Bulletin of the American Physical Society
APS March Meeting 2018
Volume 63, Number 1
Monday–Friday, March 5–9, 2018; Los Angeles, California
Session F51: Self Organization in the Cytoskeleton IFocus
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Sponsoring Units: DBIO GSOFT Chair: M. Betterton, Univ of Colorado - Boulder Room: LACC 511C |
Tuesday, March 6, 2018 11:15AM - 11:27AM |
F51.00001: Reconstitution of aster movement and cell division plane positioning in Xenopus egg extract James Pelletier, Christine Field, John Oakey, Jay Gatlin, Nikta Fakhri, Timothy Mitchison Microtubule (MT) asters are a self-organizing network of short, dynamic microtubules oriented radially outward from a microtubule organizing center (MTOC). To facilitate cell division, asters must grow and move through a crowded cytoplasm. Aster movement is thought to be driven by MT-length-dependent forces from cytoplasmic dynein opposed by hydrodynamic drag; however, it remains unclear how these forces propagate through the network of short, and short-lived, microtubules, resulting in aster movement. We reconstituted aster movement in cytoplasmic extract from Xenopus laevis frog eggs, and we imaged the growth and interaction of asters under slit-like confinement. Aster boundaries stopped growing when they interacted with neighboring asters. Boundaries between asters formed dynamic, polygonal tessellations that resembled 2D foams. MTOCs moved within asters relative to boundaries and centroids. This movement was partially inhibited by the CC1 fragment of dynactin, which inhibits dynein-dependent forces. We are modelling the dynamic mechanical properties of asters to probe whether known forces can explain the results. We conclude the short, dynamic nature of microtubules facilitates aster growth and movement through a crowded cytoplasm. |
Tuesday, March 6, 2018 11:27AM - 11:39AM |
F51.00002: Coarse-grained simulations elucidate mechanical sorting principles in the actin cytoskeleton Simon Freedman, Glen Hocky, Isabelle Bunge, Aaron Dinner In cells, actin binding proteins (ABPs) segregate to different cytoskeletal networks to perform distinct functions, such as forming filopodaie to enable cell migration, or polarizing actin bundles to enable cell division. Recent experimental work has found that mechanical principles may drive this segregation. For example, the binding of a short crosslinker protein to two actin filaments may promote the binding of other short crosslinkers, and inhibit the binding of longer crosslinkers due to the actin’s bending rigidity. We use coarse grained simulation to measure the magnitude of sorting as a function of the mechanical characteristics of actin and ABPs, and predict the energetic competition associated with this sorting mechanism. We also use simulation to determine actin network characteristics that yield material separation through emergent surface tension, such as in actin liquid crystals. Our work can therefore be used to engineer self-sorting biomimetic materials and understand how the heterogeneous distribution of molecules in cells drives biological processes. |
Tuesday, March 6, 2018 11:39AM - 11:51AM |
F51.00003: Mechanistic Basis of Spindle Size Control and Scaling Reza Farhadifar, Michael Shelley, Daniel Needleman The size and morphology of intracellular structures such as the nucleus, Golgi apparatus, and mitotic spindle dramatically vary between different cell types, yet the mechanisms that regulate the size of these structures are not understood. Interestingly, the size of most intracellular structures scales with cell size, i.e. larger cells tend to have larger nucleus and spindles. So far, many models have been proposed to explain such scaling behavior, but rigorous testing of these models inside the cells is challenging, and often not feasible. To overcome this challenge, we combined the statistical framework of quantitative genetics, with cell biology and biophysics to develop a general methodology to quantitatively examine different models of spindle size control and scaling for the first mitotic spindle in C. elegans. We also use laser ablation technique to quantitatively measure changes in forces under different genetic perturbations. The combination of quantitative genetics with cell biology and biophysics provides a systematic and unbiased method to study mechanisms that contribute to size regulation of intracellular structure and also will give us a deeper understanding of the evolution of these structures. |
Tuesday, March 6, 2018 11:51AM - 12:03PM |
F51.00004: Active Self-Organization of a Composite Actin and Microtubule Gliding System. Leila Farhadi, Carline Do Rosario Fermino, Jennifer Ross Active matter is the field that studies non equilibrium system in different scales, from schools of fish to cytoskeleton of living cells. Playing an important role in cell functions, microtubules and actin are two types of filaments in cytoskeleton that interact with their associated motor proteins, myosin and kinesin respectively. Prior work has inspected with phase diagram for active filaments of either type of cytoskeletal filament separately. Here, we study both filaments in a composite active system where both kinesin-1 and myosin are adhered to the cover glass to create gliding filaments. We observe that the isotropic to aligned phase transition is almost coincident for actin and microtubules. |
