Congenital heart defects affect over 1% of all live births and represent the most common birth defect. Much of the complexity of heart development is underpinned by the interaction of multiple different cell types (second heart field, neural crest and endothelial cells) to orchestrate formation of the fully functional heart.  Nedd4^-/- mice display heart outflow tract defects reminiscent of those observed in children, including double outlet right ventricle (DORV) and persistent truncus arteriosis (PTA). Given we have previously demonstrated a role for the ubiquitin ligase Nedd4 in cranial neural crest cell development (Wiszniak, 2013, Dev. Biol. 15:186), we hypothesised that Nedd4 may also play a role in cardiac neural crest cells, underpinning the outflow tract defects observed.

Removal of Nedd4 specifically in neural crest cells (Wnt1-Cre; Nedd4^fl/fl embryos) resulted in outflow tract defects reminiscent of the complete knockout, suggesting a specific role for Nedd4 in neural crest cells. Interestingly, genetic lineage tracing of neural crest cells with GFP in Wnt1-Cre; Nedd4^fl/flmutants revealed no deficiency of neural crest cells within the outflow tract.  However, cardiac precursors of the second heart field, which sit adjacent to the migrating cardiac neural crest cells, exhibited premature cardiomyocyte differentiation, leading to a reduced progenitor pool able to contribute to the correct growth and lengthening of the outflow tract. This suggests cardiac neural crest cells signal to the second heart field to maintain these cells in a progenitor state.  Inadequate outflow tract lengthening lead to incomplete rotation of the outflow tract and subsequent incorrect artery-ventricle alignment.

Current work is focussed on dissecting the interaction between neural crest cells and the second heart field. We have identified potential ubiquitinated target proteins of Nedd4 using a SILAC-labelling mass-spectroscopy approach, as well as assessed altered transcriptional programs in the second heart field using RNA sequencing. Using these complementary approaches, we have identified several candidate signalling pathways that neural crest cells use to instruct second heart field development.