Oxidative Stress Induced by Arsenite is Involved in YTHDF2 Phase Separation

YTH N-6-methyladenosine RNA binding protein 2 (YTHDF2) undergoes phase separation in response to the stimulation of high concentration of arsenite, suggesting that oxidative stress, the major mechanism of arsenite toxicity, may play a role in YTHDF2 phase separation. However, whether arsenite-induced oxidative stress is involved in phase separation of YTHDF2 has yet to be established. To explore the effect of arsenite-induced oxidative stress on YTHDF2 phase separation, the levels of oxidative stress, YTHDF2 phase separation, and N-6-methyladenosine (m(6)A) in human keratinocytes were detected after exposure to various concentrations of sodium arsenite (0-500 mu M; 1 h) and antioxidant N-acetylcysteine (0-10 mM; 2 h). We found that arsenite promoted oxidative stress and YTHDF2 phase separation in a concentration-dependent manner. In contrast, pretreatment with N-acetylcysteine significantly relieved arsenate-induced oxidative stress and inhibited YTHDF2 phase separation. As one of the key factors to YTHDF2 phase separation, N-6-methyladenosine (m(6)A) levels in human keratinocytes were significantly increased after arsenite exposure, accompanied by upregulation of m(6)A methylesterase levels and downregulation of m(6)A demethylases levels. On the contrary, N-acetylcysteine mitigated the arsenite-induced increase of m(6)A and m(6)A methylesterase and the arsenite-induced decrease in m(6)A demethylase. Collectively, our study firstly revealed that oxidative stress induced by arsenite plays an important role in YTHDF2 phase separation driven by m(6)A modification, which provides new insights into the arsenite toxicity from the phase-separation perspective.