Signaling Roles of Intermediate Filament Isoforms in Neural Development

该项目旨在通过解决生物学中一个悬而未决的问题来推进科学知识：创造不同类型细胞的分子原理是什么？研究的重点是一类以细胞类型特异性模式表达的蛋白质，中间丝（IF）蛋白。该项目的重点是大脑中的神经元，这些神经元在发育成熟的过程中会切换IF亚型，包括将长轴突延伸到大脑中的正确位置。这项研究旨在确定为什么神经元在生长过程中会从一种IF亚型切换到另一种。该项目还将通过促进预期的社会成果来扩大影响：1)扩大STEM领域的教育，以支持更知情、更有科学素养的公众；2)增加STEM劳动力的多样性和包容性。特别是，该实验室旨在让女性、残疾人和未被充分代表的少数族裔充分参与STEM的各个层面。该项目每年夏天为一名代表性不足的少数民族学生提供本科生（REU）研究经验。由该项目资助的REU通过调查生长轴突中各种中间细丝的作用，提供实验室研究的实践经验。此外，入选的REU学生将参加弗吉尼亚大学设立的暑期研究实习计划（SRIP），该计划为在个人和团体环境中从事STEM职业提供专业技能培训和指导。SRIP有成功地将参与的本科研究人员安置到STEM职业的历史。本项目旨在发现神经元中发育中的IF亚型从巢蛋白到神经丝的转换如何影响微管细胞骨架的信号传导，从而调节生长锥的结构和运动。PI的实验室最近发现IF蛋白巢蛋白调节生长锥形态。此外，巢蛋白改变了生长锥对引导信号Sema3a的响应。它通过结合激酶Cdk5及其底物，微管相关蛋白DCX，从而增加DCX的磷酸化。随着神经元的成熟，它们将IF亚型从巢蛋白转换为神经丝，后者结合DCX而不结合Cdk5。该项目将验证神经元if通过Sema3a信号下游的DCX调节轴突生长锥的微管动力学和捆绑的新假设。目的1将发现实验中频开关如何影响轴突微管和生长锥行为。目的2将发现实验中频开关如何影响皮层神经元的Sema3a反应。该项目将利用最先进的成像方法，结合不同中频亚型的下调和过表达，在培养的皮质神经元中揭示不同中频亚型在轴突生长过程中调节生长锥微管动力学的机制。生长锥在发育过程中对细胞外信号的响应不仅受可用信号、表面表达的受体、细胞内底物的影响，还受表达的IF亚型的影响。PI提出神经元IF亚型可以作为无处不在的激酶级联的放大器或抑制器，以提供局部和细胞类型特异性的效应反应调节。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This project aims to advance scientific knowledge by addressing an unanswered question in biology: what are the molecular principles that create different types of cells? The research focuses on a class of proteins which are expressed in strikingly cell-type specific patterns, the intermediate filament (IF) proteins. The project focuses on neurons in the brain which are known to switch IF subtype as they develop their mature shape, including extending long axons to the correct location in the brain. The research aims to determine why neurons switch from one IF subtype to another as they grow. The project will also broaden the impact by contributing to desired societal outcomes: 1) broadening education in STEM fields to support a better informed and more scientifically literate public, and 2) increasing the diversity and inclusivity of the STEM workforce. Especially, the lab aims for full participation of women, persons with disabilities, and underrepresented minorities in STEM at all levels. The project funds research experiences for undergraduates (REU) for one underrepresented minority student during each summer. The REU funded by this project provides hands-on experience in laboratory research by investigating the roles of diverse intermediate filaments in growing axons. In addition, the selected REU student will participate in the institutionally established SRIP (summer research internship program) at the University of Virginia which provides professional skill training and guidance for pursuing a career in STEM in individual and group settings. SRIP has a history of successfully placing participating undergraduate researchers into STEM careers.This project seeks to discover how developmental IF subtype switching from nestin to neurofilaments in neurons affects signaling to the microtubule cytoskeleton to regulate growth cone structure and motility. The PI’s lab recently discovered that the IF protein nestin regulates growth cone morphology. In addition, nestin changes the response of growth cones to the guidance cue Sema3a. It does so by binding the kinase Cdk5 and its substrate, the microtubule associated protein DCX, thereby increasing DCX phosphorylation. As neurons mature, they switch IF subtype from nestin to neurofilaments, which bind DCX but not Cdk5. The project will test the novel hypothesis that neuronal IFs regulate microtubule dynamics and bundling in axonal growth cones via DCX downstream of Sema3a signaling. Aim 1 will discover how experimental IF switching affects axonal microtubules and growth cone behavior. Aim 2 will discover how experimental IF switching affects Sema3a responses in cortical neurons. The project will use state-of-the-art imaging approaches in cultured cortical neurons in combination with downregulation and overexpression of different IF subtypes to uncover the mechanisms by which different IF subtypes modulate microtubule dynamics in growth cones during axon outgrowth. Responsiveness to extracellular cues by growth cones during development is influenced not only by the cue available, the receptors expressed on the surface, the intracellular substrates present, but also by the IF subtypes expressed. The PI proposes that neuronal IF subtypes can act as either amplifiers or dampeners of ubiquitous kinase cascades to provide local and cell-type specific regulation of effector responses.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.