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Identifying synergistic regulation involving c-Myc and sp1 in human tissues.

Parisi F, Wirapati P, Naef F - Nucleic Acids Res. (2007)

Bottom Line: Dual sites show several distinct features compared to the single regulator sites: specifically, they exhibit overall higher degree of conservation between human and rodents, stronger correlation with TFIID-bound promoters, and preference for permissive chromatin state.Namely, the correlation with c-Myc expression in promoters harboring dual-sites is increased for stronger sp1 sites by strong sp1 binding and the effect is largest in proliferating tissues.Our approach shows how integrated functional analyses can uncover tissue-specific and combinatorial regulatory dependencies in mammals.

View Article: PubMed Central - PubMed

Affiliation: Swiss Institute for Experimental Cancer Research (ISREC) and NCCR Molecular Oncology, Lausanne, Switzerland.

ABSTRACT
Combinatorial gene regulation largely contributes to phenotypic versatility in higher eukaryotes. Genome-wide chromatin immuno-precipitation (ChIP) combined with expression profiling can dissect regulatory circuits around transcriptional regulators. Here, we integrate tiling array measurements of DNA-binding sites for c-Myc, sp1, TFIID and modified histones with a tissue expression atlas to establish the functional correspondence between physical binding, promoter activity and transcriptional regulation. For this we develop SLM, a methodology to map c-Myc and sp1-binding sites and then classify sites as sp1-only, c-Myc-only or dual. Dual sites show several distinct features compared to the single regulator sites: specifically, they exhibit overall higher degree of conservation between human and rodents, stronger correlation with TFIID-bound promoters, and preference for permissive chromatin state. By applying regression models to an expression atlas we identified a functionally distinct signature for strong dual c-Myc/sp1 sites. Namely, the correlation with c-Myc expression in promoters harboring dual-sites is increased for stronger sp1 sites by strong sp1 binding and the effect is largest in proliferating tissues. Our approach shows how integrated functional analyses can uncover tissue-specific and combinatorial regulatory dependencies in mammals.

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Related in: MedlinePlus

Summary of expression levels for genes with binding sites across tissues. The , c-Myc-only, sp1-only and dual groups are as in Table 1. They are represented by 31 (c-Myc), 57 (sp1), 57 (dual), 600 () probesets in the tissue atlas. The gene expression matrix is condition centered. (A) In all three lanes (c-Myc and sp1 expression, ChIP group and tissue) the horizontal axis represents the 79 tissue conditions from the SymAtlas tissue atlas (40), each represented for the sp1-only, c-Myc-only and dual groups. Ordering is according to increasing mean expression (from left to right) per group and tissue. In the lane ‘ChIP group’, the dual sites cluster at the right end of the scale, and this correlates with high c-Myc expression (blue track, top). Sp1-only sites have generally lower expression followed by c-Myc-only sites which are interspersed. The ‘tissue’ lane emphasizes that blood samples (20/79 samples, shown in red for the lymphoid and orange for the myeloid lineage) are enriched at the high expression end. (B) Quantification of data in (A). The dots show for each condition the mean expression level in each group. The lines show the correlation between the group means and c-Myc mRNA level. Slopes and adjusted R-squared are reported; all correlation are significant (P < 10−9 for the M group, P < 10−10 for the B group and P < 0.0001 for the  group). The mean expressions in c-Myc-only and dual groups correlate well with c-Myc expression levels. The dual group shows the highest slope (0.13) and clear positive outliers in the blood lineage. The weak correlation between the  group and c-Myc mRNA probably reflects indirect regulation. The low outliers in the c-Myc and dual groups coincide with terminally differentiated tissues, e.g. skin, uterus and tongue.
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Figure 3: Summary of expression levels for genes with binding sites across tissues. The , c-Myc-only, sp1-only and dual groups are as in Table 1. They are represented by 31 (c-Myc), 57 (sp1), 57 (dual), 600 () probesets in the tissue atlas. The gene expression matrix is condition centered. (A) In all three lanes (c-Myc and sp1 expression, ChIP group and tissue) the horizontal axis represents the 79 tissue conditions from the SymAtlas tissue atlas (40), each represented for the sp1-only, c-Myc-only and dual groups. Ordering is according to increasing mean expression (from left to right) per group and tissue. In the lane ‘ChIP group’, the dual sites cluster at the right end of the scale, and this correlates with high c-Myc expression (blue track, top). Sp1-only sites have generally lower expression followed by c-Myc-only sites which are interspersed. The ‘tissue’ lane emphasizes that blood samples (20/79 samples, shown in red for the lymphoid and orange for the myeloid lineage) are enriched at the high expression end. (B) Quantification of data in (A). The dots show for each condition the mean expression level in each group. The lines show the correlation between the group means and c-Myc mRNA level. Slopes and adjusted R-squared are reported; all correlation are significant (P < 10−9 for the M group, P < 10−10 for the B group and P < 0.0001 for the group). The mean expressions in c-Myc-only and dual groups correlate well with c-Myc expression levels. The dual group shows the highest slope (0.13) and clear positive outliers in the blood lineage. The weak correlation between the group and c-Myc mRNA probably reflects indirect regulation. The low outliers in the c-Myc and dual groups coincide with terminally differentiated tissues, e.g. skin, uterus and tongue.

