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      The Helical Superstructure of Intermediate Filaments

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          Abstract

          Intermediate filaments are the least explored among the large cytoskeletal elements. We show here that they display conformational anomalies in narrow microfluidic channels. Their unusual behavior can be understood as the consequence of a previously undetected, large scale helically curved superstructure. Confinement in a channel orders the otherwise soft, strongly fluctuating helical filaments and enhances their structural correlations, giving rise to experimentally detectable, strongly oscillating tangent correlation functions. We propose an explanation for the detected intrinsic curving phenomenon - an elastic shape instability that we call autocoiling. The mechanism involves self-induced filament buckling via a surface stress located at the outside of the cross-section. The results agree with ultrastructural findings and rationalize for the commonly observed looped intermediate filament shapes.

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          The pivot algorithm: A highly efficient Monte Carlo method for the self-avoiding walk

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            The statistics and dynamics of confined or entangled stiff polymers

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              Deconstructing the Late Phase of Vimentin Assembly by Total Internal Reflection Fluorescence Microscopy (TIRFM)

              Quantitative imaging of intermediate filaments (IF) during the advanced phase of the assembly process is technically difficult, since the structures are several µm long and therefore they exceed the field of view of many electron (EM) or atomic force microscopy (AFM) techniques. Thereby quantitative studies become extremely laborious and time-consuming. To overcome these difficulties, we prepared fluorescently labeled vimentin for visualization by total internal reflection fluorescence microscopy (TIRFM). In order to investigate if the labeling influences the assembly properties of the protein, we first determined the association state of unlabeled vimentin mixed with increasing amounts of labeled vimentin under low ionic conditions by analytical ultracentrifugation. We found that bona fide tetrameric complexes were formed even when half of the vimentin was labeled. Moreover, we demonstrate by quantitative atomic force microscopy and electron microscopy that the morphology and the assembly properties of filaments were not affected when the fraction of labeled vimentin was below 10%. Using fast frame rates we observed the rapid deposition of fluorescently labeled IFs on glass supports by TIRFM in real time. By tracing their contours, we have calculated the persistence length of long immobilized vimentin IFs to 1 µm, a value that is identical to those determined for shorter unlabeled vimentin. These results indicate that the structural properties of the filaments were not affected significantly by the dye. Furthermore, in order to analyze the late elongation phase, we mixed long filaments containing either Alexa 488- or Alexa 647-labeled vimentin. The ‘patchy’ structure of the filaments obtained unambiguously showed the elongation of long IFs through direct end-to-end annealing of individual filaments.
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                Author and article information

                Journal
                13 March 2018
                Article
                1803.04691
                32c1bf95-e3c0-44b9-b905-0fe489e157fc

                http://arxiv.org/licenses/nonexclusive-distrib/1.0/

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                cond-mat.soft

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