Mechanics and Motility of Epithelial Cells: From Single Cell Behavior to Collective Migration
von Mark Skamrahl
Datum der mündl. Prüfung:2022-07-05
Erschienen:2023-01-13
Betreuer:Prof. Dr. Andreas Janshoff
Gutachter:Prof. Dr. Andreas Janshoff
Gutachter:Prof. Dr. Sarah Köster
Gutachter:Prof. Dr. Jörg Großhans
Gutachter:Prof. Dr. Silvio O. Rizzoli
Gutachter:Prof. Dr. Holger Bastians
Gutachter:Prof. Dr. Timo Betz
Zum Verlinken/Zitieren:
http://dx.doi.org/10.53846/goediss-9668
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Zusammenfassung
Englisch
Mechanical behavior of cells plays a crucial role in a plethora of biological processes. Despite its importance, this aspect of life science has only more recently gained increasing interest. Yet, without mechanical force the most basic motion cannot be initiated at any length scale, ranging from single molecules to cells, organs, and the whole organism. Similar to human motion, cells employ mechanical force to move directedly, accomplishing diverse tasks. The interplay of motility and mechanics of single cells and force transmission between tightly connected neighbor cells shapes epithelial behavior, giving rise to collective phenomena. In this dissertation, three main aspects of epithelial cell mechanics and motility are discussed. In the first project (Chapter 3), a simultaneous combination of atomic force microscopy (AFM) and fluorescence recovery after photobleaching (FRAP) was established, calibrated and then applied to cells. Here, AFM was used to provide a mechanical stimulus to single cells. In a simultaneous manner, FRAP was applied as the main readout tool, quantifying the turnover of actin in cellular stress fibers. Systematic application of forces ranging from 100 pN to 10 nN revealed a mechanical adaptation which scaled in an exponential manner: With increasing force application, actin turnover was downregulated to yield longer filaments, which could potentially withstand the external stress better. This novel combination of classic biophysical methods and its first application may enable promising insights into diverse mechanoadaptive processes, such as cell migration. Chapter 4 discusses collective migration of epithelial cell layers. Wildtype (WT) cells and ones that lack tight junction (TJ) components (ZO1/2 double knockdown, dKD) were compared. dKD cell layers were found to be immobile and jammed. This was attributed to an extreme upregulation of actomyosin contractility upon dKD. However, not all cells were able to contract: In a tug-of-war mechanism, only some cells were able to contract, thereby pulling on their neighbors, which in turn were stretched laterally. The laterally contracted cells lacked directed motion and, thus, were particularly responsible for the jamming. In contrast, the larger, stretched cells remained more mobile. This mechanism was confirmed in co-cultures of WT and dKD cells, as the contractile dKD cells slowed down the whole layer. Overall, collective migration was abolished upon TJ disruption. Interestingly, single dKD cells remained motile, rendering the described mechanism a highly collective effect. This work demonstrates that TJs are vital for collective cell migration and tissue fluidity. Building on results from Chapter 4, the third research project in Chapter 5 focused on co-cultures of dKD and WT cells. In initial migration measurements of these co-cultures a distinct demixing into clusters was observed but its origin remained unknown. Therefore, new mixing experiments were streamlined to specifically address the governing mechanism of this clustering. Here, dKD and WT cells demixed significantly compared with WT/WT controls. To explain this behavior, the tug-of-war from the previous project was examined further, this time with all dKD cells able to contract, stretching out WT neighbors. The WT cells mainly responded by excess surface area dilatation. The tug-of-war resulted in tension increase at all junctions connected to dKD cells. In addition, the dKD cells exhibited weaker cell-cell adhesion. Finally, to assess the relative impact of differential adhesion and contractility on demixing, the latter was specifically decreased. This resulted in a slowing of the initial segregation but the final demixed state remained unchanged. Accordingly, differential contractility is needed for efficient early cell segregation but adhesion dominates on longer time scales. This well-controlled case of demixing provides novel insights into how physical forces govern cell sorting, which is crucial during development.
Keywords: biophysics; cell mechanics; tight junctions; migration; cell sorting; tension; contractility; adhesion; actin; atomic force microscopy; jamming; collective behavior; epithelial cells; collective migration; myosin; fluorescence recovery after photobleaching; mechanobiology