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Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA 17033
2To whom correspondence should be addressed. E-mail: jjefferson{at}psu.edu.
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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KEY WORDS: • leucine • liver • ribosomal protein mRNA translation • rats
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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]. This step requires a multisubunit complex, referred
to as eukaryotic initiation factor
(eIF)3
4F. Assembly
of the eIF4F complex is regulated by the phosphorylation state of the
translational repressor, eIF4E-binding protein 1 (4E-BP1). 4E-BP1
prevents formation of the eIF4F complex by sequestering the mRNA cap
binding protein, eIF4E, into an inactive complex under conditions of
hypophosphorylation. Hyperphosphorylation of 4E-BP1 allows elF4E to
bind elF4G, a large protein that serves as a scaffold for elF4F complex
assembly.
Alterations in the rate of protein synthesis reflect the translation of
all mRNAs in the cell, although not necessarily to the same extent. The
translation of a single or subset of mRNAs can be discordant with
overall rates of protein synthesis. One class of mRNAs that demonstrate
specific regulation includes the ribosomal proteins (rp), a family of
80 members that associate in equimolar amounts with the ribosomal RNAs
to form a mature 80S ribosome. Mammalian rp mRNAs are characterized
structurally by the presence of a Terminal
OligoPyrimidine tract (TOP) at the 5'-end of the
transcript, consisting of a C residue adjacent to the m7GTP
cap site, followed by an uninterrupted sequence of 4–20 pyrimidines
(2)
. Genes that contain a TOP tract in their 5'
untranslated region code for proteins involved in the production and
function of the translational apparatus (3)
.
Apportionment of rp mRNAs across polysome profiles suggests two
discrete populations, i.e., translationally active (and thus, polysome
associated) under growth conditions and translationally inactive (and
thus, resident in subpolysomal fractions) under conditions of growth
inhibition (4)
. Recruitment of TOP mRNAs into polysomes is
associated with hyperphosphorylation of the 70-kDa ribosomal protein S6
kinase (S6K1). Phosphorylation of rp S6 by S6K1 is thought to
facilitate the initiation process by enhancing the affinity of the
ribosome for binding TOP mRNAs (5)
. In
mitogen-stimulated cells, the efficiency of translation of TOP
mRNAs is mediated by the activity of S6K1 (6)
. This effect
is specific for TOP mRNAs because treatment of cells with rapamycin, an
inhibitor of S6K1 phosphorylation, selectively suppresses translation
of TOP mRNAs (7)
.
Previous work in our laboratory demonstrated that oral administration
of the branched-chain amino acid (BCAA) leucine alone stimulates
the synthesis of total mixed proteins in muscle concomitant with
increased eIF4F assembly and S6K1 phosphorylation (8
,9)
.
However, it is not known whether feeding leucine stimulates translation
initiation and rates of protein synthesis in tissues other than
skeletal muscle. The purpose of this study was to investigate the role
of leucine in regulating total vs. specific protein synthesis in the
liver. Particular emphasis was placed on the regulation of rp mRNA
translation and the role of S6K1 in mediating enhanced translation of
TOP-containing mRNAs in vivo.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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The animal facilities and protocol were reviewed and approved by the
Institutional Animal Care and Use Committee of The Pennsylvania State
University College of Medicine. Male Sprague-Dawley rats (
200 g)
were maintained on a 12-h light:dark cycle with food (Harlan-Teklad
Rodent Chow, Madison, WI) and water provided freely. Rats were deprived
of food for 18 h and then randomly assigned to continue as
food-deprived (Con) or to receive one of three dietary treatments
by oral gavage as follows: L-leucine (Leu),
L-isoleucine (Ile) or L-valine (Val). The
amount of each amino acid administered was 135 mg/100 g body, prepared
as 54.0 g/L in distilled water. Food-deprived rats (Con) received
an equal volume of saline (0.155 mol/L) by oral gavage.
Sample collection.
