Developmental Biology 295 (2006) 580– 588
Bmp2 instructs cardiac progenitors to form the heart-valve-inducing field
José Rivera-Feliciano, Clifford J. Tabin
Department of Genetics, Harvard Medical School, Boston, MA 02115, USA
Received for publication 29 November 2005; revised 23 March 2006; accepted 30 March 2006 Available online 4 April 2006
A hallmark of heart-valve development is the swelling and deposition of extracellular matrix in the heart-valve region. Only myocardium overlying this region can signal to underlying endothelium and cause it to lose cell–cell contacts, delaminate, and invade the extracellular space abutting myocardium and endocardium to form endocardial cushions (EC) in a process known as epithelial to mesenchymal transformation (EMT). The heart-valve myocardium expresses bone morphogenetic protein-2 (Bmp2) coincident with development of valve mesenchyme. BMPs belong to the transforming growth factor beta superfamily (TGF-β) and play a wide variety of roles during development. We show that conditional ablation of Bmp2 in cardiac progenitors results in cell fate changes in which the heart-valve region adopts the identity of differentiated chamber myocardium. Moreover, Bmp2-deficient hearts fail to induce production and deposition of matrix at the heart-valve-forming region, resulting in the inability of the endothelium to swell and impairing the development of ECs. Furthermore, in collagen invasion assays, Bmp2 mutant endothelium is incapable of undergoing EMT, and addition of BMP2 protein to mutant heart explants rescues this phenotype. Our results demonstrate that Bmp2 is both necessary and sufficient to specify a field of cardiac progenitor cells as the heart-valve-inducing region amid developing atria and ventricles. © 2006 Elsevier Inc. All rights reserved.
Keywords: Heart development; Endocardial cushion; epithelial–mesenchymal transition, EMT; Bmp2; Atrioventricular canal, AVC; Cardiac morphogenesis; Cardiac development; Bone morphogenetic protein
Proper heart chamber and valve development is essential for life, and problems in the ontogeny of the heart valves and septa account for a preponderance of cases of congenital heart disease (Fishman and Chien, 1997; McFadden and Olson, 2002; Pierpont et al., 2000). The heart-valve progenitors originate adjoining the developing atria and ventricles in a region known as the atrioventricular canal (AVC), in an arrangement that prefigures the architecture of the adult cardiac chambers.
Several genes show restricted spatial expression along the antero-posterior axis of the developing heart (Mjaatvedt, 1999). They divide the heart into five regions: outflow tract (OT), right ventricle, left ventricle, atria, and inflow tract (IT). Concurrent with looping and growth of the ventricles, the ECs form and remodel to become the OT and AV valves (Bolender and Markwald, 1979; Fishman and Chien, 1997; McFadden and Olson, 2002). Inductive signals from the myocardium stimulate
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endothelial cells in these regions to undergo EMT (Mjaatvedt et al., 1987). The potential to respond to these signals is unique within the ECs, as atrial or ventricular endothelia cannot undergo transformation in vivo or in vitro even when exposed to suitable stimuli (Runyan and Markwald, 1983). Hence, mesenchyme formation within the heart depends on both a localized myocardial-inducing activity and an endothelial cell competence.
BMPs are part of the transforming growth factor beta superfamily (TGF-β) and have a broad variety of roles during development (Hogan, 1996). Bmp2, Bmp4 and Bmp5 are localized temporally and spatially to the myocardium overlying the AVC (Cai et al., 2005; Jiao et al., 2003; Lyons et al., 1990). Conventional generation of Bmp2 null embryos results in death by embryonic day 8.5 (Zhang and Bradley, 1996), prior to cushion formation. This has precluded the examination of Bmp2 loss-of-function during specification of AVC and EC morpho- genesis. Conditional ablation of Bmp4 in the myocardium reveals that it regulates AV septation after cushions are formed (Jiao et al., 2003). In Bmp5-deficient embryos, heart develop- ment is normal (Kingsley et al., 1992). BMPs bind two serine/ threonine kinase receptors, type I and type II. Conditional