Cytoskeleton disruption and plasma membrane damage determine methuosis of normal and malignant cells

Abstract

Methuosis represents a novel cell death modality characterized by catastrophic cytoplasmic vacuolization in normal and malignant cells. However, the critical role and the underlying mechanism of cytoskeleton and plasma membrane damage in methuotic cells are largely unknown. Herein, maduramicin-treated myocardial cells (H9c2) and indole chalcone-exposed glioma cells (U251) were used as methuosis model to uncover this secret. We found that cytoskeleton protein F-actin, α-tubulin, β-tubulin and filamin A/B were disrupted in a reversible-dependent manner. In addition, RhoA-ROCK1 signaling pathway mediated cytoskeleton disruption in methuotic cells. Excessive cytoplasmic vacuolization triggered cellular plasma membrane damage and the release of DAMPs, including LDH, ATP and CRT. Furthermore, at the end phase of methuotic cells, plasma membrane was damaged independent of pore-forming protein p-MLKL and GSDMD. Endosomal sorting complex required for transport (ESCRT)-Ⅲ especially its subunit CHMP3 and CHMP5 negatively regulated excessive vacuolization-induced plasma membrane damage in cells undergoing methuosis. In conclusion, for the first time, the critical role and potential mechanism of cytoskeleton and plasma membrane damage in methuotic cells are known, which would facilitate the employment of methuosis in life science and pharmacology.