Tumour cells are sensitised to ferroptosis via RB1CC1-mediated transcriptional reprogramming

Background: Ferroptosis, a form of regulated cell death, is an important topic in the field of Cancer research. However, the signalling pathways and factors that sensitise tumour cells to Ferroptosis remain elusive.

Methods: We determined the level of Ferroptosis in cells by measuring cell death and lipid Reactive Oxygen Species (ROS) production. The expression of RB1-inducible coiled-coil 1 (RB1CC1) and related proteins was analyzed by immunoblotting and immunohistochemistry. Immunofluorescence was used to determine the subcellular localization of RB1CC1. We investigated the mechanism of RB1CC1 nuclear translocation by constructing a series of RB1CC1 variants. To examine the ferroptosis- and RB1CC1-dependent transcriptional program in tumour cells, chromatin immunoprecipitation Sequencing was performed. To assess the effect of c-Jun N-terminal kinase (JNK) agonists on strenthening imidazole ketone erastin (IKE) therapy, we constructed cell-derived xenograft mouse models. Mouse models of hepatocellular carcinoma to elucidate the importance of Rb1cc1 in IKE-based therapy of liver tumourigenesis.

Results: RB1CC1 is upregulated by lipid ROS and that nuclear translocation of phosphorylation of RB1CC1 at Ser537 was essential for sensitising Ferroptosis in tumour cells. Upon Ferroptosis induction, nuclear RB1CC1 sharing forkhead box (FOX)-binding motifs recruits elongator acetyltransferase complex subunit 3 (ELP3) to strengthen H4K12Ac histone modifications within enhancers linked to Ferroptosis. This also stimulated transcription of ferroptosis-associated genes, such as coiled-coil-helix-coiled-coil-helix domain containing 3 (CHCHD3), which enhanced mitochondrial function to elevate mitochondrial ROS early following induction of Ferroptosis. FDA-approved JNK activators reinforced RB1CC1 nuclear translocation and sensitised cells to Ferroptosis, which strongly suggested that JNK is upstream of RB1CC1. Nuclear localisation of RB1CC1 correlated with lipid peroxidation in clinical lung Cancer specimens. Rb1cc1 was essential for Ferroptosis agonists to suppress liver tumourigenesis in mice.

Conclusions: Our findings indicate that RB1CC1-associated signalling sensitises tumour cells to Ferroptosis and that targeting RB1CC1 may be beneficial for tumour treatment.