Amino-acid limitation induces transcription from the human C/EBPβ gene via an enhancer activity located downstream of the protein coding sequence

Chin Chen, Elizabeth Dudenhausen, Hong Chen, Yuan Xiang Pan, Altin Gjymishka, Michael S. Kilberg

Research output: Contribution to journalArticle

Abstract

For animals, dietary protein is critical for the nutrition of the organism and, at the cellular level, protein nutrition translates into amino acid availability. Amino acid deprivation triggers the AAR (amino acid response) pathway, which causes enhanced transcription from specific target genes. The present results show that C/EBPβ (CCAAT/enhancer-binding protein β) mRNA and protein content were increased following the deprivation of HepG2 human hepatoma cells of a single amino acid. Although there was a modest increase in mRNA half-life following histidine limitation, the primary mechanism for the elevated steady-state mRNA was increased transcription. Transient transfection documented that C/EBPβ genomic fragments containing the 8451 bp 5′ upstream of the transcription start site did not contain amino-acid-responsive elements. However, deletion analysis of the genomic region located 3′ downstream of the protein coding sequence revealed that a 93 bp fragment contained an amino-acid-responsive activity that functioned as an enhancer. Exogenous expression of ATF4 (activating transcription factor 4), known to activate other genes through amino acid response elements, caused increased transcription from reporter constructs containing the C/EBPβ enhancer in cells maintained in complete amino acid medium. Chromatin immunoprecipitation demonstrated that RNA polymerase II is bound at the C/EBPβ promoter and at the 93 bp regulatory region in vivo, whereas ATF4 binds to the enhancer region only. Immediately following amino acid removal, the kinetics of binding for ATF4, ATF3, and C/EBPβ itself to the 93 bp regulatory region were similar to those observed for the amino-acid-responsive asparagine synthetase gene. Collectively the findings show that expression of C/EBPβ, which contributes to the regulation of amino-acid-responsive genes, is itself controlled by amino acid availability through transcription.

Original languageEnglish (US)
Pages (from-to)649-658
Number of pages10
JournalBiochemical Journal
Volume391
Issue number3
DOIs
StatePublished - Nov 1 2005
Externally publishedYes

Fingerprint

CCAAT-Enhancer-Binding Proteins
Transcription
Genes
Amino Acids
Proteins
Activating Transcription Factor 4
Nucleic Acid Regulatory Sequences
Nutrition
Messenger RNA
Aspartate-Ammonia Ligase
Availability
Dietary Proteins
RNA Polymerase II
Chromatin Immunoprecipitation
Transcription Initiation Site
Response Elements
Histidine
Chromatin
Transfection
Half-Life

Keywords

  • Activating transcription factor 3 (ATF3)
  • Activating transcription factor 4 (ATF4)
  • Amino acid response element (AARE)
  • Basic leucine-zipper (bZIP)
  • CCAAT/enhancer-binding protein (C/EBP)
  • nutrient starvation

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Amino-acid limitation induces transcription from the human C/EBPβ gene via an enhancer activity located downstream of the protein coding sequence. / Chen, Chin; Dudenhausen, Elizabeth; Chen, Hong; Pan, Yuan Xiang; Gjymishka, Altin; Kilberg, Michael S.

In: Biochemical Journal, Vol. 391, No. 3, 01.11.2005, p. 649-658.

Research output: Contribution to journalArticle

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T1 - Amino-acid limitation induces transcription from the human C/EBPβ gene via an enhancer activity located downstream of the protein coding sequence

AU - Chen, Chin

AU - Dudenhausen, Elizabeth

AU - Chen, Hong

AU - Pan, Yuan Xiang

AU - Gjymishka, Altin

AU - Kilberg, Michael S.

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AB - For animals, dietary protein is critical for the nutrition of the organism and, at the cellular level, protein nutrition translates into amino acid availability. Amino acid deprivation triggers the AAR (amino acid response) pathway, which causes enhanced transcription from specific target genes. The present results show that C/EBPβ (CCAAT/enhancer-binding protein β) mRNA and protein content were increased following the deprivation of HepG2 human hepatoma cells of a single amino acid. Although there was a modest increase in mRNA half-life following histidine limitation, the primary mechanism for the elevated steady-state mRNA was increased transcription. Transient transfection documented that C/EBPβ genomic fragments containing the 8451 bp 5′ upstream of the transcription start site did not contain amino-acid-responsive elements. However, deletion analysis of the genomic region located 3′ downstream of the protein coding sequence revealed that a 93 bp fragment contained an amino-acid-responsive activity that functioned as an enhancer. Exogenous expression of ATF4 (activating transcription factor 4), known to activate other genes through amino acid response elements, caused increased transcription from reporter constructs containing the C/EBPβ enhancer in cells maintained in complete amino acid medium. Chromatin immunoprecipitation demonstrated that RNA polymerase II is bound at the C/EBPβ promoter and at the 93 bp regulatory region in vivo, whereas ATF4 binds to the enhancer region only. Immediately following amino acid removal, the kinetics of binding for ATF4, ATF3, and C/EBPβ itself to the 93 bp regulatory region were similar to those observed for the amino-acid-responsive asparagine synthetase gene. Collectively the findings show that expression of C/EBPβ, which contributes to the regulation of amino-acid-responsive genes, is itself controlled by amino acid availability through transcription.

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