Journal: Nucleic Acids Research

Article Title: PPARG-centric transcriptional re-wiring during differentiation of human trophoblast stem cells into extravillous trophoblasts

doi: 10.1093/nar/gkaf669

Figure Lengend Snippet: PPARG-centric transcriptional networks with cell-type-specific TFs and cofactors. ( A ) Heatmap showing the ChIP-seq signals of PPARG, RXRA, TSC TFs (MSX2 and GATA2), EP300, and H3K27ac, along with ATAC-seq signals at PPARG-bound TSC-specific, common, and EVT-specific loci in TSCs. ( B ) Heatmap showing the ChIP-seq signals of PPARG, RXRA, EVT TFs (DLX5 and DLX6), EP300, and H3K27ac, along with ATAC-seq signals at PPARG-bound TSC-specific, common, and EVT-specific loci in EVTs. ( C ) Model: in TSCs, PPARG associates with RXRA and TSC-specific TFs to maintain self-renewal independently of LBD-mediated transcriptional activity. In contrast, during EVT differentiation, PPARG cooperates with RXRA, LBD-associated cofactors, and EVT-specific TFs to activate gene expression via LBD-mediated transcriptional regulation. ( D ) Heatmap showing the log2-normalized gene expression upon T007 treatment on EVT day 3 (left). Genes were ordered according to their relative expression levels in T007-treated cells versus control. The PPARG occupancy signals on EVT day 3 for these ordered genes were plotted as the moving window average (window size, 100; bin size, 1) (right). ( E ) Histograms showing ChIP-seq read enrichment for PPARG, RXRA, EP300, NCOA3, MED1, MED12, MED15, and H3K27ac in control cells (blue) and T007-treated cells (red) on EVT day 3, around selected PPARG summits associated with genes significantly down-regulated by T007. Data are shown as mean normalized read counts from two biological replicates. Statistical comparisons were performed using two-tailed Student's t -tests; *, **, and *** indicate P -value < 0.05, 0.01, and 0.001, respectively; ns, not significant.

Article Snippet: The antibodies used for chromatin immunoprecipitation sequencing (ChIP-seq) were PPARG (Cell Signaling Technology, 2443S, 10 μl), RXRA (Cell Signaling Technology, 3085S, 10 μl), EP300 (Abcam, ab10485, 10 μl), NCOA3 (Cell Signaling Technology, 2126S, 10 μl), MED1 (Novus Biologicals, NB100-2574, 10 μl), MED12 (Bethyl Laboratories, A300-774A, 10 μl), MED15 (Proteintech, 11566-1-AP, 10 μl), and H3K27ac (Cell Signaling Technology, 8173S, 10 μl).

Techniques: ChIP-sequencing, Activity Assay, Gene Expression, Expressing, Control, Two Tailed Test