equibiaxial-stretching-device-for-high-magnification-live-cell

Equibiaxial Stretching Device for High Magnification Live-Cell Confocal Fluorescence Microscopy

Cells are continuously exposed to mechanical forces in physiological and pathological situations, including a variety of tensile and compressive stresses. ...

Barcelona Institute of Science and Technology (BIST)

King's College London

YAP is a mechanosensitive transcriptional activator with a critical role in cancer, regeneration, and organ size control. Here, we show that force applied to the nucleus directly drives YAP nuclear translocation by decreasing the mechanical restriction of nuclear pores to molecular transport. Exposure to a stiff environment leads cells to establish a mechanical connection between the nucleus and the cytoskeleton, allowing forces exerted through focal adhesions to reach the nucleus. Force transmission then leads to nuclear flattening, which stretches nuclear pores, reduces their mechanical resistance to molecular transport, and increases YAP nuclear import. The restriction to transport is further regulated by the mechanical stability of the transported protein, which determines both active nuclear transport of YAP and passive transport of small proteins. Our results unveil a mechanosensing mechanism mediated directly by nuclear pores, demonstrated for YAP but with potential general applicability in transcriptional regulation.

Force Triggers YAP Nuclear Entry by Regulating Transport across Nuclear Pores.

The cellular microenvironment is highly heterogeneous and dynamic. Therefore, cells must be equipped with molecular tools to adapt and respond to constantly fluctuating inputs. One such input is mechanical force, which activates signalling and regulates cell behaviour in the process of mechanotransduction. Whereas the mechanisms activating mechanotransduction are well studied, the reversibility of this process, whereby cells disassemble and reverse force-activated signalling pathways upon cessation of mechanical stimulation is far less understood. In this review we will outline some of the key experimental techniques to investigate the reversibility of mechanical signalling, and key discoveries arising from them.

The reversibility of cellular mechano-activation.

The mechanical properties of the extracellular matrix dictate tissue behaviour. In epithelial tissues, laminin is a very abundant extracellular matrix component and a key supporting element. Here we show that laminin hinders the mechanoresponses of breast epithelial cells by shielding the nucleus from mechanical deformation. Coating substrates with laminin-111-unlike fibronectin or collagen I-impairs cell response to substrate rigidity and YAP nuclear localization. Blocking the laminin-specific integrin β4 increases nuclear YAP ratios in a rigidity-dependent manner without affecting the cell forces or focal adhesions. By combining mechanical perturbations and mathematical modelling, we show that β4 integrins establish a mechanical linkage between the substrate and keratin cytoskeleton, which stiffens the network and shields the nucleus from actomyosin-mediated mechanical deformation. In turn, this affects the nuclear YAP mechanoresponses, chromatin methylation and cell invasion in three dimensions. Our results demonstrate a mechanism by which tissues can regulate their sensitivity to mechanical signals.

The laminin-keratin link shields the nucleus from mechanical deformation and signalling.

The cell nucleus is continuously exposed to external signals, of both chemical and mechanical nature. To ensure proper cellular response, cells need to regulate not only the transmission of these signals, but also their timing and duration. Such timescale regulation is well described for fluctuating chemical signals, but if and how it applies to mechanical signals reaching the nucleus is still unknown. Here we demonstrate that the formation of fibrillar adhesions locks the nucleus in a mechanically deformed conformation, setting the mechanical response timescale to that of fibrillar adhesion remodelling (~1 hour). This process encompasses both mechanical deformation and associated mechanotransduction (such as via YAP), in response to both increased and decreased mechanical stimulation. The underlying mechanism is the anchoring of the vimentin cytoskeleton to fibrillar adhesions and the extracellular matrix through plectin 1f, which maintains nuclear deformation. Our results reveal a mechanism to regulate the timescale of mechanical adaptation, effectively setting a low pass filter to mechanotransduction.

Article	Year
Force Triggers YAP Nuclear Entry by Regulating Transport across Nuclear Pores. Cell\| PubMed ID: 29107331	2017
The reversibility of cellular mechano-activation. Current opinion in cell biology\| PubMed ID: 37633090	2023
The laminin-keratin link shields the nucleus from mechanical deformation and signalling. Nature materials\| PubMed ID: 37709930	2023
Fibrillar adhesion dynamics govern the timescales of nuclear mechano-response via the vimentin cytoskeleton. bioRxiv : the preprint server for biology\| PubMed ID: 37986921	2023

Amy E. M. Beedle

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