Bulletin of the American Physical Society
APS March Meeting 2022
Volume 67, Number 3
Monday–Friday, March 14–18, 2022; Chicago
Session D07: Intracellular Transport I: Motor-driven Transport and Cytoskeletal ActivityFocus Recordings Available
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Sponsoring Units: DBIO Chair: Ajay Gopinathan, University of California Merced Room: McCormick Place W-179A |
Monday, March 14, 2022 3:00PM - 3:36PM |
D07.00001: Mechanics of Cytoplasmic Dynein Invited Speaker: Ahmet Yildiz Cytoplasmic dynein is responsible for motility and force generation functions towards the microtubule minus end. In comparison to kinesin and myosin motors, the mechanism of dynein motility was not well understood due to the complexity of its structure and massive size. Using single-molecule imaging methods, we presented a robust mechanistic model of how dynein steps along microtubules and generates force. We reversed the direction of dynein motility by protein engineering, which explained why all dyneins move towards the microtubule minus-end. We have also studied the activation and motility of mammalian dynein-dynactin complexes, and how the dynein transport machinery is regulated by dynein-associated factors, microtubule-associated proteins, and cargo adaptors. These results alter our view on how dynein functions as a motor and transports cargos in cells. |
Monday, March 14, 2022 3:36PM - 3:48PM |
D07.00002: Effects of Motor Mobility in Cargo Transport. Nimisha Krishnan, Jennifer L Ross Long-range intracellular transport is mediated by motor proteins, kinesin and dynein, transporting cargos along a complex microtubule network. Here, we investigate the effects of network complexity and cargo surface properties on a cargos ability to get across a network. We compare single kinesin motor transport ability to these cargos: small cargos (quantum dots) with rigid motor attachments, large cargos (beads) with rigid motor attachments, and large cargos (emulsion droplets) with fluid motor attachments. The network complexity is altered from single microtubules, to microtubule intersections and random networks of filaments. These studies will help us understand how motors can intrinsically navigate complex networks of filamentsin live cells. |
Monday, March 14, 2022 3:48PM - 4:00PM |
D07.00003: VESICLE TRANSPORTATION OF ZYMOGEN GRANULE BY MYOSIN; FORCE MEASURING THE STEPPING BY OPTICAL TWEEZER Takeshi Sakaomto, Takeshi Sakaomto Zymogen granule is an enzymatic vesicle in the pancreas. The surface of the zymogen granules (ZGs) has several different myosin classes, such as myosin Ib, VI, VIIb, and Vc. These molecular motors may contribute ZG’s transportation in alpha-cells. To understand what the molecular motors involved in the vesicle trafficking, we observed the in vitro motility of the purified ZGs from rat pancreas and exam the stepping manner and force that is used by a single beam optical tweezer. The average force and maximum force from myosins on the ZGs are 0.3 ± 0.1 pN and 0.5 ± 0.2 pN, respectively. A typical force of a single molecular motor was a range of 1 pN to 5pN. The average step-size is 35 nm, which coincides with the actin-helical repeat. A stepping traces were detected within the range of force. We expect that each step was exerted by a single motor. However, different myosins may be able to generate force simultaneously. To understand which molecular motor exerted a force on ZGs, myosin inhibitor, PCIP and TIP, were used in this study. Less than 10 mM of and more than 100 mM of PCIP inhibits motor activities of myosin Ic and Vc, respectively, but does not inhibit myosin VI and VIIb. TIP (10~100 mM) inhibits Myosin VI activity, but does not inhibit other myosin's activity. Using only PCIP or TIP or a combination of PCIP/TIP, stepping movement of ZGs by using the optical trap has been observed. Successful results will present about the directionality of the movement as well as stepping kinetics. |
Monday, March 14, 2022 4:00PM - 4:12PM |
D07.00004: Heterogeneity in single motor velocity leads to enhanced velocity of membrane-bound cargoes carried by teams of molecular motors Niranjan Sarpangala, Ajay Gopinathan Intracellular cargoes like vesicles and organelles are observed to be transported by teams of molecular motors. These teams are often composed of motors with heterogeneous single motor properties. But the relevance of heterogeneity on the multi-motor functioning is unclear, especially in the in-vivo context where the motors are coupled through a lipid membrane. Previous experiments have shown that membrane-bound cargoes move with a higher velocity than membrane-free cargoes when carried by teams of motors. This higher velocity is attributed to the asymmetric off rate of motors and recentering of fluid cargoes upon detachment of lagging motors. Here we show using Brownian dynamics simulations that the underlying heterogeneity in single motor velocities is essential for the above mechanism and hence the cargo speedup. We also explore whether there is an optimum motor heterogeneity that maximizes the flux of cargoes. Our results explain previous experimental results with testable predictions for future experiments on membrane-bound cargo transport. |
Monday, March 14, 2022 4:12PM - 4:24PM |
