PP2A in the regulation of cell motility and invasion

Cell motility is a very critical phenomenon that plays an important role in the development of eukaryotic organisms. One of the well studied cell motility phenomena is chemotaxis, which is described as a directional movement of cell in response to changes in external chemotactic gradient. Numerous studies conducted both in unicellular organism and in mammalian cells have demonstrated the importance of phosphatidylionositol-3 kinase (PI3K) in this process. In addition, it is now well established that although PI3K plays an activation role in chemotaxis, the role of phosphatases is also critical to maintain this dynamic cyclical process. Protein phosphatase 2A (PP2A) is a major serine/threonine phosphatase that is a key player in regulating PI3K signaling. PP2A is abundantly and ubiquitously expressed and has been highly conserved during the evolution of eukaryotes. PP2A is composed of three protein subunits, A, B, and C. Subunit 'A' is a 60-65 kDa structural component, 'C' is a 36-38 kDa catalytic subunit, and 'B' is a 54-130 kDa regulatory subunit. The core complex of PP2A is comprised of the A and C subunits, which are tightly associated and this dimeric core complexes with the regulatory B subunit. The B subunit determines the substrate specificity as well as the spatial and temporal functions of PP2A. PP2A plays an important role in regulating multiple signal transduction pathways, including cell-cycle regulation, cell-growth and development, cytoskeleton dynamics, and cell motility. This review focuses on the role of PP2A in regulating motility of normal and transformed cells.     

Rab5 in the regulation of cell motility and invasion

Cellular invasion requires careful regulation of the cell migration and apoptotic signaling cascades, allowing cell movement and survival of the emigrating populations. Components of the endosomal machinery are involved in these processes, and in particular the role of small GTPases of the Rab family has become appreciated. Rab5 is best known for its role in regulating the trafficking of early endosomes, however, it has recently been appreciated to associate with and regulate the routing of complexes containing integrins, the primary cellular receptors for the extracellular matrix. The association regulates the spatio temporal activation of signals of downstream growth factors and integrins. Rab proteins have also been linked to apoptosis mediated by cell surface death receptors, which elicit the activation of the death cascade via activation of caspase 8. Recently, the link between trafficking, apoptosis and cell migration was strengthened, as Rab5 was determined to work in conjunction with caspase 8 in promoting tumor cell motility and metastasis by regulating β1 integrin traffic. The capacity to connect and regulate these pathways identifies Rab5 as a key player in future studies of cell migration and tumor dissemination.     

Modular design of micropattern geometry achieves combinatorial enhancements in cell motility

Basic micropattern shapes, such as stripes and teardrops, affect individual facets of cell motility, such as migration speed and directional bias, respectively. Here, we test the idea that these individual effects on cell motility can be brought together to achieve multidimensional improvements in cell behavior through the modular reconstruction of the simpler "building block" micropatterns. While a modular design strategy is conceptually appealing, current evidence suggests that combining environmental cues, especially molecular cues, such as growth factors and matrix proteins, elicits a highly nonlinear, synergistic cell response. Here, we show that, unlike molecular cues, combining stripe and teardrop geometric cues into a hybrid, spear-shaped micropattern yields combinatorial benefits in cell speed, persistence, and directional bias. Furthermore, cell migration speed and persistence are enhanced in a predictable, additive manner on the modular spear-shaped design. Meanwhile, the spear micropattern also improved the directional bias of cell movement compared to the standard teardrop geometry, revealing that combining geometric features can also lead to unexpected synergistic effects in certain aspects of cell motility. Our findings demonstrate that the modular design of hybrid micropatterns from simpler building block shapes achieves combinatorial improvements in cell motility. These findings have implications for engineering biomaterials that effectively mix and match micropatterns to modulate and direct cell motility in applications, such as tissue engineering and lab-on-a-chip devices.     

