Regulating the scaling of growth and pattern during neural development

组织的形成需要两个基本的细胞过程的协调：生长和图案化。在神经系统中，生长和模式化的最初驱动力是称为形态发生素的信号因子。它们激活基因调控网络，以指定发育中的中枢神经系统（CNS）内不同的细胞命运，并在发育过程中塑造组织和器官的大小和边界。因此，清楚地了解形态发生素如何完成组织工程的这些壮举，对发育生物学至关重要。调节组织生长和存活的三个关键形态发生途径是由分泌蛋白Sonic Hedgehog（Shh）、Wingless/Int-related（Wnt）和骨形态发生蛋白（BMP）调节的那些。我们小组的工作揭示了颅面和神经发育过程中Shh和Wnt信号之间的复杂关系。我们产生了一系列独特的小鼠突变体，这些突变体在发育过程中仔细滴定了Shh信号的水平。我们发现过度的Shh信号导致颅面原基中Wnt信号和生长促进途径的限制，导致发育不良。我们还表明，如果Shh水平过高，细胞死亡发生在迁移的神经嵴细胞，并防止其整合与基板细胞在感觉神经节形成。因此，需要适当水平的Shh信号传导以确保生长信号与发育组织中的细胞命运获取的协调。然而，目前尚不清楚Shh水平如何实现这种平衡，回答这个问题将揭示脊椎动物的生长和规格如何根据广泛不同的身体大小进行缩放。拟议的研究计划将利用我们的Shh通路突变体来生成一个“可扩展的”脊髓类器官模型系统，以测试生长和模式之间的关系。生长动力学将与使用单细胞RNA测序技术绘制的神经细胞亚型的发育轨迹相关联。一种补充方法涉及在改变Shh信号水平的背景下去除关键的生长促进检查点控制基因，并评估其对神经系统生长和模式的影响。这种遗传方法还将用绿色荧光蛋白标记Shh阳性组织，从而允许通过Shh形态发生体梯度分析支撑生长控制和模式化的动态细胞和分子事件。然后，我们将确定基因网络，调节决策分支点参与协调生长控制和细胞规格的神经组织。在这样做的过程中，我们的研究计划将解决进化发育生物学中的一个长期存在的问题，即“生长和形式”的缩放，并通过优化用于生成适合替代疗法的功能组织的方法，为加拿大人的生活做出贡献。

Tissue formation requires the coordination of two fundamental cellular processes: growth and patterning. Within the nervous system the initial driver of growth and patterning are signaling factors called morphogens. They activate gene regulatory networks to specify distinct cell fates within the developing central nervous system (CNS) and sculpt the size and boundaries to tissues and organs during development. Thus, a clear understanding of how morphogens can accomplish these feats of tissue engineering is of central importance to developmental biology. Three key morphogenetic pathways that regulate tissue growth and survival are those regulated by the secreted proteins Sonic Hedgehog (Shh), Wingless/Int-related (Wnt), and Bone Morphogenetic Protein (BMP). Work from our group has revealed the intricate relationship between Shh and Wnt signaling during craniofacial and nerve development. We generated a series of unique mouse mutants that carefully titrated the levels Shh signaling during development. We discovered that excessive Shh signaling leads to a restriction Wnt signaling and growth promoting pathways in craniofacial primordia, resulting in hypoplasia. We also showed that if Shh levels are too high, cell death occurs in migrating neural crest cells and prevents their integration with placode cells during sensory ganglia formation. Therefore, the appropriate levels of Shh signaling is required to ensure the coordination of growth signals with cell fate acquisition in developing tissues. It is however unclear how Shh levels act to achieves this balance, and answering this question will shed light on how growth and specification can scale according to the widely divergent body sizes in vertebrates. The proposed research program will take advantage of our Shh pathway mutants to generate a 'scalable' spinal organoid model system to test the relationship between growth and patterning. Growth dynamics will be correlated with a mapping of the developmental trajectories of neural cell subtypes using single cell RNA sequencing technology. A complimentary approach involves removing a key growth promoting checkpoint control gene within the context of altered Shh signaling levels and evaluate its effect on the growth and pattering of the nervous system. This genetic approach will also label Shh-positive tissues with green fluorescent protein, allowing for an analysis of dynamic cellular and molecular events underpinning growth control and patterning by the Shh mophogen gradient. We will then identify the gene networks that regulate the decision branch points involved in coordinating growth control and cell specification in the neural tissues. In so doing our research program will address a long-standing problem in evolutionary developmental biology, the scaling of 'growth and form', and will also make contributions to the lives of Canadians by optimizing methodologies for the generation of functional tissues suitable for replacement therapies.