Effective Cell Membrane Tension is Independent of Substrate Stiffness

Most animal cells are surrounded by a cell membrane and an underlying actomyosin cortex. Both structures are linked with each other, and they are under tension. Membrane tension and cortical tension both influence many cellular processes, including cell migration, division, and endocytosis. However, while actomyosin tension is regulated by substrate stiffness, how membrane tension responds to mechanical substrate properties is currently poorly understood. Here, we probed the effective membrane tension of neurons and fibroblasts cultured on glass and polyacrylamide substrates of varying stiffness using optical tweezers. In contrast to actomyosin-based traction forces, both peak forces and steady state tether forces of cells cultured on hydrogels were independent of substrate stiffness and did not change after blocking myosin II activity using blebbistatin, indicating that tether and traction forces are not directly linked with each other. Peak forces on hydrogels were about twice as high in fibroblasts if compared to neurons, indicating stronger membrane-cortex adhesion in fibroblasts. Finally, tether forces were generally higher in cells cultured on hydrogels compared to cells cultured on glass, which we attribute to substrate-dependent alterations of the actomyosin cortex and an inverse relationship between tension along stress fibres and cortical tension. Our results provide new insights into the complex regulation of membrane tension, and they pave the way for a deeper understanding of biological processes instructed by it. ### Competing Interest Statement The authors have declared no competing interest.