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Optogenetics Tool Brings Dynamics of Cellular and Tissue Mechanics to Light

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OKAZAKI, Japan, Jan. 10, 2022 — A research team at the National Institute for Basic Biology (NIBB) developed an optogenetic tool that can reduce cellular contractile force to better understand how contractile forces generated by cells — those that affect an array of biological processes, including cell motility, cytokinesis, and tissue morphogenesis — influence cell dynamics. The tool, called OptoMYPT, uses blue light to induce relaxation of actomyosin contractility at the subcellular level; it inactivates nonmuscle myosin II, an actin-binding protein that generates cellular contractility in coordination with actin filaments.

The researchers found that blue-light illumination via OptoMYPT was sufficient to cause a decrease in actomyosin contractile force in mammalian cells and Xenopus embryos.

According to Kazuhiro Aoki, a professor at NIBB, the team believes that the tool will be useful for understanding various embryological and cell biological phenomena involving the actomyosin cytoskeleton.

“In the future, we expect that it can be used for freely designing the shapes of cells and tissues and for forming artificial organs,” Aoki said.

The researchers focused on myosin phosphatase target subunit 1 (MYPT1), a protein required for myosin inactivation, for the development of OptoMYPT. MYPT1 brings protein phosphatase 1c (PP1c) in close proximity to phosphorylated myosin, which results in dephosphorylation and inactivation of myosin.

OptoMYPT uses the PP1c binding domain of MYPT1 to optically manipulate the localization of PP1c originally existing in cells.

Design of the OptoMYPT system (top). Blue light exposure induces translocation of SspB-PP1BD and endogenous PP1c from the cytoplasm to the membrane, leading to the inactivation of myosin near the membrane. The images below show membrane recruitment of SspB-PP1BD upon blue light exposure. Courtesy of Aoki Lab.
Design of the OptoMYPT system (top). Blue light exposure induces translocation of SspB-PP1BD and endogenous PP1c from the cytoplasm to the membrane, leading to the inactivation of myosin near the membrane. The images below show membrane recruitment of SspB-PP1BD upon blue light exposure. Courtesy of Aoki Lab.
The researchers combined a PP1c-binding domain of MYPT1 with an optogenetic dimerizer, to allow light-dependent recruitment of endogenous PP1c to the plasma membrane. They used a photoactivatable protein called iLID (improved Light-Induced Dimer) to control the localization and activity of the proteins with light. Blue light irradiation caused the iLID protein to bind to single-stranded binding proteins (SspB).

“First, the iLID protein is localized to the cell membrane, while SspB fused with PP1BD of MYPT1 is expressed within the cytoplasm,” researcher Kei Yamamoto said. “Blue light exposure then induces the translocation of SspB-PP1BD from the cytoplasm to the membrane through binding to iLID, leading to the co-recruitment of endogenous PP1c to the membrane. Finally, the membrane-recruited PP1c dephosphorylates and inactivates myosin near the cell membrane.”

When the OptoMYPT-expressing cells were exposed to blue light irradiation, PP1c was translocated to the cell membrane, and actin- and myosin-mediated contractile force was reduced.

When the researchers shined blue light on both poles of the dividing cell to weaken the tensile force generated at the cell cortex, the result was an acceleration in the ingression speed of the cell cleavage furrow. When the tension of the cell cortex was weakened on only one side, the researchers further found that an oscillatory cytoplasmic flow occurred between the two daughter cells.

Local light illumination experiments during cell division (left) and its illustrated summary (right). Blue light illumination on both poles accelerated ingression speed (middle). Blue light illumination on the single-pole induced shape oscillation (bottom). Courtesy of Aoki Lab.
Local light illumination experiments during cell division (left) and its illustrated summary (right). Blue light illumination on both poles accelerated ingression speed (middle). Blue light illumination on the single-pole induced shape oscillation (bottom). Courtesy of Aoki Lab.
By applying local force perturbations using OptoMYPT, the research team demonstrated that the optimal strength and symmetry of the forces generated at the cell surface are essential for the normal progression of cell division.

The OptoMYPT system could provide opportunities to better understand the mechanics of morphogenesis and to shape the morphology of cells and tissues with precision and flexibility. By combining red light-responsive optogenetic tools with blue light-responsive tools such as OptoMYPT, it could be possible for scientists to create more sophisticated morphology with an increase or decrease in contractile force in the same cells and tissues.

The research was published in Nature Communications (www.doi.org/10.1038/s41467-021-27458-3).

Photonics.com
Jan 2022
GLOSSARY
optogenetics
A discipline that combines optics and genetics to enable the use of light to stimulate and control cells in living tissue, typically neurons, which have been genetically modified to respond to light. Only the cells that have been modified to include light-sensitive proteins will be under control of the light. The ability to selectively target cells gives researchers precise control. Using light to control the excitation, inhibition and signaling pathways of specific cells or groups of...
plasma
A gas made up of electrons and ions.
Biophotonicscell analysisoptogeneticsbiomedical imagingAsia PacificResearch & Technologyeducationcell morphologycell mechanicsplasmaflowbiofluorescencedimerizationphotodimerizationNational Institute for Basic Biologymolecularproteincytokinesisphotoactivation

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