Abstract
BackgroundCoactivator associated arginine methyltransferase 1 (CARM1) has been identified as a regulator of breast cancer (BC) progression, yet the underlying mechanisms remain elusive. MethodsQuantitative real-time polymerase chain reaction (qRT-PCR) was employed to assess the mRNA expression of CARM1 and solute carrier family 7 member 11 (SLC7A11). Western blotting was conducted to detect the protein expressions of CARM1, ubiquitin specific peptidase 4 (USP4), and SLC7A11. Cell viability, apoptosis, invasion, and migration were evaluated using CCK-8 assay, flow cytometry, transwell assay, and wound-healing assay, respectively. Fe2+ and GSH levels were determined by colorimetric assay. Fluorescence microscopy and flow cytometry were utilized to quantify reactive oxygen species (ROS) production. Co-immunoprecipitation (Co-IP) assay and cycloheximide (CHX) assay were performed to investigate the relationship between USP4 and CARM1. Xenograft mouse model assay was conducted to validate the effects of USP4 silencing and CARM1 overexpression on the malignant phenotypes of BC cells. ResultsCARM1 and SLC7A11 expression was upregulated in BC tissues and cells when compared with normal breast tissues and cells. Silencing of CARM1 inhibited the malignant phenotypes of BC cells, including decreased cell viability, invasion, and migration and increased cell apoptosis, ferroptosis and oxidative stress. In addition, USP4 stabilized CARM1 protein expression through its deubiquitinating activity. Overexpression of CARM1 attenuated the effects of USP4 silencing in both MCF-7 and MDA-MB-231 cells. Furthermore, silencing of CARM1 reduced SLC7A11 expression, and SLC7A11 overexpression relieved the CARM1 silencing-induced effects. Further, overexpression of CARM1 counteracted the inhibitory effects of USP4 silencing on tumor growth in vivo. ConclusionOur study reveals a novel mechanism by which USP4-dependent CARM1 promotes the malignant growth of BC cells by interacting with SLC7A11. Targeting this axis may provide a potential therapeutic strategy for BC. Micro AbstractBackground: Previous evidence has suggested the participation of coactivator associated arginine methyltransferase 1 (CARM1) in breast cancer (BC) progression, yet the underlying mechanisms remain elusive.Methods: Ubiquitin assay and cycloheximide (CHX) assay were performed to investigate the relationship between USP4 and CARM1. The study also performed various assays including CCK-8 assay, flow cytometry, transwell assay, wound-healing assay, colorimetric assay, fluorescence microscopy, and xenograft mouse model assay to analyze the effects of USP4 and CARM1 on the malignant progression of BC cells.Results: Silencing of CARM1 inhibited cell viability, invasion, and migration and promoted cell apoptosis, ferroptosis and oxidative stress. USP4 stabilized CARM1 protein expression, and overexpression of CARM1 relieved the effects of USP4 silencing on BC cell malignancy and tumor growth. In addition, SLC7A11 overexpression relieved the CARM1 silencing-induced effects.Conclusion: USP4-dependent CARM1 promoted the malignant growth of BC cells by interacting with SLC7A11.
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