1 School of Kinesiology, University of Illinois Chicago, Chicago, Illinois 60608; and 2 Department of Health Sciences, Boston University, Boston, Massachusetts 02215
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In the rat, denervation
and hindlimb unloading are two commonly employed models used to study
skeletal muscle atrophy. In these models, muscle atrophy is generally
produced by a decrease in protein synthesis and an increase in protein
degradation. The decrease in protein synthesis has been suggested to
occur by an inhibition at the level of protein translation. To better
characterize the regulation of protein translation, we investigated the
changes that occur in various translation initiation and elongation
factors. We demonstrated that both hindlimb unloading and denervation
produce alterations in the phosphorylation and/or total amount of the 70-kDa ribosomal S6 kinase, eukaryotic initiation factor 2 -subunit, and eukaryotic elongation factor 2. Our findings indicate that the
regulation of these protein translation factors differs between the
models of atrophy studied and between the muscles evaluated (e.g.,
soleus vs. extensor digitorum longus).
elongation factor; initiation factor; protein kinase; protein synthesis
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
HYPODYNAMIA is
defined as reduced load-bearing or locomotor activity by skeletal
muscle and generally results in muscle atrophy. Hypodynamia-induced
atrophy has been shown to occur following a variety of situations, such
as spaceflight (33), prolonged bed rest (18),
and cast immobilization (6). One model commonly used to
study hypodynamia is hindlimb unloading, in which the hindlimbs are
suspended off the ground to remove the normal gravitational load on the
muscles. As a result, the soleus muscle, which is composed primarily of
slow-twitch fibers in the rat, undergoes substantial atrophy, while the
primarily fast-twitch extensor digitorum longus (EDL) muscle remains
relatively unaffected (3). The atrophy of the soleus has
been partly attributed to a decrease in protein synthesis
(34). In fact, rates of protein synthesis in the soleus
may decrease as early as 6 h after unloading (34). The inhibition of protein synthesis most likely occurs at the translational level, since the synthesis of proteins such as -myosin heavy chain decreases, while mRNA levels remain unchanged
(34). To date, the mechanisms that regulate translation in
this model remain largely undefined.
Denervation is another model of hypodynamia that has been used to study muscle atrophy. In this model, similar to hindlimb unloading, the soleus muscle undergoes greater atrophy than the EDL. Another similarity to the hindlimb unloading model is a rapid decrease in protein synthesis of the soleus muscle (8). Surprisingly, the rate of protein synthesis in the EDL shows a gradual increase and eventually exceeds basal levels by 7 days (8), an effect that has not been observed in the EDL during hindlimb unloading. It has been suggested that the regulation of protein synthesis rates during denervation also occurs at the level of translation (23). However, the mechanisms that regulate the changes in protein translation following denervation have not been determined.
The process of protein translation can be divided into three stages: initiation, elongation, and termination. Each of these stages has been shown to be regulated by numerous protein factors termed initiation, elongation, and release factors (22, 27). It has been well established that initiation and elongation are the steps of translation that are regulated with little regulation occurring during termination. More specifically, most studies to date suggest that initiation is the primary site of regulation for the majority of mRNAs in the cell (31).
There are two major points of control that can regulate the rate of
initiation during translation. The first major control point is the
binding of the initiator methionyl-tRNA to the 40S ribosomal subunit.
This step is regulated by eukaryotic initiation factor 2 (eIF-2; a
heterodimer composed of -,
-, and
-subunits), which mediates
ribosomal binding of the methionyl-tRNA in a GTP-dependent manner
(26). As a product of this initiation step, eIF-2 is released in its GDP-bound state. To return eIF-2 to its active GTP-bound state, the GDP must be recycled for another GTP in a reaction
catalyzed by the guanine exchange factor eIF-2B (24). The
recycling of GTP by eIF-2B can be inhibited by phosphorylation of eIF-2
on the
-subunit (29). Therefore, an increase in
eIF-2
subunit phosphorylation can lead to an inhibition of
translation initiation.
