Biology subjects

Beta, C.

Publications and source records attributed to Beta, C..

7 recordsLinked to original sources

bioRxiv2026

Coiled-coil domain kinking controls laminin-332 cleavage by elastase

Laminins are trimeric glycoproteins and essential components of basement membranes, where they provide structural support and mediate cell-matrix adhesion. Their -, {beta}-, and {gamma}-chains combine into a flexible trimeric coiled-coil domain forming the laminin long arm, which links the N-terminal short arms and the C-terminal globular domains. However, ultrastructural insight into the laminin coiled-coil domain remains limited, and the function of coiled-coil flexibility remains unclear. Using high-speed atomic force microscopy (HS-AFM) imaging, we previously investigated the ultrastructure of laminin-332 and observed dynamic kinking around a putative molecular hinge in the center of the coiled-coil domain. In this study, we integrate HS-AFM time-lapse imaging with AlphaFold structure prediction and normal-mode flexible fitting (NMFF) to generate dynamic models of the laminin-332 hinge during coiled-coil domain kinking, providing atomic-scale insight into the underlying molecular rearrangements. Furthermore, we use HS-AFM to visualize, in real time, the digestion of individual laminin-332 molecules by pancreatic elastase and show that coiled-coil kinking directs elastase cleavage to the hinge site, thereby ensuring reliable generation of the elastase 8 (E8) fragment containing the integrin binding site. In contrast, laminin molecules with extended coiled-coil conformations are cleaved at arbitrary sites, resulting in complete coiled-coil removal and degradation. Our results thus identify a novel conformational control mechanism directing proteolytic processing of laminin-332 through defined coiled-coil kinking. Besides expanding the understanding of dynamic laminin coiled-coil function, the gained ultrastructural insight into trimeric coiled-coil bending may aid the rational design of flexible coiled-coil modules for future protein engineering applications.

cell biology↗

bioRxiv2026

CaMKIIα holoenzymes self-organize into worm-like mesoscale clusters

Ca2+/calmodulin-dependent protein kinase II (CaMKII) is highly enriched in dendritic spines at concentrations comparable to those of cytoskeletal proteins and plays a central role in synaptic plasticity. During long-term potentiation (LTP), CaMKII further accumulates in spines. However, the mechanisms governing its higher-order organization remain poorly understood. Here, we use high-speed atomic force microscopy to visualize inter-holoenzyme interaction of CaMKII at mesoscopic scales (5-500 nm). Under freely diffusible conditions, CaMKII holoenzymes do not form stable clusters. In contrast, when spatially confined, they assemble into worm-like chain clusters mediated by kinase-domain interactions. These clusters expand upon activation, concomitant with the dissociation of the regulatory segment. Notably, the CaMKII P212L mutant associated with neurodevelopmental disorders, forms extensive clusters even in the basal state. Together, our findings demonstrate that CaMKII-CaMKII interactions drive mesoscale cluster formation and that precise regulation of cluster size and activation-dependent growth might be critical for synaptic signaling.

neuroscience↗

bioRxiv2024

Modelling how lamellipodia-driven cells maintain persistent migration and interact with external barriers

Cell motility is fundamental to many biological processes, and cells exhibit a variety of migration patterns. Many motile cell types follow a universal law that connects their speed and persistency, a property that can originate from the intracellular transport of polarity cues due to the global actin retrograde flow. This mechanism was termed the "Universal Coupling between cell Speed and Persistency"(UCSP). Here we implemented a simplified version of the UCSP mechanism in a coarse-grained "minimal-cell" model, which is composed of a three-dimensional vesicle that contains curved active proteins. This model spontaneously forms a lamellipodia-like motile cell shape, which is however sensitive and can depolarize into a non-motile form due to random fluctuations or when interacting with external obstacles. The UCSP implementation introduces long-range inhibition, which stabilizes the motile phenotype. This allows our model to describe the robust polarity observed in cells and explain a large variety of cellular dynamics, such as the relation between cell speed and aspect ratio, cell-barrier scattering, and cellular oscillations in different types of geometric confinements. Significance StatementCoupling curved membrane proteins to active protrusive forces that arise from recruited actin polymerization, can lead, in the presence of adhesion, to self-organization of a leading-edge cluster and a motile "minimal-cell". However, this polarized and motile shape can become unstable, and due to fluctuations or interactions with external perturbations transform to an immotile, symmetric shape. Here we couple the spatial organization of the curved active proteins to a global advection of a polarity cue along the cells activity axis. Introducing long-range inhibition, the resultant gradient of the polarity-cue stabilizes the motile, polarized "minimal-cell" vesicle. We thereby present a robust model of cell motility that can explain a variety of cellular shape-migration relations, cell-barrier scattering and spontaneous oscillations of confined cells.

biophysics↗

bioRxiv2024

Chemically induced dislocation and mutation of essential cytoskeletal proteins: A case study focusing on Coronin-A

