Cycles Of Detachment Research Articles

Spatial organization of the cytoskeleton enhances cargo delivery to specific target areas on the plasma membrane of spherical cells

Intracellular transport is vital for the proper functioning and survival of a cell. Cargo (proteins, vesicles, organelles, etc) is transferred from its place of creation to its target locations via molecular motor assisted transport along cytoskeletal filaments. The transport efficiency is strongly affected by the spatial organization of the cytoskeleton, which constitutes an inhomogeneous, complex network. In cells with a centrosome microtubules grow radially from the central microtubule organizing center towards the cell periphery whereas actin filaments form a dense meshwork, the actin cortex, underneath the cell membrane with a broad range of orientations. The emerging ballistic motion along filaments is frequently interrupted due to constricting intersection nodes or cycles of detachment and reattachment processes in the crowded cytoplasm. In order to investigate the efficiency of search strategies established by the cell’s specific spatial organization of the cytoskeleton we formulate a random velocity model with intermittent arrest states. With extensive computer simulations we analyze the dependence of the mean first passage times for narrow escape problems on the structural characteristics of the cytoskeleton, the motor properties and the fraction of time spent in each state. We find that an inhomogeneous architecture with a small width of the actin cortex constitutes an efficient intracellular search strategy.

Durable adhesives based on electrospun poly(vinylidene fluoride) fibers

ABSTRACTHerein, the fabrication of poly(vinylidene fluoride) (PVDF) fibrous membrane using electrospinning is reported and its use for dry‐adhesive applications is demonstrated. The shear and normal adhesion performance of the samples was investigated using an Instron tensile tester and an atomic force microscope (AFM) respectively. For shear adhesion measurements, the electrospun membrane was finger pressed on to a glass slide and pulled in shear mode using a tensile tester. The thickness of the electrospun membrane was varied and the effect of thickness on shear adhesion was investigated. The shear adhesion strength increased when the thickness of the samples was reduced. Shear adhesion strength of a 200 µm thick sample was determined to be approximately 0.165 N/cm. For normal adhesion measurements, a flat tipless cantilever was used to indent the sample and then retract back to measure the pull‐off force. High shear adhesion strength and normal pull‐off force recorded are attributed to the fine size of the fibers that conform to the asperities present on the surfaces of the glass slide and the AFM cantilever. The durability of the adhesive was also verified by repeating the AFM adhesion measurements over 1000 consecutive attachment–detachment cycles. The pull‐off force was seen to be constant over 1000 attachment–detachment cycles. Our results indicate that these electrospun fibrous membranes can potentially be used as reusable dry‐adhesives. © 2016 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2017, 134, 44393.

Copper resistance, motility and the mineral dissolution behavior were assessed as novel factors involved in bacterial adhesion in bioleaching

A study was carried out on the adhesion to sulfide minerals of chemolithoautotrophic acidophilic bacteria obtained from industrial copper bioleaching operation. For this purpose, a mixed culture obtained from an industrial process and two metabolically different pure strains of Acidithiobacillus: A. ferrooxidans and A. ferridurans were used. These microorganisms showed significant differences in adhesion with respect to pyrite and chalcocite in terms of the temporal dynamics patterns and preference. A complex dynamics that involve cycles of attachment and detachment can only be explained by considering both, the intrinsic characteristics of the microorganisms, such as hydrophobicity, resistance and motility, and the properties of each mineral like their hydrophobicity, dissolution behavior and ionic contribution.

Dynein motors: How AAA+ ring opening and closing coordinates microtubule binding and linker movement.

Dyneins are a family of motor proteins that move along the microtubule. Motility is generated in the motor domain, which consists of a ring of six AAA+ (ATPases associated with diverse cellular activities) domains, the linker and the microtubule-binding domain (MTBD). The cyclic ATP-hydrolysis in the AAA+ ring causes the remodelling of the linker, which creates the necessary force for movement. The production of force has to be synchronized with cycles of microtubule detachment and rebinding to efficiently create movement along the microtubule. The analysis of four dynein motor domain crystal structures in the essay presented here provides evidence that this crucial coordination is carried out by open/closed AAA+ ring conformations.