Mentions: We assess the functionality of the identified ChIP sites by considering the expression profiles of all c-Myc and sp1 sites in a tissue expression compendium (40). We are thus implicitly testing whether binding sites measured in Jurkat cells are functional in other cell types. While this is not expected for all regulators, it may hold here. First, there are many lymphoid-related conditions in the gene expression atlas where we expect similarity in the chromatin states. Second, c-Myc and sp1 are basic transcription factors that mediate generic or conserved functions. Comparing the mean expression levels in all three groups and tissues we find that these are highly correlated with c-Myc mRNA level which probably reflects the connection between c-Myc levels and proliferation (Figure 3A). Moreover while the sp1-only sites have the lowest expression, followed by the c-Myc-only sites, the dual sites are generally expressed at highest levels, noticeably in lymphoid lineages which are closest to Jurkat cells (Figure 3A, tissue track). The association between blood lineage, c-Myc expression and high expression of the dual site targets is quantified in Figure 3B. The mean expression levels of genes with c-Myc sites, or those with dual c-Myc and sp1 sites, are significantly correlated with c-Myc mRNA levels across conditions. For sp1 this correlation is not significant (Figure S7). Interestingly, the dual group is correlated with c-Myc expression with a slope that is ∼30% larger that for c-Myc-only sites, indicating that sp1 may contribute synergistically to the induction by c-Myc. As expected the genes without sites show much weaker correlation. It is also apparent from the conditions with lowest c-Myc mRNA expression that the genes with c-Myc sites, either single or accompanied by sp1 sites, have higher baseline expression than genes without sites, or genes with sp1 only sites (Figure S7). Given that it is highly unlikely that c-Myc sites would systematically hit high affinity probes, this presumably reflects that c-Myc sites are frequent in promoters of housekeeping genes that can be induced by multiple other regulators. A few terminally differentiated conditions appear uncorrelated despite intermediate to high c-Myc expression levels.Figure 3.


Identifying synergistic regulation involving c-Myc and sp1 in human tissues.

Parisi F, Wirapati P, Naef F - Nucleic Acids Res. (2007)

Summary of expression levels for genes with binding sites across tissues. The , c-Myc-only, sp1-only and dual groups are as in Table 1. They are represented by 31 (c-Myc), 57 (sp1), 57 (dual), 600 () probesets in the tissue atlas. The gene expression matrix is condition centered. (A) In all three lanes (c-Myc and sp1 expression, ChIP group and tissue) the horizontal axis represents the 79 tissue conditions from the SymAtlas tissue atlas (40), each represented for the sp1-only, c-Myc-only and dual groups. Ordering is according to increasing mean expression (from left to right) per group and tissue. In the lane ‘ChIP group’, the dual sites cluster at the right end of the scale, and this correlates with high c-Myc expression (blue track, top). Sp1-only sites have generally lower expression followed by c-Myc-only sites which are interspersed. The ‘tissue’ lane emphasizes that blood samples (20/79 samples, shown in red for the lymphoid and orange for the myeloid lineage) are enriched at the high expression end. (B) Quantification of data in (A). The dots show for each condition the mean expression level in each group. The lines show the correlation between the group means and c-Myc mRNA level. Slopes and adjusted R-squared are reported; all correlation are significant (P < 10−9 for the M group, P < 10−10 for the B group and P < 0.0001 for the  group). The mean expressions in c-Myc-only and dual groups correlate well with c-Myc expression levels. The dual group shows the highest slope (0.13) and clear positive outliers in the blood lineage. The weak correlation between the  group and c-Myc mRNA probably reflects indirect regulation. The low outliers in the c-Myc and dual groups coincide with terminally differentiated tissues, e.g. skin, uterus and tongue.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC1851645&req=5