Role of mTOR signaling in tumor cell motility, invasion and metastasis

Tumor cell migration and invasion play fundamental roles in cancer metastasis. The mammalian target of rapamycin (mTOR), a highly conserved and ubiquitously expressed serine/threonine (Ser/Thr) kinase, is a central regulator of cell growth, proliferation, differentiation and survival. Recent studies have shown that mTOR also plays a critical role in the regulation of tumor cell motility, invasion and cancer metastasis. Current knowledge indicates that mTOR functions as two distinct complexes, mTORC1 and mTORC2. mTORC1 phosphorylates p70 S6 kinase (S6K1) and eukaryotic initiation factor 4E (eIF4E) binding protein 1 (4E-BP1), and regulates cell growth, proliferation, survival and motility. mTORC2 phosphorylates Akt, protein kinase C α (PKCα) and the focal adhesion proteins, and controls the activities of the small GTPases (RhoA, Cdc42 and Rac1), and regulates cell survival and the actin cytoskeleton. Here we briefly review recent knowledge of mTOR complexes and the role of mTOR signaling in tumor cell migration and invasion. We also discuss recent efforts about the mechanism by which rapamycin, a specific inhibitor of mTOR, inhibits cell migration, invasion and cancer metastasis.     

Geometric determinants of directional cell motility revealed using microcontact printing

Mammalian cells redirect their movement in response to changes in the physical properties of their extracellular matrix (ECM) adhesive scaffolds, including changes in available substrate area, shape, or flexibility. Yet, little is known about the cell's ability to discriminate between different types of spatial signals. Here we utilize a soft-lithography-based, microcontact printing technology in combination with automated computerized image analysis to explore the relationship between ECM geometry and directional motility. When fibroblast cells were cultured on fibronectin-coated adhesive islands with the same area (900 micrometers2) but different geometric forms (square, triangle, pentagon, hexagon, trapezoid, various parallelograms) and aspect ratios, cells preferentially extended new lamellipodia from their corners. In addition, by imposing these simple geometric constraints through ECM, cells were directed to deposit new fibronectin fibrils in these same corner regions. These data indicate that mammalian cells can sense edges within ECM patterns that exhibit a wide range of angularity and that they use these spatial cues to guide where they will deposit ECM and extend new motile processes during the process of directional migration.     

Shape and motility of a model cell: a computational study

We have investigated the shape, size, and motility of a minimal model of an adherent biological cell using the Monte Carlo method. The cell is modeled as a two dimensional ring polymer on the square lattice enclosing continuously polymerizing and depolymerizing actin networks. Our lattice model is an approximate representation of a real cell at a resolution of one actin molecule, 5 nm. The polymerization kinetics for the actin network are controlled by appropriate reaction probabilities which correspond to the correct experimental reaction rates. Using the simulation data we establish various scaling laws relating the size of the model cell to the concentration of polymerized and unpolymerized actin molecules and the length of the enclosing membrane. The computed drift velocities, which characterize the motility of the cell, exhibit a maximum at a certain fraction of polymerized actin which agrees with physiological fractions observed in experiments. The appearance of the maximum is related to the competition between the polymerization-induced protrusion of the membrane and the concomitant suppression of membrane fluctuations.     

A role for confined water in chaperonin function

Chaperonins engulf other proteins and accelerate their folding by an unknown mechanism. Here, we combine all-atom molecular dynamics simulations with data from experimental assays of the activity of the bacterial chaperonin GroEL to demonstrate that a chaperonin's ability to facilitate folding is correlated with the affinity of its interior surface for water. Our results suggest a novel view of the behavior of confined water for models of in vivo protein folding scenarios.     

A chemical inhibitor reveals the role of Raf kinase inhibitor protein in cell migration

Raf kinase inhibitor protein (RKIP) is a modulator of cell signaling that functions as an endogenous inhibitor of multiple kinases. We demonstrate here a positive role for RKIP in the regulation of cell locomotion. We discovered that RKIP is the relevant cellular target of locostatin, a cell migration inhibitor. Locostatin abrogates RKIP's ability to bind and inhibit Raf-1 kinase, and it acts by disrupting a protein-protein interaction, an uncommon mode of action for a small molecule. Small interfering RNA-mediated silencing of RKIP expression also reduces cell migration rate. Overexpression of RKIP converts epithelial cells to a highly migratory fibroblast-like phenotype, with dramatic reduction in the sensitivity of cells to locostatin. RKIP is therefore the compound's valid target and a key regulator of cell motility.     

