Mammalian cells respond to nutritional limitation of protein/amino acids by increasing the expression of a wide spectrum of proteins via a signaling pathway that will be referred to as the Amino Acid Response (AAR). Although it is known that amino acid availability can modulate protein expression, the mechanisms by which these events occur are not well understood. Using human HepG2 hepatoma cells, we have documented previously that amino acid limitation increases the expression of the bZIP transcription factor C/EBPb through transcriptional control and that, in turn, an elevated level of C/EBPb induces transcription from amino acid responsive genes. Initial studies for this application have yielded the novel observation that there is amino acid response element (AARE) activity in a 93 bp fragment from the human C/EBPb gene 3' to the protein coding sequence. Preliminary experiments have also shown that amino acid limitation causes an increase in the total abundance of C/EBPb protein and an increase in the nuclear translocation of C/EBPb phosphorylated on Thr235. Our global hypothesis is that transcriptional control of human C/EBPb expression and post-translational control of C/EBPb function represent important regulatory steps in the AAR pathway. Using protein restricted diets in rats and amino acid limitation of HepG2 cells the Specific Aims of the proposed studies are to: 1) identify the genomic AARE responsible for amino acid-dependent transcription of the C/EBPb gene by using deletion analysis, in vivo footprinting, and single nucleotide mutagenesis;2) identify the AARE binding proteins and characterize their role in the AAR pathway;3) determine if there are changes in synthesis or turnover of one of the three C/EBPb protein isoforms (LAP*, LAP, LIP);and 4) investigate whether or not phosphorylation of C/EBPb is important in regulating the role that it plays in signaling amino acid availability. Our long-term goal is to understand how mammalian cells respond to their nutritional environment through gene expression, using amino acid availability as the model.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
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Integrative Nutrition and Metabolic Processes Study Section (INMP)
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Maruvada, Padma
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University of Florida
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Su, Nan; Thiaville, Michelle M; Awad, Keytam et al. (2009) Protein or amino acid deprivation differentially regulates the hepatic forkhead box protein A (FOXA) genes through an activating transcription factor-4-independent pathway. Hepatology 50:282-90
Kilberg, Michael S; Shan, Jixiu; Su, Nan (2009) ATF4-dependent transcription mediates signaling of amino acid limitation. Trends Endocrinol Metab 20:436-43
Palii, S S; Kays, C E; Deval, C et al. (2009) Specificity of amino acid regulated gene expression: analysis of genes subjected to either complete or single amino acid deprivation. Amino Acids 37:79-88
Abbatiello, Susan E; Pan, Yuan-Xiang; Zhou, Mi et al. (2008) Mass spectrometric quantification of asparagine synthetase in circulating leukemia cells from acute lymphoblastic leukemia patients. J Proteomics 71:61-70
Thiaville, Michelle M; Pan, Yuan-Xiang; Gjymishka, Altin et al. (2008) MEK signaling is required for phosphorylation of eIF2alpha following amino acid limitation of HepG2 human hepatoma cells. J Biol Chem 283:10848-57
Su, Nan; Kilberg, Michael S (2008) C/EBP homology protein (CHOP) interacts with activating transcription factor 4 (ATF4) and negatively regulates the stress-dependent induction of the asparagine synthetase gene. J Biol Chem 283:35106-17
Thiaville, Michelle M; Dudenhausen, Elizabeth E; Awad, Keytam S et al. (2008) Activated transcription via mammalian amino acid response elements does not require enhanced recruitment of the Mediator complex. Nucleic Acids Res 36:5571-80
Aiken, Kimberly J; Bickford, Justin S; Kilberg, Michael S et al. (2008) Metabolic regulation of manganese superoxide dismutase expression via essential amino acid deprivation. J Biol Chem 283:10252-63
Thiaville, Michelle M; Dudenhausen, Elizabeth E; Zhong, Can et al. (2008) Deprivation of protein or amino acid induces C/EBPbeta synthesis and binding to amino acid response elements, but its action is not an absolute requirement for enhanced transcription. Biochem J 410:473-84
Pan, Yuan-Xiang; Chen, Hong; Thiaville, Michelle M et al. (2007) Activation of the ATF3 gene through a co-ordinated amino acid-sensing response programme that controls transcriptional regulation of responsive genes following amino acid limitation. Biochem J 401:299-307

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