Exactly 1 h after oral gavage, rats were killed by decapitation. Trunk blood was collected and centrifuged at 1,800 x g for 10 min to obtain serum. The whole liver was excised, blotted, weighed and divided into three parts. One portion of liver was weighed and homogenized in 3 volumes of Buffer A, consisting of (in mmol/L) 40 N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (pH 7.5), 100 potassium chloride and 5 magnesium chloride. The Buffer A homogenate was centrifuged at 3000 x g for 15 min at 4°C for subsequent polysome profile analysis, as described below. A second portion of liver was homogenized in 7 volumes of Buffer B, consisting of (in mmol/L) 20 N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (pH 7.4), 100 potassium chloride, 0.2 EDTA, 2 ethylene glycol-bis(ß-aminoethyl ether)-N,N,N',N'-tetraacetic acid, 1 dithiothreitol, 50 sodium fluoride, 50 ß-glycerophosphate, 0.1 phenylmethylsulfonyl fluoride, 1 benzamidine and 0.5 sodium vanadate. An aliquot of the Buffer B homogenate was used for the measurement of protein synthesis as described below. The remaining Buffer B homogenate was immediately centrifuged at 10,000 x g for 10 min at 4°C. The resulting supernatant was used to examine the phosphorylation of 4E-BP1, S6K1 and rpS6 as described below. The remaining portion of liver was quickly frozen in liquid nitrogen for the subsequent isolation of total RNA. All serum and tissue samples were stored at -70°C.
Measurement of liver protein synthesis.
A bolus (1.0 mL/100 g body) of
L-[2,3,4,5,6-3H] phenylalanine (150 mmol/L
containing 3.70 GBq/L, Amersham, Piscataway, NJ) was injected via the
tail vein 50 min after oral gavage for the measurement of hepatic
protein synthesis (10)
. The elapsed time from injection of
the metabolic tracer until homogenization of liver was recorded as the
actual time for incorporation of radiolabeled amino acid into protein.
Fractional rates of protein synthesis were estimated from the rate of
incorporation of radioactive phenylalanine into liver protein, using
the specific radioactivity of serum phenylalanine as an indication of
the precursor pool (11)
. The use of serum phenylalanine as
the precursor pool was validated (12)
.
Polysome profiles.
One volume of detergent (10% Triton X-100, 10% sodium deoxycholate) was mixed with nine volumes of postnuclear supernatant (Buffer A) and layered over 10–70% linear sucrose density gradients. The gradients were centrifuged at 90,000 x g for 210 min at 4°C in a Beckman SW28 rotor (Palo Alto, CA). After centrifugation, the gradients were fractionated on an Isco density gradient fractionator (Lincoln, NE). The UV absorption at 254 nm was recorded continuously and 5-mL fractions were collected for subsequent extraction of total RNA.
Total RNA extraction from whole liver and sucrose gradients.
Total RNA from frozen liver samples was isolated using Tri Reagent (13) (Molecular Research Center, Cincinnati, OH). Sucrose fractions were acid phenol/chloroform extracted twice (1:1, v/v, sucrose/acid phenol, pH 4.5, Ambion, Austin, TX) and ethanol precipitated overnight. The RNA concentration and purity of all samples were determined by measuring the UV absorbance at 260 nm and the ratio of optical densitites 260 nm/280 nm, respectively.
Preparation of RNA probes.
