The Experts below are selected from a list of 273 Experts worldwide ranked by ideXlab platform
Michael G. Vicker - One of the best experts on this subject based on the ideXlab platform.
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Dual chemotaxis signalling regulates Dictyostelium development: Intercellular cyclic AMP pulses and intracellular F-actin disassembly waves induce each other
European journal of cell biology, 2008Co-Authors: Michael G. Vicker, James F. GrutschAbstract:Aggregating Dictyostelium discoideum amoebae periodically emit and relay cAMP, which regulates their chemotaxis and morphogenesis into a multicellular, differentiated organism. Cyclic AMP also stimulates F-actin assembly and chemotactic Pseudopodium extension. We used actin-GFP expression to visualise for the first time intracellular F-actin assembly as a spatio-temporal indicator of cell reactions to cAMP, and thus the kinematics of cell communication, in aggregating streams. Every natural cAMP signal pulse induces an autowave of F-actin disassembly, which propagates from each cell's leading end to its trailing end at a linear rate, much slower than the calculated and measured velocities of cAMP diffusion in aggregating Dictyostelium. A sequence of transient reactions follows behind the wave, including anterior F-actin assembly, chemotactic Pseudopodium extension and cell advance at the cell front and, at the back, F-actin assembly, extension of a small retrograde Pseudopodium (forcing a brief cell retreat) and chemotactic stimulation of the following cell, yielding a 20s cAMP relay delay. These dynamics indicate that stream cell behaviour is mediated by a dual signalling system: a short-range cAMP pulse directed from one cell tail to an immediately following cell front and a slower, long-range wave of intracellular F-actin disassembly, each inducing the other.
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Reaction–diffusion waves of actin filament polymerization/depolymerization in Dictyostelium Pseudopodium extension and cell locomotion
Biophysical chemistry, 2000Co-Authors: Michael G. VickerAbstract:Abstract Cell surface movements and the intracellular spatial patterns and dynamics of actin filament (F-actin) were investigated in living and formalin-fixed cells of Dictyostelium discoideum by confocal microscopy. Excitation waves of F-actin assembly developed and propagated several micrometers at up to 26 μm/min in cells which had been intracellularly loaded with fluorescently labeled actin monomer. Wave propagation and extinction corresponded with the initiation and attenuation of Pseudopodium extension and cell advance, respectively. The identification of chemical waves was supported by the ring, sphere, spiral and scroll wave patterns, which were observed in the extensions of fixed cells stained with phalloidin-rhodamine, and by the similar, asymmetrical [F-actin] distribution in wavefronts in living and fixed cells. These F-actin patterns and dynamics in Dictyostelium provide evidence for a new supramolecular state of actin, which propagates as a self-organized, reaction–diffusion wave of reversible F-actin assembly and affects Pseudopodium extension. Actin’s properties of oscillation and self-organization might also fundamentally determine the nature of the eukaryotic cell’s reactions of adaptation, timing and signal response.
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reaction diffusion waves of actin filament polymerization depolymerization in dictyostelium Pseudopodium extension and cell locomotion
Biophysical Chemistry, 2000Co-Authors: Michael G. VickerAbstract:Abstract Cell surface movements and the intracellular spatial patterns and dynamics of actin filament (F-actin) were investigated in living and formalin-fixed cells of Dictyostelium discoideum by confocal microscopy. Excitation waves of F-actin assembly developed and propagated several micrometers at up to 26 μm/min in cells which had been intracellularly loaded with fluorescently labeled actin monomer. Wave propagation and extinction corresponded with the initiation and attenuation of Pseudopodium extension and cell advance, respectively. The identification of chemical waves was supported by the ring, sphere, spiral and scroll wave patterns, which were observed in the extensions of fixed cells stained with phalloidin-rhodamine, and by the similar, asymmetrical [F-actin] distribution in wavefronts in living and fixed cells. These F-actin patterns and dynamics in Dictyostelium provide evidence for a new supramolecular state of actin, which propagates as a self-organized, reaction–diffusion wave of reversible F-actin assembly and affects Pseudopodium extension. Actin’s properties of oscillation and self-organization might also fundamentally determine the nature of the eukaryotic cell’s reactions of adaptation, timing and signal response.
