RNA m6A reader YTHDF3/UBE2G2 m6A methylation/ACSL4 ubiquitination axis-facilitated cell ferroptosis to mediate benzene hematotoxicity and the protective effect of melatonin

Introduction: Long-term benzene exposure results in hematotoxicity. N6-methyladenosine (m6A) is a type of post-transcriptional modification and plays roles in various diseases. However, the role of m6A modification in benzene hematotoxicity is still unclear.

Methods: m6A epitranscriptomic and RNA-seq analyses were conducted to explore the effect of benzene on the m6A level and expression of mRNAs. Benzene-induced hematotoxicity mouse models and melatonin intervention models were constructed, and RIP, MeRIP, and Co-IP assays were used to investigate the role of m6A in benzene hematotoxicity and the protective effect of melatonin.

Results: We found that benzene altered m6A methylation of various mRNAs and identified a key m6A-methylated mRNA, ubiquitin-conjugating enzyme E2 G2 (UBE2G2), through GO analysis. m6A reading proteins recognized m6A modifications to regulate mRNA's fate. By searching an m6A target prediction website, we discovered that the m6A reader YTHDF3 bound with the UBE2G2 m6A site, and we also validated their interactions via RIP and MeRIP assays. Mechanistically, benzene exposure resulted in low expression of YTHDF3, which down-regulated the mRNA's stability and expression of UBE2G2 through the recognition of m6A modification. Moreover, overexpression of YTHDF3 mitigated UBE2G2 reduction, which further confirmed their regulatory relationship. UBE2G2 regulates proteins' ubiquitination to affect their expression. Through a Co-Immunoprecipitation assay and functional analysis, we demonstrated that ubiquitination modified ACSL4 due to UBE2G2-mediated benzene-induced lipid peroxidation and ferroptosis. Moreover, melatonin alleviated benzene-induced hematotoxicity by modulating the YTHDF3/UBE2G2/ACSL4 axis.

Conclusions: The m6A reader YTHDF3/UBE2G2 m6A methylation/ACSL4 ubiquitination axis facilitated cell ferroptosis to mediate benzene hematotoxicity, and melatonin had an alleviating effect on benzene hematotoxicity.