Vimentin filament transport and organization revealed by single-particle tracking and 3D FIB-SEM

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Journal of Cell Biology. 2025 Apr 03;224:e202406054. doi: 10.1083/jcb.202406054

Vimentin filament transport and organization revealed by single-particle tracking and 3D FIB-SEM Hess LabLippincott-Schwartz LabCellMap

Renganathan B, Moore AS, Yeo W, Petruncio A, Ackerman D, Weigel AV, Team TC, Pasolli HA, Xu CS, Shtengel G, Hess HF, Serpinskaya AS, Zhang HF, Lippincott-Schwartz J, Gelfand VI

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Abstract

Vimentin intermediate filaments (VIFs) form complex, tightly packed networks; due to this density, traditional imaging approaches cannot discern single-filament behavior. To address this, we developed and validated a sparse vimentin-SunTag labeling strategy, enabling single-particle tracking of individual VIFs and providing a sensitive, unbiased, and quantitative method for measuring global VIF motility. Using this approach, we define the steady-state VIF motility rate, showing a constant ∼8% of VIFs undergo directed microtubule-based motion irrespective of subcellular location or local filament density. Significantly, our single-particle tracking approach revealed uncorrelated motion of individual VIFs within bundles, an observation seemingly at odds with conventional models of tightly cross-linked bundles. To address this, we acquired high-resolution focused ion beam scanning electron microscopy volumes of vitreously frozen cells and reconstructed three-dimensional VIF bundles, finding that they form only loosely organized, semi-coherent structures from which single VIFs frequently emerge to locally engage neighboring microtubules. Overall, this work demonstrates single VIF dynamics and organization in the cellular milieu for the first time.

bioRxiv Preprint: https://doi.org/10.1101/2024.06.10.598346

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Vimentin filament transport and organization revealed by single-particle tracking and 3D FIB-SEM

Abstract

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