DOCK11 promotes HBV cccDNA formation through a PARP1-dependent mechanism

Hepatitis B virus (HBV) infection remains a major global health concern due to the persistence of covalently closed circular DNA (cccDNA) in the hepatocyte nucleus, which contributes to viral persistence and hepatocarcinogenesis despite antiviral therapy. Poly(ADP-ribose) polymerase 1 (PARP1) has been reported to bind to HBV relaxed circular DNA (rcDNA) and contribute to the conversion of rcDNA to cccDNA. We previously identified dedicator of cytokinesis 11 (DOCK11) as a host factor whose suppression reduced cccDNA levels in HBV infection models; however, the precise role of DOCK11 in the process of cccDNA formation remains unclear. In this study, we investigated the role of DOCK11 in HBV cccDNA formation and the underlying mechanisms. Using a minicircle HBV cccDNA reporter, we observed that luciferase activity declined significantly in DOCK11-depleted hepatoma cells over time, consistent with impaired cccDNA maintenance. In a cell-free assay using rcDNA and nuclear extracts, DOCK11 overexpression significantly enhanced cccDNA formation, while DOCK11 knockout significantly suppressed it. Co-immunoprecipitation suggested that DOCK11 forms a multiprotein complex with HBV DNA. Furthermore, liquid chromatography-mass spectrometry analysis identified PARP1 as a significant component of this complex. Notably, while DOCK11 did not affect PARP1 expression or enzymatic activity, DOCK11 knockout significantly reduced PARP1 recruitment to UV-irradiated DNA damage sites and its association with rcDNA, suggesting a supportive role for DOCK11 in PARP1 recruitment to rcDNA. In summary, DOCK11 promotes HBV cccDNA formation, possibly through its interaction with HBV DNA and PARP1. These findings position DOCK11 as a promising therapeutic target for cccDNA reduction in chronic HBV infection.IMPORTANCEHepatitis B virus (HBV) causes chronic liver infection, affecting hundreds of millions of people worldwide and increasing the risk of cirrhosis and hepatocellular carcinoma. Persistent HBV infection is maintained by covalently closed circular DNA (cccDNA), which is resistant to current antiviral therapies. Understanding cccDNA maintenance mechanisms, including cccDNA formation, is therefore critical for developing curative treatments. Poly(ADP-ribose) polymerase 1 (PARP1) is known to bind HBV relaxed circular DNA (rcDNA) and contribute to the conversion of rcDNA to cccDNA. In this study, we identified the host protein dedicator of cytokinesis 11 as a host regulator of HBV cccDNA formation through its interaction with PARP1, an enzyme involved in DNA repair. Our findings reveal a previously unrecognized mechanism by which HBV exploits host cellular machinery to persist in hepatocytes. By inhibiting this pathway, our work highlights potential targets for novel therapies aimed at eliminating cccDNA and achieving a functional cure for HBV infection.