Journal: bioRxiv
Article Title: Genome Organizer SATB1 selectively activates a defined subset of EMT genes driving metastatic breast cancer
doi: 10.64898/2026.05.29.728584
Figure Lengend Snippet: Analysis of SATB1-regulated genes. A) DAVID functional annotation analysis of 585 SATB1-upregulated genes (fc ≥2, p adj ≤0.05) in MCF10A-1 compared with control non SATB1-expressing cells determined by RNA-seq. X-axis [-log (Benjamini)] indicates the statistical significance of each gene group. B) Gene Set Enrichment Analysis (GSEA) using RNA-seq genes ranked by log2 fold change and tested gene sets defined by the top and bottom 2,000 GRO-seq–SATB1-regulated genes. GRO-seq upregulated genes are enriched among RNA-seq upregulated genes (top panel), while GRO-seq downregulated genes are enriched among RNA-seq downregulated genes (bottom panel), demonstrating strong directional concordance between nascent transcriptional regulation and steady-state RNA expression. Enrichment scores (ES) were calculated using the Broad Institute preranked GSEA algorithm (weight p = 1) with 10,000 permutations, and normalized enrichment scores (NES) were derived using same-sign null distributions. Nominal p-values and false discovery rates (FDR) were computed according to Broad Institute methodology. Positive NES indicates enrichment toward the top of the RNA-seq ranked list, whereas negative NES indicates enrichment toward the bottom. Gene-set hit positions are indicated by thin vertical tick marks in the separate “Hits” panel below.
Article Snippet: We used the MCF10A-1 cell line described in Ordinario et al .This cell line has been derived from MCF10A cells, originally obtained from the American Type Culture Collection (ATCC), by passaging more than 50 times in culture medium containing fetal bovine serum.
Techniques: Functional Assay, Control, Expressing, RNA Sequencing, RNA Expression, Derivative Assay
ATCC is a verified supplier 
ATCC manufactures this product 
![Analysis of SATB1-regulated genes. A) DAVID functional annotation analysis of 585 SATB1-upregulated genes (fc ≥2, p adj ≤0.05) in <t>MCF10A-1</t> compared with control non SATB1-expressing cells determined by RNA-seq. X-axis [-log (Benjamini)] indicates the statistical significance of each gene group. B) Gene Set Enrichment Analysis (GSEA) using RNA-seq genes ranked by log2 fold change and tested gene sets defined by the top and bottom 2,000 GRO-seq–SATB1-regulated genes. GRO-seq upregulated genes are enriched among RNA-seq upregulated genes (top panel), while GRO-seq downregulated genes are enriched among RNA-seq downregulated genes (bottom panel), demonstrating strong directional concordance between nascent transcriptional regulation and steady-state RNA expression. Enrichment scores (ES) were calculated using the Broad Institute preranked GSEA algorithm (weight p = 1) with 10,000 permutations, and normalized enrichment scores (NES) were derived using same-sign null distributions. Nominal p-values and false discovery rates (FDR) were computed according to Broad Institute methodology. Positive NES indicates enrichment toward the top of the RNA-seq ranked list, whereas negative NES indicates enrichment toward the bottom. Gene-set hit positions are indicated by thin vertical tick marks in the separate “Hits” panel below.](https://bio-rxiv-images-cdn.bioz.com/dois_ending_with_84/10__64898_slash_2026__05__29__728584/10__64898_slash_2026__05__29__728584___F1.large.jpg)
![Analysis of SATB1-regulated EMT-promoting genes . A) GSEA was performed independently using genes ranked by log2 fold change (log2FC) from RNA-seq and GRO-seq datasets. A curated gene set comprising 300 EMT-promoting genes was tested for enrichment. Enrichment scores (ES), normalized enrichment scores (NES), nominal p-values and false discovery rates (FDR) are indicated. Gene-set hit positions are indicated by thin vertical tick marks in a separate “Hits” panel below. B) DAVID gene enrichment analysis of 98 SATB1-upregulated EMT-promoting genes in SATB1- transduced MCF10A-1 compared with control cells (fc > 1.2, p < 0.07) determined by GRO-seq. X-axis [-log (Benjamini)] indicates the statistical significance of each gene group.](https://bio-rxiv-images-cdn.bioz.com/dois_ending_with_84/10__64898_slash_2026__05__29__728584/10__64898_slash_2026__05__29__728584___F2.large.jpg)
![DAVID gene enrichment analysis of B[a]P-upregulated genes compared with SATB1-upregulated genes identified by GRO-seq in SATB1-stably transduced MCF10A-1 cells. A) Gene enrichment within each DAVID functional group is shown for B[a]P-upregulated EMT-promoting genes after 3 days of treatment (72 genes, FC3; blue bars) and SATB1-upregulated EMT-promoting genes (98 genes, FC1; black bars). B) Gene enrichment is shown for B[a]P-upregulated EMT-promoting genes after 3 days of treatment (72 genes, FC3; blue bars) and after 3 hrs of treatment (54 genes; red bars), demonstrating a time-dependent increase in enrichment of B[a]P-upregulated EMT-promoting genes within each gene group. Whereas SATB1-upregulated genes are highly enriched in cellular components related to extracellular matrix (ECM) structure and function (top six gene groups), these gene groups remain weakly enriched among B[a]P-upregulated genes after 3 days of B[a]P exposure. The x-axis [−log(Benjamini)] indicates the statistical significance of each gene group.](https://bio-rxiv-images-cdn.bioz.com/dois_ending_with_84/10__64898_slash_2026__05__29__728584/10__64898_slash_2026__05__29__728584___F5.large.jpg)
![Time-dependent reduction in the effects of B[a]P on EMT gene activation in the presence of SATB1, as detected by GRO-seq. A) Genes upregulated by B[a]P after 3 hrs in SATB1-expressing MCF10A-1 cells (selected using fc > 1.2 and p < 0.07, and arranged in descending order centered on FC4; green bars) were compared with genes upregulated by B[a]P after 3 hrs in control MCF10A-1 cells (without selection; FC2, red bars). Fold changes in transcript levels are shown for each gene. B) Genes upregulated by B[a]P after 3 days in SATB1-expressing MCF10A-1 cells (selected using fc > 1.2 and p < 0.07, and arranged in descending order centered on FC5; pink bars) were compared with genes upregulated by B[a]P after 3 days in control MCF10A-1 cells (without selection; FC3, blue bars). C) Genes upregulated by B[a]P after 3 days in control MCF10A-1 cells (selected using fc > 1.2 and p < 0.07, and arranged in descending order centered on FC3; blue bars) were compared with genes upregulated by B[a]P after 3 days in SATB1-expressing MCF10A-1 cells (without selection; FC5, red bars). This analysis is similar to panel B, except that transcript levels for FC3 genes were centered instead of FC5 genes. Fold changes in transcript levels are shown for each gene.](https://bio-rxiv-images-cdn.bioz.com/dois_ending_with_84/10__64898_slash_2026__05__29__728584/10__64898_slash_2026__05__29__728584___F6.large.jpg)