Lactylation of RNA m ⁶ A demethylase ALKBH5 promotes innate immune response to DNA herpesviruses and mpox virus

RNA <i>N</i>⁶-methyladenosine (m⁶A) demethylase AlkB homolog 5 (ALKBH5) plays a crucial role in regulating innate immunity. Lysine acylation, a widespread protein modification, influences protein function, but its impact on ALKBH5 during viral infections has not been well characterized. This study investigates the presence and regulatory mechanisms of a previously unidentified lysine acylation in ALKBH5 and its role in mediating m⁶A modifications to activate antiviral innate immune responses. We demonstrate that ALKBH5 undergoes lactylation, which is essential for an effective innate immune response against DNA herpesviruses, including herpes simplex virus type 1 (HSV-1), Kaposi's sarcoma-associated herpesvirus (KSHV), and mpox virus (MPXV). This lactylation attenuates viral replication. Mechanistically, viral infections enhance ALKBH5 lactylation by increasing its interaction with acetyltransferase ESCO2 and decreasing its interaction with deacetyltransferase SIRT6. Lactylated ALKBH5 binds interferon-beta (IFN-β) messenger RNA (mRNA), leading to demethylation of its m⁶A modifications and promoting IFN-β mRNA biogenesis. Overexpression of ESCO2 or depletion of SIRT6 further enhances ALKBH5 lactylation to strengthen IFN-β mRNA biogenesis. Our results identify a posttranslational modification of ALKBH5 and its role in regulating antiviral innate immune responses through m⁶A modification. The finding provides an understanding of innate immunity and offers a potential therapeutic target for HSV-1, KSHV, and MPXV infections.