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

Wnt家族形态形成因子在动物发育过程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，细胞如何感知和解释Wnt梯度以及Wnt梯度分布对动物发育的影响程度尚不清楚。包括我们最近的工作在内的一些研究表明，wnt的梯度分布在动物发育的某些方面是必不可少的。本项目旨在以秀丽隐杆线虫（Caenorhabditis elegans）为模型系统，增加我们对Wnt信号的梯度依赖和独立机制的理解。最近，我们报道了Wnt指示神经元过程或神经突的收缩，称为神经突修剪。秀丽隐杆线虫发育中的PDB（后背B）神经元的延时成像显示，一些神经突经历了定型修剪。我们发现PDB神经突修剪依赖于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的梯度分布对于PDB神经突修剪不是必需的。有趣的是，我们发现PDB突触的位置取决于Wnt梯度。综上所述，我们的研究结果表明，单个神经元以上下文依赖的方式使用梯度依赖和独立的Wnt信号。研究目的和方法本项目的长期目标是了解Wnt信号在动物发育中的各种作用。我们的短期目标是利用秀丽隐杆线虫扩展我们在梯度依赖和独立Wnt信号在神经元发育中的知识。我们将解决以下具体的研究目标。1. 揭示梯度无关的Wnt信号在神经突修剪中的作用机制。利用遗传筛选，我们将揭示Wnt信号在神经突修剪中的下游效应。2. 揭示梯度依赖性Wnt信号在突触定位中的作用机制。通过基因筛选，我们将发现Wnt效应基因在控制突触定位中起特殊作用。3. 了解哪些神经发育事件依赖于Wnt梯度。我们将使用我们生成的“非扩散Wnt”突变体来研究已知由Wnt信号调节的各种神经发育事件。我们将揭示wnt依赖性的神经突修剪和突触定位的机制，这两者仍然知之甚少。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同样的机制存在于其他模式生物中。我们将通过我们的研究项目培养具有国际竞争力的下一代科学家，他们在遗传学、分子生物学和显微镜学方面具有广泛的专业知识。

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.