Hepatitis B virus X protein recruits methyltransferases to affect cotranscriptional N6-methyladenosine modification of viral/host RNAs.

Chronic hepatitis B virus (HBV) infections are one of the leading causes of cirrhosis and hepatocellular carcinoma. N6-methyladenosine (mA) modification of cellular and viral RNAs is the most prevalent internal modification that occurs cotranscriptionally. Previously, we reported the dual functional role of mA modification of HBV transcripts in the viral life cycle. Here, we show that viral HBV X (HBx) protein is responsible for the mA modifications of viral transcripts. HBV genomes defective in HBx failed to induce mA modifications of HBV RNAs during infection/transfection, while ectopic expression of HBx restores mA modifications of the viral RNAs but not the mutant HBx carrying the nuclear export signal. Using chromatin immunoprecipitation assays, we provide evidence that HBx and mA methyltransferase complexes are localized on the HBV minichromosome to achieve cotranscriptional mA modification of viral RNAs. HBx interacts with METTL3 and 14 to carry out methylation activity and also modestly stimulates their nuclear import. This role of HBx in mediating mA modification also extends to host phosphatase and tensin homolog (PTEN) mRNA. This study provides insight into how a viral protein recruits RNA methylation machinery to mA-modify RNAs.