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Pseudopodium extension and amoeboid locomotion in Dictyostelium discoideum : possible autowave behaviour of F-actin
Physica D: Nonlinear Phenomena, 1997Co-Authors: Michael G. Vicker, Wei Xiang, Peter Jörg Plath, Werner WosniokAbstract:Abstract Supramolecular patterns of filamentous (F-)actin up to several micrometres across were visualized within projections of locomotory amoebae after cell fixation and staining with phalloidin-rhodamine. The patterns included rings, single and double spirals, some apparently colliding and disintegrating. Cell stimulation with a pulse of the chemoattractant cyclic AMP induced damping oscillations in F-actin ring frequency with a period of 6–7 s. Ring front propagation after stimulation was modelled by Markov and Fourier methods at 3.1–17.5 μm/min, similar to actual cell speed. We argue that the dynamics and detailed morphological correspondence of these F-actin structures to wave patterns in chemical reaction-diffusion systems strongly supports the interpretation that Dictyostelium cytoplasm behaves as an unstable, excitable medium enabling the propagation of self-organized, physico-chemical relaxation oscillations, i.e. autowaves, of reversible F-actin assembly or aggregation — a new state of actin - fundamental to Pseudopodium extension, cell locomotion, chemotaxis and other cell functions.
Richard L. Klemke - One of the best experts on this subject based on the ideXlab platform.
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JCB Article Regulation of cell migration and survival by focal adhesion targeting of Lasp-1
2013Co-Authors: Yi Hsing Lin, Anar A. Brahmbhatt, Zee-yong Park, Dayin Lin, Marie-christine Rio, John R. Yates, Richard L. KlemkeAbstract:Large-scale proteomic and functional analysis of isolated pseudopodia revealed the Lim, actin, and SH3 domain protein (Lasp-1) as a novel protein necessary for cell migration, but not adhesion to, the extracellular matrix (ECM). Lasp-1 is a ubiquitously expressed actin-binding protein with a unique domain configuration containing SH3 and LIM domains, and is overexpressed in 8–12 % of human breast cancers. We find that stimulation of nonmotile and quiescent cells with growth factors or ECM proteins facilitates Lasp-1 relocalization from the cell periphery to the leading edge of the Pseudopodium, where it associate
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Proteomics method for identification of Pseudopodium phosphotyrosine proteins.
Methods in molecular biology (Clifton N.J.), 2011Co-Authors: Ying Wang, Richard L. KlemkeAbstract:Cell migration requires actin/myosin-mediated membrane protrusion of a Pseudopodium (or lamellipodium) and its attachment to the substratum. This process guides the direction of cell movement through cytoskeletal remodeling and is regulated by complex signaling networks that act spatially downstream of integrin adhesion receptors. Understanding how these regulatory networks are organized in migratory cells is important for many physiological and pathological processes, including wound healing, immune function, and cancer metastasis. Here, we describe methods for the immunoaffinity purification of phosphotyrosine proteins (pY) from pseudopodia that have been isolated from migratory cells. These methods are compatible with current mass spectrometry-based protein identification technologies and can be utilized for the large-scale identification of the Pseudopodium pY proteome in various migratory cell lines, including primary and cancer cells.
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Abstract 4464: Ciclopirox Olamine alters PEAK-1-mediated cytoskeleton organization and invasion of metastatic cancer cells
Experimental and Molecular Therapeutics, 2011Co-Authors: Jessica M. Weems, Richard L. KlemkeAbstract:The majority of cancer patients die from metastatic disease because there are no drugs available to prevent the spread of cancer and eradicate metastases in patients. The antimycotic drug ciclopirox olamine (CPX) is under investigation as a repositioned drug candidate which has anti-tumor and anti-angiogenesis activity in experimental models. Because CPX has been FDA tested and shows exceptionally low side effects in patients even at high doses, it has significant potential as both a therapeutic to prevent cancer recurrence after surgical resection and for the long term systemic treatment of cancer patients with overt metastatic disease. However, CPX has not yet been tested for its ability to inhibit metastatic cell function. In this study we examined the ability of CPX to inhibit metastatic cancer cell migration, invasion, and proliferation, and evaluated the signal transduction pathways that regulate these processes. Interestingly, CPX was observed to inhibit the expression of the tyrosine kinase, Pseudopodium-enriched atypical kinase 1 (PEAK1), which is necessary for proper remodeling of the actin cytoskeleton and focal adhesion dynamics in metastatic cancer cells (PNAS, 107(24), 10920-5). Associated with the decrease in PEAK1 expression by CPX