Chai Y, Tian D, Li W, Ou G
Tsinghua University, Beijing, China.
Methods Cell Biol. 2017;137:253-266. doi: 10.1016/bs.mcb.2016.03.040. Epub 2016 Jun 1.
Cytokinesis requires the interplay between the cytoskeleton and plasma membrane. Emerging evidence indicates that some cytokinetic components are essential for the postcytokinetic events such as epithelium organization and neural development. We have recently developed live cell imaging and conditional knockout techniques to visualize cytokinetic proteins in Caenorhabditis elegans Q neuroblasts and separate their postcytokinetic functions from cytokinetic ones. Here we describe how the fluorescent reporter strains and conditional knockout C. elegans are generated and how live cell imaging of Q neuroblast development are performed in our laboratory. Using these protocols, we uncovered a novel role of Anillin in stabilizing the actin network during neuronal migration and neurite outgrowth, and the postcytokinetic fate of midbody, which is released into the extracellular space and degraded by the adjacent macrophage using an apoptotic mimicry. These protocols could also be applicable to study other cellular processes in C. elegans or adapted to other model organisms, to provide better insights into their developmental basis.
胞质分裂需要细胞骨架和质膜之间的相互作用。新出现的证据表明，一些胞质分裂成分对于胞质分裂后的事件（如上皮组织形成和神经发育）至关重要。我们最近开发了活细胞成像和条件性基因敲除技术，以可视化秀丽隐杆线虫Q神经母细胞中的胞质分裂蛋白，并将它们的胞质分裂后功能与胞质分裂功能区分开来。在这里，我们描述了荧光报告菌株和条件性基因敲除秀丽隐杆线虫是如何产生的，以及在我们实验室中如何对Q神经母细胞发育进行活细胞成像。使用这些方案，我们发现了膜收缩蛋白在神经元迁移和神经突生长过程中稳定肌动蛋白网络的新作用，以及中体的胞质分裂后命运，中体被释放到细胞外空间，并被相邻的巨噬细胞利用凋亡模拟机制降解。这些方案也可用于研究秀丽隐杆线虫中的其他细胞过程，或适用于其他模式生物，以更好地洞察其发育基础。