D07.00005: Spiraling galaxies of microtubules Michael J Shelley, Jesse Gatlin, Gokberk Kabacaoglu, ABDULLAH BASHAR SAMI, David Stein Look inside a living cell as it prepares to divide and you will find in it arrays of stiff biopolymers -- microtubules --radiating outwards from mobile nucleating sites called centrosomes. By interacting with cell boundaries and motor-proteins, centrosomal arrays move and position genetic material in the cell. This motion takes place in the fluidic slurry -- cytoplasm -- that fills the cell. Given the complexity of real cells, understanding how centrosomes do their job is difficult. New experiments have created artificial cells enclosing artificial centrosomes that, like their wildtype counterparts, nucleate microtubule arrays and move. These experiments show microtubule arrays stably centered in its cell, arrays spinning like spiral galaxies, and rotating arrays switching back and forth like a washing machine. We recover and organize this complex dynamics in a fluid-structure model of growing microtubules pushing against the cell boundary, and against each other through the surrounding fluid. Analysis of a coarse-grained model shows that the system is controlled by a combination of microtubule density and cell size, and the collective organization of microtubule bending by hydrodynamics. Large-scale simulations show that rotation and oscillations arise from an intricate and surprising interplay between C-shaped and S-shaped microtubule bending modes. |
Monday, March 14, 2022 4:24PM - 4:36PM |
D07.00006: aLENS: towards the cellular-scale simulation of motor-driven cytoskeletal assemblies Wen Yan, Saad Ansari, Adam R Lamson, Matt Glaser, Meredith D Betterton, Michael J Shelley The cytoskeleton – a collection of polymeric filaments, molecular motors, and crosslinkers – is a foundational example of active matter, and in the cell assembles into organelles that guide basic biological functions. Simulation of cytoskeletal assemblies is an important tool for modeling cellular processes and understanding their surprising material properties. Here we present aLENS, a novel computational framework to surmount the limits of conventional simulation methods. We model molecular motors with crosslinking kinetics that adhere to a thermodynamic energy landscape, and integrate the system dynamics while efficiently and stably enforcing hard-body repulsion between filaments – molecular potentials are entirely avoided in imposing steric constraints. Utilizing parallel computing, we simulate different mixtures of tens to hundreds of thousands of cytoskeletal filaments and crosslinking motors, recapitulating self-emergent phenomena such as bundle formation and buckling, and elucidating how motor type, thermal fluctuations, internal stresses, and confinement determine the evolution of active matter aggregates. |
Monday, March 14, 2022 4:36PM - 4:48PM |
D07.00007: ATP consumption in an actin-myosin network: Response functions in an NADH Assay Francis M Cavanna Actin-myosin networks are used in a wide variety of cellular tasks, including locomotion, structural reorganization, and cell signaling. These networks consist of actin biopolymers along which myosin motors bind to and step along the actin filaments. When these actin-myosin networks are crosslinked, they typically shrink uniformly to a small cluster. Using video analysis, we extract from these networks a strain rate: ε. We also couple the actin-myosin motor to a NADH fluorescence assay and compute the energy consumption density e from the decrease in NADH concentration. Dividing the strain rate by the energy consumption density gives a response function, which we call contraction efficiency, and allows a description of the contraction kinetics of the active gel. We present the regimes of contraction, as well as identifying the optimal conditions to maximize contraction efficiency in a freely contracting actin network. |
Monday, March 14, 2022 4:48PM - 5:00PM |
D07.00008: Modeling the collective motion of microtubules driven by mobile kinesin motors Madhuvanthi Athani, Fabian Jan Schwarzendahl, Fereshteh L Memarian, Ajay Gopinathan, Linda S Hirst, Kinjal Dasbiswas, Daniel A Beller The microtubule gliding assay is a commonly used in-vitro technique to study the collective motion of microtubules as an active fluid. When computationally modeling the movement of microtubules, a spatially uniform distribution of the motor proteins is usually assumed which represents the fixed motor positions of the experiments. Inspired by recent experiments on lipid substrates, we explore a modified setup where kinesin motors are mobile and thus activity varies spatiotemporally. We use Brownian Dynamics simulations to study the collective motions of microtubules, modeled as flexible chains of particles that "self-propel" only in the vicinity of diffusing motors, which we also simulate explicitly. With these simulations, we probe the feedback between the microtubules' active phase behavior and the motors' dynamically reconfiguring activity field. We control the degree of overlap of microtubules using a modified Weeks–Chandler–Anderson potential and show that depending on the degree of overlap and the flexibility of the chains, our model predicts that gliding assays transition between an active polar state and an active nematic state. We also explore how the active phase behavior changes with the motors' diffusion constant. |
Monday, March 14, 2022 5:00PM - 5:12PM |
