Extended Data Fig. 3: CKB-mediated GPX4 S104 phosphorylation stabilizes GPX4.
From: Creatine kinase B suppresses ferroptosis by phosphorylating GPX4 through a moonlighting function
(a, b) Alignment of protein sequences spanning GPX4 S104 and the adjacent CMA-target motif from different species. The structure of GPX4 exhibiting CMA-target motif and S104. (c) Purified His-CKB WT or T133D proteins on Ni-NTA beads were incubated with the indicated GST-GPX4 proteins for an in vitro kinase assay. Then the beads were removed and the remaining GST-GPX4 proteins were incubated with His-HSC70 followed by a GST pulldown assay as indicated. (d, e) Huh7 and HCCLM3 cells expressing GPX4 shRNA with reconstituted expression of the indicated GPX4 proteins were harvested for immunoblotting and mRNA detection using quantitative PCR as indicated. Data are the mean ± SD (n = 6). N.S., not significant (two-tailed Student’s t-test).