A second control point during initiation is the binding of the mRNA to the 43S ribosomal subunit. One of the proteins that may regulate this step is the ribosomal S6 kinase (p70S6k). This kinase appears to confer selective translation of mRNAs that contain a 5' polypyrimidine tract as a common feature. The transcripts from theses mRNAs encode proteins that are generally involved in the translational apparatus, such as the ribosomal proteins (e.g., S6) (12) and elongation factors [e.g., eukaryotic elongation factor 2 (eEF-2)] (13). The translation of these mRNAs is dependent on the activity of p70S6k. The activity of p70S6k is regulated by changes in its phosphorylation state (4).
Rates of protein synthesis can also be regulated at the elongation
phase of translation. One factor that regulates elongation is eEF-2.
eEF-2 mediates the translocation step of elongation. Similar to what is
known for eIF-2, phosphorylation of eEF-2 results in inhibition of
elongation by decreasing its affinity for the ribosome by 10 to 100 times (2).
The purpose of this study was to characterize changes in known
translation factors that might regulate protein synthesis during hindlimb unloading and denervation models of muscle atrophy. In this
study we demonstrate that initiation and elongation factors are being
regulated in both models of atrophy. We show that, in general, the
factors investigated (p70S6k, eIF-2, and eEF-2) change
their state of phosphorylation, and/or quantity, in a manner that is
consistent with the changes that have previously been reported to occur
in protein synthesis rates. However, the results demonstrate that
during hypodynamia-induced atrophy, the mechanisms regulating the
translation of proteins differ dramatically between the type of muscle
and model studied.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Materials.
Rabbit polyclonal anti-p70S6k antibody was purchased from
Santa Cruz Biotechnology (Santa Cruz, CA). Mouse monoclonal
anti-eIF-2 antibody was a generous gift provided by Dr. Leonard S. Jefferson (Pennsylvania State University). Rabbit polyclonal
anti-phospho-Ser-51 eIF-2
antibody was purchased from Research
Genetics (Huntsville, AL). Rabbit polyclonal anti-eEF-2 antibody and
rabbit polyclonal anti-phospho-Thr-56 eEF-2 antibodies were generous
gifts provided by Dr. Angus C. Narin (The Rockefeller University).
Peroxidase-conjugated horse anti-mouse and goat anti-rabbit antibodies
were purchased from Vector Laboratories (Burlingame, CA).
Polyvinylidene difluoride (PVDF) membranes were purchased from
Millipore (Bedford, MA). Enhanced chemiluminescence (ECL) detection
reagent was purchased from Amersham Pharmacia Biotech (Amersham, UK).
Re-probe buffer was purchased from Geno Tech (St. Louis, MO). DC
protein assay kit was purchased from Bio-Rad (Hercules, CA).
Animal models and muscle processing.
All experimental procedures were approved by the University of Illinois
at Chicago Animal Care Committee. Animals were housed individually and
allowed free access to food and water throughout the experimental
period. Female Wistar rats (Charles River Laboratories, Wilmington,
MA), 3 mo of age, were randomly assigned to hindlimb unloading control
(CNT), sham-operated denervation control (SHM), denervated (DNV), or
hindlimb-unloaded (HU) groups. DNV and SHM animals were anesthetized
with pentobarbital sodium (40 mg/kg). The muscles of the hindlimb were
denervated by isolation and removal of ~1 cm of the sciatic nerve
immediately proximal to the division of the peroneal and tibial
branches. SHM animals underwent identical surgeries; however, the
sciatic nerve was left intact. Hindlimb unloading was performed by the
methods described previously (25). All animals were
allowed food and water ad libitum. After 12 h or 7 days of
hindlimb unloading or denervation, the soleus and EDL muscles were
removed, quickly weighed, and homogenized in a buffer containing 20 mM
Tris (pH 7.5), 150 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1% Triton X-100, 2.5 mM sodium pyrophosphate, 1 mM -glycerophosphate, 1 mM
Na3VO4, 1 µg/ml leupeptin, and 1 mM
phenylmethylsulfonyl fluoride. The homogenate was then centrifuged at
15,000 g for 10 min at 4°C, and the supernatant was used
for further analysis. The soleus and EDL muscles from SHM animals were
collected 12 h postoperation and processed as described above.
