Hand2 Is an Essential Regulator for Two Notch-Dependent Functions within the Embryonic Endocardium

The basic-helix-loop-helix (bHLH) transcription factor Hand2 plays critical roles during cardiac morphogenesis via expression and function within myocardial, neural crest, and epicardial cell populations. Here, we show that Hand2 plays two essential Notch-dependent roles within the endocardium. Endocardial ablation of Hand2 results in failure to develop a patent tricuspid valve, intraventricular septum defects, and hypotrabeculated ventricles, which collectively resemble the human congenital defect tricuspid atresia. We show endocardial Hand2 to be an integral downstream component of a Notch endocardium-to-myocardium signaling pathway and a direct transcriptional regulator of Neuregulin1. Additionally, Hand2 participates in endocardium-to-endocardium-based cell signaling, with Hand2 mutant hearts displaying an increased density of coronary lumens. Molecular analyses further reveal dysregulation of several crucial components of Vegf signaling, including VegfA, VegfR2, Nrp1, and VegfR3. Thus, Hand2 functions as a crucial downstream transcriptional effector of endocardial Notch signaling during both cardiogenesis and coronary vasculogenesis.We thank Danny Carney and Hannah Lohr for technical assistance and support. We also thank the Riley Heart Research Center Group for discussion and helpful feedback. Furthermore, we thank Thomas Coate for kindly providing EfnB2fx/fx mice. Infrastructural support at the Herman B Wells Center is partially supported by the Riley Children's Foundation and the Carleton Buehl McCulloch Chair. Grant support for this work was provided by NIH grants 1R01HL120920-01, 1R01HL122123-01, and 1R0AR061392-03 (A.B.F.) and American Heart Association predoctoral fellowship 12PRE11700006 (N.J.V.).S