Tuesday, March 6, 2018 12:03PM - 12:15PM |
F51.00005: Model of mitotic spindle self-assembly without motor proteins Adam Lamson, Christopher Edelmaier, Matthew Glaser, M. Betterton The microtubule-based mitotic spindle segregates chromosomes during eukaryotic cell division. Assembly of the mitotic spindle is an out-of-equilibrium process typically requiring interaction between dynamic microtubules (MTs), MT-associated proteins, and MT organizing centers. We study mitotic spindle formation in the fission yeast S. Pombe because of its relatively small number of components. Surprisingly, spindle formation in S. Pombe requires no force generation from motor proteins; in the absence of motors, the crosslinking molecule Ase1/PRC1 drives spindle assembly. However, it is not clear what physical properties of Ase1 are necessary for spindle bipolarity. Our coarse-grained model demonstrates formation of stable bipolar spindles with only crosslinkers present. By varying properties of our simulated Ase1 molecules, we find specific molecular features required for spindle assembly. In particular, Ase1 length, binding kinetics, and MT-bound diffusion coefficient control its ability to trap MT antiparallel overlaps and form a bipolar spindle. Our results illustrate how molecular features of a MT bundler promote self-organization and stability of the mitotic spindle. |
Tuesday, March 6, 2018 12:15PM - 12:51PM |
F51.00006: Microtubule cryptography: the effects of tubulin diversity on polymer structure, dynamics and readout by cellular effectors Invited Speaker: Antonina Roll-Mecak Microtubules are essential non-covalent polymers composed of -tubulin subunits. Deceptively uniform ultrastructurally, microtubules are mosaic and contain multiple tubulin isoforms functionalized with abundant posttranslational modifications. Tubulin isoforms and posttranslational modifications vary widely between cell types and their patterns are stereotyped, suggesting roles in spatial and temporal control. An increasing body of evidence supports the hypothesis that the combinatorial information expressed through tubulin genetic and chemical diversity controls microtubule dynamics, mechanics and interactions with microtubule effectors and thus constitutes a “tubulin code”. Although discovered over thirty years ago, a mechanistic understanding of the tubulin code has remained elusive. I will present two studies from my lab that illustrate how the tubulin code can regulate microtubule properties in cis, by directly affecting microtubule structure and dynamic instability, and in trans, by precisely controlling the activity of a microtubule effector, a microtubule severing enzyme. (1) I will discuss our recent data that reveals how varying tubulin isoform composition proportionally tunes microtubule dynamic parameters and (2) I will discuss our data that demonstrates that glutamylation, a posttranslational modification that involves the addition of variable numbers of glutamates to the intrinsically disordered tubulin C-terminal tails acts as a rheostat and tunes microtubule severing as a function of glutamate number added per tubulin. Unexpectedly, glutamylation is a non-linear biphasic tuner and becomes inhibitory beyond a threshold. Furthermore, the inhibitory effect of localized glutamylation propagates across neighboring microtubules, modulating severing in trans. This work provided the first quantitative evidence for a graded, spatially controlled response to a tubulin posttranslational modification and constitutes an essential step towards understanding how the cell interprets the tubulin code. |
Tuesday, March 6, 2018 12:51PM - 1:03PM |
F51.00007: Effects of tubulin post-translational modification on microtubule bending rigidity and C-terminal tails Kathryn Wall, Tanner Bobak, Scott Tilden, Taviare Hawkins, Loren Hough Microtubules are cytoskeletal filaments important for cellular processes including cell division, intracellular transport, and cellular movement. Tubulin is the heterodimeric building block of microtubules. Regulation of tubulin in the cell occurs primarily through post-translational modifications on the C-terminal tails. The intrinsically disordered C-terminal tails are involved in mediating binding interactions with tubulin and affecting larger scale microtubule dynamics. However, the means by which a small, flexible domain can have these large effects is unknown. We probe atomic level behavior of the C-terminal tails by NMR (Wall et al ACS Chem Bio 2016). In parallel, we used fluorescence microscopy to study microtubule thermal fluctuations, allowing us to determine the flexural rigidity of microtubules assembled from distinct tubulin pools. |
Tuesday, March 6, 2018 1:03PM - 1:15PM |
F51.00008: 3D modeling of cargo motility on microtubule networks Jared Bergman, Matthew Bovyn, Manasa Gudheti, Steven Gross, Jun Allard, Michael Vershinin Microtubules tend to form complex 3D networks in many cells. Experimentally modeling full 3D network complexity has been a challenge to date because microtubule networks are rather sparse and structured and often do not form a true hydrogel. We report a novel approach to creating 3D filament networks in vitro using holographic optical trapping and we further report the first experimental study of cargo navigation across a minimal 3D microtubule network under controlled conditions. We will discuss a theoretical model which captures the key experimental observations and the avenues for future research. |
