The Molecular Mechanism Of PKC Signaling Pathway In Ferroptosis

Ferroptosis is a novel form of programmed cell death.To systematically investigate the mechanism of ferroptosis,a genome wide RNAi screening was performed and PKC was identified as a positive candidate of ferroptosis.There are 4members in the classic PKC family,including PKCα,PKCβI,PKCβII and PKCγ.In this study,I first found PKC inhibitors can block ferroptosis in MEFs,HT1080 and MDA-MB-231 Cells.To further confirm the RNAi screening results,I knocked down PKCβ in HT1080,MEF and MDA-MB-231 cells using RNAi technology.I found that knocking down of PKC significantly blocks ferroptosis.To further confirm the specificity of PKC in ferroptosis,I overexpressed dominant-negative mutations of PKCα,PKCβI,PKCβII and PKCγ in MDA-MB-231 cells individually.The results showed that only overexpression of PKCβI,PKCβII can block ferroptosis,suggesting PKCβI and PKCβII but not PKCα or PKCγ are required for ferroptosis.I also induced PKCβ back to PKCβ knockdown MDA-MB-231 cells and found wild-type but not dominant mutant form of PKCβ can restore ferroptosis sensitivity.In summary,this study revealed that PKC is an important positive regulator of ferroptosis.PKC is an important kinase.Next,I will identify downstream targets of PKC using co-immunoprecipitation(Co-IP)coupled with mass spectrometry.