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. Author manuscript; available in PMC: 2021 Jun 14.
Published in final edited form as: Cell Rep. 2021 May 18;35(7):109145. doi: 10.1016/j.celrep.2021.109145

Figure 5. Exogenous FOXA2 expression does not restore definitive endoderm chromatin accessibility in GATA6−/− cells.

Figure 5.

(A) Doxycycline-inducible FOXA2-3× FLAG cDNA vector.

(B) FOXA2 overexpression differentiation model.

(C) Immunofluorescence analysis of GATA6, FOXA2, and DAPI in GATA6+/+ and GATA6Ex4Δ1/Δ2;ind FOXA2 cells with and without doxycycline at day 4 of differentiation. Scale bars, 100 μm. Positive cell population: flow cytometry, n = 3, mean ± SD.

(D) Graph showing ATAC-seq fragment depth GATA6+/+ and GATA6Ex4Δ1/Δ2;ind FOXA2 cells with increasing doxycycline supplementation at sites of slow-onset GATA6-dependent chromatin remodeling in definitive endoderm; n = 2,404 regions.

(E) Venn diagram of loci bound by FOXA2 in GATA6+/+ and GATA6Ex4Δ1/Δ2;ind FOXA2 (++dox) cells at day 4 of differentiation with heatmaps of FOXA2 occupancy for each identified subset.

(F) Transcription factor motifs enriched within each subset of FOXA2 bound loci. p values: hypergeometric motif enrichment over the alternate subsets of FOXA2-bound loci.

(G) Heatmaps depicting GATA6 and FOXA2 ChIP-seq intensity at sites of GATA6-dependent chromatin remodeling in GATA6+/+ and GATA6Ex4Δ1/Δ2;ind FOXA2 (++dox) in mesendoderm and definitive endoderm populations.

(H) GATA6 and FOXA2 ChIP-seq data aligned with ATAC-seq profiles during definitive endoderm formation at the RASSF10 gene loci in GATA6+/+ and GATA6Ex4Δ1/Δ2;ind FOXA2 cells (++dox); GATA6−/− refers to GATA6Ex4Δ1/Δ2;ind FOXA2 cells.