Transcriptomic and single-cell analyses reveal the prognostic value of ETV4 and its role in shaping the immune landscape of colorectal cancer

<h4>Background</h4>Colorectal cancer (CRC) remains a major health threat with poor prognosis in advanced stages. The transcription factor ETV4 (ETS translocation variant 4) has been implicated in various cancers, but its specific role, prognostic value, and mechanisms in CRC, particularly concerning the tumor immune microenvironment, are not fully understood.<h4>Methods</h4>We performed a comprehensive analysis using transcriptomic data from The Cancer Genome Atlas (TCGA) and single-cell RNA sequencing data from the GEO database (GSE231559). Bioinformatic approaches included survival analysis, immune cell infiltration estimation via CIBERSORT, gene set enrichment analysis (GSEA), gene set variation analysis (GSVA), and single-cell clustering. Experimental validation was conducted on clinical CRC tissue samples and HCT-116 cells, employing flow cytometry, immunohistochemistry, quantitative real-time PCR (Q-PCR), and Western blot.<h4>Results</h4>In the TCGA cohort, ETV4 was significantly upregulated in 650 CRC tissues vs 51 adjacent normal tissues, with high expression linked to poorer overall survival in CRC patients.Immune infiltration analysis revealed correlations between ETV4 expression and specific immune cell subsets, notably macrophages. Flow cytometry and immunohistochemistry confirmed that high ETV4 expression was linked to increased polarization of immunosuppressive M2-type macrophages in the tumor microenvironment. Furthermore, bioinformatic GSVA and subsequent wet-lab experiments demonstrated that ETV4 is a downstream target gene of the WNT/β-catenin signaling pathway. Activation of this pathway upregulated ETV4 expression, while inhibition downregulated it, establishing a functional link.<h4>Conclusion</h4>This integrative study reveals that ETV4 is a prognostic biomarker in CRC. It promotes tumor progression by reshaping the immunosuppressive microenvironment, particularly through inducing M2 macrophage polarization, and is regulated by the oncogenic WNT/β-catenin pathway. These findings suggest that ETV4 is expected to become a potential diagnostic biomarker and candidate therapeutic target for colorectal cancer, providing a novel direction for subsequent research on the diagnosis and treatment of CRC.