Role of CLASP in neuronal morphogenesis during development

CLASP 在发育过程中神经元形态发生中的作用

• 批准号: 7735703
• 负责人: Fengquan Zhou
• 金额: $ 35.88万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-03 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7735703
• 关键词: Address; Afferent Neurons; Attention; Axon; Behavior; Binding; Biological Assay; Brain; Cell Culture Techniques; Cell division; Cues; Cytoskeleton; Dendrites; Development; Drosophila genus; Electroporation; Event; Genetic; Glycogen Synthase Kinase 3; Goals; Growth; In Vitro; Injury; Link; Mediating; Mediator of activation protein; Mental disorders; Microfilaments; Microtubules; Molecular; Morphogenesis; Mutation; Natural regeneration; Nerve Regeneration; Nervous system structure; Neurodevelopmental Disorder; Neurons; Pathway interactions; Physiological; Plus End of the Microtubule; Positioning Attribute; Process; Property; Proteins; Regulation; Role; Side; Signal Transduction; Testing; axon growth; axon guidance; base; experience; extracellular; in utero; in vivo; insight; migration; nervous system development; neural circuit; neural precursor cell; novel; public health relevance; relating to nervous system; repaired; research study; response; synaptogenesis

DESCRIPTION (provided by applicant): Neuronal morphogenesis is a highly regulated process that ultimately depends on the remodeling of neuronal cytoskeleton in response to extracellular cues. Most previous studies of neuronal cytoskeleton focus on the regulation of actin filaments by extracellular cues. Very few studies have been done to investigate how neuronal microtubules (MTs) are regulated. Regulation of MTs is involved in every step of brain development, such as asymmetrical cell division of neural precursor cells, neuronal migration, axon growth and guidance, dendrite development, and synaptogenesis. Mutation of many MT regulating proteins during development is often associated with mental illnesses, underscoring the importance of MT regulation in normal brain development. Thus, our long-term goal is to understand how extracellular signals governing neuronal morphogenesis are transduced into MT reorganization in neurons that is necessary for proper axonal and dendritic development. Our preliminary study has revealed a novel mechanism by which MT plus end tracking proteins, CLASPs, regulate mammalian axonal and dendritic growth. We found that, unlike other +TIPs that only track MT plus ends, CLASPs display dual bindings to either the plus ends or along the sides of MTs (or MT lattices) in neurons. Functionally, we show that this unique dual MT binding behavior of CLASPs allows them to differentially regulate MT organization and axon growth in different neurons. In regenerating sensory neurons, CLASP mainly bind to MT plus ends and function to support fast axon growth. In contrast, CLASP in developing cortical neurons show increased binding along the side of MTs and act to restrict axon growth. In addition to restricting axon growth, we also provide evidence that CLASPs function to support the development of cortical neuron dendrites. Interestingly, axon guidance cue Slit also functions to repel axons and in the meantime promote dendritic growth. Because CLASP has been placed downstream of Slit to mediate axon repulsion in Drosophila, we hypothesize that CLASPs, with their unique dual MT binding property, may be converging targets of Slit-Robo signaling to regulate mammalian axonal and dendritic development. Thus, the overall goal of this study is to elucidate the role of CLASP in regulation of neuronal morphogenesis in response to extracellular cues during cortical development. To test this hypothesis, we will 1) elucidate the molecular mechanism by which CLASPs regulate MTs to control axon growth and dendritic development, 2) determine the roles of CLASPs in Slitmediated axon repulsion and dendritic growth using in vitro axon guidance assay and cell cultures, and 3) determine the in vivo roles of CLASPs in cortical neuron axon growth/guidance, and dendritic development during cortical development using in utero electroporation. This study will reveal novel molecular mechanisms by which extracellular cues regulate MTs to control neuronal morphogenesis. PUBLIC HEALTH RELEVANCE: Neuronal morphogenesis, including axon growth, guidance, and dendrite growth are key molecular events underlying the formation of the neural circuit during development, or neural repair after injuries. Mistakes in these processes during development are believed to cause many neurodevelopmental disorders. Therefore, our proposed study will not only help us understand how neural circuits form during development, but also provide valuable information of how to promote neural regeneration following injuries.

描述(申请人提供)：神经元形态发生是一个高度调控的过程，最终依赖于神经元细胞骨架对细胞外提示的重塑。以往对神经元细胞骨架的研究主要集中在细胞外信号对肌动蛋白细丝的调节。很少有人研究神经元微管(MTS)是如何被调节的。MTS的调控涉及大脑发育的每一个阶段，如神经前体细胞的不对称分裂、神经元迁移、轴突生长和引导、树突发育和突触形成。许多MT调节蛋白在发育过程中的突变通常与精神疾病有关，这突显了MT调节在正常大脑发育中的重要性。因此，我们的长期目标是了解控制神经元形态发生的细胞外信号如何转化为神经元中的MT重组，这是正常的轴突和树突发育所必需的。我们的初步研究揭示了一种新的机制，通过MT+末端跟踪蛋白CLAP来调节哺乳动物的轴突和树突生长。我们发现，与其他只追踪MT+末端的+TIPS不同，CLASPS在神经元中显示出与正末端或沿着MT(或MT晶格)两侧的双重结合。在功能上，我们表明这种独特的双重MT结合行为使它们能够不同地调节不同神经元中的MT组织和轴突生长。在再生感觉神经元中，CLAP主要与MT+末端结合，起到支持轴突快速生长的作用。相反，发育中的皮质神经元中的CALAP显示出沿MTS一侧的结合增加，并限制轴突的生长。除了限制轴突生长外，我们还提供了CLASPS支持皮质神经元树突发育的证据。有趣的是，轴突引导线索狭缝也有排斥轴突的功能，同时也促进树突的生长。由于在果蝇中，CLASP被放置在SILT下游以调节轴突排斥，因此我们推测，CLASP凭借其独特的双重MT结合特性，可能是Sit-Robo信号调节哺乳动物轴突和树突发育的汇聚靶点。因此，本研究的总体目标是阐明CLAP在调节神经元形态发生中的作用，以响应皮质发育过程中的细胞外线索。为了验证这一假说，我们将1)阐明CALPS调节MTS以控制轴突生长和树突发育的分子机制，2)利用体外轴突引导试验和细胞培养来确定CLASPS在狭缝介导的轴突排斥和树突生长中的作用，以及3)利用宫内电穿孔技术在体内确定CALPS在皮质神经元轴突生长/引导和树突发育中的作用。这项研究将揭示细胞外信号调节MTS以控制神经元形态发生的新的分子机制。公共卫生相关性：神经元形态发生，包括轴突生长、引导和树突生长，是发育过程中神经回路形成或损伤后神经修复的关键分子事件。发育过程中的错误被认为会导致许多神经发育障碍。因此，我们提出的研究不仅将帮助我们了解神经回路在发育过程中是如何形成的，而且还将为如何促进损伤后的神经再生提供有价值的信息。