The full-length cDNAs for ribosomal proteins S4, S8 and L26 were
kindly provided by Dr. Ira Wool (Department of Biochemistry and
Molecular Biology, University of Chicago, Chicago, IL). The plasmid
pUC8 was digested with PstI to produce 250-, 212- and 185-bp fragments
for L26, S4 and S8, respectively. The cDNA for albumin was in the
pBR322 vector and the plasmid pAlb576 (14)
was digested
with EcoRV followed by AccI to produce a 280-bp albumin fragment. The
four fragments were subcloned individually into the pBluescript II SK +
vector (Stratagene, La Jolla, CA) and transformed into bacteria (TOP10
One-Shot cells, Invitrogen, Carlsbad, CA). Colonies were isolated
and DNA was purified (Wizard Plus Mini and Maxi-Preps DNA
Purification System, Promega, Madison, WI). Plasmid stocks of
pBluescript-L26, pBluescript-S4, pBluescript-S8 and pBluescript-Alb
were then linearized with EcoRI, BamHI, HindIII and AccI, respectively,
to produce DNA template stocks of each probe for in vitro
transcription. A linearized ß-actin DNA template (126 bp) was
purchased from Ambion. One microgram of each DNA template was mixed
with 25 µmol/L [32P]UTP (800 Ci/mmol, 30
TBq/mmol; Amersham) and limiting (0.1 mmol/L) unlabeled UTP in a
20-µL transcription reaction according to the
manufacturer’s instructions (MAXIscript In Vitro Transcription Kit,
Ambion) to produce single-stranded RNA probes. The resulting
reactions were treated with DNase I, heat denatured and gel purified by
loading onto a 5% acrylamide-8 mol/L urea mini-slab gel. After
electrophoresis, the full-length probes were located by
short-term (20 s) exposure of the gel to X-ray film. The
full-length RNA probes were subsequently excised from the gel with
a clean blade and incubated overnight in probe elution buffer (0.5
mol/L ammonium acetate, 1 mmol/L EDTA, 0.2% SDS; RPA III, Ambion). The
gel was briefly reexposed to X-ray film after excision to ensure
that the correct bands were isolated.
Ribonuclease protection assay.
Total RNA (5 µg) isolated from either whole liver or sucrose density gradient fractions was coprecipitated with L26, S4, S8 and ß-actin RNA probes in a single tube. Albumin mRNA expression was detected in a separate ribonuclease protection assay. Samples were heat denatured and allowed to hybridize to the RNA probes overnight in a 42°C water bath in hybridization buffer (RPA III, Ambion). Samples were digested with RNaseA/TI the next day, and the protected, double-stranded mRNA fragments were ethanol precipitated, heat denatured and loaded onto 5% acrylamide-8 mol/L urea slab gels. After electrophoresis, gels were wrapped in plastic and exposed to X-ray film at -70°C for up to 4 h.
Examination of 4E-BP1 phosphorylation state.
An aliquot of the 10,000 x g supernatant (Buffer
B) was boiled for 10 min and then centrifuged at 10,000 x g for 30 min at 4°C. The resulting supernatant was
mixed with an equal volume of 2X sample buffer and then subjected to
protein immunoblot analysis as described previously (15)
.
Quantitation of eIF4G · eIF4E complexes.
eIF4E was immunoprecipitated from 10,000 x g (Buffer B) supernatants of liver homogenates using a monoclonal antibody to eIF4E (15). Next, samples were subjected to immunoblot analysis using a polyclonal antibody to eIF4G to determine the association of eIF4G with eIF4E (15). Results were normalized to the amount of eIF4E in the immunoprecipitates.
Phosphorylation of S6K1 and ribosomal protein S6.
Phosphorylation of S6K1 and rpS6 was determined in 10,000 x g (Buffer B) supernatants by protein immunoblot analysis
as previously described (16)
. Phosphorylation of S6K1 at
Thr389, a site whose phosphorylation is associated with maximal
activation of the kinase, was determined using an anti-phospho-S6K1
(Thr389) antibody (Cell Signaling Technology, Beverly, MA). The
anti-rpS6 antibody was raised against a phosphopeptide that included
five phosphorylation sites in the C-terminus of the protein
(generously provided by Dr. M. Birnbaum, University of Pennsylvania).
Statistical analysis.