was decreased total cellular tyrosine phosphoprotein content, Pseudopodium protrusion, cancer cell spreading, movement, and proliferation on extracellular matrix proteins. Collectively, our findings indicate that at therapeutic doses, CPX potently alters cytoskeletal signaling, cancer cell movement, and proliferation by modulating PEAK1 activity. These findings are important in light of the fact that PEAK1 amplification and cytoskeleton modulation have been linked to anchorage-independent cell growth, cell migration, and tumor progression in human gastrointestinal cancers. The exceptional safety record of CPX and our findings support the notion that CPX has significant potential as a repositioned therapeutic agent to combat cancer recurrence and spread in patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4464. doi:10.1158/1538-7445.AM2011-4464
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Methods for Pseudopodia Purification and Proteomic Analysis
Science's STKE : signal transduction knowledge environment, 2007Co-Authors: Ying Wang, Shi-jian Ding, Wei Wang, Feng Yang, Jon M. Jacobs, David G. Camp, Richard D. Smith, Richard L. KlemkeAbstract:Directional cell migration requires the formation of a dominant Pseudopodium in the direction toward which the cell migrates. When a migratory cell is stimulated with a chemoattractant or extracellular matrix (ECM) gradient, it responds with localized amplification of signals on the side facing the gradient. The signals mediate reorganization of the actin-myosin cytoskeleton, leading to morphological polarization of the cell and Pseudopodium extension. To identify these signals, we developed an approach to biochemically isolate the Pseudopodium from the cell body using 3.0-micrometer porous filters for large-scale quantitative proteomic and phosphoproteomic analysis. Here, we detail the methodology for Pseudopodium purification and proteomic analysis. This model system should be widely applicable for the analysis of the Pseudopodium proteome from various migratory cell lines, including primary and cancer cell lines stimulated with a diverse array of chemoattractants, ECM proteins, or both.
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Biochemical purification of pseudopodia from migratory cells.
Methods in molecular biology (Clifton N.J.), 2007Co-Authors: Ying Wang, Richard L. KlemkeAbstract:Cell migration requires the formation of a leading Pseudopodium (lamellipodium) in the direction of movement. This process requires signal amplification to facilitate directional sensing mechanisms that lead to actin-mediated membrane extension. However, it has been difficult to study pseudopodia formation because it has not been possible to purify this structure for biochemical analysis. Here we describe a method to biochemically purify the protruding Pseudopodium from the cell body compartment using polycarbonate microporous filters. Cells are cultured on top of 3.0-microm porous filters and allowed to extend pseudopodia through the small pores to the undersurface in response to a gradient of either chemokine or extracellular matrix (ECM) protein. Pseudopodia and cell bodies are then differentially scraped from the filter surface into lysis buffer for biochemical analysis. Using this method, it is possible to identify novel Pseudopodium and cell body proteins as well as study the spatiotemporal organization of signaling processes that regulate Pseudopodium formation and cell polarity. This method will help facilitate our understanding of how cells protrude pseudopodia through small openings in the ECM and vasculature during cancer cell invasion, immune cell surveillance, and embryonic development.
Ken-ichiro Ishida - One of the best experts on this subject based on the ideXlab platform.
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Fine-structural Observations on Siliceous Scale Production and Shell Assembly in the Testate Amoeba Paulinella chromatophora.
Protist, 2016Co-Authors: Mami Nomura, Ken-ichiro IshidaAbstract:The fine structure of shell formation was observed in P. chromatophora. Scales were formed one by one in silica deposition vesicles (SDVs) that were supported by an array of microtubules, which are probably involved in determining the shape and size of scales. The timing of silicic acid transport into an SDV was shown to be at an early stage of scale production because silicon was detected within SDVs containing immature scales. During the shell construction process, vesicles containing two types of dense materials were observed. One type of vesicle contains lower-density material and is located at the front edge of the branched, thick Pseudopodium, extending from the maternal shell to the newly formed shell. The other type of vesicle, which contains higher-density material, was also observed in the thick Pseudopodium. It appears that microtubules are involved in the shell construction process.
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Detailed process of shell construction in the photosynthetic testate amoeba Paulinella chromatophora (euglyphid, Rhizaria).
