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The chromatin Remodeler CHD8 is required for activation of progesterone receptor-dependent enhancers.

Ceballos-Chávez M, Subtil-Rodríguez A, Giannopoulou EG, Soronellas D, Vázquez-Chávez E, Vicent GP, Elemento O, Beato M, Reyes JC - PLoS Genet. (2015)

Bottom Line: We also show that CHD8 is not required for H3K27 acetylation, but contributes to increase accessibility of the enhancer to DNaseI.Furthermore, CHD8 was required for RNAPII recruiting to the enhancers and for transcription of enhancer-derived RNAs (eRNAs).Taken together our data demonstrate that CHD8 is involved in late stages of PR enhancers activation.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biology Department, Centro Andaluz de Biología Molecular y Medicina Regenerativa (CABIMER), Consejo Superior de Investigaciones Científicas (CSIC), Seville, Spain.

ABSTRACT
While the importance of gene enhancers in transcriptional regulation is well established, the mechanisms and the protein factors that determine enhancers activity have only recently begun to be unravelled. Recent studies have shown that progesterone receptor (PR) binds regions that display typical features of gene enhancers. Here, we show by ChIP-seq experiments that the chromatin remodeler CHD8 mostly binds promoters under proliferation conditions. However, upon progestin stimulation, CHD8 re-localizes to PR enhancers also enriched in p300 and H3K4me1. Consistently, CHD8 depletion severely impairs progestin-dependent gene regulation. CHD8 binding is PR-dependent but independent of the pioneering factor FOXA1. The SWI/SNF chromatin-remodelling complex is required for PR-dependent gene activation. Interestingly, we show that CHD8 interacts with the SWI/SNF complex and that depletion of BRG1 and BRM, the ATPases of SWI/SNF complex, impairs CHD8 recruitment. We also show that CHD8 is not required for H3K27 acetylation, but contributes to increase accessibility of the enhancer to DNaseI. Furthermore, CHD8 was required for RNAPII recruiting to the enhancers and for transcription of enhancer-derived RNAs (eRNAs). Taken together our data demonstrate that CHD8 is involved in late stages of PR enhancers activation.

No MeSH data available.


Related in: MedlinePlus

Hormone-dependent CHD8 recruitment to PR binding sites.(A) Overlapping between CHD8 identified peaks in proliferating cells (blue) or in cells stimulated with R5020 for 5 min (red) or 45 min (green). (B) Distribution of CHD8 peaks in cells stimulated with R5020 for 5 or 45 min. Categories as in Fig 1A. (C) Enrichment of CHD8 binding in response to R5020 or vehicle, upon treatment for 5 min (R5020 5 min, EtOH 5 min, red) or 45 min (R5020 45 min, EtOH 5 min, green) or in proliferating un-induced conditions (proliferation, blue), in four regions containing progesterone-responsive genes: HSD11B2, FKBP5, NFE2L3 and IL6ST. (D) Most significant de novo motif (P-value: 4.7x10-75) identified using ChIPseeqerFIRE and MEME suite [72,73], in the CHD8-binding regions of T47D-MTVL cells stimulated with R5020 for 45 min. (E) Overlapping between progesterone-dependent CHD8 binding sites (green) and PRbs (red) [25] in T47D-MTVL cells stimulated with R5020. (F-J) CHD8 occupancy after 5 (red) or 45 (green) min of R5020 treatment, plotted as the average density of reads counted around the centre of all PRbs (F), around PRbs showing a high (G) or a low (H) nucleosome remodelling index (NRI), around p300 binding sites after R5020 (I) and around PRbs that show H3K4me1 enrichment (J). CHD8 occupancy is expressed as normalized tag density. PRbs, PR binding sites.
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pgen.1005174.g002: Hormone-dependent CHD8 recruitment to PR binding sites.(A) Overlapping between CHD8 identified peaks in proliferating cells (blue) or in cells stimulated with R5020 for 5 min (red) or 45 min (green). (B) Distribution of CHD8 peaks in cells stimulated with R5020 for 5 or 45 min. Categories as in Fig 1A. (C) Enrichment of CHD8 binding in response to R5020 or vehicle, upon treatment for 5 min (R5020 5 min, EtOH 5 min, red) or 45 min (R5020 45 min, EtOH 5 min, green) or in proliferating un-induced conditions (proliferation, blue), in four regions containing progesterone-responsive genes: HSD11B2, FKBP5, NFE2L3 and IL6ST. (D) Most significant de novo motif (P-value: 4.7x10-75) identified using ChIPseeqerFIRE and MEME suite [72,73], in the CHD8-binding regions of T47D-MTVL cells stimulated with R5020 for 45 min. (E) Overlapping between progesterone-dependent CHD8 binding sites (green) and PRbs (red) [25] in T47D-MTVL cells stimulated with R5020. (F-J) CHD8 occupancy after 5 (red) or 45 (green) min of R5020 treatment, plotted as the average density of reads counted around the centre of all PRbs (F), around PRbs showing a high (G) or a low (H) nucleosome remodelling index (NRI), around p300 binding sites after R5020 (I) and around PRbs that show H3K4me1 enrichment (J). CHD8 occupancy is expressed as normalized tag density. PRbs, PR binding sites.