THE QUASI-ROCHE LOBE OVERFLOW STATE IN THE EVOLUTION OF CLOSE BINARY SYSTEMS CONTAINING A RADIO PULSAR

We study the evolution of close binary systems formed by a normal (solar composition), intermediate mass donor star together with a neutron star. We consider models including irradiation feedback and evaporation. These non-standard ingredients deeply modify the mass transfer stages of these binaries. While models that neglect irradiation feedback undergo continuous, long standing mass transfer episodes, models including these effect suffer a number cycles of mass transfer and detachment. During mass transfer the systems should reveal themselves as low-mass X-ray binaries (LMXBs), whereas when detached they behave as a binary radio pulsars. We show that at these stages irradiated models are in a Roche lobe overflow (RLOF) state or in a quasi-RLOF state. Quasi-RLOF stars have a radius slightly smaller than its Roche lobe. Remarkably, these conditions are attained for orbital period and donor mass values in the range corresponding to a family of binary radio pulsars known as "redbacks". Thus, redback companions should be quasi-RLOF stars. We show that the characteristics of the redback system PSR J1723-2837 are accounted for by these models. In each mass transfer cycle these systems should switch from LMXB to binary radio pulsar states with a timescale of \sim million years. However, there is recent and fast growing evidence of systems switching on far shorter, human timescales. This should be related to instabilities in the accretion disc surrounding the neutron star and/or radio ejection, still to be included in the model having the quasi-RLOF state as a general condition.

Bubble activation from a hydrophobic spot at “negative” surface superheats in subcooled boiling

We present experimental results on the controlled bubble generation from a single PTFE (polytetrafluoroethylene) spot—with diameter varying from 2 mm to 6 mm—deposited on a flat polished copper surface that was submersed in subcooled pure water. The static contact angle of the PTFE coating was measured to be over 120°, which conveniently produced a clear contrast with the copper substrate in terms of wettability that ensured controlled bubble nucleation. By making use of a high-speed camera, statistical details about the bubble formation that include the departure frequency and diameter have been obtained at various surface temperatures. An interesting observation was made of repeated cycles of bubble nucleation and detachment at nominally negative surface superheats (i.e., the wall temperature being below the saturation temperature at the system pressure), which featured particularly long bubble growth time and seemingly no waiting time. The vertical temperature distribution inside the bubble, which was measured by a micro-thermocouple of about 250 μm in diameter, suggests a relatively stable bubble composition of water vapor and dissolved air. A heat-pipe analogy was drawn to describe the internal heat transfer mechanism of bubble growth on a mixed wettability surface under subcooled conditions.

Fibrillar adhesion with no clusterisation: Functional significance of material gradient along adhesive setae of insects.

It has been recently demonstrated that adhesive tarsal setae of beetles possess material gradients along their length. These gradients presumably represent an evolutionary optimization enhancing the adaptation to rough surfaces while simultaneously preventing clusterisation of the setae by lateral collapse. The numerical experiment of the present study has clearly demonstrated that gradient-bearing fibers with short soft tips and stiff bases have greater advantage in maximizing adhesion and minimizing clusterisation in multiple attachment–detachment cycles, if compared to the fibers with longer soft tips on the stiff bases and fibers with stiff tips on the soft bases. This study not only manifests the crucial role of gradients in material properties along the setae in beetle fibrillar adhesive system, but predicts that similar gradients must have been convergently evolved in various lineages of arthropods.

