Measuring microtubule polarity in spindles with second-harmonic generation

Che-Hang Yu; Noah Langowitz; Hai-Yin Wu; Reza Farhadifar; Jan Brugues; Tae Yeon Yoo; Daniel Needleman

doi:10.1016/j.bpj.2014.03.009

Measuring microtubule polarity in spindles with second-harmonic generation

Biophys J. 2014 Apr 15;106(8):1578-87. doi: 10.1016/j.bpj.2014.03.009.

Authors

Che-Hang Yu¹, Noah Langowitz², Hai-Yin Wu³, Reza Farhadifar⁴, Jan Brugues⁵, Tae Yeon Yoo², Daniel Needleman⁴

Affiliations

¹ School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts. Electronic address: chehangyu@fas.harvard.edu.
² School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts.
³ Department of Physics, Harvard University, Cambridge, Massachusetts.
⁴ School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts; Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts; FAS Center for Systems Biology, Harvard University, Cambridge, Massachusetts.
⁵ School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts; Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts; FAS Center for Systems Biology, Harvard University, Cambridge, Massachusetts; Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.

Abstract

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.

Publication types

Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Animals
Caenorhabditis elegans / cytology
Caenorhabditis elegans / embryology
Caenorhabditis elegans / metabolism
Cell Extracts
Cell Polarity*
Computer Simulation*
Embryo, Nonmammalian / cytology
Embryo, Nonmammalian / metabolism
Microtubules / metabolism*
Spindle Apparatus / metabolism*
Xenopus laevis

Substances

Cell Extracts