The Journal of eukaryotic microbiology, 2014Co-Authors: Mami Nomura, Takuro Nakayama, Ken-ichiro IshidaAbstract:Most euglyphids, a group of testate amoebae, have a shell that is constructed from numerous siliceous scales. The euglyphid Paulinella chromatophora has photosynthetic organelles (termed cyanelles or chromatophores), allowing it to be cultivated more easily than other euglyphids. Like other euglyphids, P. chromatophora has a siliceous shell made of brick-like scales. These scales are varied in size and shape. How a P. chromatophora cell makes this shell is still a mystery. We examined shell construction process in P. chromatophora in detail using time-lapse video microscopy. The new shell was constructed by a specialized Pseudopodium that laid out each scale into correct position, one scale at a time. The present study inferred that the sequence of scale production and secretion was well controlled.
Mami Nomura - One of the best experts on this subject based on the ideXlab platform.
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Fine-structural Observations on Siliceous Scale Production and Shell Assembly in the Testate Amoeba Paulinella chromatophora.
Protist, 2016Co-Authors: Mami Nomura, Ken-ichiro IshidaAbstract:The fine structure of shell formation was observed in P. chromatophora. Scales were formed one by one in silica deposition vesicles (SDVs) that were supported by an array of microtubules, which are probably involved in determining the shape and size of scales. The timing of silicic acid transport into an SDV was shown to be at an early stage of scale production because silicon was detected within SDVs containing immature scales. During the shell construction process, vesicles containing two types of dense materials were observed. One type of vesicle contains lower-density material and is located at the front edge of the branched, thick Pseudopodium, extending from the maternal shell to the newly formed shell. The other type of vesicle, which contains higher-density material, was also observed in the thick Pseudopodium. It appears that microtubules are involved in the shell construction process.
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Detailed process of shell construction in the photosynthetic testate amoeba Paulinella chromatophora (euglyphid, Rhizaria).
The Journal of eukaryotic microbiology, 2014Co-Authors: Mami Nomura, Takuro Nakayama, Ken-ichiro IshidaAbstract:Most euglyphids, a group of testate amoebae, have a shell that is constructed from numerous siliceous scales. The euglyphid Paulinella chromatophora has photosynthetic organelles (termed cyanelles or chromatophores), allowing it to be cultivated more easily than other euglyphids. Like other euglyphids, P. chromatophora has a siliceous shell made of brick-like scales. These scales are varied in size and shape. How a P. chromatophora cell makes this shell is still a mystery. We examined shell construction process in P. chromatophora in detail using time-lapse video microscopy. The new shell was constructed by a specialized Pseudopodium that laid out each scale into correct position, one scale at a time. The present study inferred that the sequence of scale production and secretion was well controlled.
Y Igarashi - One of the best experts on this subject based on the ideXlab platform.
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Sphingosine-1-phosphate inhibits actin nucleation and Pseudopodium formation to control cell motility of mouse melanoma cells.
FEBS letters, 1996Co-Authors: S Yamamura, Y Sadahira, F Ruan, S Hakomori, Y IgarashiAbstract:Sphingosine-1-phosphate (Sph-1-P), the initial product of sphingosine (Sph) catabolism, has been reported to inhibit motility of mouse melanoma B16/F1 and other types of cells at very low concentrations (10-100 nM). Sph-1-P (100 nM-1 microM) inhibited Pseudopodium formation by blocking polymerization and reorganization of actin filaments in newly formed pseudopodia, and reduced F-actin by approximately 25% in F1 cells. A pyrene-labeled actin nucleation assay revealed that Sph-1-P (100 nM) inhibits actin nucleation mediated by F1 cell plasma membranes. These results suggest that Sph-1-P interacts with molecules associated with actin nucleation to inhibit reorganization of Pseudopodium formation and cell motility.
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sphingosine 1 phosphate inhibits actin nucleation and Pseudopodium formation to control cell motility of mouse melanoma cells
FEBS Letters, 1996Co-Authors: S Yamamura, Y Sadahira, F Ruan, S Hakomori, Y IgarashiAbstract:Sphingosine-1-phosphate (Sph-1-P), the initial product of sphingosine (Sph) catabolism, has been reported to inhibit motility of mouse melanoma B16/F1 and other types of cells at very low concentrations (10–100 nM). Sph-1-P (100 nM–1 μM) inhibited Pseudopodium formation by blocking polymerization and reorganization of actin filaments in newly formed pseudopodia, and reduced F-actin by ∼ 25% in F1 cells. A pyrene-labeled actin nucleation assay revealed that Sph-1-P (100 nM) inhibits actin nucleation mediated by F1 cell plasma membranes. These results suggest that Sph-1-P interacts with molecules associated with actin nucleation to inhibit reorganization of Pseudopodium formation and cell motility.
