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Profiling of promoter occupancy by PPARalpha in human hepatoma cells via ChIP-chip analysis.

van der Meer DL, Degenhardt T, Väisänen S, de Groot PJ, Heinäniemi M, de Vries SC, Müller M, Carlberg C, Kersten S - Nucleic Acids Res. (2010)

Bottom Line: We found that GW7647 increased PPARalpha binding to 4220 binding regions.Our data furthermore demonstrate interaction between PPARalpha and STAT transcription factors in PPARalpha-mediated transcriptional repression, and suggest interaction between PPARalpha and TBP, and PPARalpha and C/EBPalpha in PPARalpha-mediated transcriptional activation.Overall, our analysis leads to important new insights into the mechanisms and impact of transcriptional regulation by PPARalpha in human liver and highlight the importance of cross-talk with other transcription factors.

View Article: PubMed Central - PubMed

Affiliation: Nutrition, Metabolism and Genomics group, Division of Human Nutrition, Wageningen University, Bomenweg 2, NL-6703 HD Wageningen, The Netherlands.

ABSTRACT
The transcription factor peroxisome proliferator-activated receptor alpha (PPARalpha) is an important regulator of hepatic lipid metabolism. While PPARalpha is known to activate transcription of numerous genes, no comprehensive picture of PPARalpha binding to endogenous genes has yet been reported. To fill this gap, we performed Chromatin immunoprecipitation (ChIP)-chip in combination with transcriptional profiling on HepG2 human hepatoma cells treated with the PPARalpha agonist GW7647. We found that GW7647 increased PPARalpha binding to 4220 binding regions. GW7647-induced binding regions showed a bias around the transcription start site and most contained a predicted PPAR binding motif. Several genes known to be regulated by PPARalpha, such as ACOX1, SULT2A1, ACADL, CD36, IGFBP1 and G0S2, showed GW7647-induced PPARalpha binding to their promoter. A GW7647-induced PPARalpha-binding region was also assigned to SREBP-targets HMGCS1, HMGCR, FDFT1, SC4MOL, and LPIN1, expression of which was induced by GW7647, suggesting cross-talk between PPARalpha and SREBP signaling. Our data furthermore demonstrate interaction between PPARalpha and STAT transcription factors in PPARalpha-mediated transcriptional repression, and suggest interaction between PPARalpha and TBP, and PPARalpha and C/EBPalpha in PPARalpha-mediated transcriptional activation. Overall, our analysis leads to important new insights into the mechanisms and impact of transcriptional regulation by PPARalpha in human liver and highlight the importance of cross-talk with other transcription factors.

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Overlap between GW7647-induced PPARα binding and GW7647-induced changes in expression. (A) Significant induction of PPARα targets by GW7647 treatment. (B) Number of genes significantly altered upon GW7647 treatment as determined by microarray analysis using criteria: fold change >1.2 and q-value <0.05. (C) Overlap between genes assigned to GW7647-induced PPARα binding regions and genes altered after treatment with GW7647 as determined by transcriptomics. (D) Percentage of GW7647-induced PPARα binding regions linked to either up- or down-regulated genes that contain at least one V$PERO site, as determined using Genomatix. Similar analysis was done for all GW7647-induced PPARα binding regions as well as a control set of promoter regions in the Genomatix promoter database with similar size range as the binding regions identified by ChIP-chip (1000–1500 bp).
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Figure 2: Overlap between GW7647-induced PPARα binding and GW7647-induced changes in expression. (A) Significant induction of PPARα targets by GW7647 treatment. (B) Number of genes significantly altered upon GW7647 treatment as determined by microarray analysis using criteria: fold change >1.2 and q-value <0.05. (C) Overlap between genes assigned to GW7647-induced PPARα binding regions and genes altered after treatment with GW7647 as determined by transcriptomics. (D) Percentage of GW7647-induced PPARα binding regions linked to either up- or down-regulated genes that contain at least one V$PERO site, as determined using Genomatix. Similar analysis was done for all GW7647-induced PPARα binding regions as well as a control set of promoter regions in the Genomatix promoter database with similar size range as the binding regions identified by ChIP-chip (1000–1500 bp).

Mentions: To investigate the relation between genes assigned to a GW7647-induced PPARα binding region and expression of that particular gene, we performed expression microarray analysis on HepG2 cells treated with GW7647 for 6 h. Confirming activation of PPARα, established PPARα targets CYP1A1, ADFP and TRIB3 were significantly induced by GW7647 (Figure 2A). Genes were considered significantly regulated if the mean fold change exceeded 1.2 and q-value <0.05. The low cut-off for fold change was used because the magnitude of induction of PPARα target genes in HepG2 cells is limited. Using these criteria 555 genes were differentially regulated after 6 h GW7647 treatment (Figure 2B). Slightly more genes were up-regulated than down-regulated. Differentially regulated genes were compared with the genes assigned to the GW7647-induced PPARα binding regions. It was found that 54 genes up-regulated by 6 h GW7647 treatment were linked to at least one PPARα binding region, representing 17.8% of all up-regulated genes (Figure 2C and Supplementary Data). In comparison, 16.2% of all genes on the expression array were linked to a PPARα binding region, which indicates that genes up-regulated by GW7647 showed minimal enrichment for PPARα binding. Surprisingly, a PPARα binding region was also linked to 46 genes down-regulated by 6 h treatment with GW7647, representing 18.3% of all down-regulated genes (Figure 2C, Supplementary Data). Expression of the far majority of genes assigned to a GW7647-induced PPARα binding region was not altered by GW7647 treatment.Figure 2.


