Jona Kayser

Slow dynamics and internal stress relaxation in bundled cytoskeletal networks

Crosslinked and bundled actin filaments form networks that are essential for the mechanical properties of living cells. Reconstituted actin networks have been extensively studied not only as a model system for the cytoskeleton, but also to understand the interplay between microscopic structure and macroscopic viscoelastic properties of network-forming soft materials. These constitute a broad class of materials with countless applications in science and industry. So far, it has been widely assumed that reconstituted actin networks represent equilibrium structures. Here, we show that fully polymerized actin/fascin bundle networks exhibit surprising age-dependent changes in their viscoelastic properties and spontaneous dynamics, a feature strongly reminiscent of out-of-equilibrium, or glassy, soft materials. Using a combination of rheology, confocal microscopy and space-resolved dynamic light scattering, we demonstrate that actin networks build up stress during their formation and then slowly relax towards equilibrium owing to the unbinding dynamics of the crosslinking molecules.

The Small Heat Shock Protein Hsp27 Affects Assembly Dynamics and Structure of Keratin Intermediate Filament Networks

The mechanical properties of living cells are essential for many processes. They are defined by the cytoskeleton, a composite network of protein fibers. Thus, the precise control of its architecture is of paramount importance. Our knowledge about the molecular and physical mechanisms defining the network structure remains scarce, especially for the intermediate filament cytoskeleton. Here, we investigate the effect of small heat shock proteins on the keratin 8/18 intermediate filament cytoskeleton using a well-controlled model system of reconstituted keratin networks. We demonstrate that Hsp27 severely alters the structure of such networks by changing their assembly dynamics. Furthermore, the C-terminal tail domain of keratin 8 is shown to be essential for this effect. Combining results from fluorescence and electron microscopy with data from analytical ultracentrifugation reveals the crucial role of kinetic trapping in keratin network formation.

Assembly kinetics determine the structure of keratin networks

Living cells exhibit an enormous bandwidth of mechanical and morphological properties that are mainly determined by the cytoskeleton. In metazoan cells this composite network is constituted of three different types of filamentous systems: actin filaments, microtubules and intermediate filaments. Keratin-type intermediate filaments are an essential component of epithelial tissues, where they comprise networks of filaments and filament bundles. However, the underlying mechanisms leading to this inherently polymorphic structure remain elusive. Here, we show that keratin filaments form kinetically trapped networks of bundles under near-physiological conditions in vitro. The network structure is determined by the intricate interplay between filament elongation and their lateral association to bundles and clusters.

Towards constructing extracellular matrix-mimetic hydrogels: An elastic hydrogel constructed from tandem modular proteins containing tenascin FnIII domains☆

Protein-based hydrogels have been developed for various biomedical applications where they provide artificial extracellular microenvironments that mimic the physical and biochemical characteristics of natural extracellular matrices (ECMs). In natural ECMs, a large number of proteins are tandem modular proteins consisting of many individually folded functional domains that confer structural and biological functionalities. Such tandem modular proteins are promising building blocks for constructing ECM-mimetic biomaterials. However, their use for such purposes has not been explored extensively. Tenascin-C (TNC) is an ECM tandem modular protein and plays an important role in mechanotransduction by regulating important cell-matrix interactions. The third FnIII domain of TNC (TNfn3) contains an RGD sequence and is known to bind integrins. Here we use the TNfn3 domain and resilin sequence-based tandem modular protein FRF4RF4R (F represents the TNfn3 domain and R represents the resilin sequence, respectively) as a building block to construct protein-based ECM-mimetic hydrogels. The tandem modular protein-based building block FRF4RF4R closely mimics the architecture of the naturally occurring tandem modular ECM protein TNC and incorporates intact RGD-containing FnIII domains. Our results demonstrate that tandem modular proteins containing TNfn3 can be readily photochemically crosslinked into elastic hydrogels, whose Youngs modulus can be tuned by the concentration of the tandem modular protein solution. In vitro studies demonstrate that none of the photochemical crosslinking reaction components are cytotoxic at the level tested, and the hydrogel supports the spread of human lung fibroblast cells. Our results demonstrate that FRF4RF4R-based hydrogel is a novel ECM-mimetic hydrogel.

Excess of mutational jackpot events in expanding populations revealed by spatial Luria–Delbrück experiments

The genetic diversity of growing cellular populations, such as biofilms, solid tumours or developing embryos, is thought to be dominated by rare, exceptionally large mutant clones. Yet, the emergence of these mutational jackpot events is only understood in well-mixed populations, where they stem from mutations that arise during the first few cell divisions. To study jackpot events in spatially structured populations, we track mutant clones in microbial populations using fluorescence microscopy and population sequencing. High-frequency mutations are found to be massively enriched in microbial colonies compared with well-shaken liquid cultures, as a result of late-occurring mutations surfing at the edge of range expansions. Thus, jackpot events can be generated not only when mutations arise early but also when they occur at favourable locations, which exacerbates their role in adaptation and disease. In particular, because spatial competition with the wild type keeps most mutant clones in a quiescent state, strong selection pressures that kill the wild type promote drug resistance.