My colleagues and I have provided two lines of evidence suggesting that defects in Platelet-Derived Growth Factor-A (PDGF-A) function result in lethality or developmental deformities during early vertebrate embryogenesis. First, mRNA for PDGF-A and its receptor are maternally-encoded in Xenopus and mice, and are expressed throughout early development. Second, mice harboring a deletion of the receptor for PDGF-A fail to develop normally past 8.5 days and usually die in utero. I would like to extend these preliminary findings in the experimentally accessible Xenopus embryo. Because the temporal and spatial expression patterns of PDGF and receptor are conserved evolutionarily, these studies should have relevance to human physiology and development. The research has two broad objectives. The first is relatively straightforward and builds on my preliminary studies on the role of PDGF during early embryogenesis. The second is more complicated and relies on my observation that PDGF-A and its receptor are markers for anterior development. Using these markers, I propose to identify novel morphogens that specify antero-posterior pattern formation. The first objective is to determine the function of PDGF-A during development. Antiserum to PDGFA and its receptor will be used immunohistologically to identify those cells producing PDGF and receptor protein. This should extend my preliminary results localizing mRNA for PDGF-A and its receptor to ectoderm and mesoderm, respectively, in the gastrula. Recombinant Xenopus PDGF-A will be evaluated for effects on differentiation of mesodermal cells in culture. Finally, PDGF-deficient embryos will be produced and monitored for developmental abnormalities. To create PDGF-deficient animals, I have already constructed mutant PDGF genes that suppress wild-type activity in a trans-dominant fashion. Embryos will be made deficient in PDGF by the injection of synthetic mRNA encoding mutant PDGF. Abnormalities of these embryos may reveal the in vivo function(s) of the growth factor. Of particular interest will be abnormalities appearing at early gastrula to late neurula stages when PDGF and receptor mRNA levels are highest. The second objective is to use PDGF-A and its receptor as markers to identify factors that control antero-posterior patterns of differentiation. Preliminary results show that mRNA encoding PDGF and its receptor are expressed in an anterior to posterior gradient in the gastrula and neurula. Thus, PDGF-A appears to be expressed in anterior ectoderm, while its receptor is localized in the anterior mesoderm. Candidate agents thought to influence axial polarity will be studied in an attempt to induce (or suppress) the differentiation of cells expressing PDGF-A and its receptor. These factors will be evaluated in conjunction with tissues (such as the organizer) for their effect on expression of PDGF-A and its receptor. These experiments may lead to the identification of morphogens that control antero-posterior polarity in vertebrates.
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