A Robust, GFP-Orthogonal Photoswitchable Inhibitor Scaffold Extends Optical Control over the Microtubule Cytoskeleton

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A Robust, GFP-Orthogonal Photoswitchable Inhibitor Scaffold Extends Optical Control over the Microtubule Cytoskeleton
 
Gao, Li; Meiring, Joyce C M; Kraus, Yvonne; Wranik, Maximilian; Weinert, Tobias; Pritzl, Stefanie D; Bingham, Rebekkah; Ntouliou, Evangelia; Jansen, Klara I; Olieric, Natacha; Standfuss, Jörg; Kapitein, Lukas C; Lohmüller, Theobald; Ahlfeld, Julia; Akhmanova, Anna; Steinmetz, Michel O; Thorn-Seshold, Oliver
(2021) Cell Chemical Biology, volume 28, issue 2, pp. 228 - 241.e6
(Article)
Abstract
Photocontrollable reagents have unique potential as high spatiotemporal precision modulators of biological systems. Here, Gao et al. demonstrate a GFP-orthogonal and metabolically stable photoswitch that allows optical control over microtubule dynamics and architecture with subcellular resolution. The photoswitch scaffold also offers new possibilities for photopharmaceutical design against other targets.
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Keywords: antimitotic, azobenzene, cell cycle, cytoskeleton, microtubule dynamics, photochromismism, photopharmacology, photoswitch, spatiotemporal control, tubulin polymerization inhibitor, Taverne, Biochemistry, Molecular Medicine, Molecular Biology, Pharmacology, Drug Discovery, Clinical Biochemistry
DOI: https://doi.org/10.1016/j.chembiol.2020.11.007
ISSN: 2451-9448
Publisher: Elsevier
Note: Funding Information: This research was supported by funds from the German Research Foundation (DFG: SFB1032 Nanoagents for Spatiotemporal Control project B09 to O.T.-S. and project A08 to T.L.; Emmy Noether grant no. 400324123 , SFB TRR 152 project P24 no. 239283807 , and SPP 1926 project no. 426018126 to O.T.-S.); the Swiss National Science Foundation ( 31003A_166608 to M.O.S.); and the Munich Centre for NanoScience initiative (CeNS). J.C.M.M. acknowledges support from an EMBO Long-Term Fellowship. X-ray diffraction data were collected at the beamline X06SA at the Swiss Light Source (Paul Scherrer Institut, Villigen PSI, Switzerland). We thank S. Schmidt (LMU) for help with synthesis; K.T. Wanner (LMU) for collegial discussions; H. Harz, I. Solvei, and C. Jung (LMU microscopy platforms), K. Bartel, A. Vollmar, H. Leonhardt, and S. Zahler (LMU) for access to general biology and microscopy facilities; and D. Hörl (LMU) for measuring microscope laser power. We dedicate this paper to GFP’s discoverer Osamu Shimomura,whose devoted research has made modern chemical biology possible. Funding Information: This research was supported by funds from the German Research Foundation (DFG: SFB1032 Nanoagents for Spatiotemporal Control project B09 to O.T.-S. and project A08 to T.L.; Emmy Noether grant no. 400324123, SFB TRR 152 project P24 no. 239283807, and SPP 1926 project no. 426018126 to O.T.-S.); the Swiss National Science Foundation (31003A_166608 to M.O.S.); and the Munich Centre for NanoScience initiative (CeNS). J.C.M.M. acknowledges support from an EMBO Long-Term Fellowship. X-ray diffraction data were collected at the beamline X06SA at the Swiss Light Source (Paul Scherrer Institut, Villigen PSI, Switzerland). We thank S. Schmidt (LMU) for help with synthesis; K.T. Wanner (LMU) for collegial discussions; H. Harz, I. Solvei, and C. Jung (LMU microscopy platforms), K. Bartel, A. Vollmar, H. Leonhardt, and S. Zahler (LMU) for access to general biology and microscopy facilities; and D. H?rl (LMU) for measuring microscope laser power. We dedicate this paper to GFP's discoverer Osamu Shimomura,whose devoted research has made modern chemical biology possible. L.G. performed synthesis, single-photon photocharacterization, in vitro studies, figure preparation, coordinated data assembly, and wrote the manuscript. J.C.M.M. performed characterization of EB3 dynamics, temporally reversible live-cell imaging studies, and subcellularly localized photoswitching. Y.K. performed in vitro studies. M.W. performed tubulin protein production, purification, crystallization, crystal handling, and X-ray data collection, processing, and refinement. T.W. performed tubulin crystal handling, data collection, and data processing and refinement. S.D.P. performed two-photon photocharacterization. R.B. performed cell-free tubulin polymerization assays. E.N. performed live-cell imaging on A549 cell lines. K.I.J. and L.C.K. performed primary neuron isolation and culture. N.O. performed tubulin protein production, purification, and crystallization. T.L. supervised two-photon photocharacterization. J.S. and M.O.S. supervised protein crystallography. A.A. supervised characterization of EB3 dynamics, temporally reversible cell imaging and subcellularly localized photoswitching. J.A. performed in vitro studies, coordinated data assembly and figure preparation, and supervised all other cell biology. O.T.-S. designed the concept and experiments, supervised all other experiments, coordinated data assembly, and wrote the manuscript with input from all authors. The authors declare no competing interests. Publisher Copyright: © 2020 Elsevier Ltd
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