Notch Signaling and the Somitogenesis Oscillator

描述（由申请人提供）：在分子、细胞和多细胞水平上，生物组织的空间和时间生成是生物网络内许多独立组分相互作用的产物。这些网络构成了复杂的系统，其属性大于组成部分的总和。在体节发生过程中，一个时间模式化机制，振荡器，产生了一个新兴的空间模式，预示着形态节的形成。该振荡器产生基因表达的条纹/波，这些条纹/波重复地穿过将要经历分割的细胞的场。我们的第一个目标是建立一个更详细的年表的基因表达和形态的周期性变化，发生在斑马鱼的体节发生使用体内成像，仔细分期胚胎和荧光原位杂交。最近的工作，包括我们自己的工作，已经发现Notch信号通路在振荡器机制和更直接调节形态分割的后期过程中具有连续的功能。在斑马鱼中，我们已经表明Notch配体aei/deltaD、bea/deltaC和Notch靶基因her 1各自在振荡器机制中起作用，但各自具有不同的突变表型。我们的第二个目的是通过两个δ突变体的比较表型分析、双突变体分析、异位表达实验和遗传镶嵌来区分两个δ同源物的功能。我们假设aei/deltaD和bea/deltaC在振荡器电路内具有不同的信号作用，并且δ信号的相对功能和强度在分割组织内的不同位置处变化。第三个目标是研究her 1在振荡器中的功能。我们将确定Her 1蛋白质水平是否振荡，以及蛋白质的稳定性是否决定振荡器的频率。最终目标是开发振荡器的实时读数，以便可视化活胚胎中的振荡。累积起来，这些实验解决了细胞与其紧邻细胞之间的局部信号相互作用如何通过Notch途径在近轴中胚层内产生紧急的，更高水平的组织。这种对支配体节发生的遗传网络的分析是我们理解脊椎动物发育的核心，对生物回路的复杂特性具有广泛的影响。

DESCRIPTION (provided by applicant): At the molecular, cellular and multicellular level, the spatial and temporal generation of biological organization is the product of the interaction of many independent components within biological networks. These networks constitute complex systems with properties greater than the sum of the constituent parts. During somitogenesis, a temporal patterning mechanism, the oscillator, gives rise to a emergent spatial pattern that presages the formation of morphological segments. This oscillator creates stripes/waves of gene expression that repeatedly travel through the field of cells about to undergo segmentation. Our first objective is to establish a more detailed chronology of the cyclical changes in gene expression and morphology that occur during zebrafish somitogenesis using vivo imaging, carefully staged embryos and fluorescent in situ hybridization. Recent work, including our own, has found that the Notch signaling pathway has successive functions in both the oscillator mechanism and in a later process that more directly regulates morphological segmentation. In the zebrafish, we have shown that the Notch ligands aei/deltaD, bea/deltaC and Notch target gene her1 each function within the oscillator mechanism, but each has a distinct mutant phenotype. Our second aim is to distinguish the functions of the two delta homologs by comparative phenotypic analysis of the two delta mutants, double mutant analysis, ectopic expression experiments and genetic mosaics. We hypothesize that aei/deltaD and beaJdeltaC have distinct signaling roles within the oscillator circuit and that the relative function and intensities of the delta signals vary at different positions within the segmenting tissue. The third aim examines the function of her1 within the oscillator. We will determine if Her1 protein levels oscillate and if the stability of the protein determines the frequency of the oscillator. The final aim is to develop a real-time readout of the oscillator in order to visualize the oscillations in live embryos. Cumulatively, these experiments address how local signaling interactions between a cell and its immediate neighbors via the Notch pathway give rise to an emergent, higher-level of organization within the paraxial mesoderm. This analysis of the genetic network that governs somitogenesis is central to our understanding of vertebrate development and has broad implications concerning the complex properties of biological circuits.