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FJ
EXPRESS SUMMARY ARTICLE The Full-length version of this article is also available, published online March 5, 2001 as doi:10.1096/fj.00-0717fje. |
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Department of Pediatrics, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-7220, USA
2Correspondence: Department of Pediatrics, Division of Endocrinology, 509 Burnett-Womack Building, CB# 7220, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7220, USA. E-mail: joseph_d'ercole{at}unc.edu
SPECIFIC AIMS
The aim of this study was to define the transcriptional programs up-regulated in liver by starvation. Understanding the molecular basis of the liver’s metabolic switch from the well-fed state to starvation will shed light on the molecular mechanisms underlying the metabolic alterations of nutritional abnormalities, ranging from undernutrition to obesity.
PRINCIPAL FINDINGS
We isolated mRNA from the livers of rats that had been starved for
48 h and from well-fed rats. We constructed a subtracted cDNA
library representing starvation up-regulated genes after differentially
expressed mRNAs were selected by suppression subtractive hybridization.
Positive clones from the library were identified by filter arrays,
followed by RNA dot blots and/or Northern blots for further
confirmation. A total of 54 genes expressed in starved rat liver were
identified and confirmed to be up-regulated by starvation (Table 1
).
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1. Starvation increases the expression of genes involved in energy
metabolism
A total of seven enzymes involved in energy metabolism were shown
to be up-regulated in liver by starvation. These enzymes are involved
in gluconeogenesis, fatty acid oxidation, and ATP biosynthesis and
hydrolysis. For instance, the expression of 3–2-trans-enoyl-CoA
isomerase was induced by starvation. Very long-chain acyl (VLCA) CoA
synthetase and VLCA CoA dehydrogenase were increased 3.2-fold and
2.2-fold, respectively.
2. Starvation increases the expression of genes involved in protein
metabolism
A total of nine genes involved in protein turnover, i.e., protein
synthesis and degradation, were found to be up-regulated during
starvation. Among these are 60S ribosomal subunits L14 (1.4-fold) and
L38 (1.9-fold), 40S ribosomal subunit S9 (1.6-fold),
glutamyl-prolyl-tRNA synthetase (4.2-fold), proteasome subunit Z
(1.9-fold), and subtilisin/kexin isozyme SKI-1 (2.1-fold).
3. Starvation increases the expression of stress-response genes
Starvation elicited a massive hepatic stress response. We found
that the expression of a total of 11 stress-response genes was
increased by starvation. Five of them are acute-phase proteins (both
type 1 and type 2) such as 
-fibrinogen (4.2-fold) and amyloid A
protein (2.4-fold). Another five are proteins involved in a variety of
detoxification systems, such as UDP-glucose dehydrogenase (4.7-fold)
and liver carboxylesterase E (4.6-fold).
4. Starvation increases the expression of nutrient transporter and
receptor genes
The expression of some nutrient transporters and receptors were
up-regulated by starvation. L-type amino acid transporter 1 was
increased 7.1-fold, and N-system amino acid transporter was increased
3.9-fold. Although the effect of nutrient deprivation on the hepatic
expression of monocarboxylate transporter MCT-2 has been controversial,
we showed that starvation increased liver MCT-2 mRNA level by 4.6-fold.
Interleukin 4 receptor, likely involved in mediating hepatic
acute-phase response, was increased twofold.
5. Starvation increases the expression of genes encoding signal
transducers
A total of six genes encoding signal transducers were identified
to be starvation up-regulated. Three of these are transducers of MAP
kinase pathway: p38 MAP kinase (twofold), MAP kinase kinase 2
(2.6-fold), and MAP kinase phosphatase 1 (2.1-fold). Rac 1, a member of
Rho family, was increased 2.2-fold. Src kinase (Csk) was increased
1.5-fold. We also found that starvation up-regulated the expression of
phosphorylated N-glycoprotein pp63 (3.3-fold), a natural inhibitor of
insulin receptor tyrosine kinase.
