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. Author manuscript; available in PMC: 2022 Jan 1.
Published in final edited form as: Nature. 2021 Jun 23;595(7867):438–443. doi: 10.1038/s41586-021-03674-1

Extended Data Figure 11: MEOX1 is a regulator of fibroblast activation.

Extended Data Figure 11:

a,b. Meox1 expression measured by qPCR in mouse primary cardiac fibroblasts (a) and immortalized cardiac fibroblasts (b) in Unstim condition, TGFβ siCtrl and TGFβ siMeox1. One-way ANOVA followed by Tukey post hoc test. c. Immunofluorescence staining of αSMA in Unstim, and TGFβ cells with a control or a Meox1-targeting siRNA. Nuclei are marked by Hoechst; scale bar =100μm. Quantification of αSMA staining is shown to the right (two independent experiments). Fold change intensity is normalized to cell number. One-way ANOVA followed by Tukey post hoc test. d. αSMA expression measured by qPCR in Unstim condition, TGFβ siCtrl and TGFβ siMeox1. One-way ANOVA followed by Tukey post hoc test. e. Representative images of Edu incorporation in Unstim condition, TGFβ siCtrl and TGFβ siMeox1. DAPI (blue), Edu (red) and CellMask (green). Quantification (two independent experiments) on the right. One-way ANOVA followed by Tukey post hoc test. f. Meox1 expression measured by qPCR in WT and Meox1 over-expression (o/e) mouse immortalized cardiac fibroblasts. Unpaired t-test (Two-tailed). g. Pearson correlation of the three replicates of MEOX1 anti-HA ChIPseq in Unstim and TGFβ-treated cells. h. MEOX1 anti-HA ChIPseq coverage in all protein coding genes (± 2kb gene body) in Unstim and TGFβ-treated fibroblasts. i. Pearson correlation of the two independent biological replicates of PROseq in TGFβ siCtrl and TGFβ siMeox1. j. PROseq coverage in Unstim, TGFβ siCtrl and TGFβ siMeox1 at the distal elements defined as more transcribed in TGFβ vs. Unstim (2101 sites, see Fig. 2a) that are either bound by MEOX1 (496 regions – top panel) or not (1,605 regions – bottom panel). k. PROseq coverage in Unstim, TGFβ siCtrl and TGFβ siMeox1 at the distal elements with high H3K27ac enrichment in Unstim bound by MEOX1 (379 regions). l. PROseq coverage of differentially transcribed genes (Wald test followed by Benjamini/Hochberg) in TGFβ-treated fibroblasts with Ctrl or Meox1 siRNA and associated top GO terms. Signal for replicate 1 & 2 is shown. m. PROseq tag density (± 5kb gene body) in TGFβ siCtrl and TGFβ siMeox1 in genes upregulated in TGFβ siCtrl vs. TGFβ siMeox1 (left); and genes upregulated in TGFβ siMeox1 vs. TGFβ siCtrl (right). n. Violin plot showing normalized expression score of genes upregulated in TGFβ siCtrl vs. TGFβ siMeox1 in PROseq that were captured in the scRNAseq. Expression of Sham and TAC fibroblast samples is depicted. o. Number of MEOX1-bound genes in MEOX1 ChIPseq (in TGFβ-treated cells) in ±2kb gene body, ±100kb gene body or ±500kb gene body in genes differentially transcribed in PROseq: upregulated in TGFβ vs. Unstim (left); downregulated in TGFβ vs. Unstim (center left); upregulated in TGFβ siCtrl vs. TGFβ siMeox1 (center right); upregulated in TGFβ siMeox1 vs. TGFβ siCtrl (right). p. Coverage of MEOX1 ChIP (TGFβ-treated cells), H3K27ac ChIPseq (Unstim and TGFβ-treated cells) and PROseq (Unstim condition, TGFβ siCtrl and TGFβ siMeox1) at the Postn locus. The Postn Peak10/11 region is highlighted in red.

a-f, Numbers above bar graphs show significant p-val. Data are shown as means ± SEM.