m5C-modified circRREB1 promotes lung cancer progression by inducing mitophagy.

BACKGROUND: Lung cancer is the most common malignant tumour and the leading cause of cancer-related death. circular RNAs (circRNAs) have important biological functions and are closely related to tumour development. The 5-methylcytosine (m5C) modification can regulate the molecular fate of RNA molecules and thus influence disease development. METHODS: High-throughput RNA sequencing was used to construct the differential expression profiles of circRNAs. The m5C modification of circRREB1 was explored through methylated RNA immunoprecipitation (MeRIP) and crosslinking-immunoprecipitation (CLIP). RNA stability experiments, fluorescence in situ hybridization (FISH), and nuclear-cytoplasmic fractionation experiments were performed to explore the effects of the m5C modification on circRREB1. A system for the silencing and overexpression of circRREB1 was established, and in vitro and in vivo experiments were conducted to study the biological functions of circRREB1. Tagged RNA affinity purification (TRAP), RNA immunoprecipitation (RIP), and coimmunoprecipitation (Co-IP) experiments were conducted to reveal the molecular mechanisms of circRREB1. RESULTS: In this study, we found that circRREB1 is highly expressed in lung cancer tissues and cells and that patients with high circRREB1 expression have a poor prognosis. We discovered that circRREB1 undergoes the m5C modification mediated by the methyltransferase NSUN2. This modification facilitates its nuclear export via the m5C reader ALYREF. Functional studies demonstrated that circRREB1 promotes lung cancer progression both in vitro and in vivo. Mechanistically, circRREB1 directly binds to HSPA8 and stabilizes it by inhibiting ubiquitin-dependent degradation, thereby inducing mitophagy through the HSPA8/PINK1/Parkin signalling axis and ultimately promoting the development of lung cancer. CONCLUSIONS: This study revealed the presence of m5C modifications on circRREB1 and showed that m5C-modified circRREB1 can induce mitophagy, ultimately promoting lung cancer. These findings provide not only a theoretical basis for further exploration of the mechanisms underlying lung cancer development but also potential targets for lung cancer therapy.