Mentions: CHD8 has been suggested to be a nuclear receptor co-activator [18,19]; however, very little is known about this function of the protein. To gain insight into its role in hormone dependent transcriptional regulation we have analyzed by ChIP-seq the distribution of CHD8 in progesterone-treated T47D-MTVL cells. For that, cells were subjected to 48 h of serum deprivation and then stimulated during 5 or 45 minutes with the synthetic progestin R5020 (10 nM) or the vehicle (ethanol) as control. A very small number of peaks were found in vehicle treated cells, suggesting that serum deprivation strongly decreases the association of CHD8 to the chromatin. These data extend our previous observation about absence of CHD8 in four G1/S transition genes in quiescent cells [17]. However, 1132 and 4532 progestin-induced CHD8 peaks were found at 5 and 45 minutes, respectively, suggesting that CHD8 is quickly recruited to the chromatin after progestin treatment. Most of the sites found after 5 minutes (73%) were also identified after 45 minutes (Fig 2A). Only 18% (832) of the hormone-specific peaks were also found under proliferating conditions (Fig 2A). A large majority of the hormone specific sites were found at intronic and intergenic regions (Fig 2B) and low enrichment was found at TSS (S3 Fig). A representative example of the ChIP-seq data close to four well-known progesterone-dependent genes (HSD11B2, FKBP5, NFE2L3 and IL6ST) is shown in Fig 2C.


The chromatin Remodeler CHD8 is required for activation of progesterone receptor-dependent enhancers.

Ceballos-Chávez M, Subtil-Rodríguez A, Giannopoulou EG, Soronellas D, Vázquez-Chávez E, Vicent GP, Elemento O, Beato M, Reyes JC - PLoS Genet. (2015)

Hormone-dependent CHD8 recruitment to PR binding sites.(A) Overlapping between CHD8 identified peaks in proliferating cells (blue) or in cells stimulated with R5020 for 5 min (red) or 45 min (green). (B) Distribution of CHD8 peaks in cells stimulated with R5020 for 5 or 45 min. Categories as in Fig 1A. (C) Enrichment of CHD8 binding in response to R5020 or vehicle, upon treatment for 5 min (R5020 5 min, EtOH 5 min, red) or 45 min (R5020 45 min, EtOH 5 min, green) or in proliferating un-induced conditions (proliferation, blue), in four regions containing progesterone-responsive genes: HSD11B2, FKBP5, NFE2L3 and IL6ST. (D) Most significant de novo motif (P-value: 4.7x10-75) identified using ChIPseeqerFIRE and MEME suite [72,73], in the CHD8-binding regions of T47D-MTVL cells stimulated with R5020 for 45 min. (E) Overlapping between progesterone-dependent CHD8 binding sites (green) and PRbs (red) [25] in T47D-MTVL cells stimulated with R5020. (F-J) CHD8 occupancy after 5 (red) or 45 (green) min of R5020 treatment, plotted as the average density of reads counted around the centre of all PRbs (F), around PRbs showing a high (G) or a low (H) nucleosome remodelling index (NRI), around p300 binding sites after R5020 (I) and around PRbs that show H3K4me1 enrichment (J). CHD8 occupancy is expressed as normalized tag density. PRbs, PR binding sites.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4403880&req=5