Protein concentration was determined by the DC protein assay (Bio-Rad Laboratories).
Quantification of p70S6k phosphorylation. Tissue supernatant (5 µg protein) was analyzed by SDS-PAGE on a 7.5% acrylamide gel. After electrophoretic separation, proteins were transferred to a PVDF membrane. The membrane was incubated in 5% blotto (5% powdered milk in 1× Tris-buffered saline, 1% Tween 20) overnight at 4°C. After the overnight incubation, the membrane was probed with anti-p70S6k antibody (1:2,500) for 1 h, followed by anti-rabbit antibody (1:5,000) for 45 min. The blots were then developed using an ECL Western blotting kit (Amersham Pharmacia Biotech). p70S6k resolves into multiple bands after electrophoretic separation, with the slower migrating bands representing states of increased phosphorylation. The percent phosphorylation was therefore quantified as previously described (1). All densitometric measurements were carried out on an Alpha Imager 2200 (Alpha Innotech).
Quantification of phosphorylated and total eIF-2.
Tissue supernatant (25 µg protein) was analyzed by SDS-PAGE on a
12.5% acrylamide gel and Western blotted as described in Quantification of p70S6k
phosphorylation. The phosphorylated form of the 36-kDa
protein was detected with an anti-phospho-Ser-51 eIF-2
antibody
(1:1,250). The membrane was stripped with Re-Probe buffer for 30 min at
room temperature, followed by overnight incubation in 5% blotto at 4°C. The total eIF-2
was detected with anti-eIF-2
antibody
(1:700). Phosphorylated and total eIF-2
measurements were normalized
for any loading differences by quantification of Coomassie blue-stained proteins in the gel that contained proteins in the 50- to 80-kDa range.
For the denervated EDL, the relative percent phosphorylation was
expressed as the ratio of amount of the phosphorylated form to the
total amount of the protein.
Quantification of phosphorylated and total eEF-2. Tissue supernatant (5 µg protein) was analyzed by SDS-PAGE on a 7.5% acrylamide gel and Western blotted as described in Quantification of p70S6k phosphorylation. The phosphorylated form of the 100-kDa protein was detected with an anti-phospho-Thr-56 eEF-2 antibody (1:500). The membrane was then stripped with Re-probe buffer for 30 min at room temperature, followed by overnight incubation in 5% blotto at 4°C. The total eEF-2 was detected with anti-eEF-2 antibody (1:250). Phosphorylated and total eEF-2 measurements were normalized for any loading differences by quantification of Coomassie blue-stained proteins from the 20- to 65-kDa range.
Statistical analysis. All data are expressed as means + SE. Unless otherwise noted, all HU samples were compared with CNT samples, and DNV samples were compared with SHM samples. Statistical analysis of the data was performed using analysis of variance (ANOVA). If differences were found, the Student-Newman-Keuls post hoc test was used to locate the source of the difference. Data were tested for normal distribution and homogeneity of variance before analysis was performed. All analyses were performed with SigmaStat statistical software (SPSS, Chicago, IL.).
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Hindlimb unloading and denervation induce similar atrophy.
Hindlimb unloading and denervation produced similar effects on muscle
weight-to-body weight ratio in the soleus muscles. Muscle weight-to-body weight ratio was not significantly altered after 12 h of hindlimb unloading or denervation. However, by 7 days, this ratio
had decreased 29% from control values after hindlimb unloading and
35% after denervation (P < 0.05) (Fig.
