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Structured illumination to spatially map chromatin motions

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abstract
We describe a simple optical method that creates structured illumination of a photoactivatable probe and apply this method to characterize chromatin motions in nuclei of live cells. A laser beam coupled to a diffractive optical element at the back focal plane of an excitation objective generates an array of near diffraction-limited beamlets with FWHM of 340  ±  30  nm, which simultaneously photoactivate a 7  ×  7 matrix pattern of GFP-labeled histones, with spots 1.70  μm apart. From the movements of the photoactivated spots, we map chromatin diffusion coefficients at multiple microdomains of the cell nucleus. The results show correlated motions of nearest chromatin microdomain neighbors, whereas chromatin movements are uncorrelated at the global scale of the nucleus. The method also reveals a DNA damage-dependent decrease in chromatin diffusion. The diffractive optical element instrumentation can be easily and cheaply implemented on commercial inverted fluorescence microscopes to analyze adherent cell culture models. A protocol to measure chromatin motions in nonadherent human hematopoietic stem and progenitor cells is also described. We anticipate that the method will contribute to the identification of the mechanisms regulating chromatin mobility, which influences most genomic processes and may underlie the biogenesis of genomic translocations associated with hematologic malignancies.
subject
diffractive optical elements
diffusion
microscopes
luminescence
cancer
contributor
Bonin, K. (author)
Smelser, A. (author)
Salvador, N. (author)
Holzwarth, G. (author)
Wang, K. (author)
Levy, P. (author)
Vidi, P. (author)
date
2020-03-09T16:04:05Z (accessioned)
2020-03-09T16:04:05Z (available)
5/15/18 (issued)
identifier
Keith D. Bonin, Amanda Smelser, Naike Salvador Moreno, George Holzwarth, Kevin Wang, Preston Levy, Pierre-Alexandre Vidi, "Structured illumination to spatially map chromatin motions," J. Biomed. Opt. 23(5) 056007 (15 May 2018) (citation)
https://doi.org/10.1117/1.JBO.23.5.056007 (doi)
http://hdl.handle.net/10339/96042 (uri)
language
en (iso)
publisher
SPIE
rights
https://creativecommons.org/licenses/by/4.0/ (uri)
source
Journal of Biomedical Optics
title
Structured illumination to spatially map chromatin motions
type
Article

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