Data for: Growth cone advance requires EB1 as revealed by genomic replacement with a light-sensitive variant

A challenge in analyzing dynamic intracellular cell biological processes is the dearth of methodologies that are sufficiently fast and specific to perturb intracellular protein activities. We previously developed a light-sensitive variant of the microtubule plus end-tracking protein EB1 by inserting a blue light-controlled protein dimerization module between functional domains. Here, we describe an advanced method to replace endogenous EB1 with this light-sensitive variant in a single genome editing step, thereby enabling this approach in human induced pluripotent stem cells (hiPSCs) and hiPSC-derived neurons. We demonstrate that acute and local optogenetic EB1 inactivation in developing cortical neurons induces microtubule depolymerization in the growth cone periphery and subsequent neurite retraction. In addition, advancing growth cones are repelled from areas of blue light exposure. These phenotypes were independent of the neuronal EB1 homolog EB3, revealing a direct dynamic role of EB1-mediated microtubule plus end interactions in neuron morphogenesis and neurite guidance.

Methods
This dataset contains raw data and analysis used to produce figures and conclusions associated with bioRxiv/eLife manuscript. Detailed description of the data collection and analysis methodology can be found in the associated manuscript.

解析动态细胞内生物学过程的核心挑战之一，是缺乏足够快速且特异性强的方法，用以扰动细胞内蛋白质的活性。此前我们通过在功能结构域之间插入蓝光调控的蛋白质二聚化模块，开发了微管正端追踪蛋白EB1的光敏感变体。本研究报道一种优化策略，可通过单步基因组编辑将内源性EB1替换为该光敏感变体，从而实现该技术在人类诱导多能干细胞（human induced pluripotent stem cells，hiPSCs）及其诱导分化神经元中的应用。我们证实，在发育中的皮层神经元内实施急性局部光遗传EB1失活，可诱导生长锥外周区域发生微管解聚，继而引发神经突回缩。此外，正向迁移的生长锥会被蓝光照射区域排斥。上述表型不受神经元EB1同源蛋白EB3的影响，由此揭示了EB1介导的微管正端相互作用在神经元形态发生与神经突导向过程中发挥的直接动态调控功能。

方法
本数据集包含用于生成bioRxiv/eLife关联论文中图表与结论的原始数据及分析结果。数据采集与分析方法的详细说明可参阅该关联论文。