Figure 3: Summary of expression levels for genes with binding sites across tissues. The , c-Myc-only, sp1-only and dual groups are as in Table 1. They are represented by 31 (c-Myc), 57 (sp1), 57 (dual), 600 () probesets in the tissue atlas. The gene expression matrix is condition centered. (A) In all three lanes (c-Myc and sp1 expression, ChIP group and tissue) the horizontal axis represents the 79 tissue conditions from the SymAtlas tissue atlas (40), each represented for the sp1-only, c-Myc-only and dual groups. Ordering is according to increasing mean expression (from left to right) per group and tissue. In the lane ‘ChIP group’, the dual sites cluster at the right end of the scale, and this correlates with high c-Myc expression (blue track, top). Sp1-only sites have generally lower expression followed by c-Myc-only sites which are interspersed. The ‘tissue’ lane emphasizes that blood samples (20/79 samples, shown in red for the lymphoid and orange for the myeloid lineage) are enriched at the high expression end. (B) Quantification of data in (A). The dots show for each condition the mean expression level in each group. The lines show the correlation between the group means and c-Myc mRNA level. Slopes and adjusted R-squared are reported; all correlation are significant (P < 10−9 for the M group, P < 10−10 for the B group and P < 0.0001 for the group). The mean expressions in c-Myc-only and dual groups correlate well with c-Myc expression levels. The dual group shows the highest slope (0.13) and clear positive outliers in the blood lineage. The weak correlation between the group and c-Myc mRNA probably reflects indirect regulation. The low outliers in the c-Myc and dual groups coincide with terminally differentiated tissues, e.g. skin, uterus and tongue.
Mentions: We assess the functionality of the identified ChIP sites by considering the expression profiles of all c-Myc and sp1 sites in a tissue expression compendium (40). We are thus implicitly testing whether binding sites measured in Jurkat cells are functional in other cell types. While this is not expected for all regulators, it may hold here. First, there are many lymphoid-related conditions in the gene expression atlas where we expect similarity in the chromatin states. Second, c-Myc and sp1 are basic transcription factors that mediate generic or conserved functions. Comparing the mean expression levels in all three groups and tissues we find that these are highly correlated with c-Myc mRNA level which probably reflects the connection between c-Myc levels and proliferation (Figure 3A). Moreover while the sp1-only sites have the lowest expression, followed by the c-Myc-only sites, the dual sites are generally expressed at highest levels, noticeably in lymphoid lineages which are closest to Jurkat cells (Figure 3A, tissue track). The association between blood lineage, c-Myc expression and high expression of the dual site targets is quantified in Figure 3B. The mean expression levels of genes with c-Myc sites, or those with dual c-Myc and sp1 sites, are significantly correlated with c-Myc mRNA levels across conditions. For sp1 this correlation is not significant (Figure S7). Interestingly, the dual group is correlated with c-Myc expression with a slope that is ∼30% larger that for c-Myc-only sites, indicating that sp1 may contribute synergistically to the induction by c-Myc. As expected the genes without sites show much weaker correlation. It is also apparent from the conditions with lowest c-Myc mRNA expression that the genes with c-Myc sites, either single or accompanied by sp1 sites, have higher baseline expression than genes without sites, or genes with sp1 only sites (Figure S7). Given that it is highly unlikely that c-Myc sites would systematically hit high affinity probes, this presumably reflects that c-Myc sites are frequent in promoters of housekeeping genes that can be induced by multiple other regulators. A few terminally differentiated conditions appear uncorrelated despite intermediate to high c-Myc expression levels.Figure 3.

Bottom Line: Dual sites show several distinct features compared to the single regulator sites: specifically, they exhibit overall higher degree of conservation between human and rodents, stronger correlation with TFIID-bound promoters, and preference for permissive chromatin state.Namely, the correlation with c-Myc expression in promoters harboring dual-sites is increased for stronger sp1 sites by strong sp1 binding and the effect is largest in proliferating tissues.Our approach shows how integrated functional analyses can uncover tissue-specific and combinatorial regulatory dependencies in mammals.

View Article: PubMed Central - PubMed

Affiliation: Swiss Institute for Experimental Cancer Research (ISREC) and NCCR Molecular Oncology, Lausanne, Switzerland.

ABSTRACT
Combinatorial gene regulation largely contributes to phenotypic versatility in higher eukaryotes. Genome-wide chromatin immuno-precipitation (ChIP) combined with expression profiling can dissect regulatory circuits around transcriptional regulators. Here, we integrate tiling array measurements of DNA-binding sites for c-Myc, sp1, TFIID and modified histones with a tissue expression atlas to establish the functional correspondence between physical binding, promoter activity and transcriptional regulation. For this we develop SLM, a methodology to map c-Myc and sp1-binding sites and then classify sites as sp1-only, c-Myc-only or dual. Dual sites show several distinct features compared to the single regulator sites: specifically, they exhibit overall higher degree of conservation between human and rodents, stronger correlation with TFIID-bound promoters, and preference for permissive chromatin state. By applying regression models to an expression atlas we identified a functionally distinct signature for strong dual c-Myc/sp1 sites. Namely, the correlation with c-Myc expression in promoters harboring dual-sites is increased for stronger sp1 sites by strong sp1 binding and the effect is largest in proliferating tissues. Our approach shows how integrated functional analyses can uncover tissue-specific and combinatorial regulatory dependencies in mammals.

Show MeSH
Related in: MedlinePlus