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ATF2 is required for amino acid-regulated transcription by orchestrating specific histone acetylation.

Bruhat A, Chérasse Y, Maurin AC, Breitwieser W, Parry L, Deval C, Jones N, Jousse C, Fafournoux P - Nucleic Acids Res. (2007)

Bottom Line: Using ATF2-deficient mouse embryonic fibroblasts, we demonstrate that ATF2 is essential in the acetylation of histone H4 and H2B in vivo.The role of ATF2 on histone H4 acetylation is dependent on its binding to the AARE and can be extended to other amino acid regulated genes.Thus, ATF2 is involved in promoting the modification of the chromatin structure to enhance the transcription of a number of amino acid-regulated genes.

View Article: PubMed Central - PubMed

Affiliation: UMR 1019, Unité de Nutrition Humaine, INRA de Theix, 63122 Saint Genès Champanelle, France. bruhat@clermont.inra.fr

ABSTRACT
The transcriptional activation of CHOP (a CCAAT/enhancer-binding protein-related gene) by amino acid deprivation involves the activating transcription factor 2 (ATF2) and the activating transcription factor 4 (ATF4) binding the amino acid response element (AARE) within the promoter. Using a chromatin immunoprecipitation approach, we report that in vivo binding of phospho-ATF2 and ATF4 to CHOP AARE are associated with acetylation of histones H4 and H2B in response to amino acid starvation. A time course analysis reveals that ATF2 phosphorylation precedes histone acetylation, ATF4 binding and the increase in CHOP mRNA. We also show that ATF4 binding and histone acetylation are two independent events that are required for the CHOP induction upon amino acid starvation. Using ATF2-deficient mouse embryonic fibroblasts, we demonstrate that ATF2 is essential in the acetylation of histone H4 and H2B in vivo. The role of ATF2 on histone H4 acetylation is dependent on its binding to the AARE and can be extended to other amino acid regulated genes. Thus, ATF2 is involved in promoting the modification of the chromatin structure to enhance the transcription of a number of amino acid-regulated genes.

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Transcription factor binding and histone acetylation to CHOP AARE in response to leucine starvation. (A) Scheme of the human CHOP gene indicating the different amplicons produced for the ChIP analysis: A (−1678 to −1478), B (−472 to −301) and C (+1163 to +1372). The AARE is boxed in grey. (B) HeLa cells were incubated 2 h either in control (+leu) or leucine-free medium (−leu) and harvested. ChIP analysis was performed as described under Materials and Methods using antibodies specific for ATF2 and ATF4 and different sets of primers to produce amplicon A, B or C. Data were plotted as the percentage of antibody binding versus the amount of PCR product obtained using a standardized aliquot of input chromatin. Each point represents the mean value of three independent experiments, and the error bars represent the standard error of the means. (C) The experiment described in (B) was also performed using antibodies specific for acetylated H3, acetylated H4 and acetylated H2B.
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Figure 2: Transcription factor binding and histone acetylation to CHOP AARE in response to leucine starvation. (A) Scheme of the human CHOP gene indicating the different amplicons produced for the ChIP analysis: A (−1678 to −1478), B (−472 to −301) and C (+1163 to +1372). The AARE is boxed in grey. (B) HeLa cells were incubated 2 h either in control (+leu) or leucine-free medium (−leu) and harvested. ChIP analysis was performed as described under Materials and Methods using antibodies specific for ATF2 and ATF4 and different sets of primers to produce amplicon A, B or C. Data were plotted as the percentage of antibody binding versus the amount of PCR product obtained using a standardized aliquot of input chromatin. Each point represents the mean value of three independent experiments, and the error bars represent the standard error of the means. (C) The experiment described in (B) was also performed using antibodies specific for acetylated H3, acetylated H4 and acetylated H2B.

Mentions: Electrophoresis mobility shift analysis has demonstrated that ATF2 and ATF4 are each capable of binding to the AARE sequence within the CHOP promoter in vitro (4,10). To investigate in vivo binding of these factors, HeLa cells were incubated in control or leucine-free medium for 2 h and ChIP assays were performed with primer sets covering either the 5′ region (amplicon A), the AARE (amplicon B) or the first intron (amplicon C) of the CHOP gene (Figure 2A). The results show an increase in ATF4 binding to the AARE following 2 h of leucine deprivation whereas the binding of ATF2 remained constitutive (Figure 2B). Furthermore, binding of ATF2 and ATF4 was not detected in the 5′ region and in the first intron of CHOP confirming that these factors bind specifically to the AARE.Figure 2.


