Gata4 drives Hh-signaling for second heart field migration and outflow tract development

Abstract

32Dominant mutations of Gata4, an essential cardiogenic transcription factor (TF), cause 33 outflow tract (OFT) defects in both human and mouse. We investigated the molecular 34 mechanism underlying this requirement. Gata4 happloinsufficiency in mice caused OFT 35 defects including double outlet right ventricle (DORV) and conal ventricular septum 36 defects (VSDs). We found that Gata4 is required within Hedgehog (Hh)-receiving second 37 heart field (SHF) progenitors for normal OFT alignment. Increased Pten-mediated cell-38 cycle transition, rescued atrial septal defects but not OFT defects in Gata4 heterozygotes. 39 SHF Hh-receiving cells failed to migrate properly into the proximal OFT cushion in Gata4 40 heterozygote embryos. We find that Hh signaling and Gata4 genetically interact for OFT 41 development. Gata4 and Smo double heterozygotes displayed more severe OFT 42 abnormalities including persistent truncus arteriosus (PTA) whereas restoration of 43 Hedgehog signaling rescued OFT defects in Gata4-mutant mice. In addition, enhanced 44 expression of the Gata6 was observed in the SHF of the Gata4 heterozygotes. These 45 results suggested a SHF regulatory network comprising of Gata4, Gata6 and Hh-signaling 46 for OFT development. This study indicates that Gata4 potentiation of Hh signaling is a 47 general feature of Gata4-mediated cardiac morphogenesis and provides a model for the 48 molecular basis of CHD caused by dominant transcription factor mutations.49 3 50Gata4 is an important protein that controls the development of the heart. Human who 52 possess a single copy of Gata4 mutation display congenital heart defects (CHD), 53 including the double outlet right ventricle (DORV). DORV is an alignment problem in 54 which both the Aorta and Pulmonary Artery originate from the right ventricle, instead of 55 originating from the left and the right ventricles, respectively. To study how Gata4 56 mutation causes DORV, we used a Gata4 mutant mouse model, which displays DORV. 57We showed that Gata4 is required in the cardiac precursor cells for the normal alignment 58 of the great arteries. Although Gata4 mutation inhibits the rapid increase in number of the 59 cardiac precursor cells, rescuing this defects does not recover the normal alignment of 60 the great arteries. In addition, there is a movement problem of the cardiac precursor cells 61 when migrating toward the great arteries during development. We further showed that a 62 specific molecular signaling, Hh-signaling, is responsible to the Gata4 action in the 63 cardiac precursor cells. Importantly, over-activating the Hh-signaling rescues the DORV 64 in the Gata4 mutant embryos. This study provides an explanation for the ontogeny of 65 CHD. 66 4 67 Introduction 68 Congenital Heart Defects (CHDs) CHDs occurr in approximately 1% of live births [1] 69 and are the most common serious birth defects in humans [2, 3]. Approximately one third 70 of the CHDs involve malformations of the outflow tract (OFT), which leads to significant 71 morbidity and mortality o...