LIVE CELLS AS OPEN NON-EQUILIBRIUM SYSTEMS

Kinesin and Myosin Motors Compete to Drive Rich Multi-Phase Dynamics in Programmable Cytoskeletal Composites

10.21203/rs.3.rs-1179494/v1 ◽

2022 ◽

Author(s):

Daisy Achiriloaie ◽

Christopher Currie ◽

Jonathan Michel ◽

Maya Hendija ◽

K Alice Lindsay ◽

...

Keyword(s):

Live Cells ◽

Active Matter ◽

Advective Flow ◽

Phase Dynamics ◽

Equilibrium Dynamics ◽

Single Force ◽

Myosin Motors ◽

Multi Phase ◽

Acting In Concert ◽

Non Equilibrium

Abstract The cytoskeleton of biological cells relies on a diverse population of motors, filaments, and binding proteins acting in concert to enable non-equilibrium processes ranging from mitosis to chemotaxis. The cytoskeleton’s versatile reconfigurability, programmed by interactions between its constituents, make it a foundational active matter platform. However, current active matter endeavors are limited largely to single force-generating components acting on a single substrate – far from the composite cytoskeleton in live cells. Here, we engineer actin-microtubule composites, driven by kinesin and myosin motors and tuned by crosslinkers, that restructure into diverse morphologies from interpenetrating filamentous networks to de-mixed amorphous clusters. Our Fourier analyses reveal that kinesin and myosin compete to delay kinesin-driven restructuring and suppress de-mixing and flow, while crosslinking accelerates reorganization and promotes actin-microtubule correlations. The phase space of non-equilibrium dynamics falls into three broad classes– slow reconfiguration, fast advective flow, and multi-mode ballistic dynamics – with structure-dynamics relations described by the relative contributions of elastic and dissipative responses to motor-generated forces.

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4-dimensional, high-resolution imaging of living cells

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100122484 ◽

1992 ◽

Vol 50 (1) ◽

pp. 416-417

Author(s):

Shinya Inoué

Keyword(s):

Light Microscope ◽

Living Cells ◽

Live Cells ◽

Video Tape ◽

Active Living ◽

High Resolution Imaging ◽

3 Dimensional ◽

Dynamic Architecture ◽

Resolution Imaging ◽

Disk Memory

This paper reports progress of our effort to rapidly capture, and display in time-lapsed mode, the 3-dimensional dynamic architecture of active living cells and developing embryos at the highest resolution of the light microscope. Our approach entails: (A) real-time video tape recording of through-focal, ultrathin optical sections of live cells at the highest resolution of the light microscope; (B) repeat of A at time-lapsed intervals; (C) once each time-lapsed interval, an image at home focus is recorded onto Optical Disk Memory Recorder (OMDR); (D) periods of interest are selected using the OMDR and video tape records; (E) selected stacks of optical sections are converted into plane projections representing different view angles (±4 degrees for stereo view, additional angles when revolving stereos are desired); (F) analysis using A - D.

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Application of analytical electron microscopy to studies of equilibrium and non-equilibrium segregation in materials

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100130705 ◽

1992 ◽

Vol 50 (2) ◽

pp. 1214-1215

Author(s):

Edward A Kenik

Keyword(s):

Electron Microscopy ◽

Analytical Electron Microscopy ◽

Extended Defects ◽

Probe Diameter ◽

Specimen Holder ◽

Analytical Electron ◽

Scanning Transmission ◽

Solute Atoms ◽

Equilibrium Segregation ◽

Non Equilibrium

Segregation of solute atoms to grain boundaries, dislocations, and other extended defects can occur under thermal equilibrium or non-equilibrium conditions, such as quenching, irradiation, or precipitation. Generally, equilibrium segregation is narrow (near monolayer coverage at planar defects), whereas non-equilibrium segregation exhibits profiles of larger spatial extent, associated with diffusion of point defects or solute atoms. Analytical electron microscopy provides tools both to measure the segregation and to characterize the defect at which the segregation occurs. This is especially true of instruments that can achieve fine (<2 nm width), high current probes and as such, provide high spatial resolution analysis and characterization capability. Analysis was performed in a Philips EM400T/FEG operated in the scanning transmission mode with a probe diameter of <2 nm (FWTM). The instrument is equipped with EDAX 9100/70 energy dispersive X-ray spectrometry (EDXS) and Gatan 666 parallel detection electron energy loss spectrometry (PEELS) systems. A double-tilt, liquid-nitrogen-cooled specimen holder was employed for microanalysis in order to minimize contamination under the focussed spot.

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Multi-mode light microscopy of microtubule assembly dynamics and chromosome movement in vivo and In vitro

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100166117 ◽

1996 ◽

Vol 54 ◽

pp. 730-731

Author(s):

E. D. Salmon ◽

J. C. Waters ◽

C. Waterman-Storer

Keyword(s):

Imaging System ◽

Ccd Camera ◽

Transmitted Light ◽

Microtubule Assembly ◽

Live Cells ◽

Optical Efficiency ◽

Multiple Band ◽

Multi Mode

We have developed a multi-mode digital imaging system which acquires images with a cooled CCD camera (Figure 1). A multiple band pass dichromatic mirror and robotically controlled filter wheels provide wavelength selection for epi-fluorescence. Shutters select illumination either by epi-fluorescence or by transmitted light for phase contrast or DIC. Many of our experiments involve investigations of spindle assembly dynamics and chromosome movements in live cells or unfixed reconstituted preparations in vitro in which photodamage and phototoxicity are major concerns. As a consequence, a major factor in the design was optical efficiency: achieving the highest image quality with the least number of illumination photons. This principle applies to both epi-fluorescence and transmitted light imaging modes. In living cells and extracts, microtubules are visualized using X-rhodamine labeled tubulin. Photoactivation of C2CF-fluorescein labeled tubulin is used to locally mark microtubules in studies of microtubule dynamics and translocation. Chromosomes are labeled with DAPI or Hoechst DNA intercalating dyes.

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Characterization of a Motile Cellular Domain Arising During the Amoebo-Flagellate Transformation of Physarum Polycephalum

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s1431927600002440 ◽

1980 ◽

Vol 38 ◽

pp. 552-553

Author(s):

K.I. Pagh ◽

M.R. Adelman

Keyword(s):

Cell Shape ◽

Physarum Polycephalum ◽

Slime Mold ◽

Live Cells ◽

Cellular Domain

Unicellular amoebae of the slime mold Physarum polycephalum undergo marked changes in cell shape and motility during their conversion into flagellate swimming cells (l). To understand the processes underlying motile activities expressed during the amoebo-flagellate transformation, we have undertaken detailed investigations of the organization, formation and functions of subcellular structures or domains of the cell which are hypothesized to play a role in movement. One focus of our studies is on a structure, termed the “ridge” which appears as a flattened extension of the periphery along the length of transforming cells (Fig. 1). Observations of live cells using Nomarski optics reveal two types of movement in this region:propagation of undulations along the length of the ridge and formation and retraction of filopodial projections from its edge. The differing activities appear to be associated with two characteristic morphologies, illustrated in Fig. 1.

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