Vinculin is required for caradiac neural crest to mediate outflow tract septation and semilunar valve development in mouse

Abstract

Vinculin (Vcl) is a key adaptor protein at the focal adhesion (FA) mediating various cellular processes and developmental events. In this project, we elucidated how Vcl regulates cardiac neural crest (CNC) development using a NC specific Vcl knockout (VclKO) mouse model. VclKO mutants died within few hours after birth, due to heart failures. VclKO mutants at E18.5 present with ventricular septal defect (VSD), interrupted aortic arches (IAA) and persistent truncus arteriosus (PTA). Formation of semilunar valves (SLV) was also found interrupted, in which the aortic and pulmonary valves of the mutants were much thicker than those of the control and of unequal size. The cardiac defects of the mutants are primarily caused by the failure of CNCs to properly populate the cardiac outflow tract (OFT) and differentiate into vascular smooth cell (VSMC). Delayed migration of CNCs was observed along the cardiac OFT and in the endocardial ridges. VclKO CNCs failed to fully populate the proximal aorta and to give rise to VSMC, resulting in the incomplete septation of aorta and pulmonary artery, leading to IAA and PTA. During the SLV development, CNCs induce the endothelial-tomesenchymal transition to generate enough mesenchymal cells for the expansion of OFT endocardial cushions (EC); regulate the myocardialization to guide myocardial cells to invade ECs; and provide instructive signals to orchestrate apoptosis and extracellular matrix production for the valve remodelling. The reduced number of CNCs in the OFT of the mutants severely interrupted the myocardialization, in which the myocardial cells failed to invade the ECs to support the SLV development. Moreover, the valve remodelling process was found defective in VclKO, such that excessive mesenchymal cells were present, resulting in the thickened valves. Ventricular septum closure relies on the coordination between the ECs of the proximal OFT and the atrioventricular canal (AVC). The defective development of the OFT ECs in VclKO disturbed the movement and remodelling of the AVC ECs, and interrupted the formation of the membranous ventricular septum, leading to VSD. In sum, Vcl is playing the pivotal roles in the migration and differentiation of CNCs during the OFT development, SLV formation and ventricular septum closure.