Abstract
Vimentin intermediate filaments, together with actin filaments and microtubules, constitute the cytoskeleton in cells of mesenchymal origin. The mechanical properties of the filaments themselves are encoded in their molecular architecture and depend on their ionic environment. It is thus of great interest to disentangle the influence of both the ion type and their concentration on vimentin assembly. We combine small angle X-ray scattering and fluorescence microscopy and show that vimentin in the presence of the monovalent ions, K+ and Na+, assembles into "standard filaments" with a radius of about 6 nm and 32 monomers per cross-section. In contrast, di- and multivalent ions, independent of type and valency, lead to the formation of thicker filaments associating over time into higher order structures. Hence, our results may indeed be of relevance for living cells, as local ion concentrations in the cytoplasm during certain physiological activities may differ considerably from average intracellular concentrations.
Highlights
Networks of Excitable Cells’’ (MBExC), University of Gottingen, Germany † Electronic supplementary information (ESI) available: Comparison of different buffer systems; azimuthal integration error; detailed description of the theoretical model; precipitation thresholds; additional data plots
To systematically investigate the influence of different ions on vimentin assembly, bundling and network formation, we study five different ions, i.e., K+ and Na+, Mg2+ and Ca2+, Co3+, as well as spermine, Sp4, a polyamine that is a tetravalent polycation in solution at physiological pH
This is in agreement with the literature, where Herrmann et al found filaments with up to two or three times more monomers per crosssection when vimentin is assembled in the presence of MgCl2.19 In particular, we show that vimentin forms single filaments in the presence of monovalent ions and networks in the presence of multivalent ions
Summary
Networks of Excitable Cells’’ (MBExC), University of Gottingen, Germany † Electronic supplementary information (ESI) available: Comparison of different buffer systems; azimuthal integration error; detailed description of the theoretical model; precipitation thresholds; additional data plots. When renatured two monomers form parallel coiled-coils and, subsequently, anti-parallel, half-staggered tetramers that are stable in low salt buffer. Tetramers have a length of 60 nm and a diameter of 5 nm.[14,15] Upon addition of salt, tetramers laterally assemble into unit length filaments (ULFs), which – in the case of vimentin – typically consist of 32 monomers. This lateral assembly step is followed by longitudinal annealing and a compaction step, after which the filament has a diameter of 11 nm and a repeat along the filament of 43 nm.
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