1A). The same pattern was
observed in the EDL, but the degree of atrophy was smaller. Hindlimb
unloading induced a 9% decrease (not statistically significant:
P = 0.19), while denervation produced a 12% reduction, in muscle weight-to-body weight ratio by 7 days (P < 0.05) (Fig. 1B).
|
Hindlimb unloading and denervation have different effects on
p70S6k phosphorylation.
In the soleus muscle, hindlimb unloading and denervation induced
similar alterations in p70S6k phosphorylation. Muscles from
HU animals showed a significant reduction in p70S6k
phosphorylation by 12 h, and this effect was still evident at 7 days compared with muscles from CNT animals (P < 0.05). Muscles from DNV animals also showed a significant reduction in
p70S6k phosphorylation at 12 h (P < 0.05) but not at 7 days compared with muscles from SHM animals (Fig.
2). It should be noted that there was a
trend for a decrease in the phosphorylation state of p70S6k
in the soleus of SHM animals compared with CNT animals at 12 h;
however, this was not statistically significant (P = 0.06). By 7 days after the sham operation, p70S6k
phosphorylation was similar to basal levels (data not shown). This
observation suggests that the sham operation may have produced a
transient decrease in p70S6k phosphorylation in the soleus
muscles. Therefore, it may be more appropriate to compare the
p70S6k phosphorylation states from DNV animals to those of
CNT animals. In this case, the p70S6k phosphorylation state
is markedly reduced at both 12 h and 7 days postdenervation
(P < 0.05).
|
|
Alterations in the phosphorylated and total amount of eIF-2
occur with denervation but not hindlimb unloading.
Hindlimb unloading and denervation had no effect on the phosphorylated
or total amount of eIF-2
in soleus muscles at either 12 h or 7 days (Fig. 4 A-F).
In addition, hindlimb unloading also had no effect on the
phosphorylated or total amount of eIF-2
in the EDL muscles at either
time points (Fig. 5, A,
C, and E). However, the denervated EDL muscles
showed a 22% reduction in the amount of phosphorylated eIF-2
(P < 0.05), and this decrease was accompanied by a
200% increase in the total amount of eIF-2
after 7 days of
denervation (P < 0.05), an effect that was not observed at 12 h. Together, these changes produced a 78%
reduction in the ratio of phosphorylated to total eIF-2
after 7 days of denervation (Fig. 5, B, D, F,
and G).
|
|
Alterations in the phosphorylated and total amount of eEF-2 occur
with hindlimb unloading and denervation.
In the soleus muscles, there was no change in the phosphorylated form
or total amount of eEF-2 at 12 h after hindlimb unloading. However, after 7 days, the soleus muscles of HU animals exhibited a
50% decrease in the amount of eEF-2 in the phosphorylated form (P < 0.05), while total eEF-2 was not altered. In
contrast, eEF-2 phosphorylation or amount did not change in the soleus
muscles of DNV animals at either 12 h or 7 days (Fig.
6, A-F).
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The effects of muscle unloading and denervation on protein
synthesis have been known for many years (7, 34). Muscle
atrophy associated with these models of hypodynamia is generally
believed to occur through a decrease in protein synthesis with
concomitant increases in protein degradation (7, 19).
However, this is not always the case, because denervation-induced
atrophy of the EDL is characterized by an increase in protein
synthesis, implying an even greater contribution from degradation
(8). The experiments described in this study focus on the
control of protein synthesis through translation factors. Our findings
demonstrate that regulation differs between models of hypodynamia as
well as between the muscle types that are studied. In general,
p70S6k, eIF-2, and eEF-2 changed their phosphorylation
state, and/or quantity (Table 1), in a
direction that is consistent with the changes in protein synthesis that
have been previously reported (8, 20, 34). These results
provide the first insight into the molecular events controlling muscle
protein synthesis during hypodynamia-induced atrophy.
|
Regulation of translation factors in the soleus after hindlimb unloading or denervation. It has previously been suggested that the rapid decrease in protein synthesis observed in the soleus muscle after 18 h of hindlimb unloading is due to inhibition at the elongation phase of translation (17). This conclusion was based on polysome profiles, which showed a shift in mRNA from the light to heavy polysome pool (17). However, based on the analysis of translation factors, our data suggest that inhibition also occurs during the initiation of translation after hindlimb unloading. This conclusion is based on the decreased phosphorylation of p70S6k.
