CDC42 is required for epicardial and pro-epicardial development by mediating FGF receptor trafficking to the plasma membrane

Abstract

The epicardium contributes to multiple cardiac lineages and is essential for cardiac development and regeneration. However, the mechanism of epicardium formation is unclear. This study aimed to establish the cellular and molecular mechanisms underlying the dissociation of pro-epicardial cells (PECs) from the pro-epicardium (PE) and their subsequent translocation to the heart to form the epicardium. We used lineage tracing, conditional deletion, mosaic analysis and ligand stimulation in mice to determine that both villous protrusions and floating cysts contribute to PEC translocation to myocardium in a CDC42-dependent manner. We resolved a controversy by demonstrating that physical contact of the PE with the myocardium constitutes a third mechanism for PEC translocation to myocardium, and observed a fourth mechanism in which PECs migrate along the surface of the inflow tract to reach the ventricles. Epicardial-specific Cdc42 deletion disrupted epicardium formation, and Cdc42 null PECs proliferated less, lost polarity and failed to form villous protrusions and floating cysts. FGF signaling promotes epicardium formation in vivo, and biochemical studies demonstrated that CDC42 is involved in the trafficking of FGF receptors to the cell membrane to regulate epicardium formation.

SRSF3 is a key regulator of epicardial formation

“…Failure to form the epicardial layer was due to a diminished source of epicardial progenitor cells within the PEO, as a result of their decreased proliferative capacity in the absence of SRSF3. However, recombination in the Tg(Gata5-Cre) model also targeted a subset of cardiomyocytes, thus, combined depletion of SRSF3, in most epicardial progenitor cells and a subset of cardiomyocytes, contributed to the severe phenotype, as embryos with impaired epicardial formation generally die at later stages (von Gise et al 2011; Li et al 2017). Early stage embryonic lethality was previously reported in cardiac-specific SRSF3 knockout mice due to impaired cardiomyocyte proliferation and survival (Ortiz-Sánchez et al 2019).…”

Epicardium in Heart Development

“…In mice, it was first reported that PE cell clusters detach from the proepicardial surface to form cysts, which then float freely across the pericardial cavity to attach to the myocardium around embryonic day 9 (E9.0) (Komiyama et al 1987). Newer studies in mice suggest that both direct contact between the PE and myocardium and adhesion of free-floating PE cell cysts to the myocardium, underlie this translocation process (Rodgers et al 2008;Li et al 2017). Mice bearing a mutation of Par3, a polarity gene, display disrupted basoapical polarity and impaired cyst formation, suggesting that cell polarity is critical for PE cyst formation (Hirose et al 2006).…”

“…Mice bearing a mutation of Par3, a polarity gene, display disrupted basoapical polarity and impaired cyst formation, suggesting that cell polarity is critical for PE cyst formation (Hirose et al 2006). Similarly, an epicardium-specific deletion of Cdc42 also disrupts cell polarity and blocks the formation of PE protrusions and floating epicardial clusters (Li et al 2017). Recent live imaging of PE development and translocation in zebrafish have also found that PE cell release and adhesion occur through both direct contact with the myocardium and with the floating clusters, which are driven by the heartbeat-induced flow of pericardial fluid (Peralta et al 2013;Plavicki et al 2014).…”

“…Apart from these mechanistic models, Li et al discovered an approach in mice in which PE cells grow from the sinus venosus toward the heart along the surface of the inflow tract (Fig. 2; Li et al 2017). Therefore, multiple concurrent mechanisms underlie the process of PE cell translocation.…”