All data were analyzed by the Statistica statistical software package for the Macintosh, volume II (StatSoft, Tulsa, OK). Data were analyzed using one-way ANOVA to assess main effects, with treatment group as the independent variable. When a significant overall effect was detected, differences among individual means were assessed with Duncan’s Multiple Range post-hoc test. The level of significance was set at P < 0.05 for all statistical tests.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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,9
,17)
, oral administration of leucine did not stimulate
fractional rates of protein synthesis in the liver of food-deprived
rats (Fig. 1
). Isoleucine and valine were additionally without effect.
|
-form) corresponding to the
most highly phosphorylated species. Hyperphosphorylation of 4E-BP1
results in the release of the mRNA cap binding protein, eIF4E, allowing
it to participate in eIF4F assembly. All three BCAA increased the
phosphorylation of 4E-BP1 relative to food-deprived controls, but
the effect was graded, with leucine > valine and isoleucine
intermediary (Fig. 2A
).To support the concept that increases in 4E-BP1 phosphorylation
promote eIF4F assembly, the association of eIF4G with eIF4E was
examined. Oral leucine administration significantly enhanced the
binding of eIF4G to eIF4F, whereas oral administration of isoleucine
and valine were not effective (Fig. 2B
).
|
). S6K1 resolves on SDS polyacrylamide gels into multiple
electrophoretic forms, with more slowly migrating species corresponding
to more highly phosphorylated forms of the protein. To determine
whether the observed phosphorylation patterns corresponded to a
potential change in activity, we examined phosphorylation of S6K1 at
Thr389, a site that has been shown to be associated with maximal
activity of the protein when phosphorylated (18)
. Similar
to 4E-BP1, phosphorylation at this site was graded, with leucine
> isoleucine > valine compared with controls (Fig. 3B
). Finally, we examined the phosphorylation of the
ribosomal protein S6. Phosphorylation of rpS6 was significantly
increased only in rats administered leucine (Fig. 3C
). These
results suggest that among the BCAA, leucine is most effective at
stimulating the activation of the S6K1 signaling pathway in liver.
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), the first two fractions were designated "subpolysomal." These
fractions contained protein, RNA, free ribosomal subunits and
monosomes. The third and fourth fractions were then designated
"polysomal." These fractions contained disomes, trisomes and
polysomes of increasing ribosome number bound to mRNA. The total RNA
from each sucrose fraction was extracted and the content of both rp and
non-rp mRNAs was determined in each fraction. The results of this
analysis provide a relative measure of how actively each message was
being translated under the experimental conditions. The distribution of
three TOP-containing messages, rpL26, rpS4 and rpS8, was compared
with that of albumin and ß-actin (both non-TOP mRNAs) (Fig. 4)
.
Total mRNA expression of all proteins examined was not altered 1 h
after BCAA administration in whole liver (Fig. 5
).
|
|
and Table 1
). The albumin gene demonstrated slight-to-moderate movement among
fractions when food deprivation was compared with BCAA administration,
but this difference was not significant (ANOVA, P < 0.06). The tendency for albumin mRNA to shift into polysomes with BCAA
administration agrees with a previous study by Yap et al.
(19)
showing that feeding a single or mixture of
amino acids to food-deprived rats increases the proportion of
albumin mRNA resident in free and membrane-bound polysomes. In any
case, the majority (61–81%) of albumin mRNA remained associated with
heavy polysomes in all treatment groups (Fig. 4
and Table 1
). In
contrast to these non-TOP transcripts, most (70–75%) of the mRNAs
encoding the ribosomal proteins were unloaded from polysomes in
food-deprived controls, residing in the subpolysomal fractions
(Fig. 4
and Table 1
). Oral administration of isoleucine or valine did
not significantly shift the proportion of these mRNAs to the
polysome fractions compared with controls (Fig. 4
and Table 1
). In
contrast, oral administration of leucine promoted significant
redistribution of ribosomal protein mRNAs, more than doubling the
proportion of transcripts existing in the polysome fractions (Fig. 4
and Table 1
). These data suggest that among the BCAA, leucine alone
favors increased translation of hepatic ribosomal proteins. This effect
appears to relate to the degree of S6K1 phosphorylation in vivo.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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,21)
. On the other
hand, essential amino acid deprivation or imbalance causes
disaggregation of polysomes and represses global rates of protein
synthesis in the liver (20
,22)
.
