TRMT6‐Mediated m ¹ A Modification of CDK9 mRNA is a Dual‐Pronged Pathogenic Driver for HBV‐Related Hepatocellular Carcinoma

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.