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STAT3 acts through pre-existing nucleosome-depleted regions bound by FOS during an epigenetic switch linking inflammation to cancer.

Fleming JD, Giresi PG, Lindahl-Allen M, Krall EB, Lieb JD, Struhl K - Epigenetics Chromatin (2015)

Bottom Line: Interestingly, STAT3 directly regulates the expression of NFKB1, which encodes a subunit of NF-κB, and IL6, a cytokine that stimulates STAT3 activity.Lastly, many STAT3 binding sites are also bound by FOS and the expression of several AP-1 factors is altered during transformation in a STAT3-dependent manner, suggesting that STAT3 may cooperate with AP-1 proteins.These observations uncover additional complexities to the inflammatory feedback loop that are likely to contribute to the epigenetic switch.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115 USA.

ABSTRACT

Background: Transient induction of the Src oncoprotein in a non-transformed breast cell line can initiate an epigenetic switch to a cancer cell via a positive feedback loop that involves activation of the signal transducer and activator of transcription 3 protein (STAT3) and NF-κB transcription factors.

Results: We show that during the transformation process, nucleosome-depleted regions (defined by formaldehyde-assisted isolation of regulatory elements (FAIRE)) are largely unchanged and that both before and during transformation, STAT3 binds almost exclusively to previously open chromatin regions. Roughly, a third of the transformation-inducible genes require STAT3 for the induction. STAT3 and NF-κB appear to drive the regulation of different gene sets during the transformation process. Interestingly, STAT3 directly regulates the expression of NFKB1, which encodes a subunit of NF-κB, and IL6, a cytokine that stimulates STAT3 activity. Lastly, many STAT3 binding sites are also bound by FOS and the expression of several AP-1 factors is altered during transformation in a STAT3-dependent manner, suggesting that STAT3 may cooperate with AP-1 proteins.

Conclusions: These observations uncover additional complexities to the inflammatory feedback loop that are likely to contribute to the epigenetic switch. In addition, gene expression changes during transformation, whether driven by pre-existing or induced transcription factors, occur largely through pre-existing nucleosome-depleted regions.

No MeSH data available.


Related in: MedlinePlus

Open chromatin regions identified by FAIRE before and during transformation. (A) The pie chart represents the approximately 100,000 non-redundant FAIRE sites identified throughout transformation. A minority (17%) changed during transformation and is shown as the rows of the heatmap on the right. The columns of the heatmap display the replicates for FAIRE-seq at each of the indicated time points. For each FAIRE site, the normalized read count was calculated and the data in the heatmap was median centered by rows (yellow = increased and blue = decreased signal). (B) Distribution of FAIRE sites relative to RefSeq gene transcription start site (TSS) annotations. (C) A select set of TF motifs are enriched at FAIRE sites associated with genes whose RNA levels are differentially regulated during transformation. HOMER identified TF binding site motifs enriched at FAIRE sites within ±500 kb of the transcription start site of genes differentially expressed at the indicated time points (rows). Gray indicates an enriched motif (*P < 0.005).
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Fig1: Open chromatin regions identified by FAIRE before and during transformation. (A) The pie chart represents the approximately 100,000 non-redundant FAIRE sites identified throughout transformation. A minority (17%) changed during transformation and is shown as the rows of the heatmap on the right. The columns of the heatmap display the replicates for FAIRE-seq at each of the indicated time points. For each FAIRE site, the normalized read count was calculated and the data in the heatmap was median centered by rows (yellow = increased and blue = decreased signal). (B) Distribution of FAIRE sites relative to RefSeq gene transcription start site (TSS) annotations. (C) A select set of TF motifs are enriched at FAIRE sites associated with genes whose RNA levels are differentially regulated during transformation. HOMER identified TF binding site motifs enriched at FAIRE sites within ±500 kb of the transcription start site of genes differentially expressed at the indicated time points (rows). Gray indicates an enriched motif (*P < 0.005).

Mentions: We used FAIRE-seq [17, 18] to identify nucleosome-depleted regions in MCF-10A-ER-Src cells that were untreated (0-h control) or treated with tamoxifen (TAM) for 4, 12, or 36 h. Across all samples, we identified ~100,000 non-redundant FAIRE regions (Figure 1A) that correspond well to those observed in other breast cell lines and align with known TF binding sites [19] (Additional file 1: Figures S1 and S2). Coupled to this, we used DNA microarrays to identify genes that were differentially expressed during the transformation time course and identified 116 and 1,483 genes at 4 and 24 h post TAM treatment, respectively. Despite the major phenotypic and transcriptional changes observed during transformation, only ~17% of FAIRE regions exhibit an amplitude change in at least one time point during transformation (Figure 1A). Moreover, the majority of these changes are not the result of de novo formation of FAIRE regions, but rather due to an increase in signal at sites that already had low-amplitude FAIRE enrichment (Additional file 1: Figures S3 and S4). Such differential FAIRE regions are observed mainly at the 36-h time point (Figure 1A), yield no significant gene ontology (GO) terms (Methods), and are located mostly at regions greater than 10 kb from the transcription start site (TSS) (Figure 1B).Figure 1


STAT3 acts through pre-existing nucleosome-depleted regions bound by FOS during an epigenetic switch linking inflammation to cancer.

