Yu Che-Hang, Langowitz Noah, Wu Hai-Yin, Farhadifar Reza, Brugues Jan, Yoo Tae Yeon, Needleman Daniel
School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts.
School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts.
Biophys J. 2014 Apr 15;106(8):1578-87. doi: 10.1016/j.bpj.2014.03.009.
The spatial organization of microtubule polarity, and the interplay between microtubule polarity and protein localization, is thought to be crucial for spindle assembly, anaphase, and cytokinesis, but these phenomena remain poorly understood, in part due to the difficulty of measuring microtubule polarity in spindles. We develop and implement a method to nonperturbatively and quantitatively measure microtubule polarity throughout spindles using a combination of second-harmonic generation and two-photon fluorescence. We validate this method using computer simulations and by comparison to structural data on spindles obtained from electron tomography and laser ablation. This method should provide a powerful tool for studying spindle organization and function, and may be applicable for investigating microtubule polarity in other systems.
微管极性的空间组织，以及微管极性与蛋白质定位之间的相互作用，被认为对纺锤体组装、后期和胞质分裂至关重要，但这些现象仍知之甚少，部分原因是难以测量纺锤体中的微管极性。我们开发并实施了一种方法，通过结合二次谐波产生和双光子荧光，对整个纺锤体中的微管极性进行无扰动定量测量。我们使用计算机模拟并与通过电子断层扫描和激光烧蚀获得的纺锤体结构数据进行比较，验证了该方法。该方法应为研究纺锤体组织和功能提供一个强大的工具，并且可能适用于研究其他系统中的微管极性。