Local immune cell contributions to fracture healing in aged individuals – A novel role for interleukin 22

With increasing age, the risk of bone fractures increases while regenerative capacity decreases. This variation in healing potential appears to be linked to adaptive immunity, but the underlying mechanism is still unknown. This study sheds light on immunoaging/inflammaging, which impacts regenerative processes in aging individuals. In an aged preclinical model system, different levels of immunoaging were analyzed to identify key factors that connect immunoaged/inflammaged conditions with bone formation after long bone fracture. Immunological facets, progenitor cells, the microbiome, and confounders were monitored locally at the injury site and systemically in relation to healing outcomes in 12-month-old mice with distinct individual levels of immunoaging. Bone tissue formation during healing was delayed in the immunoaged group and could be associated with significant changes in cytokine levels. A prolonged and amplified pro-inflammatory reaction was caused by upregulated immune cell activation markers, increased chemokine receptor availability and a lack of inhibitory signaling. In immunoaged mice, interleukin-22 was identified as a core cell signaling protein that played a central role in delayed healing. Therapeutic neutralization of IL-22 reversed this specific immunoaging-related disturbed healing. Immunoaging was found to be an influencing factor of decreased regenerative capacity in aged individuals. Furthermore, a novel therapeutic strategy of neutralizing IL-22 may successfully rejuvenate healing in individuals with advanced immune experiences.Subject terms: Trauma, Mechanisms of disease, Interleukins, Osteoimmunology     

The role of aqueous interfaces in the cell

The cell is rich with biopolymeric surfaces. Yet, the role of these surfaces and attendant surface-water interfaces has received little attention among biologists, most of whom consider water as a neutral carrier. This review aims to begin bridging the gap between biology and interface science-to show that a surface-oriented approach has power to bring fresh insights into an otherwise impenetrably complex maze. In this approach the cell is treated as a polymer gel. If the cell is a gel, then a logical approach to the understanding of cell function is through an understanding of gel function. Great strides have been made recently in understanding the principles of polymer-gel dynamics, and particularly the role of the polymer-water interface. It has become clear that a central mechanism in biology is the phase-transition-a major structural change prompted by a subtle change of environment. Phase-transitions are capable of doing work and such work could be responsible for much of the work of the cell. Here, we pursue this approach. We set up a polymer-gel-based foundation for cell behavior, and explore the extent to which this foundation explains how the cell achieves its everyday tasks.     

Novel protein kinase D inhibitors cause potent arrest in prostate cancer cell growth and motility

Background  

Protein kinase D (PKD) has been implicated in a wide range of cellular processes and pathological conditions including cancer. However, targeting PKD therapeutically and dissecting PKD-mediated cellular responses remains difficult due to lack of a potent and selective inhibitor. Previously, we identified a novel pan-PKD inhibitor, CID755673, with potency in the upper nanomolar range and high selectivity for PKD. In an effort to further enhance its selectivity and potency for potential in vivo application, small molecule analogs of CID755673 were generated by modifying both the core structure and side-chains.     

7-Acetylcytochalasin B: differential effects on sugar transport and cell motility.

Cytochalasin B (CB) is a potent inhibitor of sugar transport and cell motility in animal cells. We have synthesized and characterized the CB derivative 7-acetylcytochalasin B (CBAc) and have found that it has differential effects on transport and motile processes in fibroblasts. The derivative inhibited sugar transport in human red cells, 3T3 cells, and chicken embryo fibroblasts at micromolar concentrations, although it was less potent than its parent compound. Unlike CB, which causes fibroblasts to round up and arborize at less than 10 microM, CBAc had no effect on fibroblast morphology and membrane ruffling at concentrations as high as 90 microM. Competitive binding experiments using [3H] CB showed that the affinity of CBAc for sites related to sugar transport in the red cell membrane is about one-fourth of that of CB. In contrast, similar experiments using [3H] dihydrocytochalasin B (a derivative which inhibits cell motility but not sugar transport) showed that the affinity of CBAc for sites associated with red cell spectrin and actin is only about 1/20 of that of dihydrocytochalasin B. This study demonstrates that acetylation of the C-7 hydroxyl group of CB reduces its effect on cell morphology and motility much more than its ability to inhibit sugar transport. This observation, together with our earlier work with dihydrocytochalasin B, establishes that the pharmacologic effects of CB on fibroblasts result from the binding of the drug to two distinct classes of receptors and that these receptors interact with different parts of the cytochalasin molecule.     

