mTORC2/Akt axis promotes proteotoxic stress and mitochondrial Ca overload during celastrol-induced paraptosis.

Celastrol, a triterpenoid with anticancer potential, induces paraptosis in breast cancer cells-a non-apoptotic form of cell death characterized by vacuolization of the endoplasmic reticulum (ER) and mitochondria. Although celastrol shows therapeutic promise, the signaling mechanisms mediating this pathway remain poorly defined. Here, we report that celastrol transiently activates both mTORC1 and mTORC2; however, only the mTORC2/Akt axis is essential for executing paraptotic cell death. Genetic or pharmacological inhibition of mTORC2 or Akt attenuated celastrol-induced cell death, reduced proteotoxic stress-evident from diminished polyubiquitinated protein accumulation and CHOP expression-and suppressed mitochondrial Ca overload by downregulating the mitochondrial calcium uniporter (MCU) and MICU1. Knockdown of Raptor (mTORC1 component) did not affect these processes, indicating a specific role for mTORC2. These findings define mTORC2/Akt signaling as a pro-death regulator in paraptosis, highlighting its unexpected role in driving proteotoxic and mitochondrial stress.