Classical experiments performed with Xenopus and chick embryos have suggested that the initial steps of cardiac development involve an interplay between cardiogenic mesoderm and adjacent endoderm. Our recent studies with mutant mouse embryos have shown that transcription factor GATA-4 plays an essential role in this interaction between mesoderm and endoderm. GATA-4 is expressed in pre-cardiac mesoderm and adjoining endoderm during the simultaneous processes of heart tube formation and foregut invagination. GATA-4 knockout mice die before 9.5 days p.c. and exhibit defects in ventral morphogenesis, including abnormal foregut formation and a failure of fusion of the bilateral myocardial primordia. In contrast, high percentage Gata4-/- ES cell <> Gata4+/+ chimeric mouse embryos in which Gata4+/+ cells populate only endoderm do not exhibit abnormalities in ventral development, indicating the presence of wild type cells in visceral and/or definitive endoderm is sufficient to allow normal ventral development in adjoining Gata4- - mesodermal tissues. We postulate that GATA-4 participates in the migration and fusion of the myocardial primordia by regulation the expression of cortical target genes in endoderm. We will take advantage of the inherent genetic strengths of this mouse to characterize the nature of the interactions between endoderm and cardiogenic mesoderm. To distinguish whether expression of GATA-4 in visceral or definitive endoderm cells is essential for cardiac morphogenesis, we will analyze the phenotype of chimeric mice produced by aggregating tetraploid cells with either Gata4-/- ES cells or embryos. In addition, we will examine heart development in chimeras derived from host embryos with deficiencies in visceral endoderm function (e.g., mutants in HNF-4). Differential display and in situ hybridization will be utilized to identify genes under-expressed in Gata4-/- endoderm. We will complement these experiments on mouse Gata4 with studies of patients with chromosomal deletions near the human GATA4 gene on 8p23.1, a deletion/duplication locus association with a high incidence of congenital heart disease. The exact size and boundaries of the deletions in a series of patients will be characterized using standard techniques to test the hypothesis that haploinsufficiency of GATA4 is associated with congenital heart disease. These experiments should provide insight into the mechanisms underlying both normal and pathological cardiac development in vertebrates.
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