The Mechanisms Of Bcl-2 Family Proteins In The Regulation Of Ferroptosis

Ferroptosis is a form of cell death driven by iron-dependent accumulation of lipid peroxidation.As a regulated cell death,ferroptosis is considered to be a new target of tumor therapy.Previous studies found that ferroptosis was regulated by apoptosis signaling pathway and Bcl-2 family proteins,the core regulators of apoptosis,were involved in the interaction between apoptosis and ferroptosis.However,the specific effect and mechanisms of Bcl-2 family proteins on ferroptosis are still unclear.In this study,using a series of Bcl-2 family members deficient cell lines together with clone survival,cell viability and lipid peroxidation assays,we determined that the anti-apoptotic protein Bcl-xL inhibited ferroptosis and the pro-apoptotic protein Bax promoted ferroptosis.The above results preliminary showed that Bcl-xL and Bax played an important role in ferroptosis.Mechanistically,by detecting the levels of target proteins,we found that deletion of Bcl-xL not only promoted the expression of ACSL4 and LPCAT3,which provided substrate for lipid peroxidation,but also increased the expression of lipoxygenase cofactor PEBP1,which promoted lipid peroxidation;Next,the mass spectrometry and immunoprecipitation assay confirmed that Bcl-xL combined with Hsp90,knockout of Bcl-xL enhanced the binding of GPX4 to Hsp90 and LAMP2A,which promoted the lysosomal degradation of GPX4 and ferroptosis.These results suggested that anti-apoptotic protein Bcl-xL participated in ferroptosis by regulating lipid metabolism and GPX4.At the same time,RT-qPCR and Western Blot showed that Bak/Bax double deletion promoted the expression of Nrf2,and the highly expressed Nrf2 increased the mRNA and protein levels of GPX4,which promoted the antioxidant capacity and inhibited ferroptosis.Furthermore,using Bak or Bax defective cells,we found that Bax regulated GPX4 function.Bax deletion promoted GPX4 protein expression,while Bak deletion had no significant effect on GPX4 protein.The above results suggested that Bax but not Bak participated in ferroptosis by improving cellular antioxidant capacity.In conclusion,this study clarified the role and molecular mechanisms of anti-apoptotic protein Bcl-xL and pro-apoptotic protein Bax in ferroptosis.This study broadens the function of Bcl-2 family proteins and provides a new perspective for the clinical use of drugs designed for Bcl-2 family proteins.