Fig. 6

Fig. 6 Single-cell RNA-sequencing reveals border zone-specific gene expression signatures in the regenerating heart.

A UMAP plot of scRNA-seq from 4596 cells isolated from dissected border zone regions of regenerating zebrafish hearts at 7 dpci. The microdissected region of 7 dpci ventricles used for scRNA-seq is depicted in the upper left. Created in BioRender. Beisaw, A. (2025) https://BioRender.com/zek72tx. B Gene ontology (GO) analysis of biological processes (BP) in differentially expressed genes enriched in BZ CMs compared to rCMs. log₁₀padj was calculated in comparison to all genes in the zebrafish genome using Fisher’s exact test and corrected for multiple comparisons using the Bonferroni method. log_size corresponds to the log₁₀(number of annotations for the GO Term ID in zebrafish from the EBI GOA Database). C Differentially expressed genes (padj < 0.05) arising from pseudo-bulk comparisons of gene expression in rCM versus BZ CMs (left) and mac2 versus mac3 (ECM, right) clusters. P-values were calculated using unpaired two-sided t-tests and corrected for multiple comparisons using the Benjamini-Hochberg method. D Violin plots of expression of genes involved in actin cytoskeleton and CM regulation in rCMs vs. BZ CMs clusters from scRNA-seq at 7 dpci. E Schematic illustrating the experimental set-up in Tg(myl7:Cre-ERT2); Tg(ubb:loxP-EGFP-loxP-AFos-P2A-tagBFP) (CM:AFos, top). Control CMs (n = 3 independent pools of 6 ventricles each) and CM:Afos CMs (n = 3 independent pools of 6 ventricles each) were isolated at 7 dpci and RT-qPCR analysis of actin cytoskeletal and ECM regulators enriched in BZ CMs was performed. Data are presented as mean ± SD. P-values were calculated using unpaired two-sided t-tests or a two-sided Mann–Whitney test (pfn1). Source data are presented in the Source Data file. Created in BioRender. Beisaw, A. (2025) https://BioRender.com/qqhzxg5. F Immunostaining of F-actin and Vinculin in control and CM:A-Fos ventricles at 7 dpci. Yellow arrowheads denote protruding CMs with Vinculin enrichment at the leading edge of protrusions. G Violin plots of expression of genes involved in ECM composition/regulation in mac3 (ECM) vs. mac2 clusters from scRNA-seq at 7 dpci. Scale bars: 20 μm.