Optical control of fast and processive engineered myosins in vitro and in living cells

Ruijgrok, Paul V.; Ghosh, Rajarshi P.; Zemsky, Sasha; Nakamura, Morimasa; Gong, Rui; Lin, Ning; Chen, Robert; Vachharajani, Vipul T.; Chu, Alexander E.; Anand, Namrata; Eguchi, Raphael R.; Huang, Po-Ssu; Lin, Michael Z.; Alushin, Gregory M.; Liphardt, Jan; Bryant, Zev

doi:10.1038/s41589-021-00740-7

Cited by 21 publications

(33 citation statements)

References 60 publications

(98 reference statements)

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“…The velocity of Cc XI was then estimated from the velocity measured using these constructs. The estimated velocity of Cc XI was about 20 µm/s −1 or less at 25 °C ( 10 , 22 – 26 ), which is less than about one-third of the velocity of cytoplasmic streaming observed in Chara cells. Three possibilities have been suggested as to why the velocities of Cc XI obtained using the recombinant constructs were different from that expected from cytoplasmic streaming ( 1 ).…”

mentioning

confidence: 72%

“…The amino acid sequences of Cb XI-1 MD and Cc XI MD are similar (identity and similarity are 65% and 89%, respectively). Thus, we built a homology model of acto- Cb XI-1 rigor structure based on the recently reported acto- Cc XI rigor structure (PDB: 7KCH) ( 26 ). We then compared the actin-binding modes of six myosins ( Cb XI-1, Cc XI, NM2c, Myosin VI, Myosin 1b, and Pf MyoA) using acto-myosin rigor structures ( SI Appendix , Fig.…”

Section: Resultsmentioning

confidence: 99%

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Discovery of ultrafast myosin, its amino acid sequence, and structural features

Haraguchi

Tamanaha

Suzuki

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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Cytoplasmic streaming with extremely high velocity (∼70 μm s−1) occurs in cells of the characean algae (Chara). Because cytoplasmic streaming is caused by myosin XI, it has been suggested that a myosin XI with a velocity of 70 μm s−1, the fastest myosin measured so far, exists in Chara cells. However, the velocity of the previously cloned Chara corallina myosin XI (CcXI) was about 20 μm s−1, one-third of the cytoplasmic streaming velocity in Chara. Recently, the genome sequence of Chara braunii has been published, revealing that this alga has four myosin XI genes. We cloned these four myosin XI (CbXI-1, 2, 3, and 4) and measured their velocities. While the velocities of CbXI-3 and CbXI-4 motor domains (MDs) were similar to that of CcXI MD, the velocities of CbXI-1 and CbXI-2 MDs were 3.2 times and 2.8 times faster than that of CcXI MD, respectively. The velocity of chimeric CbXI-1, a functional, full-length CbXI-1 construct, was 60 μm s−1. These results suggest that CbXI-1 and CbXI-2 would be the main contributors to cytoplasmic streaming in Chara cells and show that these myosins are ultrafast myosins with a velocity 10 times faster than fast skeletal muscle myosins in animals. We also report an atomic structure (2.8-Å resolution) of myosin XI using X-ray crystallography. Based on this crystal structure and the recently published cryo-electron microscopy structure of acto-myosin XI at low resolution (4.3-Å), it appears that the actin-binding region contributes to the fast movement of Chara myosin XI. Mutation experiments of actin-binding surface loops support this hypothesis.

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mentioning

confidence: 72%

Section: Resultsmentioning

confidence: 99%

Discovery of ultrafast myosin, its amino acid sequence, and structural features

Haraguchi

Tamanaha

Suzuki

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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“…The synthetic motor used for the experiment in Fig. 4 E and F is MyLOVChar4∼1R∼TET (71), referred to in this study as "gear-shifting" motor, which is the same construct used for experiments in ref. 27 including the data from that study analyzed in Fig.…”

Section: Experimental Methodsmentioning

confidence: 99%

“…The fixed-gear and gear-shifting motors were purified, flash frozen, and stored at -80 • C as described in ref. 71. Timevarying activity movies were stimulated over the entire sample via a 491-nm solid-state laser, while spatially varying activation was achieved by targeting a 470-nm light-emitting diode to one location in the sample using a digital micromirror array (27).…”

Section: Experimental Methodsmentioning

confidence: 99%

Machine learning active-nematic hydrodynamics

Colen

Han

Zhang

et al. 2021

Proc. Natl. Acad. Sci. U.S.A.

