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Fig. 7 A gnai3-linked TREE directs injury-induced epicardial gene expression. (A) Browser tracks of the genomic region near gnai3 showing the transcripts and chromatin accessibility profiles in the epicardium, the zCNE sites and the whole-ventricle H3K27Ac profile. Gray boxes, red arrows and numbers indicate candidate TREEs. (B) Larval expression of three gnai3-E1:EGFP reporter lines. Scale bar: 500 µm. (C) Whole-mount image of a 6 dpf heart showing no GFP expression. tcf21:H2A-mCherry (red) labels the epicardial cells. V, ventricle. Scale bar: 20 µm. (D) In situ hybridization results showing gnai3 expression in presumed epicardial cells (arrows) at 3 dpa. Dashed line indicates the injury site. Scale bar: 100 µm. (E) Whole-mount images (maximum projections) of the ventricular surface showing gnai3-E1:EGFP expression in tcf21:H2A-mCherry+ cells. An enlarged view of the outlined region is displayed at the bottom with single-channel image shown in grayscale (single optical sections). White dashed line indicates the injury sites. Arrows indicate representative EGFP+mCherry+ cells. Scale bar: 100 µm. (F) Section images demonstrating expression of gnai3-E1:EGFP (green, anti-EGFP antibody staining) in tcf21:H2A-mCherry+ (magenta) cells. Arrows indicate representative double-positive cells. Scale bar: 50 µm. (G) HCR staining of gnai3 (magenta dots) on ventricular sections (ctrl, 3 dpa) showing gnai3-E1:EGFP expression (green, anti-EGFP antibody staining) in gnai3+ cells. The outlined region is enlarged on the right. Arrows indicate an EGFP+gnai3+ cell. Scale bar: 25 µm.