As mentioned previously, a decrease in p70S6k phosphorylation has been implicated in regulating the translation of mRNAs that contain a polypyrimidine tract in their 5' untranslated region (11). A decrease in p70S6k phosphorylation was observed after only 12 h of hindlimb unloading, a time point at which the rate of protein synthesis has been shown to be inhibited (34). Additionally, the decrease in p70S6k phosphorylation was still evident after 7 days of hindlimb unloading, again a time point at which the rate of protein synthesis is inhibited (34). The temporal changes in p70S6k phosphorylation are consistent with the temporal changes in protein synthesis rates; therefore, this factor may be an important mediator in the inhibition of protein synthesis. Additionally, many of the mRNAs that are known to be translationally regulated by p70S6k encode proteins such as ribosomal subunits and elongation factors (13). Therefore, it seems possible that a decrease in p70S6k phosphorylation would lead to a decrease in global protein synthesis via a decrease in ribosomal protein components of the translational machinery. This concept of decreased synthetic machinery is also supported by the rapid changes seen in total RNA content after hindlimb unloading and denervation in the soleus muscle (21, 32). The changes we observed in p70S6k phosphorylation suggest that inhibition is occurring at initiation of translation, but this may not be the case for all mRNAs. A more general inhibition of initiation can occur through a decrease in eIF-2B activity. One of the mechanisms that can decrease eIF-2B activity is an increase in the phosphorylation of eIF-2Regulation of translation factors in the EDL after hindlimb unloading or denervation. In our study, the EDL muscle does not atrophy significantly after hindlimb unloading. Consistent with this lack of EDL atrophy were unchanged levels of p70S6k phosphorylation. In contrast, there was a significant atrophy of the EDL after denervation, but surprisingly, at 7 days there was an increase in p70S6k phosphorylation levels. The increase in p70S6k phosphorylation after 7 days of denervation is consistent with an increase in the rate of protein synthesis that has been reported to occur at this time point (8). While unexpected, the EDL-specific increase in p70S6k phosphorylation at 7 days could be due to an increased incidence of stretch as a consequence of the rat maintaining its denervated limb in a plantar-flexed position (8). In support of this concept we observed a similar increase in p70S6k phosphorylation in the tibialis anterior muscle after 7 days of denervation, while no change was seen in the denervated plantaris muscle, a fast fiber-type muscle in the posterior compartment of the hindlimb (data not shown).
An increase in p70S6k phosphorylation is not the only factor that could enhance the rate of protein synthesis in the denervated EDL muscle. In addition to the changes in p70S6k phosphorylation, the quantity of eEF-2 and eIF-2 ![]() |
ACKNOWLEDGEMENTS |
---|
We thank Joshua Lang and Drs. Shann Kim and Mark Fedele for critical review of the manuscript.
![]() |
FOOTNOTES |
---|
This work was supported by National Institute of Arthritis and Musculoskeletal and Skin Diseases Grants AR-45617 (to K. A. Esser) and AR-41705 (to S. C. Kandarian).
Address for reprint requests and other correspondence: K. Esser, School of Kinesiology, 901 W. Roosevelt, Chicago, IL 60608 (E-mail: mlc25{at}uic.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 26 October 2000; accepted in final form 16 February 2001.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Baar, K,
and
Esser K.
Phosphorylation of p70S6k correlates with increased skeletal muscle mass following resistance exercise.
Am J Physiol Cell Physiol
276:
C120-C127,
1999
2.
Carlberg, U,
Nilsson A,
and
Nygard O.
