THE ROLE OF TGF-BETA3 IN PALATOGENESIS

这项建议的长期目标是确定转化生长因子-β3诱导的腭裂融合的具体分子机制。在发育过程中，转化生长因子-β3的表达在空间和时间上都受到限制。特别是在灌流的腭部上皮中发现了异常高的表达水平。与转化生长因子-β3表达激增相一致的是，纯合子转化生长因子-β3缺陷小鼠患有双侧继发性腭裂。有研究表明，在上皮融合过程中，转化生长因子-β3触发上皮-间充质转分化和基底膜的降解，这是成功的腭裂融合所必需的过程。综合各种数据，包括转化生长因子-β3在再灌流的腭架中的表达模式和水平，转化生长因子-β3基因缺失突变小鼠的完全性腭裂外露，以及转化生长因子-β3基因缺陷的中缘上皮细胞未能从上皮细胞向间充质细胞转化分化，从而得出了这样的结论或假设：转化生长因子β3是一个主开关，能够在腭裂发生过程中启动一系列分子事件，导致成功的中线上皮融合。为了验证这一假设，我们将利用转化生长因子-β3缺失突变小鼠来研究转化生长因子-β3信号和下游生物反应。这些研究被组织成三个不同的目标：转化生长因子-βI型受体及其下游信号分子的表达和功能，AIM1中Smads的表达和功能，AIM2中上皮主基因和关键分子开关在上皮-间充质转分化中的作用，以及AIM3中金属蛋白酶及其抑制物在基底膜相关降解中的作用。这些研究最终将提高我们对导致腭裂的发病机制的理解。

The long-term goal of this proposal is to define specific molecular mechanisms of TGF-beta3-induced palatal fusion. During development TGF-beta3 expression is both spatially and temporally restricted. Exceptionally high expression levels have been found specifically in prefusion palatal epithelium. Concordant with this surge of TGF-beta3 expression, homozygous TGF-beta3-deficient mice suffer from bilateral clefting of the secondary palate. It has been suggested that during epithelial fusion, TGF-beta3 triggers epithelio-mesenchymal transdifferentiation and associated degradation of the basement membrane, processes necessary for successful palatal fusion. The combined data, including expression pattern and level of TGF-beta3 in prefusion palatal shelves, complete penetrance of cleft palate in TGF- beta3 null mutant mice and failure of TGF-beta3-deficient medial edge epithelial cells to transdifferentiate from epithelial cells to mesenchymal cells lead to the formulation or the following hypotheses: TGF beta3 is a master switch, capable of initiating a cascade of molecular events leading to successful midline epithelial fusion during palatogenesis. To test this hypothesis we will utilize TGF-beta3 null mutant mice to investigate TGF-beta3 signaling and downstream biological responses. The proposed studies have been organized to three different Aims: expression and function of TGF-beta type I receptors and their down- stream signaling molecules, Smads in Aim 1, role of epithelial mastergenes and key molecular switches in epithelio-mesenchymal transdifferentiation during epithelial fusion in Aim2, and function of metalloprotemases and their inhibitors during associated degradation of the basement membrane in Aim 3. These studies will eventually improve our understanding of the pathogenetic mechanisms that lead to formation of cleft palate, one of the most common congenital birth defects in human.