Elucidating the mechanisms of gradient dependent and independent Wnt signaling in neuronal development

背景：Wnt家族在动物发育中起着至关重要的作用。已知WNT蛋白在组织内产生梯度分布。然而，目前还不清楚细胞如何感知和解释Wnt的梯度，以及Wnt的梯度分布在多大程度上有助于动物的发育。一些研究，包括我们最近的工作，表明WNT的梯度分布在动物发育的某些方面是不必要的。这个项目是为了增加我们对Wnt信号梯度依赖和独立机制的理解，以秀丽线虫为模型系统。最近，我们报道了Wnt指导神经元突起或神经突起的收缩，称为神经突起修剪。对线虫发育中的PDB(后背部B)神经元的时间推移成像显示，一些神经突起经历了刻板的修剪。我们发现PDB神经突起的修剪依赖于Wnt信号。引人注目的是，我们没有在‘非扩散Wnt’突变体中观察到修剪缺陷。这一观察结果表明，Wnt的梯度分布不是PDB轴突修剪所必需的。有趣的是，我们发现PDB突触的位置取决于WNT梯度。综上所述，我们的结果表明，单个神经元以一种上下文相关的方式使用梯度依赖和独立的Wnt信号。研究目标和方法本项目的长期目标是了解Wnt信号在动物发育中的不同作用。我们的短期目标是利用线虫扩大我们在神经元发育中梯度依赖和独立的Wnt信号的知识。我们将针对以下具体研究目标进行研究。揭示Wnt信号在轴突修剪中的梯度非依赖性机制。利用遗传筛选，我们将揭示Wnt信号在轴突修剪中的下游效应。揭示Wnt信号在突触定位中的梯度依赖机制。利用基因筛选，我们将发现Wnt效应基因，这些基因特定地控制突触定位。3.了解哪些神经发育事件与WNT梯度相关。我们将使用我们产生的‘不可扩散的WNT’突变体来研究已知的受Wnt信号调控的各种神经发育事件。影响我们将揭示Wnt依赖的轴突修剪和突触定位的机制，这两个机制仍然知之甚少。我们还将描述哪些神经发育过程依赖于Wnt梯度。从我们的研究中获得的知识将有助于更好地理解Wnt信号在动物发育中的复杂作用。它还将帮助其他研究人员测试是否在其他模式生物中存在相同的机制。我们将通过我们的研究项目培养在遗传学、分子生物学和显微镜方面具有广泛专业知识的具有国际竞争力的下一代科学家。

Background The Wnt family of morphogens play crucial roles in animal development. Wnt proteins are known to generate gradient distributions within a tissue. However, it is not well understood how cells sense and interpret the Wnt gradients and to what extent the gradient distribution of Wnt contributes to animal development. Several studies, including our recent work, suggest that the gradient distribution of Wnts is dispensable in some aspects of animal development. This project is designed to increase our understanding of the gradient dependent and independent mechanisms of Wnt signaling using roundworm, Caenorhabditis elegans, as a model system. Progress Recently, we reported that Wnt instructs retraction of the neuronal processes or neurites, called neurite pruning. Time-lapse imaging of the developing PDB (posterior dorsal B) neuron in C. elegans revealed that some neurites undergo stereotyped pruning. We found the PDB neurite pruning depends on the Wnt signal. Strikingly, we did not observe pruning defects in the `non-diffusible Wnt' mutants. This observation suggests the gradient distribution of Wnt is not necessary for PDB neurite pruning. Interestingly, we found the position of PDB synapses depends on the Wnt gradient. Taken together, our results showed that a single neuron uses gradient dependent and independent Wnt signaling in a context-dependent manner. Research Objectives and Methods The long-term objective of this program is to understand the diverse actions of Wnt signaling in animal development. Our short-term objective is to expand our knowledge in gradient dependent and independent Wnt signaling in neuronal development using C. elegans. We will address the following specific research objectives.  1. Uncovering the mechanism of gradient independent Wnt signaling in neurite pruning. Using genetic screenings, we will uncover the downstream effectors of Wnt signaling in neurite pruning.  2. Uncovering the mechanisms of gradient dependent Wnt signaling in synapse positioning. Using genetic screenings, we will uncover Wnt effector genes that specifically function in controlling synapse positioning.  3. Understanding which neurodevelopmental events are Wnt gradient dependent. We will examine the various neurodevelopmental events known to be regulated by Wnt signaling using `non-diffusible wnt' mutants we generate. Impact We will reveal the mechanisms of Wnt-dependent neurite pruning and synapse positioning, both of which are still poorly understood. We will also characterize which neurodevelopmental processes depend on the Wnt gradient. Knowledge gained from our research will help better understand the complex actions of Wnt signaling in animal development. It will also help other researchers to test if the same mechanisms are present in other model organisms. We will raise internationally competitive next-generation scientists with broad expertise in genetics, molecular biology, and microscopy through our research program.