pgen.1005174.g002: Hormone-dependent CHD8 recruitment to PR binding sites.(A) Overlapping between CHD8 identified peaks in proliferating cells (blue) or in cells stimulated with R5020 for 5 min (red) or 45 min (green). (B) Distribution of CHD8 peaks in cells stimulated with R5020 for 5 or 45 min. Categories as in Fig 1A. (C) Enrichment of CHD8 binding in response to R5020 or vehicle, upon treatment for 5 min (R5020 5 min, EtOH 5 min, red) or 45 min (R5020 45 min, EtOH 5 min, green) or in proliferating un-induced conditions (proliferation, blue), in four regions containing progesterone-responsive genes: HSD11B2, FKBP5, NFE2L3 and IL6ST. (D) Most significant de novo motif (P-value: 4.7x10-75) identified using ChIPseeqerFIRE and MEME suite [72,73], in the CHD8-binding regions of T47D-MTVL cells stimulated with R5020 for 45 min. (E) Overlapping between progesterone-dependent CHD8 binding sites (green) and PRbs (red) [25] in T47D-MTVL cells stimulated with R5020. (F-J) CHD8 occupancy after 5 (red) or 45 (green) min of R5020 treatment, plotted as the average density of reads counted around the centre of all PRbs (F), around PRbs showing a high (G) or a low (H) nucleosome remodelling index (NRI), around p300 binding sites after R5020 (I) and around PRbs that show H3K4me1 enrichment (J). CHD8 occupancy is expressed as normalized tag density. PRbs, PR binding sites.
Mentions: CHD8 has been suggested to be a nuclear receptor co-activator [18,19]; however, very little is known about this function of the protein. To gain insight into its role in hormone dependent transcriptional regulation we have analyzed by ChIP-seq the distribution of CHD8 in progesterone-treated T47D-MTVL cells. For that, cells were subjected to 48 h of serum deprivation and then stimulated during 5 or 45 minutes with the synthetic progestin R5020 (10 nM) or the vehicle (ethanol) as control. A very small number of peaks were found in vehicle treated cells, suggesting that serum deprivation strongly decreases the association of CHD8 to the chromatin. These data extend our previous observation about absence of CHD8 in four G1/S transition genes in quiescent cells [17]. However, 1132 and 4532 progestin-induced CHD8 peaks were found at 5 and 45 minutes, respectively, suggesting that CHD8 is quickly recruited to the chromatin after progestin treatment. Most of the sites found after 5 minutes (73%) were also identified after 45 minutes (Fig 2A). Only 18% (832) of the hormone-specific peaks were also found under proliferating conditions (Fig 2A). A large majority of the hormone specific sites were found at intronic and intergenic regions (Fig 2B) and low enrichment was found at TSS (S3 Fig). A representative example of the ChIP-seq data close to four well-known progesterone-dependent genes (HSD11B2, FKBP5, NFE2L3 and IL6ST) is shown in Fig 2C.

Bottom Line: We also show that CHD8 is not required for H3K27 acetylation, but contributes to increase accessibility of the enhancer to DNaseI.Furthermore, CHD8 was required for RNAPII recruiting to the enhancers and for transcription of enhancer-derived RNAs (eRNAs).Taken together our data demonstrate that CHD8 is involved in late stages of PR enhancers activation.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biology Department, Centro Andaluz de Biología Molecular y Medicina Regenerativa (CABIMER), Consejo Superior de Investigaciones Científicas (CSIC), Seville, Spain.

ABSTRACT
While the importance of gene enhancers in transcriptional regulation is well established, the mechanisms and the protein factors that determine enhancers activity have only recently begun to be unravelled. Recent studies have shown that progesterone receptor (PR) binds regions that display typical features of gene enhancers. Here, we show by ChIP-seq experiments that the chromatin remodeler CHD8 mostly binds promoters under proliferation conditions. However, upon progestin stimulation, CHD8 re-localizes to PR enhancers also enriched in p300 and H3K4me1. Consistently, CHD8 depletion severely impairs progestin-dependent gene regulation. CHD8 binding is PR-dependent but independent of the pioneering factor FOXA1. The SWI/SNF chromatin-remodelling complex is required for PR-dependent gene activation. Interestingly, we show that CHD8 interacts with the SWI/SNF complex and that depletion of BRG1 and BRM, the ATPases of SWI/SNF complex, impairs CHD8 recruitment. We also show that CHD8 is not required for H3K27 acetylation, but contributes to increase accessibility of the enhancer to DNaseI. Furthermore, CHD8 was required for RNAPII recruiting to the enhancers and for transcription of enhancer-derived RNAs (eRNAs). Taken together our data demonstrate that CHD8 is involved in late stages of PR enhancers activation.

No MeSH data available.


Related in: MedlinePlus