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Genome-wide landscape of liver X receptor chromatin binding and gene regulation in human macrophages.

Pehkonen P, Welter-Stahl L, Diwo J, Ryynänen J, Wienecke-Baldacchino A, Heikkinen S, Treuter E, Steffensen KR, Carlberg C - BMC Genomics (2012)

Bottom Line: The liver X receptors (LXRs) are oxysterol sensing nuclear receptors with multiple effects on metabolism and immune cells.Our results show that LXR actions are focused on 112 genomic regions that contain up to 11 T09 target genes per region under the control of highly stringent LXR binding sites with individual constellations for each region.We could confirm that LXR controls lipid metabolism and transport and observed a strong association with apoptosis-related functions.

View Article: PubMed Central - HTML - PubMed

Affiliation: School of Medicine, Institute of Biomedicine, University of Eastern Finland, FIN-70210 Kuopio, Finland.

ABSTRACT

Background: The liver X receptors (LXRs) are oxysterol sensing nuclear receptors with multiple effects on metabolism and immune cells. However, the complete genome-wide cistrome of LXR in cells of human origin has not yet been provided.

Results: We performed ChIP-seq in phorbol myristate acetate-differentiated THP-1 cells (macrophage-type) after stimulation with the potent synthetic LXR ligand T0901317 (T09). Microarray gene expression analysis was performed in the same cellular model. We identified 1357 genome-wide LXR locations (FDR < 1%), of which 526 were observed after T09 treatment. De novo analysis of LXR binding sequences identified a DR4-type element as the major motif. On mRNA level T09 up-regulated 1258 genes and repressed 455 genes. Our results show that LXR actions are focused on 112 genomic regions that contain up to 11 T09 target genes per region under the control of highly stringent LXR binding sites with individual constellations for each region. We could confirm that LXR controls lipid metabolism and transport and observed a strong association with apoptosis-related functions.

Conclusions: This first report on genome-wide binding of LXR in a human cell line provides new insights into the transcriptional network of LXR and its target genes with their link to physiological processes, such as apoptosis.The gene expression microarray and sequence data have been submitted collectively to the NCBI Gene Expression Omnibus http://www.ncbi.nlm.nih.gov/geo under accession number GSE28319.

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LXR target genes with DR4-type REs. A Analysis of LXR peak versus T09 target gene pairs by relating the log2 FC of target genes with the similarity of the DR4-type REs within the closest LXR peak (± 100 kb limit). For each peak, the gene with the smallest expression P-value in that region is represented. Similarity is indicated as -log10 of the P-value obtained from the RSAT motif screening tool [26]. Pie diagrams on the top show the proportions of differentially expressed gene-peak pairs among the all gene-peak pairs within the similarity intervals 3-4.5, 4.5-6, 6-7.5, 7.5-9 and > 9. In the DR4-type RE consensus sequence R = A or G, M = A or C, and N any nucleotide. B Example region of the T09 target gene SMPDL3A containing two DR4-type REs close to its TSS. Under each sample track the arrows indicate LXR binding locations of high stringency (red) or FDR < 5% (grey). Extra lanes indicate the location of LXR in homologous sequences of the mouse genome as previously published [18], de novo LXR binding sites within the peak area and insulator barrier regions identified via CTCF binding sites observed in CD4+ T cells [32].
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Figure 5: LXR target genes with DR4-type REs. A Analysis of LXR peak versus T09 target gene pairs by relating the log2 FC of target genes with the similarity of the DR4-type REs within the closest LXR peak (± 100 kb limit). For each peak, the gene with the smallest expression P-value in that region is represented. Similarity is indicated as -log10 of the P-value obtained from the RSAT motif screening tool [26]. Pie diagrams on the top show the proportions of differentially expressed gene-peak pairs among the all gene-peak pairs within the similarity intervals 3-4.5, 4.5-6, 6-7.5, 7.5-9 and > 9. In the DR4-type RE consensus sequence R = A or G, M = A or C, and N any nucleotide. B Example region of the T09 target gene SMPDL3A containing two DR4-type REs close to its TSS. Under each sample track the arrows indicate LXR binding locations of high stringency (red) or FDR < 5% (grey). Extra lanes indicate the location of LXR in homologous sequences of the mouse genome as previously published [18], de novo LXR binding sites within the peak area and insulator barrier regions identified via CTCF binding sites observed in CD4+ T cells [32].

