Wnt5a and Wnt11 are essential for second heart field progenitor development.

Wnt/β-catenin has a biphasic effect on cardiogenesis, promoting the induction of cardiac progenitors but later inhibiting their differentiation. Second heart field progenitors and expression of the second heart field transcription factor Islet1 are inhibited by the loss of β-catenin, indicating that Wnt/β-catenin signaling is necessary for second heart field development. However, expressing a constitutively active β-catenin with Islet1-Cre also inhibits endogenous Islet1 expression, reflecting the inhibitory effect of prolonged Wnt/β-catenin signaling on second heart field development. We show that two non-canonical Wnt ligands, Wnt5a and Wnt11, are co-required to regulate second heart field development in mice. Loss of Wnt5a and Wnt11 leads to a dramatic loss of second heart field progenitors in the developing heart. Importantly, this loss of Wnt5a and Wnt11 is accompanied by an increase in Wnt/β-catenin signaling, and ectopic Wnt5a/Wnt11 inhibits β-catenin signaling and promotes cardiac progenitor development in differentiating embryonic stem cells. These data show that Wnt5a and Wnt11 are essential regulators of the response of second heart field progenitors to Wnt/β-catenin signaling and that they act by restraining Wnt/β-catenin signaling during cardiac development.