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Molecular mechanisms of the mechanical interaction between the cell nucleus and the actin cytoskeleton

Periodic Reporting for period 1 - MiMEtiC (Molecular mechanisms of the mechanical interaction between the cell nucleus and the actin cytoskeleton)

Reporting period: 2017-03-01 to 2019-02-28

Cell survival and tissue integrity depend on the ability to maintain structural and functional integrity even under conditions of stress. Cells are constantly exposed to dynamic changes in their physical microenvironment, and these mechanical forces is a source of physiologically and clinically relevant stress. Mechanical stress protection operates during cell differentiation, adhesion and migration, and is of particular importance for maintaining tissues such as skeletal muscle, heart, lung, vasculature, and skin. Yet, underlying molecular mechanisms have not been comprehensively studied. However, what is already obvious is that impairment mechanical stress protection and/or mechanoadaptation processes gives rise to diverse diseases, including myopathies, heart and kidney failure, leukocyte adhesion deficiencies, progeroid diseases and cancer. Thus, there is a critical research need for detailed molecular understanding of mechanical stress responses in order to design diagnostic tools and therapeutic interventions.
Epithelial tissues such as the skin epidermis are critical load-bearing elements of the body. They undergo large-scale, force-driven deformations during morphogenesis and in the adult organism. Strikingly, in contrast to cancer cells where mechanical deformations have been shown to induce nuclear rupture and DNA damage, such effects have not been observed in untransformed epithelial cells. On the contrary, epithelial cells have been shown to be capable of undergoing extreme deformation without damage, and to withstand long-term mechanical strain without signs of genomic damage or loss of viability. This project aimed to understand how nuclei respond to mechanical stress and how the genome is protected under extreme nuclear deformations
I hypothesized that non-cancerous cells such as skin epidermal stem/progenitor cells (EPCs) that are exposed to large-scale, dynamic mechanical force in their natural microenvironment, have acquired mechanisms and/or material properties that protect their genome against mechanical stress and dynamic changes in their force environment.
EPCs were chosen because they can be maintained in a stem cell state as well as differentiated in vitro to obtain epidermal lineages, and thus represent excellent model to identify and study mechanically controlled genetic programs that drive cell fate decisions.
To identify nuclear mechanotransduction pathways, I exposed EPCs to diverse regimes of cyclic uniaxial mechanical strain (5%, 20%, 40%, frequency 0.1 Hz). By using a large panel of experimental tools, including super resolution and regular confocal imaging, electron microscopy, quantitative phosphoproteomics, atomic force microscopy and a number of biochemical techniques, I observed that uniaxial strain, even at low strain levels, triggers nuclear deformation that catalyzes remodeling of the nuclear lamina and the lamina-associated heterochromatin. This remodeling facilitated an increase in nuclear elasticity. Intriguingly, blocking this response induced DNA damage upon strain, demonstrating that the remodeling of the lamina-associated heterochromatin is required for genome protection in the presence of strain. I further observed that high levels of strain induced a slower response where cells aligned their long axis and cytoskeletal fibers perpendicular to the direction of strain. This allowed the cells to dissipate the mechanical force before it reached the nucleus, allowing the cells to restore their steady state nuclear architecture. This project highlighted a novel, active role for chromatin as an element that can dynamically adapt to exogenous mechanical stresses by altering its own state to protect the genetic material
This interdisciplinary project provided significant advances in stem cell mechanobiology and unraveled a novel, active role for chromatin as an element that can dynamically adapt to exogenous mechanical stresses by altering its own state to protect the genetic material. Stem cell machanobiology is a field of biomedicine with great potential for societal and economic impact due to fundamental implications in degenerative disease and physiological aging. In addition to uncovering fundamental mechanisms of stem cell and tissue degeneration, this project findings on stem cell genome protection may open completely novel strategies to identify early changes in tissue degeneration to be used in diagnosis, as well as therapeutic tools to prevent degeneration or facilitate the renewal of aging tissues.