The FUS/circEZH2/KLF5/feedback loop contributes to CXCR4-induced liver metastasis of breast cancer by enhancing epithelial-mesenchymal transition

Background Metastasis of breast cancer have caused the majority of cancer-related death worldwide. The circRNAs are associated with tumorigenesis and metastasis in breast cancer according to recent research. However, the biological mechanism of circRNAs in liver metastatic breast cancer remains ambiguous yet. Methods Microarray analysis of three pairs of primary BC tissues and matched hepatic metastatic specimens identified circEZH2. We used RT-qPCR and FISH assays to confirm circEZH2 existence, characteristics, and expression. Both in vivo and in vitro, circEZH2 played an oncogenic role which promoted metastasis as well. A range of bioinformatic analysis, Western blot, RNA pull-down, RIP, ChIP, and animal experiments were used to define the feedback loop involving FUS, circEZH2, miR-217-5p, KLF5, FUS, CXCR4 as well as epithelial and mesenchymal transition. Results In our research, circEZH2 was proved to be upregulated in liver metastases in BC and predicted the worse prognosis in breast cancer patients. Overexpression of circEZH2 notably accentuated the vitality and invasion of BC cells, whereas knockdown of circEZH2 elicited the literally opposite effects. Besides, overexpressed circEZH2 promoted tumorigenesis and liver metastasis in vivo. Moreover, circEZH2 could adsorb miR-217-5p to upregulate KLF5 thus leading to activate FUS transcription which would facilitate the back-splicing program of circEZH2. Meanwhile, KLF5 could upregulated CXCR4 transcriptionally to accelerate epithelial and mesenchymal transition of breast cancer. Conclusions Consequently, a novel feedback loop FUS/circEZH2/KLF5/CXCR4 was established while circEZH2 could be novel biomarker and potential target for BC patients' therapy.

Automated summary

Metastasis of breast cancer remains a major cause of cancer-related death globally. Recent research indicates that circRNAs are associated with tumorigenesis and metastasis in breast cancer. However, the biological mechanism of circRNAs in liver metastatic breast cancer is still unclear. In this study, we identified circEZH2 through microarray analysis and confirmed its existence and expression in liver metastases of breast cancer. We demonstrated that circEZH2 is upregulated in liver metastases and predicts poor prognosis in breast cancer patients. Functionally, circEZH2 promotes the vitality and invasion of breast cancer cells, and its knockdown has the opposite effects. Furthermore, circEZH2 overexpression promotes tumorigenesis and liver metastasis in vivo. Mechanistically, circEZH2 acts as a sponge for miR-217-5p, leading to upregulation of KLF5, which activates FUS transcription and facilitates the back-splicing program of circEZH2. Additionally, KLF5 transcriptionally upregulates CXCR4, thereby accelerating epithelial-mesenchymal transition in breast cancer. In conclusion, we have identified a novel feedback loop involving FUS/circEZH2/KLF5/CXCR4, and circEZH2 may serve as a potential biomarker and therapeutic target for breast cancer patients.