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. Author manuscript; available in PMC: 2022 May 19.
Published in final edited form as: Cell Rep. 2019 Nov 26;29(9):2659–2671.e6. doi: 10.1016/j.celrep.2019.10.074

Figure 5. Gc Depletion Recapitulates MLL1i Function in EPSCs.

Figure 5.

(A) Expression of 131 MLL1 direct targets upon MLL1i (x axis) and Mll1KO (y axis). These genes also had concomitant reduction of H3K4me1 (not shown). Representative genes are highlighted. Gray line, nonlinear correlation of RNA-seq signals in MLL1i and Mll1−/− cells.

(B) GO term enrichment for downregulated genes upon MLL1i and Mll1KO.

(C) qRT-PCR validation for Gc expression as indicated. Average Gc expression from biological triplicates is presented after normalization against its expression in Mll1f/f cells, which was arbitrarily set as 1. ***p < 0.001, Mann-Whitney test (unpaired, nonparametric, two tailed).

((D) Fold enrichment of H3K4me1 and H3K4me2 at Gc by ChIP-qPCR.

(E) qRT-PCR for Vdr expression in cells as indicated. Average Vdr expression from triplicate experiments is presented after normalization against its expression in Mll1f/f cells, which was arbitrarily set as 1. For (C)–(E), error bars are SD from biological triplicates. ***p < 0.001, Mann-Whitney test (unpaired, nonparametric, two tailed).

(F) Left, principal-component analysis (PCA) plot for expression of MLL1 targets in different cells. Right, clustering analysis for gene expression (same input as PCA) in cells indicated on the left.

(G) Induction of spontaneous 2C+ cells in ESC culture upon siGc. Two small interfering RNAs (siRNAs), as well as their equal molar mixture, were used to deplete Gc. Average fold increase relative to mock-treated ESCs is presented. Error bars are SD from three independent experiments. For mock, error bar is SD from nine independent experiments.

(H) Immunofluorescence for CDX2 in chimeric blastocyst-derived siGc or siControl GFP+ Nagy R1 ESCs. Scale bars, 50 µm. * indicates GFP+/CDX2+ cells.

See also Figure S7 and Table S3.