Hepatitis B virus (HBV) regulatory protein HBx is required to initiate and maintain productive virus replication. HBx is thought to achieve these functions by hijacking Cullin-RING ubiquitin E3 ligase 4 (CRL4) to target the SMC5/6 complex, a restriction factor for extrachromosomal viral DNA degradation. Studies also indicated that DNA damage-binding protein 1 (DDB1), an adaptor protein within CRL4 complex, may stimulate HBV transcription via a mechanism that does not involve an interaction with HBx. In the present study, using a model system of recombinant covalently closed circular DNA (rcccDNA) of HBV, it was confirmed that CRL4 could upregulate the secretion of HBV antigens in the absence of HBx (rcccDNA^X-null) in cultured hepatoma cell lines. However, this effect of CRL4 was abolished by serum deprivation in the cell culture. In addition, it was found that CRL4 failed to stimulate the expression of rcccDNA^X-null in non-proliferating hepatocytes in a mouse model with DNA hydrodynamic injection. Thus, the HBx-independent, CRL4-upregulated HBV expression is likely to be closely related to rapid proliferation of cultured hepatoma cell lines. The study thus revealed a previously unappreciated role of CRL4 in HBV virology, and would be helpful for better understanding of the biological significance of HBx.