Mentions: Next we compared altered expression of LXR target genes to the occurrence of DR4-type REs within the proximal LXR peaks (Figure 4A). Although DR4-type REs with very high similarity scores are not very common (Figure 1B), they seem to be enriched to the peaks in the vicinity of DE genes, especially when located nearby up-regulated genes (Figure 5A). This is consistent with the principal activation mechanism of LXR. The six up-regulated genes ABCA1, ABCG1, SMPDL3A (sphingomyelin phosphodiesterase, acid-like 3A), NR1H3, SCD (stearoyl-CoA desaturase) and TATDN2 (TatD DNase domain containing 2) and the three down-regulated genes CNNM4 (cyclin M4), HARS (histidyl-tRNA synthetase) and PUF60 (poly-U binding splicing factor 60 KDa) have LXR binding location with a motif highly similar (P < 10-6) to a DR4-type RE. The ABCA1 gene is one of the best-known LXR target genes and contains even three DR4-type REs within its regulatory region (Figure 1C). Another example, the SMPDL3A gene, carries two DR4-type REs within an LXR twin peak very close to its TSS (Figure 5B). Using quantitative real-time PCR (qPCR) and RNA from independently performed stimulation experiments of PMA-differentiated THP-1 cells with the synthetic LXR ligands T09 and GW3965 (GW), we validated nine representative LXR targets genes: the known targets ABCA1, ABCG1, MYLIP (myosin regulatory light chain interact), NR1H3 and SCD and the novel targets PPARG, SMPDL3A, ADM (adrenomedullin) and ACSL3 (acyl-CoA synthetase long-chain family member 3) (see additional file 8: Figure S5). After 4 h of ligand treatment all nine genes were significantly up-regulated by both LXR ligands.


Genome-wide landscape of liver X receptor chromatin binding and gene regulation in human macrophages.

Pehkonen P, Welter-Stahl L, Diwo J, Ryynänen J, Wienecke-Baldacchino A, Heikkinen S, Treuter E, Steffensen KR, Carlberg C - BMC Genomics (2012)

