Pushing from within: Control of cell shape, integrity and motility by cytoskeletal pushing forces

Project Information

PushingCell

Grant agreement ID: 101071793

DOI

10.3030/101071793

EC signature date 26 April 2023

Start date 1 May 2023

End date 30 April 2029

Funded under

• European Research Council (ERC)

Total cost

€ 9 813 625,00

EU contribution

€ 9 813 625,00

Coordinated by

INSTITUTE OF SCIENCE AND TECHNOLOGY AUSTRIA
Austria

Periodic Reporting for period 1 - PushingCell (Pushing from within: Control of cell shape, integrity and motility by cytoskeletal pushing forces)

Reporting period: 2023-05-01 to 2024-10-31

Summary of the context and overall objectives of the project

The ability of cells to sense environmental cues and respond to them by adjusting their shape and motion
is fundamental for biological processes ranging from animal development to immune responses and cancer metastasis. Currently, a lot is known about how cells adhere to and pull on their surroundings. Very little research has been done on the biology and physics of pushing forces exerted by cells. The goal of the project is to fill this knowledge gap.
We investigate how cells respond to obstacles they encounter during their migratory path without binding to them. Using micro-engineered substrates, tissue mimics, in-vitro reconstitution and physical modelling, we study the interface between the obstacle, the plasma membrane of the cell and the underlying cytoskeleton. Our work provides new fundamental insights into biological and physical principles underlying the control of cell shape, integrity and motility, which are key to many physiological processes from development and homeostasis to cancer, immune responses and regeneration.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

We found that the cytoskeleton of migrating leukocytes simultaneously pushes in two different directions and uncovered how the machinery generating these pushing forces is coordinated to prevent that the cell loses its physical coherence. One pushing component is in the direction of the migratory path and therefore advances the movement of the cell front. The other is in the cell center, pushes perpendicular to the direction of migration and serves to dilate the extracellular environment to make space for bulky organelles like the nucleus. We describe a novel molecular pathway that coordinates these two pushing functions of the cytoskeleton. We are also setting-up an optical tweezer system to measure the pushing forces exerted by the cellular protrusions at the cell front.
Furthermore, we developed a physical model based on the theory of active gels to describe the actin cytoskeleton growing on a curved substrate mimicking the cell membrane deformed by external obstacles. We calculated how the pushing force developed by the cytoskeleton depends on the actin polymerisation dynamics. We found that this force, whichh is qualitatively affected by proteins sensing the membrane curvature, can affect the actin dynamics, possibly leading to a "curvature instability" leading to the formation of spontaneous cellular protrusions. In parallel, we are developing an in vitro assay using purified components to test this theoretical model.
Finally, we are developing tools that will allow us to specifically perturb different components of the cytoskeleton to evaluate their contribution to pushing forces. We have generated tools that allow to rapidly remove Intermediate filaments from a specific cell region and to rapidly disassemble a specific subpopulation of stable, post-translationally modified microtubules.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

We developed new technology to measure the pushing forces exerted by migrating cells. We established microfluidic channel systems that are physically very soft and therefore allow the cells to deform them while they pass through. The deformations of the channels by the passing cells can be visualized and quantified to infer the pushing forces that the cells apply onto the substrate. We have also generated optogenetic and chemogenetic tools to rapidly and specifically manipulate cytoskeletal components, such intermediate filaments and specific microtubule subpopulations. Finally, we developed a new model for active gels growing on curved substrates.