Actin is one of the most highly conserved proteins--the actin cytoskeleton or actin-like structures are found in all kingdoms of life. In this work, Coronin-A, a central player involved in actin dynamics, is considered as an example to compare the effect of four mutation strategies on its function in Dictyostelium discoideum. Two distinct genetic knockout methods (selection-linked integration and classical gene disruption), RNAi knockdown, and sudden loss-of-function generated by chemically induced dislocation (CID) are compared and the cell lines are characterized according to actin dependent processes: appearance, development, and motility. These phenotypic features are affected to different degrees in the mutant cell lines, providing a basis to discuss the strengths and weaknesses of each approach. This comparative study highlights the advantages of the newly introduced selection-linked integration method to knock out genes over commonly used gene disruption, and for expressing a CID-based knock-sideways system under the native promotor of the protein of interest.

cell biology↗

bioRxiv2023

A minimal physical model for curvotaxis driven by curved protein complexes at the cell's leading edge

Cells often migrate on curved surfaces inside the body, such as curved tissues, blood vessels or highly curved protrusions of other cells. Recent in-vitro experiments provide clear evidence that motile cells are affected by the curvature of the substrate on which they migrate, preferring certain curvatures to others, termed "curvotaxis". The origin and underlying mechanism that gives rise to this curvature sensitivity are not well understood. Here, we employ a "minimal cell" model which is composed of a vesicle that contains curved membrane protein complexes, that exert protrusive forces on the membrane (representing the pressure due to actin polymerization). This minimal-cell model gives rise to spontaneous emergence of a motile phenotype, driven by a lamellipodia-like leading edge. By systematically screening the behaviour of this model on different types of curved substrates (sinusoidal, cylinder and tube), we show that minimal ingredients and energy terms capture the experimental data. The model recovers the observed migration on the sinusoidal substrate, where cells move along the grooves (minima), while avoiding motion along the ridges. In addition, the model predicts the tendency of cells to migrate circumferentially on convex substrates and axially on concave ones. Both of these predictions are verified experimentally, on several cell types. Altogether, our results identify the minimization of membrane-substrate adhesion energy and binding energy between the membrane protein complexes as key players of curvotaxis in cell migration.

biophysics↗

bioRxiv2022

Role of the two flagellar stators in swimming motility of Pseudomonas putida

In the soil bacterium Pseudomonas putida, the motor torque for flagellar rotation is generated by the two stators MotAB and MotCD. Here, we construct mutant strains, in which one or both stators are knocked out and investigate their swimming motility in fluids of different viscosity and in heterogeneous structured environments (semisolid agar). Besides bright-field imaging of single cell trajectories and spreading cultures, dual color fluorescence microscopy allows us to quantify the role of the stators in forming P. putidas three different swimming modes, where the flagellar bundle pushes, pulls, or wraps around the cell body. The MotAB stator is essential for swimming motility in liquids, while spreading in semisolid agar is not affected. Moreover, if the MotAB stator is knocked out, wrapped mode formation under low viscosity conditions is strongly impaired and only partly restored for increased viscosity and in semisolid agar. In contrast, when the MotCD stator is missing, cells are indistinguishable from the wild-type in fluid experiments, but spread much slower in semisolid agar. Analysis of the microscopic trajectories reveals that the MotCD knockout strain forms sessile clusters thereby reducing the number of motile cells, while the swimming speed is unaffected. Together, both stators ensure a robust wild-type that swims efficiently under different environmental conditions. IMPORTANCEBecause of its heterogeneous habitat, the soil bacterium Pseudomonas putida needs to swim efficiently under very different environmental conditions. In this paper, we knocked out the stators MotAB and MotCD to investigate their impact on swimming motility of P. putida. While the MotAB stator is crucial for swimming in fluids, in semisolid agar both stators are sufficient to sustain a fast swimming phenotype and increased frequencies of the wrapped mode, which is know to be beneficial for escaping mechanical traps. However, in contrast to the MotAB knock-out, a culture of MotCD knock-out cells spreads much slower in the agar as it forms non-motile clusters that reduce the amount of motile cells.

microbiology↗

bioRxiv2022

Versatile patterns in the actin cortex of motile cells: Self-organized pulses can coexist with macropinocytic ring-shaped waves

Self-organized patterns in the actin cytoskeleton are essential for eukaryotic cellular life. They are the building blocks of many functional structures that often operate simultaneously to facilitate, for example, nutrient uptake and movement of cells. However, to identify how qualitatively distinct actin patterns can coexist remains a challenge. Here, we use bifurcation theory to reveal a generic mechanism of pattern coexistence, showing that different types of wave patterns can simultaneously emerge in the actin system. Our theoretical analysis is complemented by live-cell imaging experiments revealing that narrow, planar, and fast-moving excitable pulses may indeed coexist with ring-shaped macropinocytic actin waves in the cortex of motile amoeboid cells.

biophysics↗