E2F1-mediated transcriptional regulation of TMEM33 promotes colorectal cancer proliferation, metastasis, and EMT.

Colorectal cancer (CRC) remains one of the most lethal malignancies worldwide, with tumor metastasis representing the primary driver of treatment failure and poor prognosis. Transmembrane Protein 33 (TMEM33) has been implicated in tumor progression across several cancer types, yet its expression pattern, biological function, and upstream transcriptional regulation in CRC remain undefined. In this study, TMEM33 expression in CRC and matched adjacent normal tissues was assessed by quantitative reverse transcription polymerase chain reaction (qRT-PCR), Western blot, and immunohistochemistry (IHC), with prognostic associations analyzed via Kaplan-Meier survival curves. TMEM33 was significantly upregulated in CRC tissues, and high TMEM33 expression correlated with advanced tumor stage, lymph node metastasis, and unfavorable overall survival. TMEM33 knockdown in HCT116 and LOVO CRC cell lines, achieved via lentiviral infection, significantly suppressed cell proliferation as demonstrated by Cell Counting Kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) incorporation assays. TMEM33 silencing also reduced cell migration and invasion in Transwell assays, and reversed epithelial-mesenchymal transition (EMT). Mechanistically, bioinformatics screening and dual-luciferase reporter assays identified E2F transcription factor 1 (E2F1) as a transcriptional activator of TMEM33, binding its promoter to drive expression. Rescue experiments further demonstrated that E2F1 overexpression partially restored proliferation, migration, invasion, and EMT in TMEM33-knockdown cells. In vivo, TMEM33 suppression significantly inhibited subcutaneous tumor growth and reduced lung metastatic burden in nude mouse models. Collectively, these findings establish E2F1 as an upstream transcriptional regulator of TMEM33 and demonstrate that the E2F1/TMEM33 axis promotes CRC malignant progression by driving proliferation, metastasis, and EMT, highlighting this axis as a candidate therapeutic target for CRC.