002; 43(12): 3758 - 3764. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Yoshiko, K. Hirao, and N. Maeda Differentiation in C2C12 myoblasts depends on the expression of endogenous IGFs and not serum depletion Am J Physiol Cell Physiol, October 1, 2002; 283(4): C1278 - C1286. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Mauro, C. Ciccarelli, P. De Cesaris, A. Scoglio, M. Bouche, M. Molinaro, A. Aquino, and B. M. Zani PKC{alpha}-mediated ERK, JNK and p38 activation regulates the myogenic program in human rhabdomyosarcoma cells J. Cell Sci., September 15, 2002; 115(18): 3587 - 3599. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. R. Adams Exercise Effects on Muscle Insulin Signaling and Action: Invited Review: Autocrine/paracrine IGF-I and skeletal muscle adaptation J Appl Physiol, September 1, 2002; 93(3): 1159 - 1167. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Carrasco, J. Canicio, M. Palacin, A. Zorzano, and P. Kaliman Identification of Intracellular Signaling Pathways that Induce Myotonic Dystrophy Protein Kinase Expression during Myogenesis Endocrinology, August 1, 2002; 143(8): 3017 - 3025. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
X. Zhang, L. Shu, H. Hosoi, K. G. Murti, and P. J. Houghton Predominant Nuclear Localization of Mammalian Target of Rapamycin in Normal and Malignant Cells in Culture J. Biol. Chem., July 26, 2002; 277(31): 28127 - 28134. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Pallafacchina, E. Calabria, A. L. Serrano, J. M. Kalhovde, and S. Schiaffino A protein kinase B-dependent and rapamycin-sensitive pathway controls skeletal muscle growth but not fiber type specification PNAS, July 9, 2002; 99(14): 9213 - 9218. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Haddad and G. R. Adams Exercise Effects on Muscle Insulin Signaling and Action: Selected Contribution: Acute cellular and molecular responses to resistance exercise J Appl Physiol, July 1, 2002; 93(1): 394 - 403. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. E. Spangenburg and F. W. Booth Multiple signaling pathways mediate LIF-induced skeletal muscle satellite cell proliferation Am J Physiol Cell Physiol, July 1, 2002; 283(1): C204 - C211. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. I. Kontaridis, X. Liu, L. Zhang, and A. M. Bennett Role of SHP-2 in Fibroblast Growth Factor Receptor-Mediated Suppression of Myogenesis in C2C12 Myoblasts Mol. Cell. Biol., June 1, 2002; 22(11): 3875 - 3891. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. M. Scicchitano, L. Spath, A. Musaro, M. Molinaro, S. Adamo, and C. Nervi AVP Induces Myogenesis through the Transcriptional Activation of the Myocyte Enhancer Factor 2 Mol. Endocrinol., June 1, 2002; 16(6): 1407 - 1416. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Q. Xu, L. Yu, L. Liu, C. F. Cheung, X. Li, S.-P. Yee, X.-J. Yang, and Z. Wu p38 Mitogen-activated Protein Kinase-, Calcium-Calmodulin-dependent Protein Kinase-, and Calcineurin-mediated Signaling Pathways Transcriptionally Regulate Myogenin Expression Mol. Biol. Cell, June 1, 2002; 13(6): 1940 - 1952. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Shu, X. Zhang, and P. J. Houghton Myogenic Differentiation Is Dependent on Both the Kinase Function and the N-terminal Sequence of Mammalian Target of Rapamycin J. Biol. Chem., May 3, 2002; 277(19): 16726 - 16732. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. R. Barton, L. Morris, A. Musaro, N. Rosenthal, and H. L. Sweeney Muscle-specific expression of insulin-like growth factor I counters muscle decline in mdx mice J. Cell Biol., April 1, 2002; 157(1): 137 - 148. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| All ASBMB Journals | Molecular and Cellular Proteomics |
| Journal of Lipid Research | ASBMB Today |