Bulletin of the American Physical Society
APS March Meeting 2016
Volume 61, Number 2
Monday–Friday, March 14–18, 2016; Baltimore, Maryland
Session A41: Cellular Biophysics |
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Sponsoring Units: DBIO GSOFT Chair: Bo Sun, Oregon State University Room: 344 |
Monday, March 14, 2016 8:00AM - 8:12AM |
A41.00001: Effects of Matrix Alignment and Mechanical Constraints on Cellular Behavior in 3D Engineered Microtissues. Prasenjit Bose, Jeroen Eyckmans, Christopher Chen, Daniel Reich The adhesion of cells to the extracellular matrix (ECM) plays a crucial role in a variety of cellular functions. The main building blocks of the ECM are 3D networks of fibrous proteins whose structure and alignments varies with tissue type. However, the impact of ECM alignment on cellular behaviors such as cell adhesion, spreading, extension and mechanics remains poorly understood. We present results on the development of a microtissue-based system that enables control of the structure, orientation, and degree of fibrillar alignment in 3D fibroblast-populated collagen gels. The tissues self-assemble from cell-laden collagen gels placed in micro-fabricated wells containing sets of elastic pillars. The contractile action of the cells leads to controlled alignment of the fibrous collagen, depending on the number and location of the pillars in each well. The pillars are elastic, and are utilized to measure the contractile forces of the microtissues, and by incorporating magnetic material in selected pillars, time-varying forces can be applied to the tissues for dynamic stimulation and measurement of mechanical properties. Results on the effects of varying pillar shape, spacing, location, and stiffness on microtissue organization and contractility will be presented. [Preview Abstract] |
Monday, March 14, 2016 8:12AM - 8:24AM |
A41.00002: Dynamics of Cancer Cell near Collagen Fiber Chain Jihan Kim, Bo Sun Cell migration is an integrated process that is important in life. Migration is essential for embryonic development as well as homeostatic processes such as wound healing and immune responses. When cell migrates through connective extracellular matrix (ECM), it applies cellular traction force to ECM and senses the rigidity of their local environment. We used human breast cancer cell (MDA-MB-231) which is highly invasive and applies strong traction force to ECM. As cancer cell applies traction force to type I collage-based ECM, it deforms collagen fibers near the surface. Patterns of deforming collagen fibers are significantly different with pairs of cancer cells compared to a single cancer cell. While a pair of cancer cells within 60 um creates aligned collagen fiber chains between them permanently, a single cancer cell does not form any fiber chains. In this experiment we measured a cellular response and an interaction between a pair of cells through the chain. Finally, we analyzed correlation of directions between cancer cell migration and the collagen chain alignment. [Preview Abstract] |
Monday, March 14, 2016 8:24AM - 8:36AM |
A41.00003: The Characteristics of Force Production of Kinesin-5 on MCF7 Microtubules Mitra Shojania Feizabadi Unlike neural mammalian microtubules with class II of beta tubulin as the major beta tubulin in their compositions, MCF7 microtubules composed of 0{\%} class II beta tubulin isotype, 39.1{\%} class I beta tubulin isotype, 2.5{\%} class III beta tubulin isotype and 58.4{\%} class IV beta tubulin isotype. Recent studies have revealed that function of some of motor proteins can be affected by the structural composition of microtubules. In this work, we will show how the function of mitotic kinesin ( Kin-5) under external load changed when moving along bovine versus MCF7 microtubules. Along MCF7 microtubules, the detachment force was reduced and the force-velocity curve was different as compared to those related to bovine brain. We will also show that the elimination of the C-terminal tails made the transport almost similar to the two sets of microtubules. This suggests that the C-terminal tails of tubulin plays a regulatory role in Kinesin-5's function. [Preview Abstract] |
Monday, March 14, 2016 8:36AM - 8:48AM |
A41.00004: On the robustness of SAC silencing in closed mitosis Donovan Ruth, Jian Liu Mitosis equally partitions sister chromatids to two daughter cells. This is achieved by properly attaching these chromatids via their kinetochores to microtubules that emanate from the spindle poles. Once the last kinetochore is properly attached, the spindle microtubules pull the sister chromatids apart. Due to the dynamic nature of microtubules, however, kinetochore-microtubule attachment often goes wrong. When this erroneous attachment occurs, it locally activates an ensemble of proteins, called the spindle assembly checkpoint proteins (SAC), which halts the mitotic progression until all the kinetochores are properly attached by spindle microtubules. The timing of SAC silencing thus determines the fidelity of chromosome segregation. We previously established a spatiotemporal model that addresses the robustness of SAC silencing