Li Xin, Yan Changjiao, Yun Jun, Xu Xin, Wei Hongliang, Xu Xiaolong, Li Yike, Yi Jun
Department of Thyroid, Breast and Vascular Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi, China.
Department of Thyroid, Breast and Vascular Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi, China.
Clin Breast Cancer. 2025 Feb;25(2):e196-e207. doi: 10.1016/j.clbc.2024.10.001. Epub 2024 Oct 16.
Coactivator associated arginine methyltransferase 1 (CARM1) has been identified as a regulator of breast cancer (BC) progression, yet the underlying mechanisms remain elusive.
Quantitative 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. Fe 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.
CARM1 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.
Our 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.
共激活因子相关精氨酸甲基转移酶1（CARM1）已被确定为乳腺癌（BC）进展的调节因子，但其潜在机制仍不清楚。
采用定量实时聚合酶链反应（qRT-PCR）评估CARM1和溶质载体家族7成员11（SLC7A11）的mRNA表达。进行蛋白质印迹法检测CARM1、泛素特异性肽酶4（USP4）和SLC7A11的蛋白表达。分别使用CCK-8法、流式细胞术、Transwell法和伤口愈合试验评估细胞活力、凋亡、侵袭和迁移。通过比色法测定铁和谷胱甘肽水平。利用荧光显微镜和流式细胞术定量活性氧（ROS）的产生。进行免疫共沉淀（Co-IP）试验和放线菌酮（CHX）试验以研究USP4与CARM1之间的关系。进行异种移植小鼠模型试验以验证USP4沉默和CARM1过表达对BC细胞恶性表型的影响。
与正常乳腺组织和细胞相比，BC组织和细胞中CARM1和SLC7A11表达上调。CARM1沉默抑制了BC细胞的恶性表型，包括细胞活力、侵袭和迁移降低以及细胞凋亡、铁死亡和氧化应激增加。此外，USP4通过其去泛素化活性稳定CARM1蛋白表达。CARM1过表达减弱了USP4沉默对MCF-7和MDA-MB-231细胞的影响。此外，CARM1沉默降低了SLC7A11表达，而SLC7A11过表达缓解了CARM1沉默诱导的效应。此外，CARM1过表达抵消了USP4沉默对体内肿瘤生长的抑制作用。
我们的研究揭示了一种新机制，即USP4依赖的CARM1通过与SLC7A11相互作用促进BC细胞的恶性生长。靶向该轴可能为BC提供一种潜在的治疗策略。
