How to be a Heartbreaker: Podosome-Associate Proteins Fgd1, Src, and Tks5 are Necessary for Proper Heart Morphogenesis

Abstract

Congenital heart defects, which are diagnosed in roughly 1% of children and have an increasing rate of diagnosis in both adults and children, can be caused by aberrant cell migration. Two of the major events in D. rerio heart morphogenesis are the left-right asymmetric processes of cardiac jogging and looping, both of which are driven by cell migration. Nodal signaling drives asymmetric cell migration in the jogging heart, but the mechanisms underlying this are unknown. Our lab preliminarily identified Fgd1 as a Nodal pathway target in the zebrafish heart. Fgd1 is known to be involved in actin reorganization and cell migration, but its contribution to cardiac cell migration has not been explored. These proteins are also implicated in the formation of podosomes; actin-rich structures that our lab observes to form during cardiac jogging. Podosomes have not been well characterized, but they are hypothesized to play a role in cell migration due to their up-regulation in metastatic cell lines. We also investigated the role of Tks5 in heart morphogenesis, which is a scaffolding protein that has been demonstrated to be necessary for podosome formation in vitro. Src, a non-receptor tyrosine kinase, is responsible for activating both of these proteins, and has been implicated in the formation of podosomes. We used morpholinos to knockdown Fgd1 and Tks5, which resulted in defects in heart laterality and delayed cardiac jogging, as well as abnormal heart morphology, respectively. We confirmed the morpholino results by generating mutations in Fgd1 and Tks5 with CRISPR/Cas9, which resulted in embryos with similar phenotypes to the morpholino treated embryos. These results indicate that both proteins play essential roles in heart morphogenesis, suggesting that podosomes may play an important role in the process of heart development.