Engineering myosins for long-range transport on actin filaments

Cytoskeletal motors act as cargo transporters in cells1 and may be harnessed for directed transport applications in molecular detection and diagnostic devices2. High processivity — the ability to take many steps along a track before dissociating3 — is often a desirable characteristic because it allows nanoscale motors to transport cargoes over distances of microns in vivo and in vitro. Natural processive myosins4,5 are dimeric and use internal tension to coordinate the detachment cycles of the two heads6–8. Here, we show that processivity can be enhanced in engineered myosins using two non-natural strategies designed to optimize the effectiveness of random, uncoordinated stepping: (i) formation of three-headed and four-headed myosins; and (ii) introduction of flexible elements between heads. We quantify improvements using systematic single-molecule characterization of a panel of engineered motors. To test the modularity of our approach, we design a controllably bidirectional myosin that is robustly processive in both the forward and backward direction, and also produce the fastest processive cytoskeletal motor measured to date, reaching a speed of 10 μm/s.

Instantly switchable adhesion of bridged fibrillar adhesive via gecko-inspired detachment mechanism and its application to a transportation system

Inspired by the exceptional climbing ability of gecko lizards, artificial fibrillar adhesives have been extensively studied over the last decade both experimentally and theoretically. Therefore, a new leap towards practical uses beyond the academic horizon is timely and highly anticipated. To this end, we present a fibrillar adhesive in the form of bridged micropillars and its application to a transportation system with the detachment mechanism inspired by the climbing behaviour of gecko lizards. The adhesive shows strong normal attachment (~30 N cm(-2)) as well as easy and fast detachment within 0.5 s without involving complex dynamic mechanisms or specific stimulus-responsive materials. The fabrication of the bridged micropillars consists of replica moulding of polydimethylsiloxane (PDMS) micropillars, transfer of the PDMS precursor to the heads of the micropillars, and inverse placement on an inert Teflon-coated surface. Owing to the spontaneous interconnections of low viscosity PDMS precursor, bridged micropillars with a uniform capping nanomembrane (~800 nm thickness) are formed over a large area. Interestingly, macroscopic adhesion in the normal direction can be immediately switched between on and off states by changing the two detachment modes of pulling and peeling, respectively. To prove the potential of the fibrillar adhesive for practical use, an automated transportation system is demonstrated for lifting and releasing a mass of stacked glass slides over 1000 cycles of attachment and detachment.

Fabrication and analysis of enforced dry adhesives with core–shell micropillars

We present a simple method for fabricating robust dry adhesives by coating a soft polydimethyl siloxane (PDMS) thin layer on rigid backbone micropillars of polyurethane acrylate (PUA). These core–shell type micropillars demonstrated enhanced durability both in normal and shear adhesion over more than 100 cycles of attachment and detachment. Relatively strong normal (∼11.4 N cm−2) and shear (∼15.3 N cm−2) adhesion forces were observed, which were similar to or even larger than those of homogeneous PDMS micropillars. A simple theoretical model based on beam deflection theory was used to explain the experimental results.

E-020 Initial multi-center international experience with the target detachable coil

BackgroundAneurysm coil technology has evolved to require near instant detachment times and near perfect detachment performance. We evaluated the new Target coil to determine device safety and acute performance.Materials and...

Mechanism of actin filament nucleation by Vibrio VopL and implications for tandem W domain nucleation

Pathogen proteins targeting the actin cytoskeleton often serve as model systems to understand their more complex eukaryotic analogs. We show that the strong actin filament nucleation activity of Vibrio VopL depends on its three W domains and dimerization through a unique VopL C-terminal domain (VCD). The VCD displays a novel all-helical fold and interacts with the pointed end of the actin nucleus, contributing to the nucleation activity directly and through duplication of the W domain repeat. VopL promotes rapid cycles of filament nucleation and detachment, but generally has no effect on elongation. Profilin inhibits VopL-induced nucleation by competing for actin binding to the W domains. Combined, the results suggest that VopL stabilizes a hexameric double-stranded pointed end nucleus. Analysis of hybrid constructs of VopL and the eukaryotic nucleator Spire suggest that Spire may also function as a dimer in cells.