in open mitosis for the first time. Here, we focus on closed mitosis by examining yeast mitosis as a model system. Though much experimental work has been done to study the SAC in cells undergoing closed mitosis, the processes responsible are not well understood. We leverage and extend our previous model to study SAC silencing mechanism in closed mitosis. We show that a robust signal of the SAC protein accumulation at the spindle pole body can be achieved. This signal is a nonlinear increasing function of number of kinetochore-microtubule attachments, and can thus serve as a robust trigger to time the SAC silencing. Together, our mechanism provides a unified framework across species that ensures robust SAC silencing and fidelity of chromosome segregation in mitosis. [Preview Abstract] |
Monday, March 14, 2016 8:48AM - 9:00AM |
A41.00005: Stochastic model of profilin-actin polymerization Brandon Horan, Dimitrios Vavylonis A driving factor in cell motility and other processes that involve changes of cell shape is the rapid polymerization of actin subunits into long filaments. This process is regulated by profilin, a protein which binds to actin subunits and regulates elongation of actin filaments. Whether profilin stimulates polymerization by coupling to hydrolysis of ATP-bound actin is debated. ~Previous studies have proposed indirect coupling to ATP hydrolysis using rate equations, but did not include the effects of fluctuations that are important near the critical concentration. ~We developed stochastic simulations using the Gillespie algorithm to study single filament elongation at the barbed end in the presence of profilin. We used recently measured rate constants and estimated the rate of profilin binding to the barbed end such that detailed balance is satisfied. Fast phosphate release at the tip of the filament was accounted for. The elongation rate and length diffusivity as functions of profilin and actin concentration were calculated and used to extract the critical concentrations of free actin and of total actin. We show under what conditions profilin leads to an increase in the critical concentration of total actin but a decrease in the critical concentration of free actin. [Preview Abstract] |
Monday, March 14, 2016 9:00AM - 9:12AM |
A41.00006: Surface deformation and shear flow in ligand mediated cell adhesion Sarthok Sircar, Anthony Roberts We present a unified, multiscale model to study the attachment/detachment dynamics of two deforming, near spherical cells, coated with binding ligands and subject to a slow, homogeneous shear flow in a viscous fluid medium. The binding ligands on the surface of the cells experience attractive and repulsive forces in an ionic medium and exhibit finite resistance to rotation via bond tilting. The microscale drag forces and couples describing the fluid flow inside the small separation gap between the cells, are calculated using a combination of methods in lubrication theory and previously published numerical results. For a select range of material and fluid parameters, a hysteretic transition of the sticking probability curves (i.e., the function g*) between the adhesion phase (when g*>0.5) and the fragmentation phase (when g*<0.5) is attributed to a nonlinear relation between the total nanoscale binding forces and the separation gap between the cells. We show that adhesion is favored in highly ionic fluids, increased deformability of the cells, elastic binders and a higher fluid shear rate (until a critical value). Continuation of the limit points (i.e., the turning points where the slope of the function g* changes sign within a select range of critical shear [Preview Abstract] |
Monday, March 14, 2016 9:12AM - 9:24AM |
A41.00007: Nanoparticle Distributions in Cancer and other Cells from Light Transmission Spectroscopy Alison Deatsch, Nan Sun, Jeffery Johnson, Sharon Stack, Carol Tanner, Steven Ruggiero We have measured the optical properties of whole cells and lysates using light transmission spectroscopy (LTS). LTS provides both the optical extinction coefficient in the wavelength range~from 220 to 1100 nm and (by spectral inversion using a Mie model) the particle distribution density in the size range from 1 to 3000 nm. Our current work involves whole cells and lysates of cultured human oral cells and other plant and animal cells. We have found systematic differences in the optical extinction between cancer and normal whole cells and lysates, which translate to different particle size distributions (PSDs) for these materials. We have also found specific power-law dependences of particle density with particle diameter for cell lysates. This suggests a universality of the packing distribution in cells that can be compared to ideal Apollonian packing, with the cell modeled as a fractal body comprised of spheres on all size scales. [Preview Abstract] |
Monday, March 14, 2016 9:24AM - 9:36AM |
