The METTL3/TRAP1 axis as a key regulator of 5-fluorouracil chemosensitivity in colorectal cancer

Colorectal cancer (CRC) remains a significant clinical challenge, with 5-Fluorouracil (5-FU) being the frontline chemotherapy. However, chemoresistance remains a major obstacle to effective treatment. METTL3, a key methyltransferase involved in RNA methylation processes, has been implicated in CRC carcinogenesis. However, its role in modulating CRC sensitivity to 5-FU remains elusive. In this study, we aimed to investigate the role and mechanisms of METTL3 in regulating 5-FU chemosensitivity in CRC cells. Initially, we observed that 5-FU treatment inhibited cell viability and induced apoptosis, accompanied by a reduction in METTL3 expression in HCT-116 and HCT-8 cells. Subsequent assays including drug sensitivity, EdU, colony formation, TUNEL staining, and flow cytometry revealed that METTL3 depletion enhanced 5-FU sensitivity and increased apoptosis induction both in vitro and in vivo. Conversely, METTL3 overexpression conferred resistance to 5-FU in both cell lines. Moreover, knockdown of METTL3 in 5-FU-resistant CRC cell lines HCT-116/FU and HCT-15/FU significantly decreased 5-FU tolerance and induced apoptosis upon 5-FU treatment. Mechanistically, we found that METTL3 regulated 5-FU sensitivity and apoptosis induction by modulating TRAP1 expression. Further investigations using m6A colorimetric ELISA, dot blot, MeRIP-qPCR and RNA stability assays demonstrated that METTL3 regulated TRAP1 mRNA stability in an m6A-dependent manner. Additionally, overexpression of TRAP1 mitigated the cytotoxic effects of 5-FU on CRC cells. In summary, our study uncovers the pivotal role of the METTL3/TRAP1 axis in modulating 5-FU chemosensitivity in CRC. These findings provide new insights into the mechanisms underlying CRC resistance to 5-FU and may offer potential targets for future therapeutic interventions.

Graphical abstract

Proposed working model of METTL3 regulating 5-FU sensitivity and apoptosis induction in CRC cells. METTL3 is frequently upregulated in CRC cells and is mainly localized in the nucleus of tumor cells [41]. In this study, we observed that down-regulating METTL3 levels led to a decrease in m6A modification on TRAP1 mRNA in both HCT-116 and HCT-8 cells. This reduction in m6A modification resulted in decreased stability of TRAP1 mRNA, ultimately facilitating 5-FU-induced apoptosis and heightening sensitivity to the drug. Our findings suggest a potential mechanism wherein elevated METTL3 expression in CRC cells may regulate TRAP1 expression in an m6A-dependent manner, thereby enabling cells to evade 5-FU-induced apoptosis and contribute to resistance against 5-FU chemotherapy.