Growth Factor Signaling in Intestinal Development

描述（由申请人提供）：上皮细胞，例如衬在胃肠道的上皮细胞，能够进行连续的自我更新，因为它们含有一群未分化的干细胞，这些干细胞具有很大的自我更新能力，并且还可以产生子细胞，这些子细胞能够分化成上皮层正常功能所需的所有成熟细胞类型。在正常胃肠道发育过程中，介导上皮细胞建立这种分层组织的信号的性质尚未确定。成纤维细胞生长因子（FGF）是至少23种相关的间充质来源的肽生长因子的家族，其调节多种组织的正常个体发育期间发生的多种形态发生和分化事件。我们最近发现，一种FGF受体基因FGFR- 3的表达仅限于下三分之二的肠隐窝上皮中的未分化细胞，并且在隐窝形态发生期间最大程度地表达。此外，FGFR-3-/-小鼠肠道发育的初步研究表明，FGFR-3调节乳鼠肠道中新生隐窝形成的速率和复制隐窝转运细胞的数量。该提议的中心假设是，通过FGFR-3的信号传导调节正常肠个体发育期间多能上皮干细胞的命运和/或增殖。目的1是确定FGFR-3介导的信号传导是否通过对干细胞增殖和/或程序性细胞死亡的影响直接调节正常肠发育期间上皮干细胞群体的扩增。将使用在FGFR 3受体基因中具有靶向突变的小鼠来确定FGFR 3介导的信号传导对克隆发生干细胞的数量和在不同发育时间点对细胞凋亡的影响。目的2将检查FGFR 3对隐窝形态发生的作用是否通过β-连环蛋白/TCF-4依赖性机制介导。文献中的证据表明HMG转录因子TCF-4和Lef-1是隐窝上皮中增殖和凋亡事件的重要下游调节介质。细胞培养和动物模型将用于确定通过FGFR 3的信号传导是否可以调节TCF-4活性。目的3的目标是定义FGFR 3用于调节肠发育期间的形态发生事件的中间信号级联。将使用生物化学和分子方法的组合来研究肠上皮细胞系中的操作性FGFR 3信号传导途径，特别是那些撞击TCF-4的细胞系。

DESCRIPTION (provided by applicant): Epithelia, such as those lining the gastrointestinal tract, are able to undergo continuous self-renewal because they contain a population of undifferentiated stem cells that have a large capacity for self renewal and can also give rise to daughter cells which are able to differentiate into all of the mature cell types needed for normal function of the epithelial layer. The nature of the signals that mediate the establishment of this hierarchical organization of the epithelium during normal gastrointestinal development has not yet been defined. Fibroblast growth factors (FGFs) are a family of at least 23 related mesenchymally derived peptide growth factors that modulate a wide array of morphogenic and differentiation events occurring during normal ontogeny of a variety of tissues. We have recently found that expression of one FGF receptor gene, FGFR- 3, is restricted to undifferentiated cells in the lower two-thirds of the intestinal crypt epithelium and is maximally expressed during crypt morphogenesis. Additionally, preliminary studies of intestinal development in FGFR3-/- mice, demonstrate that FGFR-3 regulates both the rate of nascent crypt formation and the number of replicating crypt transit cells in the suckling mouse intestine. The central hypothesis of this proposal is that signaling through FGFR-3 regulates the fate and/or proliferation of the multipotent epithelial stem cells during normal intestinal ontogeny. Aim 1 is to determine whether FGFR-3-mediated signaling directly regulates expansion of the epithelial stem cell population during normal intestinal development through effects on stem cell proliferation and/or programmed cell death. Mice with targeted mutations in the FGFR3 receptor gene will be used to determine the effects of FGFR3 mediated signaling on the number of clonogenic stem cells and on apoptosis at various developmental time points. Aim 2 will examine whether the effects of FGFR3 on crypt morphogenesis are mediated through a beta-catenin/TCF-4 dependent mechanism. Evidence in the literature suggests that the HMG transcription factors TCF-4 and Lef-1 are important downstream regulatory mediators of proliferative and apoptotic events in the crypt epithelium. Both cell culture and animal models will be used to determine whether signaling through FGFR3 can modulate TCF-4 activity. The goal of aim 3 is to define the intermediate signaling cascades that FGFR3 uses to regulate morphogenic events during intestinal development. The operant FGFR3 signaling pathways in intestinal epithelial cell lines, especially those impinging on TCF-4, will be investigated using a combination of biochemical and molecular approaches.