An in vivo genome-wide CRISPR-Cas9 screen identifies FER as a tumor suppressor inhibiting the oncogenic transformation

Abstract Background Tumorigenesis is a complex and multistep process characterized by the progressive acquisition of various hallmarks, including unlimited proliferation, resistance to apoptosis, and increased invasiveness and metastasis. However, the molecular mechanisms underlying tumorigenesis remain poorly understood. Methods An in vivo genome-wide CRISPR–Cas9 screen was employed to identify tumor suppressor genes (TSG). The expression correlation analysis for candidate TSGs was performed in normal and cancer cells using TCGA database. To evaluate the role of FER in tumorigenesis, we firstly used publicly single-cell RNA sequencing data to investigate the association of FER expression and normal cell malignant transformation. Next, we established FER -knockout and -knockdown models in BEAS-2B and MCF10A cell lines. Colony formation assay, cell proliferation assay, EdU assay and apoptosis assay were conducted to determine the role of FER in tumorigenesis. Then RNA-seq was performed to explore the mechanism underlying the role of FER in inhibiting tumorigenesis. Additionally, Pan-Cancer analysis was used to analysis the role of FER in tumor progression. Results In our CRISPR–Cas9 screen, we identified 20 candidate genes, among which FER exhibited the strongest negative correlation with tumorigenesis. Normal cells with low FER expression exhibited elevated malignant transformation potential and stemness properties. FER knockout promoted the tumorigenesis of differentiated epithelial cells by reprogramming them into a cancer stem cell (CSC)-like state, characterized by high colony-forming efficiency and suspension growth ability, increased metabolic activity, dedifferentiation properties, and immune evasion. Furthermore, tumors with low FER expression exhibited poor prognosis and a noticeable CSC-like state. Conclusion Taken together, our findings not only provide insights into the essential role of FER as a stemness barrier in malignant cells during tumor initiation and progression but also highlight its potential as a target for future clinical diagnosis.