Profiling of promoter occupancy by PPARalpha in human hepatoma cells via ChIP-chip analysis.

van der Meer DL, Degenhardt T, Väisänen S, de Groot PJ, Heinäniemi M, de Vries SC, Müller M, Carlberg C, Kersten S - Nucleic Acids Res. (2010)

Overlap between GW7647-induced PPARα binding and GW7647-induced changes in expression. (A) Significant induction of PPARα targets by GW7647 treatment. (B) Number of genes significantly altered upon GW7647 treatment as determined by microarray analysis using criteria: fold change >1.2 and q-value <0.05. (C) Overlap between genes assigned to GW7647-induced PPARα binding regions and genes altered after treatment with GW7647 as determined by transcriptomics. (D) Percentage of GW7647-induced PPARα binding regions linked to either up- or down-regulated genes that contain at least one V$PERO site, as determined using Genomatix. Similar analysis was done for all GW7647-induced PPARα binding regions as well as a control set of promoter regions in the Genomatix promoter database with similar size range as the binding regions identified by ChIP-chip (1000–1500 bp).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2875002&req=5

Figure 2: Overlap between GW7647-induced PPARα binding and GW7647-induced changes in expression. (A) Significant induction of PPARα targets by GW7647 treatment. (B) Number of genes significantly altered upon GW7647 treatment as determined by microarray analysis using criteria: fold change >1.2 and q-value <0.05. (C) Overlap between genes assigned to GW7647-induced PPARα binding regions and genes altered after treatment with GW7647 as determined by transcriptomics. (D) Percentage of GW7647-induced PPARα binding regions linked to either up- or down-regulated genes that contain at least one V$PERO site, as determined using Genomatix. Similar analysis was done for all GW7647-induced PPARα binding regions as well as a control set of promoter regions in the Genomatix promoter database with similar size range as the binding regions identified by ChIP-chip (1000–1500 bp).
Mentions: To investigate the relation between genes assigned to a GW7647-induced PPARα binding region and expression of that particular gene, we performed expression microarray analysis on HepG2 cells treated with GW7647 for 6 h. Confirming activation of PPARα, established PPARα targets CYP1A1, ADFP and TRIB3 were significantly induced by GW7647 (Figure 2A). Genes were considered significantly regulated if the mean fold change exceeded 1.2 and q-value <0.05. The low cut-off for fold change was used because the magnitude of induction of PPARα target genes in HepG2 cells is limited. Using these criteria 555 genes were differentially regulated after 6 h GW7647 treatment (Figure 2B). Slightly more genes were up-regulated than down-regulated. Differentially regulated genes were compared with the genes assigned to the GW7647-induced PPARα binding regions. It was found that 54 genes up-regulated by 6 h GW7647 treatment were linked to at least one PPARα binding region, representing 17.8% of all up-regulated genes (Figure 2C and Supplementary Data). In comparison, 16.2% of all genes on the expression array were linked to a PPARα binding region, which indicates that genes up-regulated by GW7647 showed minimal enrichment for PPARα binding. Surprisingly, a PPARα binding region was also linked to 46 genes down-regulated by 6 h treatment with GW7647, representing 18.3% of all down-regulated genes (Figure 2C, Supplementary Data). Expression of the far majority of genes assigned to a GW7647-induced PPARα binding region was not altered by GW7647 treatment.Figure 2.

Bottom Line: We found that GW7647 increased PPARalpha binding to 4220 binding regions.Our data furthermore demonstrate interaction between PPARalpha and STAT transcription factors in PPARalpha-mediated transcriptional repression, and suggest interaction between PPARalpha and TBP, and PPARalpha and C/EBPalpha in PPARalpha-mediated transcriptional activation.Overall, our analysis leads to important new insights into the mechanisms and impact of transcriptional regulation by PPARalpha in human liver and highlight the importance of cross-talk with other transcription factors.

View Article: PubMed Central - PubMed

Affiliation: Nutrition, Metabolism and Genomics group, Division of Human Nutrition, Wageningen University, Bomenweg 2, NL-6703 HD Wageningen, The Netherlands.

ABSTRACT
The transcription factor peroxisome proliferator-activated receptor alpha (PPARalpha) is an important regulator of hepatic lipid metabolism. While PPARalpha is known to activate transcription of numerous genes, no comprehensive picture of PPARalpha binding to endogenous genes has yet been reported. To fill this gap, we performed Chromatin immunoprecipitation (ChIP)-chip in combination with transcriptional profiling on HepG2 human hepatoma cells treated with the PPARalpha agonist GW7647. We found that GW7647 increased PPARalpha binding to 4220 binding regions. GW7647-induced binding regions showed a bias around the transcription start site and most contained a predicted PPAR binding motif. Several genes known to be regulated by PPARalpha, such as ACOX1, SULT2A1, ACADL, CD36, IGFBP1 and G0S2, showed GW7647-induced PPARalpha binding to their promoter. A GW7647-induced PPARalpha-binding region was also assigned to SREBP-targets HMGCS1, HMGCR, FDFT1, SC4MOL, and LPIN1, expression of which was induced by GW7647, suggesting cross-talk between PPARalpha and SREBP signaling. Our data furthermore demonstrate interaction between PPARalpha and STAT transcription factors in PPARalpha-mediated transcriptional repression, and suggest interaction between PPARalpha and TBP, and PPARalpha and C/EBPalpha in PPARalpha-mediated transcriptional activation. Overall, our analysis leads to important new insights into the mechanisms and impact of transcriptional regulation by PPARalpha in human liver and highlight the importance of cross-talk with other transcription factors.

Show MeSH
Related in: MedlinePlus