Regulation of Axon Guidance by Second Messengers

描述(申请人提供)：神经系统的发展需要神经元的适当分化、迁移和形态发生。单个神经元的形态发生和组成人类神经系统的数万亿个神经元连接的组装，是通过轴突和树突的引导延伸发生的。我们研究的长期目标是更好地了解细胞内信号级联和效应器机制，这些机制负责发育中的大脑中轴突的生长和引导。为此，我们必须了解神经生长锥如何检测、整合和响应环境中的可溶性、细胞和基质相关的指导分子。参与检测轴突引导信息并将其转化为定向轴突生长的基因突变可能导致许多认知功能缺陷，包括自闭症、阅读障碍、心理障碍和精神发育迟缓。轴突延伸通过一个连续的过程进行，包括由肌动蛋白聚合驱动的前沿膜突起，随后是粘连和突起的稳定。没有粘连的新突起被收回，脱粘的现有突起也是如此。虽然广泛的研究集中在控制膜突起和回缩的信号上，但令人惊讶的是，人们对黏附的调节知之甚少。生长锥突起对细胞外基质(ECM)配体的稳定发生在称为点接触的特殊黏附位置。点接触是一种大分子复合体，包含结构蛋白和信号蛋白，通过跨膜整合素受体将细胞骨架与细胞外基质连接起来。我们的研究集中在了解控制点接触组装、成熟和拆卸的分子信号事件，以及轴突引导线索如何影响这些过程来控制轴突寻路。重要的是，我们的证据表明，促进生长的轴突引导线索刺激点状接触组装和周转，而抑制性线索则减缓新点状接触的组装并减少周转。我们假设轴突的导向和突触接触的稳定需要整合素依赖的点接触的调节。我们将使用各种方法和模型系统在三个特定目标上检验这一假设。在目标1中，我们将研究粘着斑激酶(FAK)在控制黏附动力学、面纱突出和丝状突尖端响应轴突引导线索的磷酸酪氨酸信号中的作用。在目标2中，我们将研究p21激活的蛋白(PAK)在调节整合素依赖的黏附、细胞突起和轴突生长中的作用。最后，在目标3中，我们将在非洲爪哇和斑马鱼胚胎的几个选择点上确定黏附位置动力学在轴突引导中的作用。 公共卫生相关性：功能神经系统的发展需要将轴突和树突准确地引导到它们的目标位置，并建立突触连接。这一建议的重点是了解轴突引导信号如何通过调节整合素依赖的粘连来调节轴突生长。由于许多认知障碍是由不正确的轴突寻径引起的，了解正常神经发育的分子基础对于设计治疗干预措施至关重要。

DESCRIPTION (provided by applicant): The development of the nervous system requires the proper differentiation, migration and morphogenesis of neurons. The morphogenesis of individual neurons and the assembly of the trillions of neuronal connections that compose the human nervous system occurs through guided extension of axons and dendrites. The long-term objective of our research is to better understand the intracellular signaling cascades and effector mechanisms that are responsible for axon outgrowth and guidance in the developing brain. For this we must understand how nerve growth cones detect, integrate and respond to soluble, as well as cell- and substratum-associated guidance molecules in their environment. Mutations in genes involved in the detection and transduction of axon guidance information into directed neurite outgrowth are likely responsible for many deficits in cognitive function, including autisms, dyslexias, psychological disorders and mental retardations. Axon extension proceeds through a sequential process that involves leading edge membrane protrusion driven by actin polymerization, followed by adhesion and protrusion stabilization. New protrusions that do not adhere are retracted, as do existing protrusions that de-adhere. While extensive research has focused on the signals that control membrane protrusion and retraction, surprisingly little is known about the regulation of adhesion. Stabilization of growth cone protrusions to extracellular matrix (ECM) ligands occurs at specialized adhesion sites called point contacts. Point contacts are macromolecular complexes, containing both structure and signaling proteins, which link the cytoskeleton to the ECM through transmembrane integrin receptors. Our research is focused on understanding the molecular signaling events that control point contact assembly, maturation and disassembly and how axon guidance cues influence these processes to control axon pathfinding. Importantly, our evidence suggests that growth promoting axon guidance cues stimulate point contact assembly and turnover, while inhibitory cues slow the assembly of new point contacts and reduce turnover. We hypothesize that guidance of axons to their proper targets and stabilization of synaptic contacts requires modulation of integrin-dependent point contacts. We will test this hypothesis using a variety of approaches and model systems in three specific aims. In Aim 1, we will examine the role of Focal Adhesion Kinase (FAK) in the control of adhesion dynamics, veil protrusion and phosphotyrosine signaling at filopodial tips in response to axon guidance cues. In Aim 2, we will examine the role of p21-Activated Kinase (PAK) proteins in the regulation of integrin-dependent adhesion, cellular protrusion and axon outgrowth. Finally, in Aim 3, we will determine the role of adhesion site dynamics in axon guidance at several choice point in both Xenopus and zebrafish embryos. PUBLIC HEALTH RELEVANCE: The development of a functional nervous system requires accurate guidance of axons and dendrites to their target locations and establishment of synaptic connections. This proposal is focused on understanding how axon guidance cues regulate axon outgrowth by modulating integrin-dependent adhesions. As a number of cognitive disorders result from improper axon pathfinding, understanding the molecular basis for normal neural development is essential for designing therapeutic interventions.