LXR target genes with DR4-type REs. A Analysis of LXR peak versus T09 target gene pairs by relating the log2 FC of target genes with the similarity of the DR4-type REs within the closest LXR peak (± 100 kb limit). For each peak, the gene with the smallest expression P-value in that region is represented. Similarity is indicated as -log10 of the P-value obtained from the RSAT motif screening tool [26]. Pie diagrams on the top show the proportions of differentially expressed gene-peak pairs among the all gene-peak pairs within the similarity intervals 3-4.5, 4.5-6, 6-7.5, 7.5-9 and > 9. In the DR4-type RE consensus sequence R = A or G, M = A or C, and N any nucleotide. B Example region of the T09 target gene SMPDL3A containing two DR4-type REs close to its TSS. Under each sample track the arrows indicate LXR binding locations of high stringency (red) or FDR < 5% (grey). Extra lanes indicate the location of LXR in homologous sequences of the mouse genome as previously published [18], de novo LXR binding sites within the peak area and insulator barrier regions identified via CTCF binding sites observed in CD4+ T cells [32].
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 5: LXR target genes with DR4-type REs. A Analysis of LXR peak versus T09 target gene pairs by relating the log2 FC of target genes with the similarity of the DR4-type REs within the closest LXR peak (± 100 kb limit). For each peak, the gene with the smallest expression P-value in that region is represented. Similarity is indicated as -log10 of the P-value obtained from the RSAT motif screening tool [26]. Pie diagrams on the top show the proportions of differentially expressed gene-peak pairs among the all gene-peak pairs within the similarity intervals 3-4.5, 4.5-6, 6-7.5, 7.5-9 and > 9. In the DR4-type RE consensus sequence R = A or G, M = A or C, and N any nucleotide. B Example region of the T09 target gene SMPDL3A containing two DR4-type REs close to its TSS. Under each sample track the arrows indicate LXR binding locations of high stringency (red) or FDR < 5% (grey). Extra lanes indicate the location of LXR in homologous sequences of the mouse genome as previously published [18], de novo LXR binding sites within the peak area and insulator barrier regions identified via CTCF binding sites observed in CD4+ T cells [32].
Mentions: Next we compared altered expression of LXR target genes to the occurrence of DR4-type REs within the proximal LXR peaks (Figure 4A). Although DR4-type REs with very high similarity scores are not very common (Figure 1B), they seem to be enriched to the peaks in the vicinity of DE genes, especially when located nearby up-regulated genes (Figure 5A). This is consistent with the principal activation mechanism of LXR. The six up-regulated genes ABCA1, ABCG1, SMPDL3A (sphingomyelin phosphodiesterase, acid-like 3A), NR1H3, SCD (stearoyl-CoA desaturase) and TATDN2 (TatD DNase domain containing 2) and the three down-regulated genes CNNM4 (cyclin M4), HARS (histidyl-tRNA synthetase) and PUF60 (poly-U binding splicing factor 60 KDa) have LXR binding location with a motif highly similar (P < 10-6) to a DR4-type RE. The ABCA1 gene is one of the best-known LXR target genes and contains even three DR4-type REs within its regulatory region (Figure 1C). Another example, the SMPDL3A gene, carries two DR4-type REs within an LXR twin peak very close to its TSS (Figure 5B). Using quantitative real-time PCR (qPCR) and RNA from independently performed stimulation experiments of PMA-differentiated THP-1 cells with the synthetic LXR ligands T09 and GW3965 (GW), we validated nine representative LXR targets genes: the known targets ABCA1, ABCG1, MYLIP (myosin regulatory light chain interact), NR1H3 and SCD and the novel targets PPARG, SMPDL3A, ADM (adrenomedullin) and ACSL3 (acyl-CoA synthetase long-chain family member 3) (see additional file 8: Figure S5). After 4 h of ligand treatment all nine genes were significantly up-regulated by both LXR ligands.

Bottom Line: The liver X receptors (LXRs) are oxysterol sensing nuclear receptors with multiple effects on metabolism and immune cells.Our results show that LXR actions are focused on 112 genomic regions that contain up to 11 T09 target genes per region under the control of highly stringent LXR binding sites with individual constellations for each region.We could confirm that LXR controls lipid metabolism and transport and observed a strong association with apoptosis-related functions.

View Article: PubMed Central - HTML - PubMed

Affiliation: School of Medicine, Institute of Biomedicine, University of Eastern Finland, FIN-70210 Kuopio, Finland.

ABSTRACT

Background: The liver X receptors (LXRs) are oxysterol sensing nuclear receptors with multiple effects on metabolism and immune cells. However, the complete genome-wide cistrome of LXR in cells of human origin has not yet been provided.

Results: We performed ChIP-seq in phorbol myristate acetate-differentiated THP-1 cells (macrophage-type) after stimulation with the potent synthetic LXR ligand T0901317 (T09). Microarray gene expression analysis was performed in the same cellular model. We identified 1357 genome-wide LXR locations (FDR < 1%), of which 526 were observed after T09 treatment. De novo analysis of LXR binding sequences identified a DR4-type element as the major motif. On mRNA level T09 up-regulated 1258 genes and repressed 455 genes. Our results show that LXR actions are focused on 112 genomic regions that contain up to 11 T09 target genes per region under the control of highly stringent LXR binding sites with individual constellations for each region. We could confirm that LXR controls lipid metabolism and transport and observed a strong association with apoptosis-related functions.

Conclusions: This first report on genome-wide binding of LXR in a human cell line provides new insights into the transcriptional network of LXR and its target genes with their link to physiological processes, such as apoptosis.The gene expression microarray and sequence data have been submitted collectively to the NCBI Gene Expression Omnibus http://www.ncbi.nlm.nih.gov/geo under accession number GSE28319.

Show MeSH