Beyond their use as substrates, the branched-chain amino acids, and
in particular leucine, function as signaling molecules to promote the
initiation of mRNA translation in skeletal muscle
(8
,9
,23)
. Contrary to our published findings in skeletal
muscle, oral administration of leucine, isoleucine or valine did not
enhance the synthesis of total liver protein. Supporting these
findings, perfusion of rat liver with leucine alone at a concentration
4 times that reported in plasma of food-deprived rats resulted in
liver protein synthesis rates that were not different from those of
rats perfused with medium containing all amino acids at 1X
concentrations (20)
. Additionally, in the only published
report examining the short-term effect of force-feeding a
single BCAA on liver polysome profiles and protein synthesis,
isoleucine did not stimulate polysome aggregation or liver protein
synthesis in rats (24)
. Collectively, the data indicate
that the BCAA by themselves are not sufficient to stimulate synthesis
of total liver protein.
Studies in cells and in vivo demonstrate that amino acids stimulate
protein synthesis congruous with the hyperphosphorylation of 4E-BP1 and
S6K1, two proteins important in the selective control of mRNA
translation (8
,17
,21
,23)
. Hyperphosphorylation of S6K1
appears to augment the translation of a specific class of genes
characterized by the presence of an oligopyrimidine tract at the
immediate 5' end of the transcript (referred to as TOP)
(2)
. The relationship between S6K1 activation and the
selective translation of TOP mRNA was established using the
immunosuppressant drug rapamycin, an inhibitor of S6K1 phosphorylation
and activation (7
,25)
. Treatment of serum-stimulated
NIH 3T3 cells with rapamycin blocks S6K1 phosphorylation and
recruitment of rp mRNA into polysomes (7)
. These
inhibitory effects are nullified upon conversion of the principal
rapamycin-sensitive phosphorylation site on S6K1 to a residue
refractory to the macrolide (7)
. Additionally, disruption
of the 5'TOP sequence also prevents the inhibitory effects of rapamycin
on rp mRNA translation, highlighting the importance of the
cis-regulatory element in the selective control of this
class of genes.
In this study, we examined the relationship between BCAA-induced
activation of the S6K1 signaling pathway and the selective movement of
rp mRNAs into polysomes. S6K1 activation has been linked to leucine
availability in hepatoma cells and perfused liver (20
, 26)
, and the connection between S6K1, rpS6 and the translation
of TOP mRNAs has been described (4)
. However, there are no
reports testing the effects of amino acid availability on rp mRNA
translation in vivo. Herein we show for the first time the unique role
of leucine in stimulating redistribution of rp mRNAs into polysomes,
suggesting increased translation of these proteins. This type of
regulation may provide temporal control of gene expression under
conditions of altered growth rate.
In this study, leucine promoted the hyperphosphorylation of 4E-BP1 and
enhanced eIF4F assembly, implying increased translation initiation in
the liver. Studies in perfused rat liver similarly demonstrate leucine
alone to enhance eIF4F assembly without altering the synthesis of total
mixed protein (20)
. This apparent paradox may be resolved
by considering that the translation of mRNAs with highly structured
5'-untranslated regions may have been selectively upregulated under the
experimental conditions and that the proportion of total mRNA
represented by this subset of mRNAs is low. Changes in eIF4E
availability are proposed to specifically modulate the synthesis of a
family of mRNAs that possess significant secondary structure in their
5'-untranslated regions (23
,27)
. Although examples of this
class of genes (such as ornithine decarboxylase) were not examined in
the current investigation, on the basis of our eIF4F assembly results,
it is likely that leucine would have affected the translation of this
class of genes to some degree. On the other hand, TOP mRNAs do not
contain highly structured 5'-untranslated regions (2)
, and
therefore, are unlikely candidates for direct regulation via 4E-BP1
hyperphosphorylation. Studies using S6K1 mutants resistant to rapamycin
demonstrate the drug to repress 4E-BP1 phosphorylation despite
continued S6K1 activation and rp mRNA translation (7
,28)
.