A sorting strategy for C. elegans based on size-dependent motility and electrotaxis in a micro-structured channel

Caenorhabditis elegans (C. elegans) is a model organism widely utilized in various fundamental studies in developmental, neural and behavioural biology. The worm features four distinct larval stages, and many research questions are stage-specific; therefore, it is necessary to sort worms by their developmental stages, which are typically represented by different size ranges. However, manually synchronizing large populations of worms is time-consuming and labour-intensive, and the commercially available automated sorter is massive and expensive. Realizing the need for a cost-effective and simple micro-platform for sorting, we report an inexpensive and novel method to accomplish this goal. The proposed micro-platform features hexagonally arrayed microstructures with geometric dimensions optimized for the maximum motility of the worms based on their sizes. In each of the optimized micro-structured platforms, only the worms with the targeted size swim continuously with the maximum undulation frequency. Additionally, the persistent and directed movement of the worms can be achieved by applying an electric field along the channel. Based on the optimally spaced microstructures and the electrotaxis behaviour of the worms, we demonstrate the feasibility of a sorting strategy of C. elegans based on their size-dependent swimming behaviour. This micro-platform can also be used for other applications, such as behavioural studies of normal and locomotion-defective mutant worms in complex structures.     

On the role of intermembrane contact in cell electrofusion

The question: is cell electrofusion mediated by a long-lived fusogenic state or does the electric field fuse the membranes directly, was investigated by a new centrifugal approach. Mouse L-cells were brought into contact by a special centrifuge device allowing high voltage pulses to be applied upon the cell pellet during centrifugation. Both stages, membrane contact and electrical breakdown of cell membranes, were controlled. The degree of cell-to-cell compression and corresponding intermembrane contact area were estimated by measuring the low-voltage resistance R of the cell pellet which grows sharply with the increase of centripetal acceleration G. The extent of electrical membrane poration and critical pulse parameters were detected by recording the breakdown current. Supercritical pulse delivery to a cell pellet compressed by intensive centrifugation (400–600 g) leads to polycaryon mass formation. The pulse amplitude required for efficient fusion (2–3 kV/cm, 20–50 μs: fusion index F ∼ 25–35%) was found to be several times higher than the amplitude sufficient to induce noticeable breakdown (300 V/cm). The shapes of the F(G) and R(G) dependences were similar, which revealed a correlation between the area of intermembrane contact and fusion probability. Fusion was negligible if the moment of pulsation and the period of intensive centrifugation were separated in time. The data obtained allow us to conclude that in the case of fusion of L-cells the action of the electric field is not mediated by any long-lived fusogenic state. The process of the common membrane surface formation driven directly by the electric field is discussed.     

A cell for dielectric measurements in fluorinated atmospheres

A device is described which is suitable for dielectric measurements from 20 to 600°C in various fluorinated atmospheres. The cell has been tested with series of natural and synthetic oxide fluorides and fluorides derived from (NH₄)₃FeF₆, chiolite and weberite structural types.     

A new cell for electrochemical processes

A simple electrochemical reactor is described that fulfills most of the desired goals normally required. It is easy to construct and scale up. The main feature of the design is an electrode sandwich in which cloth separators are used to provide electrical insulation. They provide an inter-electrode space for the electrolyte and give good mass transfer characteristics. The ohmic drop in the solution is very low and the ratio of active electrode area to electrode volume is high. Operating properties have been determined and show that the cell may be applied to systems that might at first seem unfavourable for an electrochemical process, viz.: multi-phase (especially gas/liquid) mixtures and systems with very low concentrations of electroactive species. These proposals have been tested with some organic synthesis and with waste water purification (Cr(VI) reduction). The cell has been shown to perform well and to be inexpensive - both in construction and in operation.