EphrinB/EphB signaling controls embryonic germ layer separation by contact-induced cell detachment.

Author SummaryThe formation and maintenance of tissue boundaries is an essential feature of multicellular animals, including humans. Using the frog embryo as a model system, we describe a mechanism of tissue separation that involves repeated cycles of cell attachment and detachment at the boundary between two adjacent tissues. Molecularly, this mechanism is based on a signal exchange across the boundary by a system of receptors and ligands—EphB receptors and ephrinB ligands—that are both integral cell membrane proteins, and thus require direct cell contact for signaling. In this way, cell attachment-dependent contact induces signaling which leads to a temporary detachment, followed by reattachment and a next round of signaling. Such an attachment-detachment mechanism allows for cell migration along the boundary, while at the same time preventing invasion of the stationary tissue by the migrating one.

Theoretical Analysis of the Stochastic Behaviour of Skeletal Actomyosin Complex

The skeletal muscle models are usually based on the hypotheses proposed by AF Huxley [1,2], but only recently an experimental evidence of them has been obtained: in [3] a single Myosin II shows several 5.5 nm steps on an actin filament in one preferred direction per ATP cycle, in [4] the probability for the actomyosin complex to switch from a weakly to a strongly attached state increases when the myosin is stretched (SS).We have computed the steady state solution of 2D Fokker-Plank equation associated to a system of a motor bound to a needle in the presence of an actin filament. We show how the dwell time of the stable states of the actomyosin complex is affected by the drag coefficient of the needle and use the data in [3] to estimate the bias of the energy toward the direction of the motion.Based on this analysis, we have developed a mathematical model to analyse quantitatively the feasibility of the hypothesis proposed in [3]. The stepping behaviour is view as a diffusive process in a locally tilted but globally flat potential, while the SS affects the jump probability in the attachment-detachment process. The response of the model is analysed by a stochastic simulation of the associated Langevin equations and succeeds to reproduce the behaviour of the muscle in its short time scale, related to the power stroke, and in its long time scale, related to the actin-myosin attachment detachment cycle.[1] Huxley. Prog Biophys Biophys Chem (1957)[2] Huxley, Simmons Nature (1971)[3] Kitamura et al. Biophysics (2005)[4] Iwaki et al. Nat Chem Biol (2009)

Electron microscopic evidence for the myosin head lever arm mechanism in hydrated myosin filaments using the gas environmental chamber

Muscle contraction results from an attachment–detachment cycle between the myosin heads extending from myosin filaments and the sites on actin filaments. The myosin head first attaches to actin together with the products of ATP hydrolysis, performs a power stroke associated with release of hydrolysis products, and detaches from actin upon binding with new ATP. The detached myosin head then hydrolyses ATP, and performs a recovery stroke to restore its initial position. The strokes have been suggested to result from rotation of the lever arm domain around the converter domain, while the catalytic domain remains rigid. To ascertain the validity of the lever arm hypothesis in muscle, we recorded ATP-induced movement at different regions within individual myosin heads in hydrated myosin filaments, using the gas environmental chamber attached to the electron microscope. The myosin head were position-marked with gold particles using three different site-directed antibodies. The amplitude of ATP-induced movement at the actin binding site in the catalytic domain was similar to that at the boundary between the catalytic and converter domains, but was definitely larger than that at the regulatory light chain in the lever arm domain. These results are consistent with the myosin head lever arm mechanism in muscle contraction if some assumptions are made.