Therefore, it is unlikely that redistribution of rp mRNAs in the
current study is directly linked to 4E-BP1 phosphorylation or eIF4F
assembly.
In summary, we report the unique action of leucine to promote the movement of hepatic ribosomal protein mRNAs into polysomes. This event is disjointed from overall rates of liver protein synthesis and is associated with increased phosphorylation of S6K1 and rpS6. These results suggest that leucine is both necessary and sufficient as a signaling molecule to promote ribosomal protein mRNA translation in the liver via the S6K1 signaling pathway. On the other hand, increases in the synthesis of total liver proteins require the activation of other signaling pathways that may be contingent on other factors, such as a balanced supply of amino acids.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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3 Abbreviations used: BCAA, branched-chain amino acids; Con, food-deprived rats; 4E-BP1, eIF4E-binding protein 1; eIF, eukaryotic initiation factor; Ile, food-deprived rats orally administered 270 mg isoleucine; Leu, food-deprived rats orally administered 270 mg leucine; rp, ribosomal protein; S6K1, 70-kDa ribosomal protein S6 kinase; TOP, terminal oligopyrimidine tract; Val, food-deprived rats orally administered 270 mg valine.
Manuscript received October 16, 2000. Initial review completed November 28, 2000. Revision accepted January 23, 2001.
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 C. A. Toerien and J. P. Cant Abundance and Phosphorylation State of Translation Initiation Factors in Mammary Glands of Lactating and Nonlactating Dairy Cows J Dairy Sci, June 1, 2007; 90(6): 2726 - 2734. [Abstract] [Full Text] [PDF]
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 M. P. Engelen, E. P. Rutten, C. L. De Castro, E. F. Wouters, A. M. Schols, and N. E. Deutz Supplementation of soy protein with branched-chain amino acids alters protein metabolism in healthy elderly and even more in patients with chronic obstructive pulmonary disease Am. J. Clinical Nutrition, February 1, 2007; 85(2): 431 - 439. [Abstract] [Full Text] [PDF]
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T. G. Anthony, B. J. McDaniel, P. Knoll, P. Bunpo, G. L. Paul, and M. A. McNurlan Feeding Meals Containing Soy or Whey Protein after Exercise Stimulates Protein Synthesis and Translation Initiation in the Skeletal Muscle of Male Rats J. Nutr., February 1, 2007; 137(2): 357 - 362. [Abstract] [Full Text] [PDF]
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 R. Ramakrishnan Studying apolipoprotein turnover with stable isotope tracers: correct analysis is by modeling enrichments J. Lipid Res., December 1, 2006; 47(12): 2738 - 2753. [Abstract] [Full Text] [PDF]
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 J. M. Rhoads, X. Niu, J. Odle, and L. M. Graves Role of mTOR signaling in intestinal cell migration Am J Physiol Gastrointest Liver Physiol, September 1, 2006; 291(3): G510 - G517. [Abstract] [Full Text] [PDF]
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M. D. Sans, M. Tashiro, N. L. Vogel, S. R. Kimball, L. G. D'Alecy, and J. A. Williams Leucine Activates Pancreatic Translational Machinery in Rats and Mice through mTOR Independently of CCK and Insulin J. Nutr., July 1, 2006; 136(7): 1792 - 1799. [Abstract] [Full Text] [PDF]
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 M. I. Lewis, S. C. Bodine, N. Kamangar, X. Xu, X. Da, and M. Fournier Effect of severe short-term malnutrition on diaphragm muscle signal transduction pathways influencing protein turnover J Appl Physiol, June 1, 2006; 100(6): 1799 - 1806. [Abstract] [Full Text] [PDF]
