ECM dimensionality tunes actin tension to modulate endoplasmic reticulum function and spheroid phenotypes of mammary epithelial cells

• Published in: The EMBO journal Vol. 41; no. 17; pp. e109205 - n/a
• Main Authors: Kai, FuiBoon, Ou, Guanqing, Tourdot, Richard W, Stashko, Connor, Gaietta, Guido, Swift, Mark F, Volkmann, Niels, Long, Alexandra F, Han, Yulong, Huang, Hector H, Northey, Jason J, Leidal, Andrew M, Viasnoff, Virgile, Bryant, David M, Guo, Wei, Wiita, Arun P, Guo, Ming, Dumont, Sophie, Hanein, Dorit, Radhakrishnan, Ravi, Weaver, Valerie M
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01.09.2022; John Wiley and Sons Inc
• Subjects: Actin; actin tension; Actins - metabolism; Basements; Binding; Calcium homeostasis; Cell viability; Cellular stress response; Curvature; Endoplasmic reticulum; Endoplasmic Reticulum - metabolism; Endoplasmic Reticulum Stress; Epithelial cells; Epithelial Cells - metabolism; Epithelium; Extracellular matrix; Filamins - metabolism; Homeostasis; Kinases; Mammary gland; membrane contact sites; Membrane proteins; Membranes; Organoids; Phenotype; Phenotypes; Plasma; Proteins; Resilience; Secretion; Spheroids; Tension; Topology; spheroids; membrane contact sites; endoplasmic reticulum; extracellular matrix; actin tension
• ISSN: 0261-4189; 1460-2075
• DOI: 10.15252/embj.2021109205

Patient‐derived organoids and cellular spheroids recapitulate tissue physiology with remarkable fidelity. We investigated how engagement with a reconstituted basement membrane in three dimensions (3D) supports the polarized, stress resilient tissue phenotype of mammary epithelial spheroids. Cells interacting with reconstituted basement membrane in 3D had reduced levels of total and actin‐associated filamin and decreased cortical actin tension that increased plasma membrane protrusions to promote negative plasma membrane curvature and plasma membrane protein associations linked to protein secretion. By contrast, cells engaging a reconstituted basement membrane in 2D had high cortical actin tension that forced filamin unfolding and endoplasmic reticulum (ER) associations. Enhanced filamin‐ER interactions increased levels of PKR‐like ER kinase effectors and ER‐plasma membrane contact sites that compromised calcium homeostasis and diminished cell viability. Consequently, cells with decreased cortical actin tension had reduced ER stress and survived better. Consistently, cortical actin tension in cellular spheroids regulated polarized basement membrane membrane deposition and sensitivity to exogenous stress. The findings implicate cortical actin tension‐mediated filamin unfolding in ER function and underscore the importance of tissue mechanics in organoid homeostasis. Synopsis How three‐dimensional spheroid cultures of mammary epithelial cells retain superior long‐term viability and stress resilience remains unclear. This work uncovers coupling between extracellular matrix (ECM) topology, cortical actin tension and endoplasmic reticulum (ER) function, which serves to safeguard protein secretion and cellular stress response. High cortical actin tension in 2D mammary epithelial cell cultures facilitates filamin‐PERK interaction and compromises ER function. In 3D spheroid cultures, mammary epithelial cell ‐ ECM interactions reduce cortical actin tension and filamin binding to PERK, maintaining ER homeostasis. Reduced cortical actin tension increases plasma membrane protrusion size and duration, favoring recruitment of negative curvature‐binding proteins that facilitate protein secretion. Three‐dimensional ECM supports ER homeostasis and protein secretion via a cortical actin‐filamin‐PERK axis in mammary epithelial spheroids.