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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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Rapid induction of CHOP mRNA by leucine starvation. (A) HeLa cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested after the indicated incubation times. Total RNA was extracted and real time RT-PCR was performed as described in Materials and Methods. The mRNA induction level is defined as the ratio of the relative mRNA level of leucine-starved cells to that of non-starved cells. (B) Sequence of the CHOP AARE (−313 to −295). The minimum AARE core sequence is boxed in grey. (C) ATF4 +/+ or ATF4 −/− MEF or (D) ATF2 +/+ or ATF2 −/− MEF. In (C) and (D) cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested after the indicated incubation times. Total RNA was extracted and real time RT-PCR was performed as described in Materials and Methods. The mRNA induction level is defined as the ratio of the relative mRNA level of leucine starved cells to that of non-starved cells. (E) ATF4 +/+ or ATF4 −/− MEF (left panel) or ATF2 +/+ or ATF2 −/− MEF (right panel) were incubated either in control (+) or leucine-free medium (−) for 4 h and western blot analysis of ATF2, ATF4 and phospho-ATF2 (Thr-71) was performed from nuclear extracts.
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Figure 1: Rapid induction of CHOP mRNA by leucine starvation. (A) HeLa cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested after the indicated incubation times. Total RNA was extracted and real time RT-PCR was performed as described in Materials and Methods. The mRNA induction level is defined as the ratio of the relative mRNA level of leucine-starved cells to that of non-starved cells. (B) Sequence of the CHOP AARE (−313 to −295). The minimum AARE core sequence is boxed in grey. (C) ATF4 +/+ or ATF4 −/− MEF or (D) ATF2 +/+ or ATF2 −/− MEF. In (C) and (D) cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested after the indicated incubation times. Total RNA was extracted and real time RT-PCR was performed as described in Materials and Methods. The mRNA induction level is defined as the ratio of the relative mRNA level of leucine starved cells to that of non-starved cells. (E) ATF4 +/+ or ATF4 −/− MEF (left panel) or ATF2 +/+ or ATF2 −/− MEF (right panel) were incubated either in control (+) or leucine-free medium (−) for 4 h and western blot analysis of ATF2, ATF4 and phospho-ATF2 (Thr-71) was performed from nuclear extracts.

Mentions: In previous studies, ATF2 and ATF4 were shown to have critical roles for CHOP induction in response to 4–16 h of leucine starvation (4,10). To further dissect the early molecular events involved in the induction of CHOP transcription upon amino acid starvation, we first examined the kinetics of the increase in CHOP mRNA content in response to a short period of leucine starvation in human HeLa cells. CHOP mRNA was increased 2.5-fold after 1 h of leucine starvation and reached a maximum level (9–10-fold) after 4–6 h (Figure 1A).Figure 1.


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)

Rapid induction of CHOP mRNA by leucine starvation. (A) HeLa cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested after the indicated incubation times. Total RNA was extracted and real time RT-PCR was performed as described in Materials and Methods. The mRNA induction level is defined as the ratio of the relative mRNA level of leucine-starved cells to that of non-starved cells. (B) Sequence of the CHOP AARE (−313 to −295). The minimum AARE core sequence is boxed in grey. (C) ATF4 +/+ or ATF4 −/− MEF or (D) ATF2 +/+ or ATF2 −/− MEF. In (C) and (D) cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested after the indicated incubation times. Total RNA was extracted and real time RT-PCR was performed as described in Materials and Methods. The mRNA induction level is defined as the ratio of the relative mRNA level of leucine starved cells to that of non-starved cells. (E) ATF4 +/+ or ATF4 −/− MEF (left panel) or ATF2 +/+ or ATF2 −/− MEF (right panel) were incubated either in control (+) or leucine-free medium (−) for 4 h and western blot analysis of ATF2, ATF4 and phospho-ATF2 (Thr-71) was performed from nuclear extracts.
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Figure 1: Rapid induction of CHOP mRNA by leucine starvation. (A) HeLa cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested after the indicated incubation times. Total RNA was extracted and real time RT-PCR was performed as described in Materials and Methods. The mRNA induction level is defined as the ratio of the relative mRNA level of leucine-starved cells to that of non-starved cells. (B) Sequence of the CHOP AARE (−313 to −295). The minimum AARE core sequence is boxed in grey. (C) ATF4 +/+ or ATF4 −/− MEF or (D) ATF2 +/+ or ATF2 −/− MEF. In (C) and (D) cells were incubated either in control (+leu) or leucine-free medium (−leu) and harvested after the indicated incubation times. Total RNA was extracted and real time RT-PCR was performed as described in Materials and Methods. The mRNA induction level is defined as the ratio of the relative mRNA level of leucine starved cells to that of non-starved cells. (E) ATF4 +/+ or ATF4 −/− MEF (left panel) or ATF2 +/+ or ATF2 −/− MEF (right panel) were incubated either in control (+) or leucine-free medium (−) for 4 h and western blot analysis of ATF2, ATF4 and phospho-ATF2 (Thr-71) was performed from nuclear extracts.
Mentions: In previous studies, ATF2 and ATF4 were shown to have critical roles for CHOP induction in response to 4–16 h of leucine starvation (4,10). To further dissect the early molecular events involved in the induction of CHOP transcription upon amino acid starvation, we first examined the kinetics of the increase in CHOP mRNA content in response to a short period of leucine starvation in human HeLa cells. CHOP mRNA was increased 2.5-fold after 1 h of leucine starvation and reached a maximum level (9–10-fold) after 4–6 h (Figure 1A).Figure 1.

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