Ubiquitination of α5β1 Integrin Controls Fibroblast Migration through Lysosomal Degradation of Fibronectin-Integrin Complexes

Cell migration requires endocytosis and recycling of integrins, but it is not known whether degradation of these membrane proteins is involved. Here we demonstrate that in migrating cells, a fraction of the endocytosed fibronectin receptor, alpha 5 beta 1 integrin, is sorted into multivesicular endosomes together with fibronectin and degraded in lysosomes. This sorting requires fibronectin-induced ubiquitination of the alpha 5 subunit, and the activity of the endosomal sorting complex required for transport (ESCRT) machinery, which interacts with alpha 5 beta 1 integrin. Importantly, we demonstrate that both alpha 5 ubiquitination and ESCRT functions are required for proper migration of fibroblasts. We propose that ligand-mediated degradation of alpha 5 beta 1 integrin via the ESCRT pathway is required in order to prevent endosomal accumulation of ligand-bound integrins that might otherwise form nonproductive adhesion sites. Fibronectin and alpha 5 beta 1 integrin therefore are trafficked to lysosomes in a similar way to growth factors and their receptors.

Stretchable, Adhesion-Tunable Dry Adhesive by Surface Wrinkling

We introduce a simple yet robust method of fabricating a stretchable, adhesion-tunable dry adhesive by combining replica molding and surface wrinkling. By utilizing a thin, wrinkled polydimethyl siloxane (PDMS) sheet with a thickness of 1 mm with built-in micropillars, active, dynamic control of normal and shear adhesion was achieved. Relatively strong normal (approximately 10.8 N/cm(2)) and shear adhesion (approximately 14.7 N/cm(2)) forces could be obtained for a fully extended (strained) PDMS sheet (prestrain of approximately 3%), whereas the forces could be rapidly reduced to nearly zero once the prestrain was released (prestrain of approximately 0.5%). Moreover, durability tests demonstrated that the adhesion strength in both the normal and shear directions was maintained over more than 100 cycles of attachment and detachment.

Membrane scission by the ESCRT-III complex

The endosomal sorting complex required for transport (ESCRT) system is essential for multivesicular body biogenesis, in which cargo sorting is coupled to the invagination and scission of intralumenal vesicles. The ESCRTs are also needed for budding of enveloped viruses including human immunodeficiency virus 1, and for membrane abscission in cytokinesis. In Saccharomyces cerevisiae, ESCRT-III consists of Vps20, Snf7, Vps24 and Vps2 (also known as Did4), which assemble in that order and require the ATPase Vps4 for their disassembly. In this study, the ESCRT-III-dependent budding and scission of intralumenal vesicles into giant unilamellar vesicles was reconstituted and visualized by fluorescence microscopy. Here we show that three subunits of ESCRT-III, Vps20, Snf7 and Vps24, are sufficient to detach intralumenal vesicles. Vps2, the ESCRT-III subunit responsible for recruiting Vps4, and the ATPase activity of Vps4 were required for ESCRT-III recycling and supported additional rounds of budding. The minimum set of ESCRT-III and Vps4 proteins capable of multiple cycles of vesicle detachment corresponds to the ancient set of ESCRT proteins conserved from archaea to animals.

CAMP Inhibits Cell Migration by Interfering with Rac-induced Lamellipodium Formation

Cell migration is critical for animal development and physiological as well as pathological responses. One important step during cell migration is the formation of lamellipodia at the leading edge of migrating cells. Here we report that the second messenger cAMP inhibits the migration of mouse embryonic fibroblast cells and mouse breast tumor cells. cAMP acts downstream of the small GTPase Rac and interferes with the formation of lamellipodia. Moreover, cAMP decreases the phosphorylation of the myosin light chain at the leading edge of cells and increases the phosphorylation of the vasodilator-stimulated phosphoprotein. Together with our previous report of a positive role of another second messenger, cGMP, in lamellipodium formation, our data indicate that cAMP and cGMP play opposite roles in modulating lamellipodium formation.

Surface Wrinkles for Smart Adhesion

to develop synthetic analogs as“smart” adhesives, that is, where the geometry of the patternscan tailor the adhesion of the material. However, there arecurrently two limitations in the design of patterned adhesives.First, because of either materials selection or geometric de-sign, the adhesive properties are not always reversible overthe course of multiple attachment–detachment cycles.