Abstract

Hepatocellular carcinoma (HCC) is a leading cause of death worldwide, with hepatitis B virus (HBV) infection being the major risk factor. Dysregulation of mRNA methylation contributes to tumorigenesis and virus replication. However, the association of N 1 -methyladenosine (m 1 A) modification with HCC progression and HBV replication remains unclear. Here, single-nucleus RNA sequencing (snRNA-seq) of 4 HCC and 7 adjacent tissues (2 from this study and 5 from the GSE242889) revealed elevated mRNA methylation in HCCs, with increased expression of m 1 A “writers” and “readers” and decreased expression of m 1 A “erasers”. Among them, m 1 A writer TRMT6 was up-regulated in HCC and correlated with poor patient prognosis. TRMT6 knockdown strikingly restrained the malignant phenotypes and tumorigenicity of HCC cells as well as HBV replication. Mechanistically, TRMT6-mediated m 1 A modification enhanced the stability and translation efficiency of cyclin-dependent kinase 9 ( CDK9 ) mRNA. Elevated CDK9 facilitated HCC progression by up-regulating its downstream oncogenic effectors, and stimulated HBV replication via TARDBP phosphorylation at Ser254 to enhance pgRNA transcription and repress pgRNA splicing. CDK9 inhibitor FIT-039 abrogated these effects without obvious toxicity. Thus, TRMT6-mediated m 1 A modification dually drives HCC malignancy and HBV replication, representing a promising therapeutic target, and CDK9 inhibition may constitute an effective strategy for HBV-related HCC.