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：生长和模式。在神经系统中，生长和模式形成的最初驱动力是被称为形态因子的信号因子。它们激活基因调控网络，在发育中的中枢神经系统（CNS）内指定不同的细胞命运，并在发育过程中塑造组织和器官的大小和边界。因此，清楚地了解形态因子如何完成这些组织工程的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Bone morphogenetic Protein （BMP）调节的。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，仔细地滴定了发育过程中Shh信号的水平。我们发现过多的Shh信号导致颅面原基中Wnt信号和生长促进通路受限，导致发育不全。我们还发现，如果Shh水平过高，迁移的神经嵴细胞会发生细胞死亡，并阻止它们在感觉神经节形成过程中与基板细胞结合。因此，在发育组织中，需要适当水平的Shh信号来确保生长信号与细胞命运获取的协调。然而，目前尚不清楚Shh水平是如何达到这种平衡的，回答这个问题将揭示脊椎动物的生长和规格是如何根据不同的体型进行调整的。拟议的研究计划将利用我们的Shh通路突变体来产生一个“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测序技术绘制神经细胞亚型的发育轨迹相关联。一种互补的方法包括在Shh信号水平改变的背景下去除一个关键的促进生长的检查点控制基因，并评估其对神经系统生长和模式的影响。这种遗传方法也将用绿色荧光蛋白标记Shh阳性组织，允许分析动态细胞和分子事件，支持Shh形态因子梯度的生长控制和模式。然后，我们将确定调节神经组织中涉及协调生长控制和细胞规范的决策分支点的基因网络。通过这样做，我们的研究项目将解决进化发育生物学中一个长期存在的问题，即“生长和形态”的缩放，并将通过优化适合替代疗法的功能性组织的生成方法，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.