Imaging of native transcription factors and histone phosphorylation at high resolution in live cells

Sascha Conic; Dominique Desplancq; Alexia Ferrand; Veronique Fischer; Vincent Heyer; Bernardo Reina San Martin; Julien Pontabry; Mustapha Oulad-Abdelghani; Kishore Babu N.; Graham D. Wright; Nacho Molina; Etienne Weiss; László Tora

doi:10.1083/jcb.201709153

DOI: 10.1083/jcb.201709153 | Published February 12, 2018

Graphical Abstract

Abstract

Fluorescent labeling of endogenous proteins for live-cell imaging without exogenous expression of tagged proteins or genetic manipulations has not been routinely possible. We describe a simple versatile antibody-based imaging approach (VANIMA) for the precise localization and tracking of endogenous nuclear factors. Our protocol can be implemented in every laboratory allowing the efficient and nonharmful delivery of organic dye-conjugated antibodies, or antibody fragments, into different metazoan cell types. Live-cell imaging permits following the labeled probes bound to their endogenous targets. By using conventional and super-resolution imaging we show dynamic changes in the distribution of several nuclear transcription factors (i.e., RNA polymerase II or TAF10), and specific phosphorylated histones (γH2AX), upon distinct biological stimuli at the nanometer scale. Hence, considering the large panel of available antibodies and the simplicity of their implementation, VANIMA can be used to uncover novel biological information based on the dynamic behavior of transcription factors or posttranslational modifications in the nucleus of single live cells.

Submitted: 28 September 2017
Revision received 15 December 2017
Accepted: 18 January 2018

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© 2018 Conic et al. http://www.rupress.org/terms/https://creativecommons.org/licenses/by-nc-sa/4.0/This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms/). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/).