Cytoskeletal networks in living cells under strain

by Ruth Meyer
Doctoral thesis
Date of Examination:2025-05-23
Date of issue:2025-07-15
Advisor:Prof. Dr. Sarah Köster
Referee:Prof. Dr. Sarah Köster
Referee:Prof. Dr. Timo Betz
Referee:Prof. Dr. Claudia Steinem
Referee:Prof. Dr. Alexander Egner
Referee:Prof. Dr. Stefan Klumpp
Referee:Dr. Peter Lenart
Persistent Address: http://dx.doi.org/10.53846/goediss-11385

 

 

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Abstract

English

The eukaryotic cytoskeleton consists of three types of filamentous proteins: actin filaments, microtubules and intermediate filaments (IFs). In contrast to actin and tubulin, IF proteins are expressed in a cell-type specific manner, and keratins are found in epithelial cells. In certain cell types, the keratins form a layer close to the membrane which may be referred to as an “IF-cortex”. It is hypothesized that this IF-cortex arranges with radial bundles in a “rim-and-spokes” structure in epithelia. Based on this hypothesis, IFs and actin filaments might add complementary mechanical properties to the cortex. It was previously shown that single IFs in vitro remain undamaged at high strains. We now ask the question of whether this unique force-extension behavior of single IFs is also relevant in the filament network within a cell. In this thesis, we combine cell stretching experiments on MDCK II wild-type and keratin-deficient cells with atomic force microscopy to measure the viscoelastic properties and determine the role of keratin for the cell cortex. Furthermore, we compare the influence of different stretching modes on the cell and nucleus shape to investigate the mechanical link from the periphery of the cell to the nucleus through the keratin network. Additionally, we stretch three-dimensional MDCK II cysts to high strains providing another point of view by enabling imaging of the cross section of the cells. Finally, we compare the structure of the actin and keratin networks in the cells under strain. We find that keratin-deficient cells compensate for the missing keratin, but are nevertheless very sensitive to external strain, whereas the intricate interplay between the actin and keratin cortices provides a protective mechanism to the cell by preserving the mechanical state and cell stability. Hence, this thesis provides intriguing insights on cells under strain from a mechanical to a structural level.
Keywords: cytoskeleton; keratin; actin; cell stretching; atomic force microscopy; fluorescence microscopy; cell mechanics; intermediate filaments