Cellular protrusions are typically considered as distinct structures associated with specific regulators. However, we found that these regulators coordinately localize as propagating cortical waves, ...suggesting a common underlying mechanism. These molecular events fell into two excitable networks, the signal transduction network STEN and the cytoskeletal network CEN with different wave substructures. Computational studies using a coupled‐network model reproduced these features and showed that the morphology and kinetics of the waves depended on strengths of feedback loops. Chemically induced dimerization at multiple nodes produced distinct, coordinated alterations in patterns of other network components. Taken together, these studies indicate: STEN positive feedback is mediated by mutual inhibition between Ras/Rap and PIP2, while negative feedback depends on delayed PKB activation; PKBs link STEN to CEN; CEN includes positive feedback between Rac and F‐actin, and exerts fast positive and slow negative feedbacks to STEN. The alterations produced protrusions resembling filopodia, ruffles, pseudopodia, or lamellipodia, suggesting that these structures arise from a common regulatory mechanism and that the overall state of the STEN‐CEN system determines cellular morphology.
Synopsis
Theoretical and experimental analyses indicate that signal transduction (STEN) and cytoskeletal (CEN) excitable networks interact to control dynamic wave patterns at the cell cortex. Network perturbations change wave speed and range and create distinct protrusions.
Cell cortical wave patterns are brought about by coupled Rap/Ras and Rac/F‐actin centric networks that are linked via PIP3/PKBs.
Increases in Rap/Ras or decreases in PIP2 activate while PIP3/PKBs negatively regulate STEN. Increasing Rac/F‐actin activates CEN and provides feedback to STEN.
Feedback strengths of the coupled excitable networks control speed and range of wave patterns.
Protrusive form and dynamic cell morphology are determined by the overall state of the coupled networks rather than single regulators.
Theoretical and experimental analyses indicate that signal transduction (STEN) and cytoskeletal (CEN) excitable networks interact to control dynamic wave patterns at the cell cortex. Network perturbations change wave speed and range and create distinct protrusions.
Electrotaxis, the directional migration of cells in a constant electric field, is important in regeneration, development, and wound healing. Electrotaxis has a slower response and a smaller dynamic ...range than guidance by other cues, suggesting that the mechanism of electrotaxis shares both similarities and differences with chemical-gradient-sensing pathways. We examine a mechanism centered on the excitable system consisting of cortical waves of biochemical signals coupled to cytoskeletal reorganization, which has been implicated in random cell motility. We use electro-fused giant
cells to decouple waves from cell motion and employ nanotopographic surfaces to limit wave dimensions and lifetimes. We demonstrate that wave propagation in these cells is guided by electric fields. The wave area and lifetime gradually increase in the first 10 min after an electric field is turned on, leading to more abundant and wider protrusions in the cell region nearest the cathode. The wave directions display 'U-turn' behavior upon field reversal, and this switch occurs more quickly on nanotopography. Our results suggest that electric fields guide cells by controlling waves of signal transduction and cytoskeletal activity, which underlie cellular protrusions. Whereas surface receptor occupancy triggers both rapid activation and slower polarization of signaling pathways, electric fields appear to act primarily on polarization, explaining why cells respond to electric fields more slowly than to other guidance cues.
We report the development and optimisation of an assay for quantitating iron from iron oxide nanoparticles in biological matrices by using ferene-s, a chromogenic compound. The method is accurate, ...reliable and can be performed with basic equipment common to many laboratories making it convenient and inexpensive. The assay we have developed is suited for quantitation of iron in cell culture studies with iron oxide nanoparticles, which tend to manifest low levels of iron. The assay was validated with standard reference materials and with inductively coupled plasma-mass spectrometry (ICP-MS) to accurately measure iron concentrations ∼1 × 10
g in about 1 × 10
cells (∼1 × 10
g Fe per cell). The assay requires preparation and use of a working solution to which samples can be directly added without further processing. After overnight incubation, the absorbance can be measured with a standard UV/Vis spectrophotometer to provide iron concentration. Alternatively, for expedited processing, samples can be digested with concentrated nitric acid before addition to the working solution. Optimization studies demonstrated significant deviations accompany variable digestion times, highlighting the importance to ensure complete iron ion liberation from the nanoparticle or sample matrix to avoid underestimating iron concentration. When performed correctly, this method yields reliable iron ion concentration measurements to ∼2 × 10
M (1 × 10
g/ml sample).
Cell migration requires the coordination of an excitable signal transduction network involving Ras and PI3K pathways with cytoskeletal activity. We show that expressing activated Ras GTPase-family ...proteins in cells lacking PTEN or other mutations which increase cellular protrusiveness transforms cells into a persistently activated state. Leading- and trailing-edge markers were found exclusively at the cell perimeter and the cytosol, respectively, of the dramatically flattened cells. In addition, the lifetimes of dynamic actin puncta were increased where they overlapped with actin waves, suggesting a mechanism for the coupling between these two networks. All of these phenotypes could be reversed by inhibiting signal transduction. Strikingly, maintaining cells in this state of constant activation led to a form of cell death by catastrophic fragmentation. These findings provide insight into the feedback loops that control excitability of the signal transduction network, which drives migration.
Nonlinear biomolecular interactions on membranes drive membrane remodeling crucial for biological processes including chemotaxis, cytokinesis, and endocytosis. The complexity of biomolecular ...interactions, their redundancy, and the importance of spatiotemporal context in membrane organization impede understanding of the physical principles governing membrane mechanics. Developing a minimal in vitro system that mimics molecular signaling and membrane remodeling while maintaining physiological fidelity poses a major challenge. Inspired by chemotaxis, we reconstructed chemically regulated actin polymerization inside vesicles, guiding membrane self-organization. An external, undirected chemical input induced directed actin polymerization and membrane deformation uncorrelated with upstream biochemical cues, suggesting symmetry breaking. A biophysical model incorporating actin dynamics and membrane mechanics proposes that uneven actin distributions cause nonlinear membrane deformations, consistent with experimental findings. This protocellular system illuminates the interplay between actin dynamics and membrane shape during symmetry breaking, offering insights into chemotaxis and other cell biological processes.
A cell-like system was constructed that senses an external chemical and rearranges its internal mechanical structures in response.
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