A41.00008: Simulation of the effect of confinement in actin ring formation Maral Adeli Koudehi, Dimitrios Vavylonis Actin filaments are vital for different network structures in living cells. During cytokinesis, they form a contractile ring containing myosin motor proteins and actin filament cross-linkers to separate one cell into two cells. Recent experimental studies have quantified the bundle, ring, and network structures that form when actin filaments polymerize in confined environments in vitro, in the presence of varying concentrations of cross-linkers. In this study, we performed numerical simulations to investigate the effect of actin spherical confinement and cross-linking in ring formation. We used a spring-bead model and Brownian dynamics to simulate semiflexible actin filaments that polymerize in a confining sphere with a rate proportional to the monomer concentration. Applying the model for different size of the confining spheres shows that the probability of ring formation decreases by increasing the radius (at fixed initial monomer concentration), in agreement with prior experimental data. We describe the effect of persistence length, orientation-dependent cross-linking, and initial actin monomer concentration. Simulations show that equilibrium configurations can be reached through zipping and unzipping of actin filaments in bundles and transient ring formation. [Preview Abstract] |
Monday, March 14, 2016 9:36AM - 9:48AM |
A41.00009: Signaling and Dynamic Actin Responses of B Cells on Topographical Substrates Christina Ketchum, Xiaoyu Sun, John Fourkas, Wenxia Song, Arpita Upadhyaya B cells become activated upon physical contact with antigen on the surface of antigen presenting cells, such as dendritic cells. Binding of the B cell receptor with antigen initiates actin-mediated spreading of B cells, signaling cascades and eventually infection fighting antibodies. Lymphocytes, including B cells and T cells, have been shown to be responsive to the physical parameters of the contact surface, such as antigen mobility and substrate stiffness. However the roll of surface topography on lymphocyte function is unknown. Here we investigate the degree to which substrate topography controls actin-mediated spreading and B cell activation using nano-fabricated surfaces and live cell imaging. The model topographical system consists of 600 nanometer tall ridges with spacing varying between 800 nanometers and 5 micrometers. Using TIRF imaging we observe actin dynamics, B cell receptor motion and calcium signaling of B cells as they spread on the ridged substrates. We show that the spacing between ridges had a strong effect on the dynamics of actin and calcium influx on B cells. Our results indicate that B cells are highly sensitive to surface topography during cell spreading and signaling activation. [Preview Abstract] |
Monday, March 14, 2016 9:48AM - 10:00AM |
A41.00010: Mechanisms of T Lymphocyte Activation Exposed by Super Resolution Microscopy Leonard Campanello, Wolfgang Losert, Maria Traver, Brian Schaefer, Andrew York, Hari Schroff In order to avoid the deleterious consequences of an uncontrolled immune response, tight regulatory control of positive and negative regulators during lymphocyte activation is needed. Utilizing cutting-edge super-resolution imaging technologies in combination with quantitative image analysis we explore the interplay between positive and negative regulation in activated T lymphocytes and investigate whether intercellular signaling is possibly governed by the degradation of a complex intracellular structure called the POLKADOTS signalosome. In segmenting the POLKADOTS signalosome structure by the betweenness centrality of its 3D medial axis skeleton, it was discovered that autophagosomes, small degradative intracellular organelles, localize preferentially to the ends of the filamentous POLKADOTS signalosome. These results provide new insight into the mechanisms behind the complex regulatory process that govern T lymphocyte activation. [Preview Abstract] |
Monday, March 14, 2016 10:00AM - 10:12AM |
A41.00011: Single molecule analysis of B cell receptor motion during signaling activation Ivan Rey Suarez, Peter Koo, Simon Mochrie, Wenxia Song, Arpita Upadhyaya B cells are an essential part of the adaptive immune system. They patrol the body looking for signs of infection in the form of antigen on the surface of antigen presenting cells. The binding of the B cell receptor (BCR) to antigen induces signaling cascades that lead to B cell activation and eventual production of high affinity antibodies. During activation, BCR organize into signaling microclusters, which are platforms for signal amplification. The physical processes underlying receptor movement and aggregation are not well understood. Here we study the dynamics of single BCRs on activated murine primary B cells using TIRF imaging and single particle tracking. The tracks obtained are analyzed using perturbation expectation-maximization (pEM) a systems-level analysis that allows the identification of different short-time diffusive states from a set of single particle tracks. We identified five different diffusive states on wild type cells, which correspond to different molecular states of the BCR. By using actin polymerization inhibitors and mutant cells lacking important actin regulators we were able to identify the BCR molecule configuration associated with each diffusive state. [Preview Abstract] |
