The ezrin Gene Regulates Early Cardiac Morphogenesis and Contractile Function in Zebrafish Through the Coordinated Regulation of Apoptosis, Calcium Homeostasis, and the MAPK Signaling Pathway

Ezrin, expressed by the EZR gene, is a member of the ERM protein family that connects the plasma membrane to the actin cytoskeleton, participating in processes such as cell adhesion, migration, and signaling. However, its role in cardiac morphogenesis remains incompletely understood. In zebrafish (Danio rerio), two ezrin homologs, ezra and ezrb, are present. CRISPR/Cas9 gene editing technology was used to generate ezra knockout lines, and the simultaneous knockdown of ezra and ezrb was induced via morpholino oligonucleotides (MOs). To investigate the molecular mechanisms, transcriptome sequencing and bioinformatic analysis were conducted on 48 h post-fertilization (hpf) ezrin–MO embryos, with subsequent validation using a real-time quantitative polymerase chain reaction (RT-qPCR) and whole-mount in situ hybridization (WISH) experiment. The results showed that ezra−/− exhibited a compensatory upregulation of ezrb without overt developmental defects, whereas ezrin–MO embryos presented with pericardial edema, reduced cardiac chamber size, and atrioventricular valve malformations at 48 hpf. RNA-seq revealed that myocardial contraction-related genes were significantly dysregulated and apoptotic signaling pathways were activated in ezrin–MO embryos. These findings demonstrate that ezra and ezrb are functionally redundant in cardiac development and that the loss of ezrin function may lead to cardiac developmental defects and impaired myocardial contractility via the activation of apoptotic signaling pathways.