Functional properties of phosphorylated elongation factor 2.
Eur J Biochem
191:
639-645,
1990[Abstract].
3.
Criswell, DS,
Booth FW,
DeMayo F,
Schwartz RJ,
Gordon SE,
and
Fiorotto ML.
Overexpression of IGF-I in skeletal muscle of transgenic mice does not prevent unloading-induced atrophy.
Am J Physiol Endocrinol Metab
275:
E373-E379,
1998
4.
Dufner, A,
and
Thomas G.
Ribosomal S6 kinase signaling and the control of translation.
Exp Cell Res
253:
100-109,
1999[ISI][Medline].
5.
Durante, W,
Liao L,
Reyna SV,
Peyton KJ,
and
Schafer AI.
Physiological cyclic stretch directs L-arginine transport and metabolism to collagen synthesis in vascular smooth muscle.
FASEB J
14:
1775-1783,
2000
6.
Gardiner, PF,
and
Lapointe MA.
Daily in vivo neuromuscular stimulation effects on immobilized rat hindlimb muscles.
J Appl Physiol
53:
960-966,
1982
7.
Goldberg, AL.
Protein turnover in skeletal muscle. II. Effects of denervation and cortisone on protein catabolism in skeletal muscle.
J Biol Chem
244:
3223-3229,
1969
8.
Goldspink, DF.
The effects of denervation on protein turnover of rat skeletal muscle.
Biochem J
156:
71-80,
1976[ISI][Medline].
9.
Herbert, TP,
Kilhams GR,
Batty IH,
and
Proud CG.
Distinct signalling pathways mediate insulin and phorbol ester-stimulated eukaryotic initiation factor 4F assembly and protein synthesis in HEK 293 cells.
J Biol Chem
275:
11249-11256,
2000
10.
Ikeda, M,
Kito H,
and
Sumpio BE.
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].
11.
Jefferies, HB,
Fumagalli S,
Dennis PB,
Reinhard C,
Pearson RB,
and
Thomas G.
Rapamycin suppresses 5'TOP mRNA translation through inhibition of p70s6k.
EMBO J
16:
3693-3704,
1997
12.
Jefferies, HB,
Reinhard C,
Kozma SC,
and
Thomas G.
Rapamycin selectively represses translation of the "polypyrimidine tract" mRNA family.
Proc Natl Acad Sci USA
91:
4441-4445,
1994[Abstract].
13.
Kawasome, H,
Papst P,
Webb S,
Keller GM,
Johnson GL,
Gelfand EW,
and
Terada N.
Targeted disruption of p70s6k defines its role in protein synthesis and rapamycin sensitivity.
Proc Natl Acad Sci USA
95:
5033-5038,
1998
14.
Kim, S,
Jung Y,
Kim D,
Koh H,
and
Chung J.
Extracellular zinc activates p70 S6 kinase through the phosphatidylinositol 3-kinase signaling pathway.
J Biol Chem
275:
25979-25984,
2000
15.
Kimball, SR,
Shantz LM,
Horetsky RL,
and
Jefferson LS.
Leucine regulates translation of specific mRNAs in L6 myoblasts through mTOR-mediated changes in availability of eIF4E and phosphorylation of ribosomal protein S6.
J Biol Chem
274:
11647-11652,
1999
16.
Klemperer, HG.
Lowered proportion of polysomes and decreased amino acid incorporation by ribosomes from denervated muscle.
FEBS Lett
28:
169-172,
1972[ISI][Medline].
17.
Ku, Z,
and
Thomason DB.
Soleus muscle nascent polypeptide chain elongation slows protein synthesis rate during non-weight-bearing activity.
Am J Physiol Cell Physiol
267:
C115-C126,
1994
18.
LeBlanc, A,
Gogia P,
Schneider V,
Krebs J,
Schonfeld E,
and
Evans H.
Calf muscle area and strength changes after five weeks of horizontal bed rest.
Am J Sports Med
16:
624-629,
1988[Abstract].