Fleming JD, Giresi PG, Lindahl-Allen M, Krall EB, Lieb JD, Struhl K - Epigenetics Chromatin (2015)

Open chromatin regions identified by FAIRE before and during transformation. (A) The pie chart represents the approximately 100,000 non-redundant FAIRE sites identified throughout transformation. A minority (17%) changed during transformation and is shown as the rows of the heatmap on the right. The columns of the heatmap display the replicates for FAIRE-seq at each of the indicated time points. For each FAIRE site, the normalized read count was calculated and the data in the heatmap was median centered by rows (yellow = increased and blue = decreased signal). (B) Distribution of FAIRE sites relative to RefSeq gene transcription start site (TSS) annotations. (C) A select set of TF motifs are enriched at FAIRE sites associated with genes whose RNA levels are differentially regulated during transformation. HOMER identified TF binding site motifs enriched at FAIRE sites within ±500 kb of the transcription start site of genes differentially expressed at the indicated time points (rows). Gray indicates an enriched motif (*P < 0.005).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4362815&req=5

Fig1: Open chromatin regions identified by FAIRE before and during transformation. (A) The pie chart represents the approximately 100,000 non-redundant FAIRE sites identified throughout transformation. A minority (17%) changed during transformation and is shown as the rows of the heatmap on the right. The columns of the heatmap display the replicates for FAIRE-seq at each of the indicated time points. For each FAIRE site, the normalized read count was calculated and the data in the heatmap was median centered by rows (yellow = increased and blue = decreased signal). (B) Distribution of FAIRE sites relative to RefSeq gene transcription start site (TSS) annotations. (C) A select set of TF motifs are enriched at FAIRE sites associated with genes whose RNA levels are differentially regulated during transformation. HOMER identified TF binding site motifs enriched at FAIRE sites within ±500 kb of the transcription start site of genes differentially expressed at the indicated time points (rows). Gray indicates an enriched motif (*P < 0.005).
Mentions: We used FAIRE-seq [17, 18] to identify nucleosome-depleted regions in MCF-10A-ER-Src cells that were untreated (0-h control) or treated with tamoxifen (TAM) for 4, 12, or 36 h. Across all samples, we identified ~100,000 non-redundant FAIRE regions (Figure 1A) that correspond well to those observed in other breast cell lines and align with known TF binding sites [19] (Additional file 1: Figures S1 and S2). Coupled to this, we used DNA microarrays to identify genes that were differentially expressed during the transformation time course and identified 116 and 1,483 genes at 4 and 24 h post TAM treatment, respectively. Despite the major phenotypic and transcriptional changes observed during transformation, only ~17% of FAIRE regions exhibit an amplitude change in at least one time point during transformation (Figure 1A). Moreover, the majority of these changes are not the result of de novo formation of FAIRE regions, but rather due to an increase in signal at sites that already had low-amplitude FAIRE enrichment (Additional file 1: Figures S3 and S4). Such differential FAIRE regions are observed mainly at the 36-h time point (Figure 1A), yield no significant gene ontology (GO) terms (Methods), and are located mostly at regions greater than 10 kb from the transcription start site (TSS) (Figure 1B).Figure 1

Bottom Line: Interestingly, STAT3 directly regulates the expression of NFKB1, which encodes a subunit of NF-κB, and IL6, a cytokine that stimulates STAT3 activity.Lastly, many STAT3 binding sites are also bound by FOS and the expression of several AP-1 factors is altered during transformation in a STAT3-dependent manner, suggesting that STAT3 may cooperate with AP-1 proteins.These observations uncover additional complexities to the inflammatory feedback loop that are likely to contribute to the epigenetic switch.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115 USA.

ABSTRACT

Background: Transient induction of the Src oncoprotein in a non-transformed breast cell line can initiate an epigenetic switch to a cancer cell via a positive feedback loop that involves activation of the signal transducer and activator of transcription 3 protein (STAT3) and NF-κB transcription factors.

Results: We show that during the transformation process, nucleosome-depleted regions (defined by formaldehyde-assisted isolation of regulatory elements (FAIRE)) are largely unchanged and that both before and during transformation, STAT3 binds almost exclusively to previously open chromatin regions. Roughly, a third of the transformation-inducible genes require STAT3 for the induction. STAT3 and NF-κB appear to drive the regulation of different gene sets during the transformation process. Interestingly, STAT3 directly regulates the expression of NFKB1, which encodes a subunit of NF-κB, and IL6, a cytokine that stimulates STAT3 activity. Lastly, many STAT3 binding sites are also bound by FOS and the expression of several AP-1 factors is altered during transformation in a STAT3-dependent manner, suggesting that STAT3 may cooperate with AP-1 proteins.

Conclusions: These observations uncover additional complexities to the inflammatory feedback loop that are likely to contribute to the epigenetic switch. In addition, gene expression changes during transformation, whether driven by pre-existing or induced transcription factors, occur largely through pre-existing nucleosome-depleted regions.

No MeSH data available.


Related in: MedlinePlus