Notch3 is a genetic modifier of NODAL signalling for patterning asymmetry during mouse heart looping

The TGF{beta} secreted factor NODAL is a major left determinant required for the asymmetric morphogenesis of visceral organs, including the heart. Yet, when this signalling is absent, shape asymmetry, for example of the embryonic heart loop, is not fully abrogated, indicating that there are other factors regulating left-right patterning. Here, we used a tailored transcriptomic approach to screen for genes asymmetrically expressed in the field of heart progenitors. We thus identify Notch3 as a novel left-enriched gene and validate, by quantitative in situ hybridization, its transient asymmetry in the lateral plate mesoderm and node crown, overlapping with Nodal. In mutant embryos, we analysed the regulatory hierarchy and demonstrate that Nodal in the lateral plate mesoderm amplifies Notch3 asymmetric expression. The function of Notch3 was uncovered in an allelic series of mutants. In single neonate mutants, we observe that Notch3 is required with partial penetrance for the development of ventricles, in addition to its known role in coronary arteries. In compound mutants, we reveal that Notch3 acts as a genetic modifier of Nodal, able to modulate heart looping direction and the curvature of the outflow tract. Whereas Notch3 was previously associated with the CADASIL syndrome, its contribution to asymmetric organogenesis is now relevant to severe laterality defects such as the heterotaxy syndrome.