PMC

Analysis of enrichment of TFBSs. A, Transcription factor binding motifs that are enriched in ERα (α-cell) binding sites; B, transcription factor binding motifs that are enriched in ERα (αβ-cell) binding sites; C, transcription factor binding motifs that are enriched in ERβ (β-cell) binding sites; D, transcription factor binding motifs that are enriched in ERβ (αβ-cell) binding sites.

Venn diagrams showing the overlap of ERα binding sites (PPT treatment) and ERβ binding sites (ERB-041 treatment). A, ERβ binding sites in ERβ-only cells (ERB-041 treatment) and ERα binding sites in ERα-only cells (PPT treatment); B, ERβ and ERα binding sites in cells containing both ERα and ERβ (ERB-041 vs. PPT treatment).

Presence of ERE sequences in ERα or ERβ binding sites with E₂ treatment. Binding sites were probed for the presence of full ERE, half ERE, and no ERE motifs. A, ERα binding sites in ERα-only cells; B, ERβ binding sites in ERβ-only cells; C, ERα binding sites in cells containing both ERα and ERβ; D, ERβ binding sites in cells containing both ERα and ERβ.

Effect of ER subtype partner on ER binding site distribution with E₂ treatment. A, The introduction of ERβ into the cells has a relatively minor effect on the distribution of ERα binding sites. B, ERα has a more pronounced effect on the distribution of ERβ binding sites.

Venn diagrams comparing ER binding site occupancy after cell treatment with the ERα-selective ligand (PPT) or ERβ-selective ligand (ERB-041) vs. E₂. A, ERα binding sites in ERα-only cells (E₂ vs. PPT treatment); B, ERα binding sites in cells containing ERα only or both ERα and ERβ (PPT treatment); C, ERβ binding sites in ERβ-only cells (E₂ vs. ERB-041 treatment); D, ERβ binding sites in cells containing ERβ only or both ERα and ERβ (ERB-041 treatment).

Generation of MCF-7 cells containing different complements of ERα and ERβ for ChIP-chip studies. A, MCF-7 cells were infected with control β-galactosidase-expressing adenovirus or ERβ-expressing adenovirus to generate cells containing ERα-only and ERα plus ERβ, respectively. Cells containing ERβ only were generated by knockdown of ERα by siRNA transfection of cells containing ERα plus ERβ. B, Schematic diagram showing location of tiled probes in the custom-designed tiling arrays. Each probe is 60 bp in length, and probes are tiled approximately 100 bp from each other. RNAi, RNA interference.

Venn diagrams comparing the occupancy of ER binding sites by ERα and ERβ when they are present either separately or together in cells treated with E₂. A, ERα or ERβ can each occupy many of the same sites when the other ER subtype is not present in the cells. B, When both receptors are present, ERα and ERβ share a more limited number of sites. C, Diagram showing the intersection of the intersections from A (sites in common) and B (shared sites) and how sites in common that are not shared are allocated predominantly to ERα.

Correlation between ER binding and transcriptional output in response to E₂. A, Correlation between E₂-regulated genes and binding of ERα-unique (only ERα binds), ERβ-unique (only ERβ binds), or ERα/ERβ sites (sites shared by both ERs) within ±50 kb of the transcription start site of the genes. B, FOS mRNA levels were assessed by quantitative PCR after 4 h treatment of MCF-7 cells differentially expressing ERα and/or ERβ. Data represent average fold change ± sd for three independent experiments. C, ERα and ERβ chromatin binding (by conventional ChIP assays) were measured by quantitative PCR after 45 min E₂ treatment of MCF-7 cells expressing ERα and/or ERβ. ERα and ERβ occupancy of three different ER binding sites (FOS enhancer 1, FOS enhancer 2, and FOS 3′ region) that are closest to the FOS gene are presented graphically. Enh, Enhancer.