D07.00009: Effect of Microtubule Length on Topological Defect Dynamics and Morphology in Active Nematics Derek A Hammar, Fereshteh L Memarian, Linda S Hirst Microtubules are dynamic polymers of tubulin subunits that comprise a major transport system of cells and aid in many cellular processes in conjunction with the kinesin motors that traverse the length of the microtubules. In-vivo, these motors "walk" along the tube usually carrying materials for transport. In-vitro gliding assay experiments bind one end of the motor to a substrate, allowing the microtubules to glide along a surface of motor proteins. In our work, instead the motor proteins functions as crosslinkers, with two heads connecting two microtubules instead of being anchored to a substrate. It has previously been shown that stabilized microtubules form a dynamic system in the presence of kinesin motors and ATP. Here the active matter is confined to a 2D oil-water interface and the kinesin motors promote the creation and annihilation of topological defects in the active nematic. Using fluorescence microscopy, we investigate the self-organization of GMPCPP stabilized microtubules into patterns in active nematic, looking to see how the resulting defect dynamics and morphologies vary as the length of microtubules is increased from 1 to 10 micrometers. Video image analysis allows us to measure Vrms and active length scale as a function of MT length. |
Monday, March 14, 2022 5:12PM - 5:24PM |
D07.00010: The Mammalian Meiotic Spindle: A Living Material Colm P Kelleher, Marta Venturas, Daniel J Needleman Meiosis is the specialized form of cell division that creates gametes. During meiosis, genetic material from the "mother" cell is copied and divided between several "daughter" cells. To facilitate this task, the mother cell builds a structure, the meiotic spindle, that organizes and moves chromosomes over length-scales of tens of microns and time-scales of several hours. While we have a (nearly) complete "parts list" of the dozens of bio-molecules that make up the spindle, it is far from clear how these molecules self-organize to into structures with the mechanical properties that allow the spindle to produce forces and transmit them to chromosomes over the required length- and time-scales. In this talk, I will present data and analysis that suggests that many aspects of the large-scale structure and long-time dynamics of meiotic spindles in living cells can be understood via a relatively simple continuum picture in which the spindle is modeled as an active liquid crystal. |
Monday, March 14, 2022 5:24PM - 5:36PM |
D07.00011: Self-organization of microtubules and motors drive large-scale intracellular flows in cells Reza Farhadifar, Sayantan Dutta, Gokberk Kabacaoglu, Wen Lu, Vladimir I Gelfand, Stanislav Y Shvartsman, Michael J Shelley Cytoplasmic streaming is essential for transporting and mixing nutrients, proteins, and organelles within large plant and animal cells. The large ~200um Drosophila oocyte has recently gained attention for experimental and theoretical studies of this phenomenon. We present a quantitative study of streaming in Drosophila oocytes that combines PIV of 3D time-lapse movies, with biophysical modeling and simulation. We observe a diverse family of 3D vortical flows across different oocytes, which differ in position and orientation, and which last tens of minutes. We show that a model of cytoskeletal activity at the periphery, organized by its interaction with interior fluid, explains the observed streaming structures. The emerging picture sheds light on a class of intracellular flows in large cells and highlights the wealth of questions at the interface of geometry, active matter, and basic biology. |
Monday, March 14, 2022 5:36PM - 5:48PM |
D07.00012: Steady-state directional flow in active model cytoskeletal network Jianguo Zhao, Charlie Duclut, Rahil Golipour, An Pham, Behzad Golshaei, Chonglin Guan, Mingru Li, Rudolf Oldenbourg, James L Harden, Frank Jülicher, Christoph F Schmidt Mesoscopic flow of cytoskeletal actin networks in eukaryotic cells, driven by active motor processes, is crucial in a wide variety of cellular dynamics, including intracellular transport, positioning of nuclei, cell migration and division. It remains unclear how such collective dynamics, involving subtly balanced spatiotemporal interactions of many molecular components with transient networks of polymeric actin are regulated and maintained in steady states. We here use a model system of water-in-oil emulsion droplets composed of Xenopus egg extract that contains all the ingredients for active cytoskeletal assembly. We observe conspicuous 3D radially convergent stationary flow patterns of F-actin networks driven by non-muscle myosin motors. Actin intensity and velocity profiles maintain steady-state gradients from the droplet periphery to its center while actin constantly polymerizes and depolymerizes. The contracting actin network drives a mechanical phase separation and forms of a central inclusion. In order to elucidate the mechanisms that lead to the observed patterns and to a precise and stable positioning of the steady-state inclusion in the center of the droplet, we model the network as an isotropic active gel. This model allow us to compute the steady-state actin velocity and concentration profiles as well as the magnitude of the centering force acting on the inclusion. |
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