Shear forces induce ICAM-1 nanoclustering on endothelial cells that impact on T-cell migration.

The leukocyte-specific β-integrin LFA-1 and its ligand ICAM-1, expressed on endothelial cells (ECs), are involved in the arrest, adhesion, and transendothelial migration of leukocytes. Although the role of mechanical forces on LFA-1 activation is well established, the impact of forces on its major ligand ICAM-1 has received less attention. Using a parallel-plate flow chamber combined with confocal and super-resolution microscopy, we show that prolonged shear flow induces global translocation of ICAM-1 on ECs upstream of flow direction.

Protocol for Efficient Protein Synthesis Detection by Click Chemistry in Colorectal Cancer Patient-Derived Organoids Grown .

Here, we describe a protocol to detect and visualize protein synthesis by click-chemistry-based immunofluorescence in patient-derived organoids (PDOs) . The protocol uses O-propargyl puromycin (OPP), an analog of puromycin that enters the acceptor site of ribosomes and is incorporated into nascent polypeptides. OPP can be detected by a click chemistry reaction and can be combined with conventional antibody staining.

Zonation of Ribosomal DNA Transcription Defines a Stem Cell Hierarchy in Colorectal Cancer.

Colorectal cancers (CRCs) are composed of an amalgam of cells with distinct genotypes and phenotypes. Here, we reveal a previously unappreciated heterogeneity in the biosynthetic capacities of CRC cells. We discover that the majority of ribosomal DNA transcription and protein synthesis in CRCs occurs in a limited subset of tumor cells that localize in defined niches.

Snail1-expressing fibroblasts in the tumor microenvironment display mechanical properties that support metastasis.

Crosstalk between tumor and stromal cells in the tumor microenvironment alter its properties in ways that facilitate the invasive behavior of tumor cells. Here, we demonstrate that cancer-associated fibroblasts (CAF) increase the stiffness of the extracellular matrix (ECM) and promote anisotropic fiber orientation, two mechanical signals generated through a Snail1/RhoA/αSMA-dependent mechanism that sustains oriented tumor cell migration and invasiveness. Snail1-depleted CAF failed to acquire myofibroblastic traits in response to TGFβ, including RhoA activation, αSMA-positive stress fibers, increased fibronectin fibrillogenesis, and production of a stiff ECM with oriented fibers.

The p65 subunit of NF-κB and PARP1 assist Snail1 in activating fibronectin transcription.

Snail1 is a transcriptional repressor of E-cadherin that triggers epithelial-mesenchymal transition (EMT). Here, we report assisted Snail1 interaction with the promoter of a typical mesenchymal gene, fibronectin (FN1), both in epithelial cells undergoing EMT and in fibroblasts. Together with Snail1, the p65 subunit of NF-κB and PARP1 bound to the FN1 promoter.

TFCP2c/LSF/LBP-1c is required for Snail1-induced fibronectin gene expression.

Fibronectins are cell-secreted glycoproteins that modulate cell attachment, spreading, migration, morphology, differentiation and oncogenic transformation. Fibronectin expression is activated during EMT (epithelial-mesenchymal transition) and is a hallmark of mesenchymal cells. It is shown in the present study that a transcription factor previously unrelated with EMT, TFCP2c/LSF/LBP-1c, was translocated to the nucleus and bound to the fibronectin promoter upon EMT induction by Snail1.