6. Starvation increases the expression of genes with varied
functions and genes with unknown functions
Starvation increased the expression of eight known genes with
miscellaneous functions as well as eight novel genes. For example, the
mRNA level of apolipoprotein B-100 (involved in LDL catabolism) was
increased 2.1-fold. Delta-4–3-ketosteroid 5-beta-reductase and retinal
dehydrogenase were increased 1.6-fold and 1.7-fold, respectively. One
of the insulin-like growth factor (IGF) binding proteins, IGFBP-2, was
shown to be starvation induced (SI). Among eight novel genes, a
putative peroxisomal protein was SI and a hypothetical RNA binding
protein was increased 1.4-fold.
CONCLUSIONS
Our findings represent a snapshot of genes whose expression is
up-regulated in the liver by starvation. A total of 54 genes are
reported in this study. Most of them fall into one of five major
functional categories and presumably lead to the functional alterations
summarized in Fig. 1
.
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Consistent with our current knowledge in nutritional biochemistry, we found concurrent up-regulation of genes involved in gluconeogenesis and in fatty acid oxidation. We also observed up-regulation of genes involved in protein turnover. These metabolic alterations are accompanied by a coordinate increase in expression of genes that facilitate the transport of large neutral amino acids (LAT-1), glutamine and histidine (NAT-1), and lactate and pyruvate (MCT-2). We believe that starvation-induced protein turnover in liver, as compared with that in muscle, is not likely the primary source of glucogenic amino acids required for hepatic gluconeogenesis. Rather, it is a basic cellular process that serves, among other functions, to regulate cytoplasmic content and provide amino acids for ongoing oxidative and biosynthetic reactions during nutrient deprivation.
Starvation not only causes an imbalance in energy homeostasis; it elicits an orchestrated response, similar to the acute-phase response that is characteristic of tissue injury, inflammation, and other types of insults. We observed moderate increases of at least five genes encoding acute-phase proteins believed to play a role in the restoration of homeostasis. To our knowledge, the hepatic acute-phase response during starvation has not been appreciated and thus is less well understood. Among the genes we identified as stress-response genes, five have detoxification functions. They represent a variety of detoxification systems that are responsible for degrading endogenously generated and exogenous cytotoxic compound.
A cluster of signaling transducers was increased by starvation. Our data suggest that starvation suppresses insulin signaling in part by raising pp63 expression, which in turn inhibits the activity of insulin receptor tyrosine kinase. To our surprise, this natural inhibitor of insulin receptor tyrosine kinase has not been studied extensively. We believe that an increase in pp63 together with the elevated cortisol, glucagon, catecholamines, and growth hormone blood levels is responsible in large part for insulin resistance during starvation and in the acute-phase response. This speculation is also supported by our observation that there is an increased expression of genes involved in hepatic steroid biosynthesis. We found that three major signaling pathways were up-regulated in liver during starvation; they are the MAP kinase pathway, G-protein/Rho pathway, and Src kinase pathway. We speculate that the signaling pathways represented by these genes participate in hepatic acute-phase response as well as the metabolic alterations caused by starvation.
Several other transcription programs were also shown to be increased by starvation. They include genes involved in the biosynthesis of steroids, bile acids, and retinoic acid, insulin-like growth factor binding protein production, LDL consumption, and increased oxidative activity in peroxisomes. In addition, starvation up-regulates the expression of novel genes with yet undefined functions.
In summary, starvation causes major functional changes in the liver including an acute-phase response, a metabolic shift from anabolism to catabolism aided by enhanced nutrient transportation from peripheral tissues, activation of diverse detoxification systems, and initiation of multiple signaling pathways. These findings help to define the mechanisms underlying the metabolic changes that occur in liver during starvation. They should help in probing the metabolic aberrations in the development of obesity and possibly provide clues to a feasible treatment to this health threatening condition.
FOOTNOTES
1 To read the full text of this article, go to http://www.fasebj.org/cgi/doi/10.1096/fj.00-0717fje ; to cite this
article, use FASEB J. (March 5, 2001) 10.1096/fj.00-0717fje 
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