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Hydrodynamic theories effectively describe many-body systems out of equilibrium in terms of a few macroscopic parameters. However, such parameters are difficult to determine from microscopic information. Seldom is this challenge more apparent than in active matter, where the hydrodynamic parameters are in fact fields that encode the distribution of energy-injecting microscopic components. Here, we use active nematics to demonstrate that neural networks can map out the spatiotemporal variation of multiple hydrodynamic parameters and forecast the chaotic dynamics of these systems. We analyze biofilament/molecular-motor experiments with microtubule/kinesin and actin/myosin complexes as computer vision problems. Our algorithms can determine how activity and elastic moduli change as a function of space and time, as well as adenosine triphosphate (ATP) or motor concentration. The only input needed is the orientation of the biofilaments and not the coupled velocity field which is harder to access in experiments. We can also forecast the evolution of these chaotic many-body systems solely from image sequences of their past using a combination of autoencoders and recurrent neural networks with residual architecture. In realistic experimental setups for which the initial conditions are not perfectly known, our physics-inspired machine-learning algorithms can surpass deterministic simulations. Our study paves the way for artificial-intelligence characterization and control of coupled chaotic fields in diverse physical and biological systems, even in the absence of knowledge of the underlying dynamics.

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“…Because apical constriction occurs in specific stages and areas of the developing embryo, reconstituting curved tissues requires tools to control cellular contractility in space and time. Optogenetics is a powerful methodology to gain spatiotemporal control of biological processes from the molecular to the multicellular level [9][10][11][12][13][14][15][16][17] . Izquierdo and colleagues employed optogenetics to recruit RhoGEF to the plasma membrane in Drosophila embryos.…”

Section: Introductionmentioning

confidence: 99%

Optogenetic control of apical constriction induces synthetic morphogenesis in mammalian tissues

Martínez-Ara

Taberner

Takayama

et al. 2021

Preprint

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During embryonic development, cellular forces synchronize in space and time to generate functional tissue shapes. Apical constriction is one of these force-generating processes, and it is necessary to modulate epithelial curvature in fundamental morphogenetic events, such as neural tube folding. The emerging field of synthetic developmental biology proposes bottom-up approaches to examine the contribution of each cellular process to complex morphogenesis. However, the shortage of tools to manipulate three-dimensional (3D) shapes of mammalian tissues currently hinders the progress of the field. Here we report the development of 'OptoShroom3', a new optogenetic tool that achieves fast spatiotemporal control of apical constriction in mammalian epithelia. Activation of OptoShroom3 through illumination of individual cells in an epithelial cell sheet reduced their apical surface while illumination of groups of cells caused deformation in the adjacent regions. By using OptoShroom3, we further manipulated 3D tissue shapes. Light-induced apical constriction provoked the folding of epithelial cell colonies on soft gels. Its application to murine and human neural organoids led to thickening of neuroepithelia, apical lumen reduction in optic vesicles, and flattening in neuroectodermal tissues. These results show that spatiotemporal control of apical constriction can trigger several types of 3D deformation depending on the initial tissue context.

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Optical control of fast and processive engineered myosins in vitro and in living cells

Cited by 21 publications

References 60 publications

Discovery of ultrafast myosin, its amino acid sequence, and structural features

Discovery of ultrafast myosin, its amino acid sequence, and structural features

Machine learning active-nematic hydrodynamics

Optogenetic control of apical constriction induces synthetic morphogenesis in mammalian tissues

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