Nucleocytoplasmic transport senses mechanics independently of cell density in cell monolayers.

Cells sense and respond to mechanical forces through mechanotransduction, which regulates processes in health and disease. In single adhesive cells, mechanotransduction involves the transmission of force from the extracellular matrix to the cell nucleus, where it affects nucleocytoplasmic transport (NCT) and the subsequent nuclear localization of transcriptional regulators such as YAP. However, if and how NCT is mechanosensitive in multicellular systems is unclear. Here, we characterize and use a fluorescent sensor of nucleocytoplasmic transport (Sencyt) and demonstrate that nucleocytoplasmic transport responds to mechanics but not cell density in cell monolayers. Using monolayers of both epithelial and mesenchymal phenotype, we show that NCT is altered in response both to osmotic shocks, and to the inhibition of cell contractility. Further, NCT correlates with the degree of nuclear deformation measured through nuclear solidity, a shape parameter related to nuclear envelope tension. In contrast, YAP but NCT is sensitive to cell density, showing that YAP response to cell-cell contacts is not via a mere mechanical effect of NCT. Our results demonstrate the generality of the mechanical regulation of NCT.

Membrane to cortex attachment determines different mechanical phenotypes in LGR5+ and LGR5- colorectal cancer cells.

Colorectal cancer (CRC) tumors are composed of heterogeneous and plastic cell populations, including a pool of cancer stem cells that express LGR5. Whether these distinct cell populations display different mechanical properties, and how these properties might contribute to metastasis is poorly understood. Using CRC patient derived organoids (PDOs), we find that compared to LGR5- cells, LGR5+ cancer stem cells are stiffer, adhere better to the extracellular matrix (ECM), move slower both as single cells and clusters, display higher nuclear YAP, show a higher survival rate in response to mechanical confinement, and form larger transendothelial gaps. These differences are largely explained by the downregulation of the membrane to cortex attachment proteins Ezrin/Radixin/Moesin (ERMs) in the LGR5+ cells. By analyzing single cell RNA-sequencing (scRNA-seq) expression patterns from a patient cohort, we show that this downregulation is a robust signature of colorectal tumors. Our results show that LGR5- cells display a mechanically dynamic phenotype suitable for dissemination from the primary tumor whereas LGR5+ cells display a mechanically stable and resilient phenotype suitable for extravasation and metastatic growth.