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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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Time course of ATF2 and ATF4 binding and histone acetylation during leucine starvation. (A) HeLa cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested for 0–4 h. ChIP analysis was performed as described under Materials and Methods using antibodies specific for (A) ATF4, ATF2, phospho-ATF2 (Thr-71) and (B) acetylated H3, acetylated H4 and acetylated H2B and a set of primers to amplify amplicon B (see Figure 2A). 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. The dotted line represents the increase in CHOP mRNA induction level as shown in Figure 1A.
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Figure 3: Time course of ATF2 and ATF4 binding and histone acetylation during leucine starvation. (A) HeLa cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested for 0–4 h. ChIP analysis was performed as described under Materials and Methods using antibodies specific for (A) ATF4, ATF2, phospho-ATF2 (Thr-71) and (B) acetylated H3, acetylated H4 and acetylated H2B and a set of primers to amplify amplicon B (see Figure 2A). 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. The dotted line represents the increase in CHOP mRNA induction level as shown in Figure 1A.

Mentions: We next investigated the kinetics of ATF2 and ATF4 binding to the CHOP AARE. ATF4 levels increased after 1 h of leucine deprivation (Figure 3A) and rapidly reached a maximum within 2 h. By contrast, ATF2 binding to the CHOP AARE remained constitutive throughout the 4 h period investigated. ChIP analysis using an antibody that specifically recognize ATF2 phosphorylated on threonine 71 revealed that the phosphorylation of ATF2 was detectable 30 min after removal of leucine from the medium and reached a maximum level of phosphorylation within 2 h.Figure 3.


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)

Time course of ATF2 and ATF4 binding and histone acetylation during leucine starvation. (A) HeLa cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested for 0–4 h. ChIP analysis was performed as described under Materials and Methods using antibodies specific for (A) ATF4, ATF2, phospho-ATF2 (Thr-71) and (B) acetylated H3, acetylated H4 and acetylated H2B and a set of primers to amplify amplicon B (see Figure 2A). 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. The dotted line represents the increase in CHOP mRNA induction level as shown in Figure 1A.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1851658&req=5

Figure 3: Time course of ATF2 and ATF4 binding and histone acetylation during leucine starvation. (A) HeLa cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested for 0–4 h. ChIP analysis was performed as described under Materials and Methods using antibodies specific for (A) ATF4, ATF2, phospho-ATF2 (Thr-71) and (B) acetylated H3, acetylated H4 and acetylated H2B and a set of primers to amplify amplicon B (see Figure 2A). 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. The dotted line represents the increase in CHOP mRNA induction level as shown in Figure 1A.
Mentions: We next investigated the kinetics of ATF2 and ATF4 binding to the CHOP AARE. ATF4 levels increased after 1 h of leucine deprivation (Figure 3A) and rapidly reached a maximum within 2 h. By contrast, ATF2 binding to the CHOP AARE remained constitutive throughout the 4 h period investigated. ChIP analysis using an antibody that specifically recognize ATF2 phosphorylated on threonine 71 revealed that the phosphorylation of ATF2 was detectable 30 min after removal of leucine from the medium and reached a maximum level of phosphorylation within 2 h.Figure 3.

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