Upregulation of circGDI2 inhibits tumorigenesis by stabilizing the expression of RNA m6A demethylase FTO in oral squamous cell carcinoma

Background

Oral squamous cell carcinoma (OSCC) is a malignant tumour that is difficult to identify and prone to metastasis and invasion. Circular RNAs (circRNAs) are important cancer regulators and can be used as potential biomarkers. However, OSCC-related circRNAs need to be further explored. We investigated the role of circGDI2 in OSCC and explored its downstream regulatory mechanisms.

Methods

Quantitative real-time PCR was used to detect the expression levels of circGDI2 and fat mass and obesity-associated protein (FTO) in cells. Lentiviral transfection was used to construct stable circGDI2 overexpressing cells for subsequent cell function tests. RNA pull-down, RNA Immunoprecipitation (RIP), western blotting, and protein stability assays were conducted to detect circGDI2 binding proteins and their functions. CCK8, Transwell, and wound healing assays were used to verify cell functions after overexpressing circGDI2 or suppressing FTO expression. Animal experiments were performed to verify the results in vivo.

Results

The expression of circGDI2 was markedly decreased in both OSCC cell lines and patient tissues. Overexpression of circGDI2 in OSCC cell lines led to decreased proliferation, migration, and invasion abilities. Knockdown of circGDI2 showed the opposite trend. CircGDI2 has been validated to interact with the FTO protein within cells, as evidenced by mass spectrometry and RIP assays. This interaction was found to prevent the degradation of the FTO protein. Dot blot analysis showed a reduction in N6-methyladenosine (m6A) modification after circGDI2 overexpression. Reduced FTO levels reversed the inhibitory effects of circGDI2 overexpression on cell proliferation, migration, and invasion in vitro and on tumorigenesis in vivo.

Conclusions

CircGDI2 functions as a tumour suppressor by binding to the FTO protein to reduce RNA m6A modification levels and ultimately inhibit proliferation and migration in OSCC cells. This study indicates the potential use of circGDI2 as a new target for the prevention and treatment of OSCC.