Both monovalent cations and plectin are potent modulators of mechanical properties of keratin K8/K18 networks

Abstract

Intermediate filament (IF) networks are a major contributor to cell rigidity and thus serve as vital elements to preserve the integrity of entire cell layers. Keratin K8 and K18 IFs are the basic constituents of the cytoskeleton of epithelial cells. The mechanical properties of K8/K18 networks depend on the structural arrangements of individual filaments within the network. This paper investigates the architecture of these networks in vitro under the influence of the monovalent cation potassium and that of the cytolinker protein plectin. Whereas increasing amounts of potassium ions lead to filament bundling, plectin interlinks filaments at filament intersection points but does not lead to bundle formation. The mechanics of the resulting networks are investigated by microrheology with assembled K8/K18 networks. It is shown that bundling induced by potassium ions significantly stiffens the network. Furthermore, our measurements reveal an increase in plectin-mediated keratin network rigidity as soon as an amount corresponding to more than 20% of the plectin present in cells is added to the keratin IF networks. In parallel, we investigated the influence of plectin on cell rigidity in detergentextracted epithelial vulva carcinoma derived A431 cells in situ. These cytoskeletons, containing mostly IFs, actin filaments and associated proteins, exhibit a significantly decreased stiffness, when plectin is downregulated to E10% of the normal value. Therefore, we assume that plectin, via the formation of IF-IF connections and crosslinking of IFs to actin filaments, is an important contributor to cell stiffness.

Stiffening and inelastic fluidization in vimentin intermediate filament networks

“…35 In addition it will be interesting to examine the interplay between noncovalent tail-mediated interactions and mobile crosslinkers by introducing reconstituted plectin into intermediate filament networks. 57 Figure S1: Onset stress σ 0 of nonlinearity in the shear moduli of vimentin networks (left) and linear modulus G 0 as a function of loading rate. Dashed lines indicate average values, shaded areas indicate an interval of one standard deviation.…”

Keratins determine network stress responsiveness in reconstituted actin–keratin filament systems

“…Keratin can assemble in vitro into networks in an unusually low ionic strength buffer 24 and addition of ions such as magnesium (Mg 2+ ) or potassium (K + ) at physiological concentrations is enough to induce immediate bundling, resulting in heterogeneous networks. [29][30][31] In contrast, for actin the addition of mono-and divalent ions to physiological concentrations is essential to initiate the polymerization from globular actin (G-actin) to filamentous actin (F-actin) to form entangled networks. 32 A striking difference between actin and keratin filaments is the filament persistence length, which is the length scale for the decay of the tangent-tangent correlation along the filament contour, and is proportional to the stiffness of the polymer.…”

Comparative Proteomic Analysis of Lysine Acetylation in Fish CIK Cells Infected with Aquareovirus

“…KRTs are intermediate filament proteins which act as essential cytoskeletal component involved in the maintenance of cell morphology [ 32 , 33 ]. Plectin binds to various cytoskeletal proteins including microtubules and intermediate filaments and regulates cellular survival, growth, and polarization signaling pathways, and also can modulate the mechanical properties of keratin K8/K18 networks [ 34 , 35 ]. PDIA3 is a thiol-oxidoreductase chaperone with multiple functions, including the proper folding of newly synthesized proteins and assembly of major histocompatibility complex I [ 36 ].…”