Figure 4
- ID
- ZDB-FIG-210414-46
- Publication
- Pronobis et al., 2021 - In vivo proximity labeling identifies cardiomyocyte protein networks during zebrafish heart regeneration
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(A) Timeline of experiment for inhibiting Rho A function in 14 dpa hearts. (B) Immunofluorescence images of sections of 14 dpa ventricles vehicle or Rhosin treated and stained for EdU incorporation, an indicator of cardiomyocyte proliferation. Mef2 staining marks cardiomyocyte nuclei. Dashed line, approximate resection plane. Arrowheads, Mef2+/EdU+ cardiomyocytes. (C) Quantification of Mef2/EdU assays. Inhibition of Rho A by Rhosin reduces cardiomyocyte (CM) proliferation. n = 5 animals for each condition, two independent experiments. Data show mean ± SEM. (Mann–Whitney U test). (D) Heat map of indicated proteins from the whole-cell BioID2 data set. Of these, only levels of ROCK2a changed consistently (5.1-fold; p<0.05) during regeneration. (E) Over-representation test of reactome pathways. Signaling and effectors of Rho GTPases were found to be over-represented. Fold enrichment as indicated, and protein number in red. p<0.001, FDR < 0.02%. (F) Myosin heavy chain (MHC) (green) staining ventricles from animals with induced myocardial-specific dominant-negative Rac, Rho, or Cdc42, along with vehicle-treated ventricles, at 30 dpa. Five to eight animals were assessed in each group treated with vehicle, with none of these animals displaying regeneration defects. Four of five ventricles with induced dominant-negative Rac1, and all ventricles with induced dominant-negative Rho A (n = 7/7), or Cdc42 (n = 9/9), showed obvious areas of missing myocardium. Fisher–Irwin exact test, p<0.05. Dashed line, approximate resection plane. Scale bars, 50 µm. (G) Quantification of cardiomyocyte proliferation by Mef2/Proliferating cell nuclear antigen (PCNA) staining of n = 4 animals for each condition. Data show mean ± SEM. (Mann–Whitney U test). |