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P. Anand and P. A. Gruppuso Rapamycin Inhibits Liver Growth during Refeeding in Rats via Control of Ribosomal Protein Translation but Not Cap-Dependent Translation Initiation J. Nutr., January 1, 2006; 136(1): 27 - 33. [Abstract] [Full Text] [PDF]
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S. Fujita and E. Volpi Amino Acids and Muscle Loss with Aging J. Nutr., January 1, 2006; 136(1): 277S - 280S. [Abstract] [Full Text] [PDF]
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M. Doi, I. Yamaoka, M. Nakayama, S. Mochizuki, K. Sugahara, and F. Yoshizawa Isoleucine, a Blood Glucose-Lowering Amino Acid, Increases Glucose Uptake in Rat Skeletal Muscle in the Absence of Increases in AMP-Activated Protein Kinase Activity J. Nutr., September 1, 2005; 135(9): 2103 - 2108. [Abstract] [Full Text] [PDF]
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P. J. Garlick The Role of Leucine in the Regulation of Protein Metabolism J. Nutr., June 1, 2005; 135(6): 1553S - 1556S. [Abstract] [Full Text] [PDF]
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 A. K. Reiter, D. R. Bolster, S. J. Crozier, S. R. Kimball, and L. S. Jefferson Repression of protein synthesis and mTOR signaling in rat liver mediated by the AMPK activator aminoimidazole carboxamide ribonucleoside Am J Physiol Endocrinol Metab, May 1, 2005; 288(5): E980 - E988. [Abstract] [Full Text] [PDF]
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A. K. Reiter, S. J. Crozier, S. R. Kimball, and L. S. Jefferson Meal Feeding Alters Translational Control of Gene Expression in Rat Liver J. Nutr., March 1, 2005; 135(3): 367 - 375. [Abstract] [Full Text] [PDF]
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A. Jaleel and K. S. Nair Identification of multiple proteins whose synthetic rates are enhanced by high amino acid levels in rat hepatocytes Am J Physiol Endocrinol Metab, June 1, 2004; 286(6): E950 - E957. [Abstract] [Full Text] [PDF]
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 S. Caldarola, F. Amaldi, C. G. Proud, and F. Loreni Translational Regulation of Terminal Oligopyrimidine mRNAs Induced by Serum and Amino Acids Involves Distinct Signaling Events J. Biol. Chem., April 2, 2004; 279(14): 13522 - 13531. [Abstract] [Full Text] [PDF]
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N. Hashimoto and H. Hara Dietary Amino Acids Promote Pancreatic Protease Synthesis at the Translation Stage in Rats J. Nutr., October 1, 2003; 133(10): 3052 - 3057. [Abstract] [Full Text] [PDF]
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C. J. Lynch, B. J. Patson, J. Anthony, A. Vaval, L. S. Jefferson, and T. C. Vary Leucine is a direct-acting nutrient signal that regulates protein synthesis in adipose tissue Am J Physiol Endocrinol Metab, September 1, 2002; 283(3): E503 - E513. [Abstract] [Full Text] [PDF]
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T. G. Anthony, A. K. Reiter, J. C. Anthony, S. R. Kimball, and L. S. Jefferson Deficiency of dietary EAA preferentially inhibits mRNA translation of ribosomal proteins in liver of meal-fed rats Am J Physiol Endocrinol Metab, September 1, 2001; 281(3): E430 - E439. [Abstract] [Full Text] [PDF]
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T. A. Davis, M. L. Fiorotto, D. G. Burrin, P. J. Reeds, H. V. Nguyen, P. R. Beckett, R. C. Vann, and P. M. J. O'Connor Stimulation of protein synthesis by both insulin and amino acids is unique to skeletal muscle in neonatal pigs Am J Physiol Endocrinol Metab, April 1, 2002; 282(4): E880 - E890. [Abstract] [Full Text] [PDF]
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