5B, panels g,g). on endoderm to up-regulate Hex, which, in turn, controls production of a diffusible heart-inducing factor. This novel function for Hex suggests an etiology for the cardiac malformations in Hex mutant mice and will make possible the isolation of factors that induce heart directly in the mesoderm. embryos (Marvin et al. 2001; Schneider and Mercola 2001; Tzahor and Lassar 2001). Other secreted Wnt antagonists, such as Frz-b and Szl, appear less active, probably owing to selectivity for the particular Wnts that must be prevented from signaling. The structurally distinct Dkk-1 and Crescent proteins both block signaling by preventing interaction of Wnts with receptors on the cell surface (for review, see Kawano and Kypta 2003). Intracellular inhibitors of the canonical Wnt/-catenin pathway initiate cardiogenesis (Schneider and Mercola 2001), but hearts are also induced in mesodermal explants by Wnt-11 (Pandur et al. 2002a), which stimulates noncanonical signaling through Jun N-terminal kinase (JNK) and protein kinase-C (PK-C) and might also antagonize canonical signaling through -catenin (Maye et al. 2004) in this setting. These studies indicate that inhibition of canonical Wnt/-catenin signaling and activation of noncanonical signaling are both important initiators of cardiogenesis in embryonic tissue in amphibians and amniotes, yet nearly nothing is known in any species about the genes and protein effectors that operate downstream of these pathways to initiate cardiogenesis. Their identification will be important not only for tissue engineering, but also to distinguish how heart induction differs from, and is coordinated with, other effects of Wnt signaling on cell fate and morphogenesis during embryogenesis. In embryos, Dkk1 and Crescent are produced within Spemann’s Organizer, an important signaling center of the gastrula-stage embryo that eventually gives rise to the notochord and head mesoderm and expresses other signaling proteins involved in dorsoanterior patterning, including XNr-1, a homolog of the mouse Nodal protein, and BMP antagonists noggin and chordin (for review, see Harland and Gerhart 1997). The Organizer is clearly required for heart induction, as has been shown by extirpation studies (Sater and Jacobson 1990; Nascone and Mercola 1995); however, it cannot induce either 10058-F4 native or ectopic heart tissue efficaciously unless accompanied by a small amount of underlying deep dorsoanterior endoderm (Nascone and Mercola 1995). Classical grafting studies also pointed out the heart-inducing properties of dorsoanterior endoderm in amphibians (Jacobson 1960; Jacobson and Duncan 1968; Fullilove 1970), and similar tissue extirpation and recombination experiments revealed heart-inducing activity in chick embryo anterior hypoblast (Yatskievych et al. 1997) and mouse embryonic anterior visceral endoderm (AVE) (Arai et al. 1997). The 10058-F4 latter two are both extraembryonic but share expression of certain genes with amphibian dorsoanterior endoderm suggestive of common signaling properties (for discussion, see Bouwmeester et al. 1996). Theoretically, Wnt antagonists might induce heart tissue in parallel with a signal from the dorsoanterior endoderm. One example of parallel signaling is a model (Marvin et al. 2001) based on chick embryo experiments (Sugi and Lough 1994; Schultheiss et al. 1997; Schlange et al. 2000; Marvin et al. 2001; Tzahor and Lassar 2001) in which the heart-forming region develops at the intersection where Wnt antagonists Rabbit Polyclonal to RGS10 and BMP isoforms are presumed to act. Although BMPs are clearly necessary for cardiogenesis, they are induced and needed only after the requirement for 10058-F4 Wnt antagonists and endoderm has passed (Shi et al. 2000), suggesting that another inducing signal exists in the endoderm. An alternative model is that Wnt antagonists act on the endoderm to stimulate secretion of a molecule that diffuses into adjacent mesoderm to specify heart formation. In this paper, we describe a genetic cascade that constitutes an indirect mode of action for Wnt antagonists in heart induction. We created mosaics of normally noncardiogenic ventroposterior mesendoderm consisting of cells that either express or do not.