Tuesday, March 6, 2018 1:15PM - 1:27PM |
F51.00009: Combining Computational Fluid Dynamics and Electron Tomography to Study the Mechanics of Kinetochore Microtubules Ehssan Nazockdast, Stefanie Redemann, Sebastian Fürthauer, Thomas Mueller-Reichert, Michael Shelley The accurate segregation of chromosomes, and subsequent cell division, in Eukaryotic cells is achieved by the interactions of an assembly of microtubules (MTs) and motor-proteins, known as the mitotic spindle. We use a combination of our computational platform for simulating cytoskeletal assemblies and our structural data from high-resolution electron tomography of the mitotic spindle, to study the kinetics and mechanics of MTs in the spindle, and their interactions with chromosomes during chromosome segregation in the first cell division in C.elegans embryo. We focus on kinetochore MTs, or KMTs, which have one end attached to a chromosome. KMTs are thought to be a key mechanical component in chromosome segregation. Using exploratory simulations of MT growth, bending, hydrodynamic interactions, and attachment to chromosomes, we propose a mechanical model for KMT-chromosome interactions that reproduces observed KMT length and shape distributions from electron tomography. We find that including detailed hydrodynamic interactions between KMTs is essential for agreement with the experimental observations. |
Tuesday, March 6, 2018 1:27PM - 1:39PM |
F51.00010: Density fields for branched cytoskeletal networks in confined regions Somiealo Azote, Kristian K. Müller-Nedebock
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Tuesday, March 6, 2018 1:39PM - 1:51PM |
F51.00011: Modeling F-actin Disassembly by Severing and its Effect on Stress Relaxation Sadjad Arzash, Jingchen Feng, Patrick McCall, Margaret Gardel, Fred MacKintosh Actin filaments (F-actin), ubiquitous proteins biopolymers in cells, form networks that are crucial to cell mechanics. These biopolymers not only polymerize and depolymerize from their ends but also can be severed by actin severing proteins such as Cofilin. In this work, we develop a model for the dynamics of actin filament polymerization, depolymerization, and severing. This enables us to predict stress relaxation behavior of such complex, nonequilibrium networks. We identify various regimes with distinct length-dependent relaxation behavior, including both a regime with length-independent relaxation rate, as well as a regime with a relaxation rate that increases with filament length. |
Tuesday, March 6, 2018 1:51PM - 2:03PM |
F51.00012: Substrate Mobility Produces Velocity Time Dependence in Microtubule Gliding Joseph Lopes, David Quint, Dail Chapman, Melissa Xu, Ajay Gopinathan, Linda Hirst, Jing Xu Molecular motor based transport is critical for all eukaryotic cell function. Motors often work in small teams to transport a cargo in-vivo, however understanding the factors that control and regulate the group function of multiple motors bound to a lipid membrane remains a challenge. Here we couple kinesin motors to a lipid bilayer, utilizing the microtubule gliding assay, recording and analyzing gliding velocity as a function of time. We observe a constant gliding velocity on glass that is characteristic of solid substrates, while gliding on membrane resulted in a larger than two-fold increase in velocity. When microtubules are immobilized in the absence of ATP, microtubule-bound motors are observed to build up over time. We propose an analytical model relating time dependent motor protein density to microtubule velocity, giving us a motor disassociation rate and a mechanism for the observed velocity increase. Our results provide evidence that motors coupled to a fluid-like membrane exhibit significantly different gliding behavior than observed on rigid substrates such as glass and hypothesize that motor diffusion in the membrane may play a role in biological transport processes. |
Tuesday, March 6, 2018 2:03PM - 2:15PM |
F51.00013: Coupled Shear-bending Effects on Energy Dissipation in Thermally Fluctuating Biofilaments Ameneh Maghsoodi, Noel Perkins Thermally fluctuating biofilaments including microtubules, chromosomes, and actin filaments exhibit energy dissipation from internal friction associated with viscoelastic conformational changes or fluid flow through internal filament pores, as well as external friction from hydrodynamic drag. Poirier and Marko (2002) introduced a physical model of the internal friction for thermally fluctuating biofilaments based on Langevin formulation of Euler-Bernoulli beam theory that considers deformation only due to bending. In this study, we present an extended model based on Timoshenko beam theory that explicitly considers the effects of shear deformation in addition to bending on both internal and external dissipation mechanisms. Our results reveal that shear deformation leads to dissipation dynamics on two time scales associated with internal friction (in lieu of a single time scale predicted by bending alone) and two length scales associated with external friction. This extended model successfully describes the experimental behavior for thermally fluctuating chromosomes and microtubules, and consequently, yields the superior estimates of energy dissipation deriving from internal and external frictions. |
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