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

Li Gao; Joyce C M Meiring; Yvonne Kraus; Maximilian Wranik; Tobias Weinert; Stefanie D Pritzl; Rebekkah Bingham; Evangelia Ntouliou; Klara I Jansen; Natacha Olieric; Jörg Standfuss; Lukas C Kapitein; Theobald Lohmüller; Julia Ahlfeld; Anna Akhmanova; Michel O Steinmetz; Oliver Thorn-Seshold

doi:10.1016/j.chembiol.2020.11.007

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

Li Gao, Joyce C M Meiring, Yvonne Kraus, Maximilian Wranik, Tobias Weinert, Stefanie D Pritzl, Rebekkah Bingham, Evangelia Ntouliou, Klara I Jansen, Natacha Olieric, Jörg Standfuss, Lukas C Kapitein, Theobald Lohmüller, Julia Ahlfeld, Anna Akhmanova, Michel O Steinmetz, Oliver Thorn-Seshold

Department of Pharmacy, Ludwig-Maximilians University of Munich, Munich 81377, Germany.
Paul Scherrer Institute
Chair for Photonics and Optoelectronics, Nano-Institute Munich, Department of Physics, Ludwig-Maximilians University of Munich, Munich 80539, Germany.
Department of Pharmacy, Ludwig-Maximilians University of Munich, Munich 81377, Germany. Electronic address: [email protected].

Research output: Contribution to journal › Article › Academic › peer-review

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.

Original language	English
Pages (from-to)	228-241.e6
Journal	Cell Chemical Biology
Volume	28
Issue number	2
Early online date	3 Dec 2020
DOIs	https://doi.org/10.1016/j.chembiol.2020.11.007
Publication status	Published - 18 Feb 2021

Bibliographical 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

Keywords

antimitotic
azobenzene
cell cycle
cytoskeleton
microtubule dynamics
photochromismism
photopharmacology
photoswitch
spatiotemporal control
tubulin polymerization inhibitor

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10.1016/j.chembiol.2020.11.007

PIIS2451945620304700Final published version, 4.17 MBLicence: Taverne

Cite this

Gao, L., Meiring, J. C. M., Kraus, Y., Wranik, M., Weinert, T., Pritzl, S. D., Bingham, R., Ntouliou, E., Jansen, K. I., Olieric, N., Standfuss, J., Kapitein, L. C., Lohmüller, T., Ahlfeld, J., Akhmanova, A., Steinmetz, M. O., & Thorn-Seshold, O. (2021). A Robust, GFP-Orthogonal Photoswitchable Inhibitor Scaffold Extends Optical Control over the Microtubule Cytoskeleton. Cell Chemical Biology, 28(2), 228-241.e6. https://doi.org/10.1016/j.chembiol.2020.11.007

@article{58f90478d345439e86c89d073c2f4180,

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

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.",

keywords = "antimitotic, azobenzene, cell cycle, cytoskeleton, microtubule dynamics, photochromismism, photopharmacology, photoswitch, spatiotemporal control, tubulin polymerization inhibitor",

author = "Li Gao and Meiring, {Joyce C M} and Yvonne Kraus and Maximilian Wranik and Tobias Weinert and Pritzl, {Stefanie D} and Rebekkah Bingham and Evangelia Ntouliou and Jansen, {Klara I} and Natacha Olieric and J{\"o}rg Standfuss and Kapitein, {Lukas C} and Theobald Lohm{\"u}ller and Julia Ahlfeld and Anna Akhmanova and Steinmetz, {Michel O} and Oliver Thorn-Seshold",

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. H{\"o}rl (LMU) for measuring microscope laser power. We dedicate this paper to GFP{\textquoteright}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: {\textcopyright} 2020 Elsevier Ltd",

year = "2021",

month = feb,

day = "18",

doi = "10.1016/j.chembiol.2020.11.007",

language = "English",

volume = "28",

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Gao, L, Meiring, JCM, Kraus, Y, Wranik, M, Weinert, T, Pritzl, SD, Bingham, R, Ntouliou, E, Jansen, KI, Olieric, N, Standfuss, J, Kapitein, LC, Lohmüller, T, Ahlfeld, J, Akhmanova, A, Steinmetz, MO & Thorn-Seshold, O 2021, 'A Robust, GFP-Orthogonal Photoswitchable Inhibitor Scaffold Extends Optical Control over the Microtubule Cytoskeleton', Cell Chemical Biology, vol. 28, no. 2, pp. 228-241.e6. https://doi.org/10.1016/j.chembiol.2020.11.007

TY - JOUR

T1 - A Robust, GFP-Orthogonal Photoswitchable Inhibitor Scaffold Extends Optical Control over the Microtubule Cytoskeleton

AU - Gao, Li

AU - Meiring, Joyce C M

AU - Kraus, Yvonne

AU - Wranik, Maximilian

AU - Weinert, Tobias

AU - Pritzl, Stefanie D

AU - Bingham, Rebekkah

AU - Ntouliou, Evangelia

AU - Jansen, Klara I

AU - Olieric, Natacha

AU - Standfuss, Jörg

AU - Kapitein, Lukas C

AU - Lohmüller, Theobald

AU - Ahlfeld, Julia

AU - Akhmanova, Anna

AU - Steinmetz, Michel O

AU - Thorn-Seshold, Oliver

N1 - 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

PY - 2021/2/18

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KW - antimitotic

KW - azobenzene

KW - cell cycle

KW - cytoskeleton

KW - microtubule dynamics

KW - photochromismism

KW - photopharmacology

KW - photoswitch

KW - spatiotemporal control

KW - tubulin polymerization inhibitor

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U2 - 10.1016/j.chembiol.2020.11.007

DO - 10.1016/j.chembiol.2020.11.007

M3 - Article

C2 - 33275880

SN - 2451-9448

VL - 28

SP - 228-241.e6

JO - Cell Chemical Biology

JF - Cell Chemical Biology

IS - 2

ER -

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

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Bibliographical note

Keywords

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