Deletion of ETS-1, a gene in the Jacobsen syndrome critical region, causes ventricular septal defects and abnormal ventricular morphology in mice

Abstract

Congenital heart defects comprise the most common form of major birth defects, affecting 0.7% of all newborn infants. Jacobsen syndrome (11q-) is a rare chromosomal disorder caused by deletions in distal 11q. We have previously determined that a wide spectrum of the most common congenital heart defects occur in 11q-, including an unprecedented high frequency of hypoplastic left heart syndrome (HLHS). We identified an approximately 7 Mb 'cardiac critical region' in distal 11q that contains a putative causative gene(s) for congenital heart disease. In this study, we utilized chromosomal microarray mapping to characterize three patients with 11q- and congenital heart defects that carry interstitial deletions overlapping the 7 Mb cardiac critical region. We propose that this 1.2 Mb region of overlap harbors a gene(s) that causes at least a subset of the congenital heart defects that occur in 11q-. We demonstrate that one gene in this region, ETS-1 (a member of the ETS family of transcription factors), is expressed in the endocardium and neural crest during early mouse heart development. Gene-targeted deletion of ETS-1 in mice in a C57/B6 background causes, with high penetrance, large membranous ventricular septal defects and a bifid cardiac apex, and less frequently a non-apex-forming left ventricle (one of the hallmarks of HLHS). Our results implicate an important role for the ETS-1 transcription factor in mammalian heart development and should provide important insights into some of the most common forms of congenital heart disease.

Figure 1.

Refined critical region in distal 11q. The new region is defined by the region of overlap between the previously defined ∼7 Mb terminal deletion (JS22), and that of the three patients with congenital heart defects with interstitial deletions in distal 11q.

Figure 2.

Early expression of ETS-1 in the developing mouse heart: ED8 in situ hybridization showing whole mount (A and B), coronal section and staining of endothelium using PECAM (CD31) antibody (C and D). ED8.75 in situ hybridization showing whole mount (E and F), coronal section (G), PECAM staining (H) and negative control for in situ using an ETS-1 sense probe (I and J). NF, neural fold; HT, heart tube; EC, endocardial cell; NT, neural tube; H, heart; BA1, first branchial arch; RA, right side of common atrium; LA, left side of common atrium; V, ventricle.

Figure 3.

Expression of ETS-1 in the heart in ED9.5 embryos: in situ hybridizations are shown in (A) (whole mount), and in sections: (B) (anterior coronal section) and (C) (posterior coronal section). Immunohistochemistry indicating endothelial expression using a PECAM (CD31) antibody is shown in (D) (whole mount), and in sections: (E) (anterior coronal) and (F) (posterior coronal). LacZ staining of neural crest using a Wnt1-Cre; ROSA26 LacZ indicator strain is shown in (G) (whole mount), (H) (anterior coronal) and (I) (posterior coronal). Immunofluorescence studies on cultured neural crest cells derived from ED8.5 neural crest explanted tissue: ETS-1 antibody (J); Sox10 antibody (K), DAPI staining (L); Merge of ETS-1 and Sox10 (M); Merge of ETS-1, Sox10 and DAPI (N). RA, right side of common atrium; LA, left side of common atrium; AV, atrioventricular canal; RV, right side of common ventricle; LV, left side of common ventricle; OFT, outflow tract; NCC, neural crest cell; EC, endothelial cell.

Figure 4.

In situ hybridization showing expression of ETS-1 in the region of the developing membranous interventricular septum in an ED13.5 heart (A). Higher magnification of myocardial trabeculations from an ED13.5 heart showing expression of ETS-1 in the endocardium: in situ hybridization (B) and immunofluorescence using an antibody to PECAM (C). RA, right atrium; LA, left atrium; RV, right ventricle; LV, left ventricle; VS, interventricular septum. ETS-1 expression is also detected in the endocardium of the semilunar and atrioventricular valves by ED13.5 (data not shown).

Figure 5.

Large membranous ventricular septal defect in ED16.5 ETS−/− embryos, indicated by arrowhead [wild-type is shown in (A) and (B); two mutant hearts are shown in panels (C)–(F)]. RA, right atrium; LA, left atrium; RV, right ventricle; LV, left ventricle; Pu, pulmonary artery; Ao, aorta; VSD, ventricular septal defect.