Monday, March 14, 2016 10:12AM - 10:24AM |
A41.00012: Targeting cancer cell invasiveness using homing peptide-nanocomplexes Giulia Suarato, Jillian Cathcart, Weiyi Li, Jian Cao, Yizhi Meng Matrix metalloproteinase-14 (MMP-14) plays critical roles in digesting the basement membrane and extracellular matrix and inducing cancer migration. We recently unraveled a unique role in cell invasion of the hemopexin (PEX) domain of MMP-14. The minimal motif located at the outmost strand of the fourth blade of the PEX domain was identified to form homodimers of MMP-14. A peptide (IVS4) mimicking the binding motif was shown to interrupt MMP-14 dimerization and decrease MMP-14-mediated functions. Since most invasive cancer cells express upregulated MMP-14 at the surface, IVS4 could be used as a cancer homing peptide to specifically deliver cytotoxic drugs for cancer therapy. We developed cancer homing nanocarriers by linking IVS4 to polysaccharide-based micellar nanoparticles (NPs). To determine if conjugation of IVS4 to NPs maintains the IVS4 inhibition of MMP-14 function, substrate degradation and cell migration assays were performed. IVS4-NPs efficiently prevented MMP-14-mediated substrate degradation and cell migration, and were minimally uptaken by non-cancer cells. Importantly, IVS4 confers an uptake advantage compared to the control peptide in MMP-14-expressing cells. Taken together, our findings demonstrate the potential use of IVS4-NPs as novel cancer nanotherapeutics. [Preview Abstract] |
Monday, March 14, 2016 10:24AM - 10:36AM |
A41.00013: Gating mechanosensitive channels in bacteria with an atomic force microscope. Renata Garces, Samantha Miller, Christoph F. Schmidt The regulation of growth and integrity of bacteria is critically linked to mechanical stress. Bacteria typically maintain a high difference of osmotic pressure (turgor pressure) with respect to the environment. This pressure difference (on the order of 1 atm) is supported by the cell envelope, a composite of lipid membranes and a rigid cell wall. Turgor pressure is controlled by the ratio of osmolytes inside and outside bacteria and thus, can abruptly increase upon osmotic downshock. For structural integrity bacteria rely on the mechanical stability of the cell wall and on the action of mechanosensitive (MS) channels: membrane proteins that release solutes in response to stress in the cell envelope. We here present experimental data on MS channels gating. We activate channels by indenting living bacteria with the cantilever of an atomic force microscope (AFM). We compare responses of wild-type and mutant bacteria in which some or all MS channels have been eliminated. [Preview Abstract] |
Monday, March 14, 2016 10:36AM - 10:48AM |
A41.00014: Impedance spectroscopy of micro-Droplets reveals activation of Bacterial Mechanosensitive Channels in Hypotonic Solutions Aida Ebrahimi, Muhammad A. Alam Rapid detection of bacterial pathogens is of great importance in healthcare, food safety, environmental monitoring, and homeland security. Most bacterial detection platforms rely on binary fission (i.e. cell growth) to reach a threshold cell population that can be resolved by the sensing method. Since cell division depends on the bacteria type, the detection time of such methods can vary from hours to days. In contrast, in this work, we show that bacteria cells can be detected within minutes by relying on activation of specific protein channels, i.e. mechanosensitive channels (MS channels). When cells are exposed to hypotonic solutions, MS channels allow efflux of solutes to the external solution which leads to release the excessive membrane tension. Release of the cytoplasmic solutes, in turn, results in increase of the electrical conductance measured by droplet-based impedance sensing. The approach can be an effective technique for fast, pre-screening of bacterial contamination at ultra-low concentration. [Preview Abstract] |
Monday, March 14, 2016 10:48AM - 11:00AM |
A41.00015: Physics of Bacteria During Osmotic Shock Jordan Price, William Klug Bacteria combat hypoosmotic shocks by opening mechanosensitive ion channels located within the inner membrane. These channels are believed to act as ``emergency release valves,” reducing transient pressure during the shock by regulating solute and water flux. Recent experiments have shown that cell survivability depends strongly on channel populations and the rate of osmotic shock. However, the understanding of the physical mechanisms behind osmotic protection remains unclear. We investigate how channel deletions, variations in shock rate, and cell envelope mechanics affect survivability by constructing theoretical elasticity and transport models. We find that reducing the number of channels and applying faster shocks significantly increases the time-dependent stress of the cell membrane and wall. This result provides insight into physical mechanisms that govern cell failure, including membrane rupture and wall fracture. [Preview Abstract] |
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