Specificaton and Patterning of the Animal-Vegetal Axis

项目摘要PI感兴趣的是调节海胆胚胎中动物-植物（A/V）轴规格的母体机制，以及阐明如何将此母体信息用于胚层分离。沿着A/V轴观察到的第一个分子不对称性之一是B-连环蛋白进入植物细胞核。核B-连环蛋白是经典Wnt信号的转换器，对于内中胚层的特化以及沿着A/V轴的图案形成是必不可少的。阐明典型的Wnt信号是如何在植物细胞中被激活的，将提供对指定该轴的机制的深入了解。在前一个授予期间所做的工作表明，Dishevelled（Dsh）蛋白在受精后定位于植物囊泡皮质结构。Dsh与这些膜囊泡的相互作用然后调节B-连环蛋白的核进入和经典Wnt信号传导的激活。植物细胞中B-连环蛋白的早期核靶是信号分子SpWnt 8。来自前一个授权期的结果表明，SpWnt 8是初级间充质分化和内中胚层持续特化所必需的。这些数据表明，SpWnt 8具有维持植物细胞核B-连环蛋白水平的早期功能，以及在囊胚晚期调节内中胚层基因表达和原肠胚形成的晚期功能。SpWnt 8的晚期功能似乎需要一个非经典的Wnt信号通路，也涉及Dsh。为了进一步了解Dsh和SpWnt 8在海胆胚胎中的调节和功能，我们将追求三个具体目标：1）确定在早期胚胎中调节Dsh活性的分子机制。这些研究将揭示海胆胚胎植物细胞中选择性激活经典Wnt信号的机制，并有可能揭示动物-植物轴形成的分子基础; 2）功能分析关键平面细胞极性通路（PCP）基因在动物-植物轴形成中的作用。这些研究将揭示通过这种非经典Wnt途径的信号传导是否调节早期胚胎中的内中胚层形成和原肠胚形成; 3）确定由SpWnt 8激活以调节内中胚层细胞命运的细胞内途径。这些研究将揭示SpWnt 8是否可以通过经典和非经典Wnt信号通路进行信号转导，并进一步揭示SpWnt 8激活的特定PCP信号通路。这些研究将加深我们对海胆A/V轴是如何被指定和模式化的理解，并可能为了解双侧性动物A/V轴的进化提供线索。这些研究还将包括来自科学领域代表性不足的群体的博士后、研究生和本科生。从工作的结果也将传达给学生就读于PI的发育生物学类。

Project SummaryThe PI is interested in the maternal mechanisms that regulate the specification of the animal-vegetal (A/V) axis in the sea urchin embryo, and in elucidating how this maternal information is used in germ layer segregation. One of the first molecular asymmetries seen along the A/V axis is the entry of B-catenin into vegetal cell nuclei. Nuclear B-catenin, which is the transducer of the canonical Wnt signal, is essential for specification of endomesoderm, and also for pattern formation along the A/V axis. Elucidating how canonical Wnt signaling is activated in vegetal cells will provide insight into the mechanisms that specify this axis. Work done during the previous granting period has shown that the Dishevelled (Dsh) protein becomes localized to vegetal vesicular cortical structures following fertilization. Dsh interaction with these membrane vesicles then regulates the nuclear entry of B-catenin and activation of canonical Wnt signaling. An early nuclear target of B-catenin in vegetal cells is the signaling molecule SpWnt8. Results from the previous granting period have shown that SpWnt8 is required for primary mesenchyme differentiation and the continued specification of the endomesoderm. These data suggests that SpWnt8 has an early function to maintain nuclear B-catenin levels in vegetal cells, and a late function at the late blastula stage to regulate endomesoderm gene expression and gastrulation. The late function of SpWnt8 appears to require a non-canonical Wnt signaling pathway that also involves Dsh.To further understand Dsh and SpWnt8 regulation and function in the sea urchin embryo, three specific aims will be pursued: 1) To identify molecular mechanisms that regulate Dsh activity in early embryos. These studies will reveal the mechanisms that selectively activate canonical Wnt signaling in vegetal cells of the sea urchin embryo, and have the potential to reveal the molecular basis for the animal-vegetal axis; 2) To functionally analyze the role of key planar cell polarity pathway (PCP) genes animal-vegetal axis patterning. These studies will reveal if signaling through this non-canonical Wnt pathway regulates endomesoderm formation and gastrulation in the early embryo; 3) To determine the intracellular pathways activated by SpWnt8 to regulate endomesodermal cell fates. These studies will reveal if SpWnt8 can signal via the canonical and non-canonical Wnt pathways, and further, will reveal the specific PCP pathway activated by SpWnt8.Together, these studies will increase our understanding of how the sea urchin A/V axis is specified and patterned and may provide insight into the evolution of the A/V axis in bilaterians. These studies will also include postdocs, graduate students and undergraduate students from groups underrepresented in science. Results from the work will also be communicated to students enrolled in the PI's Developmental Biology classes.