ATF2 is required for amino acid-regulated transcription by orchestrating specific histone acetylation.

Bruhat A, Chérasse Y, Maurin AC, Breitwieser W, Parry L, Deval C, Jones N, Jousse C, Fafournoux P - Nucleic Acids Res. (2007)

Transcription factor binding and histone acetylation to CHOP AARE in response to leucine starvation. (A) Scheme of the human CHOP gene indicating the different amplicons produced for the ChIP analysis: A (−1678 to −1478), B (−472 to −301) and C (+1163 to +1372). The AARE is boxed in grey. (B) HeLa cells were incubated 2 h either in control (+leu) or leucine-free medium (−leu) and harvested. ChIP analysis was performed as described under Materials and Methods using antibodies specific for ATF2 and ATF4 and different sets of primers to produce amplicon A, B or C. Data were plotted as the percentage of antibody binding versus the amount of PCR product obtained using a standardized aliquot of input chromatin. Each point represents the mean value of three independent experiments, and the error bars represent the standard error of the means. (C) The experiment described in (B) was also performed using antibodies specific for acetylated H3, acetylated H4 and acetylated H2B.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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Figure 2: Transcription factor binding and histone acetylation to CHOP AARE in response to leucine starvation. (A) Scheme of the human CHOP gene indicating the different amplicons produced for the ChIP analysis: A (−1678 to −1478), B (−472 to −301) and C (+1163 to +1372). The AARE is boxed in grey. (B) HeLa cells were incubated 2 h either in control (+leu) or leucine-free medium (−leu) and harvested. ChIP analysis was performed as described under Materials and Methods using antibodies specific for ATF2 and ATF4 and different sets of primers to produce amplicon A, B or C. Data were plotted as the percentage of antibody binding versus the amount of PCR product obtained using a standardized aliquot of input chromatin. Each point represents the mean value of three independent experiments, and the error bars represent the standard error of the means. (C) The experiment described in (B) was also performed using antibodies specific for acetylated H3, acetylated H4 and acetylated H2B.
Mentions: Electrophoresis mobility shift analysis has demonstrated that ATF2 and ATF4 are each capable of binding to the AARE sequence within the CHOP promoter in vitro (4,10). To investigate in vivo binding of these factors, HeLa cells were incubated in control or leucine-free medium for 2 h and ChIP assays were performed with primer sets covering either the 5′ region (amplicon A), the AARE (amplicon B) or the first intron (amplicon C) of the CHOP gene (Figure 2A). The results show an increase in ATF4 binding to the AARE following 2 h of leucine deprivation whereas the binding of ATF2 remained constitutive (Figure 2B). Furthermore, binding of ATF2 and ATF4 was not detected in the 5′ region and in the first intron of CHOP confirming that these factors bind specifically to the AARE.Figure 2.

Bottom Line: Using ATF2-deficient mouse embryonic fibroblasts, we demonstrate that ATF2 is essential in the acetylation of histone H4 and H2B in vivo.The role of ATF2 on histone H4 acetylation is dependent on its binding to the AARE and can be extended to other amino acid regulated genes.Thus, ATF2 is involved in promoting the modification of the chromatin structure to enhance the transcription of a number of amino acid-regulated genes.

View Article: PubMed Central - PubMed

Affiliation: UMR 1019, Unité de Nutrition Humaine, INRA de Theix, 63122 Saint Genès Champanelle, France. bruhat@clermont.inra.fr

ABSTRACT
The transcriptional activation of CHOP (a CCAAT/enhancer-binding protein-related gene) by amino acid deprivation involves the activating transcription factor 2 (ATF2) and the activating transcription factor 4 (ATF4) binding the amino acid response element (AARE) within the promoter. Using a chromatin immunoprecipitation approach, we report that in vivo binding of phospho-ATF2 and ATF4 to CHOP AARE are associated with acetylation of histones H4 and H2B in response to amino acid starvation. A time course analysis reveals that ATF2 phosphorylation precedes histone acetylation, ATF4 binding and the increase in CHOP mRNA. We also show that ATF4 binding and histone acetylation are two independent events that are required for the CHOP induction upon amino acid starvation. Using ATF2-deficient mouse embryonic fibroblasts, we demonstrate that ATF2 is essential in the acetylation of histone H4 and H2B in vivo. The role of ATF2 on histone H4 acetylation is dependent on its binding to the AARE and can be extended to other amino acid regulated genes. Thus, ATF2 is involved in promoting the modification of the chromatin structure to enhance the transcription of a number of amino acid-regulated genes.

Show MeSH