Phosphorylation of serine 177 of the small hepatitis delta antigen regulates viral antigenomic RNA replication by interacting with the processive rna polymerase II

Recent studies revealed that posttranslational modifications (e.g., phosphorylation and methylation) of the small hepatitis delta antigen (SHDAg) are required for hepatitis delta virus (HDV) replication from antigenomic to genomic RNA. The phosphorylation of SHDAg at serine 177 (Ser¹⁷⁷) is involved in this step, and this residue is crucial for interaction with RNA polymerase II (RNAP II), the enzyme assumed to be responsible for antigenomic RNA replication. This study demonstrated that SHDAg dephosphorylated at Ser ¹⁷⁷ interacted preferentially with hypophosphorylated RNAP II (RNAP IIA), which generally binds at the transcription initiation sites. In contrast, the Ser¹⁷⁷-phosphorylated counterpart (pSer¹⁷⁷-SHDAg) exhibited preferential binding to hyperphosphorylated RNAP II (RNAP IIO). In addition, RNAP IIO associated with pSer¹⁷⁷-SHDAg was hyperphosphorylated at both the Ser² and Ser⁵ residues of its carboxyl-terminal domain (CTD), which is a hallmark of the transcription elongation isoform. Moreover, the RNAP II CTD kinase inhibitor 5,6-dichloro-1-β-D-ribofuranosyl-benzimidazole (DRB) not only blocked the interaction between pSer¹⁷⁷-SHDAg and RNAP IIO but also inhibited HDV antigenomic replication. Our results suggest that the phosphorylation of SHDAg at Ser¹⁷⁷ shifted its affinity toward the RNA RNAP IIO isoform and thus is a switch for HDV antigenomic RNA replication from the initiation to the elongation stage.