Downregulation of hepatic METTL3 contributes to APAP-induced liver injury in mice

BACKGROUND & AIMS: Acetaminophen (APAP) overdose is a major cause of acute liver failure in the Western world, but its molecular mechanisms are not fully understood. Methyltransferase-like 3 (METTL3) is a core N6-methyl-adenosine (m6A) RNA methyltransferase that has been shown to regulate many physiological and pathological processes. This study aimed to investigate the role of METTL3 in APAP-induced liver injury in mice. METHODS: Hepatocyte-specific Mettl3 knockout (Mettl3-HKO) mice and adenovirus-mediated gene overexpression or knockdown were used. We assayed APAP-induced liver injury by measuring serum alanine aminotransferase/aspartate aminotransferase activity, necrotic area, cell death, reactive oxygen species levels and activation of signalling pathways. We also performed mechanistic studies using a variety of assays and molecular techniques. RESULTS: Hepatic METTL3 is downregulated in APAP-induced liver injury, and hepatocyte-specific deletion of Mettl3 accelerates APAP-induced liver injury, leading to increased mortality as a result of the dramatic activation of the mitogen-activated protein kinase kinase 4 (MKK4) / c-Jun NH2-terminal kinase (JNK) signalling pathway. Inhibition of JNK by SP600125 largely blocks APAP-induced liver injury in Mettl3-HKO mice. Hepatic deletion of Mettl3 activates the MKK4/JNK signalling pathway by increasing the protein stability of MKK4 and JNK1/2 as a result of decreased proteasome activity. Restoration of proteasome activity by overexpression of proteasome 20S subunit beta 4 (PSMB4) or proteasome 20S subunit beta 6 (PSMB6) leads to the downregulation of MKK4 and JNK in Mettl3-HKO hepatocytes. Mechanistically, METTL3 interacts with RNA polymerase II and active histone modifications such as H3K9ac, H3K27ac, and H3K36me3 to maintain the expression of proteasome-related genes. CONCLUSIONS: Our study demonstrated that downregulation of METTL3 promotes APAP-induced liver injury by decreasing proteasome activity and thereby enhancing activity of the MKK4/JNK signalling pathway. IMPACT AND IMPLICATIONS: Acetaminophen (APAP) overdose is a key cause of acute liver failure in the Western world, but its molecular mechanisms are not fully understood. We demonstrated in this study that methyltransferase-like 3 (METTL3), a core m(6)A RNA methyltransferase, is downregulated in APAP-induced liver injury, which exacerbates APAP-induced liver injury through enhancing the MKK4/JNK signalling pathway with involvement of the decreased proteasome activity.