The Prohibitin-Binding Compound Fluorizoline Activates the Integrated Stress Response through the eIF2α Kinase HRI

Abstract

Fluorizoline is a synthetic molecule that induces apoptosis, by selectively targeting prohibitins (PHBs), through induction of the BH3-only protein NOXA. This induction is transcriptionally regulated by the integrated stress response (ISR)-related transcription factors ATF3 and ATF4. Here, we evaluate the role of the four eIF2a kinases, to decipher which is responsible for the mechanism of ISR activation triggered by fluorizoline in HeLa and HAP1 cells. First, we demonstrated the involvement of the eIF2a kinases using ISR inhibitor (ISRIB) and by simultaneous downregulation of all four eIF2a kinases, as both approaches were able to increase cell resistance to fluorizoline-induced apoptosis. Furthermore, we confirmed that fluorizoline treatment results in endoplasmic reticulum (ER) stress, as evidenced by PERK activation. Despite PERK activation, this kinase was not directly involved in the ISR activation by fluorizoline. In this regard, we found that the eIF2a kinases are capable of compensating for each other's loss of function. Importantly, we demonstrated that the mitochondrial-stress-related eIF2a kinase HRI mediates ISR activation after fluorizoline treatment.