The proposed work is a continuation of a long term goal to elucidate the physiological function of vitamin A in embryonic development. Our working hypothesis is that critical early developmental events involving the specification of the cardiovascular and central nervous systems are regulated by vitamin A within a common narrow developmental window. The main focus of the present research is on the formation of the cardiovascular system.
The specific aims are: 1. Determine the spatiotemporal sequence of vitamin A-regulated heart forming and asymmetry determining morphological events. Dysregulation of morphological and cellular events caused by the lack of vitamin A during early embryogenesis will be accurately described using multiple molecular markers. These studies are essential for the subsequent analysis of molecular events. 2. Examine regulation of expression of the cardiac-specific genes GATA-4/5/6 by vitamin A. Effects of retinoids on the expression of GATAs and GATA-4 target genes will be analyzed in retinoid-deficient embryos. The involvement of retinoid receptors in GATA-4 regulation will be studied using receptor-selective ligands and oligonucleotides directed against receptors and GATAs. 3. Identify retinoid-dependent cardiac asymmetry regulating genes. Effect of vitamin A deficiency on the expression of cardiac asymmetry-linked genes will be determined, with focus on SnR-1 which is severely downregulated in absence of vitamin A. SnR-1 regulation by retinoids will be studied with retinoid receptor specific ligands and antisense oligonucleotides. The studies will utilize our novel vitamin A-deficient quail embryo model. Gene expression will be studied by whole-mount in situ hybridization. Cell and tissue diversification will be identified with molecular markers. The proposed studies offer an unique opportunity to learn about the critical role of vitamin A in cardiovascular development so as to apply this knowledge to understanding the origin of congenital heart defects that may arise as a result of marginal vitamin A nutrition during pregnancy.