19.
Loughna, P,
Goldspink G,
and
Goldspink DF.
Effect of inactivity and passive stretch on protein turnover in phasic and postural rat muscles.
J Appl Physiol
61:
173-179,
1986
20.
Loughna, PT,
Goldspink DF,
and
Goldspink G.
Effects of hypokinesia and hypodynamia upon protein turnover in hindlimb muscles of the rat.
Aviat Space Environ Med
58:
A133-A138,
1987[ISI][Medline].
21.
Maltin, CA,
Hay SM,
Delday MI,
Smith FG,
Lobley GE,
and
Reeds PJ.
Clenbuterol, a beta agonist, induces growth in innervated and denervated rat soleus muscle via apparently different mechanisms.
Biosci Rep
7:
525-532,
1987[ISI][Medline].
22.
Merrick, WC.
Mechanism and regulation of eukaryotic protein synthesis.
Microbiol Rev
56:
291-315,
1992[Abstract].
23.
Metafora, S,
Felsani A,
Cotrufo R,
Tajana GF,
Di Iorio G,
Del Rio A,
De Prisco PP,
and
Esposito V.
Neural control of gene expression in the skeletal muscle fibre: the nature of the lesion in the muscular protein-synthesizing machinery following denervation.
Proc R Soc Lond B Biol Sci
209:
239-255,
1980[ISI][Medline].
24.
Panniers, R,
and
Henshaw EC.
A GDP/GTP exchange factor essential for eukaryotic initiation factor 2 cycling in Ehrlich ascites tumor cells and its regulation by eukaryotic initiation factor 2 phosphorylation.
J Biol Chem
258:
7928-7934,
1983
25.
Peters, DG,
Mitchell-Felton H,
and
Kandarian SC.
Unloading induces transcriptional activation of the sarco(endo)plasmic reticulum Ca2+-ATPase 1 gene in muscle.
Am J Physiol Cell Physiol
276:
C1218-C1225,
1999
26.
Price, N,
and
Proud C.
The guanine nucleotide-exchange factor, eIF-2B.
Biochimie
76:
748-760,
1994[ISI][Medline].
27.
Proud, CG,
and
Denton RM.
Molecular mechanisms for the control of translation by insulin.
Biochem J
328:
329-341,
1997[ISI][Medline].
28.
Rosales, OR,
and
Sumpio BE.
Protein kinase C is a mediator of the adaptation of vascular endothelial cells to cyclic strain in vitro.
Surgery
112:
459-466,
1992[ISI][Medline].
29.
Rowlands, AG,
Panniers R,
and
Henshaw EC.
The catalytic mechanism of guanine nucleotide exchange factor action and competitive inhibition by phosphorylated eukaryotic initiation factor 2.
J Biol Chem
263:
5526-5533,
1988
30.
Ryazanov, AG,
Shestakova EA,
and
Natapov PG.
Phosphorylation of elongation factor 2 by EF-2 kinase affects rate of translation.
Nature
334:
170-173,
1988[ISI][Medline].
31.
Sonenberg, N,
Hershey JW,
and
Mathews MB.
Translation control of gene expression. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory, 2000, p. 1020.
32.
Steffen, JM,
and
Musacchia XJ.
Effect of hypokinesia and hypodynamia on protein, RNA, and DNA in rat hindlimb muscles.
Am J Physiol Regulatory Integrative Comp Physiol
247:
R728-R732,
1984
33.
Steffen, JM,
and
Musacchia XJ.
Spaceflight effects on adult rat muscle protein, nucleic acids, and amino acids.
Am J Physiol Regulatory Integrative Comp Physiol
251:
R1059-R1063,
1986
34.
Thomason, DB,
Biggs RB,
and
Booth FW.
Protein metabolism and beta-myosin heavy-chain mRNA in unweighted soleus muscle.
Am J Physiol Regulatory Integrative Comp Physiol
257:
R300-R305,
1989