Identification of two separable modules in the duck hepatitis B virus core protein.

Hepadnavirus replication requires the concerted action of the polymerase and core proteins to ensure packaging of the RNA pregenome and DNA maturation. The arginine-rich C terminus of the core protein plays an essential role in both of these steps while being dispensable for nucleocapsid formation. In an attempt to identify other functional domains of the core protein, we performed a series of trans-complementation experiments analyzing the ability of duck and human hepatitis B virus (DHBV and HBV) core protein subunits to support the replication of a core-defective DHBV genome. Plasmids expressing the N-terminal amino acids 1 to 67 or the remaining C-terminal portion, amino acids 67 to 262, of the DHBV core protein were cotransfected into LMH cells along with a replication-deficient construct coding for the DHBV pregenome and polymerase. Neither the N nor the C terminus alone yielded replication-competent core particles. However, cotransfection of plasmids that separately expressed both regions restored a normal replication pattern. Furthermore, the DHBV C terminus but not the N terminus could be replaced by the corresponding domain of the HBV core protein in this assay. Finally, coexpression of the complete HBV core protein and the N terminus from DHBV resulted in DHBV replication, while the HBV core protein alone was not functional. Taken together, these findings suggest a modular organization of the DHBV core protein in which the C terminus is functionally conserved among different hepadnaviruses.     

Distinct functions and requirements for the Cys-His boxes of the human immunodeficiency virus type 1 nucleocapsid protein during RNA encapsidation and replication.

The process of retroviral RNA encapsidation involves interaction between trans-acting viral proteins and cis-acting RNA elements. The encapsidation signal on human immunodeficiency virus type 1 (HIV-1) RNA is a multipartite structure composed of functional stem-loop structures. The nucleocapsid (NC) domain of the Gag polyprotein precursor contains two copies of a Cys-His box motif that have been demonstrated to be important in RNA encapsidation. To further characterize the role of the Cys-His boxes of the HIV-1 NC protein in RNA encapsidation, the relative efficiency of RNA encapsidation for virus particles that contained mutations within the Cys-His boxes was measured. Mutations that disrupted the first Cys-His box of the NC protein resulted in virus particles that encapsidated genomic RNA less efficiently and subgenomic RNA more efficiently than did wild-type virus. Mutations within the second Cys-His box did not significantly affect RNA encapsidation. In addition, a full complement of wild-type NC protein in virus particles is not required for efficient RNA encapsidation or virus replication. Finally, both Cys-His boxes of the NC protein play additional roles in virus replication.