Self-assembly of Epstein-Barr virus capsids.
Epstein-Barr virus (EBV), a member of the Gammaherpesvirus family, primarily infects B lymphocytes and is responsible for a number of lymphoproliferative diseases. The molecular genetics of the assembly pathway and high-resolution structural analysis of the capsid have not been determined for this lymphocryptovirus. As a first step in studying EBV capsid assembly, the baculovirus expression vector (BEV) system was used to express the capsid shell proteins BcLF1 (major capsid protein), BORF1 (triplex protein), BDLF1 (triplex protein), and BFRF3 (small capsid protein); the internal scaffold protein, BdRF1; and the maturational protease (BVRF2). Coinfection of insect cells with the six viruses expressing these proteins resulted in the production of closed capsid structures as judged by electron microscopy and sedimentation methods. Therefore, as shown for other herpesviruses, only six proteins are required for EBV capsid assembly. Furthermore, the small capsid protein of EBV (BFRF3), like that of Kaposi's sarcoma-associated herpesvirus, was found to be required for assembly of a stable structure. Localization of the small capsid protein to nuclear assembly sites required both the major capsid (BcLF1) and scaffold proteins (BdRF1) but not the triplex proteins. Mutational analysis of BFRF3 showed that the N-terminal half (amino acids 1 to 88) of this polypeptide is required and sufficient for capsid assembly. A region spanning amino acids 65 to 88 is required for the concentration of BFRF3 at a subnuclear site and the N-terminal 65 amino acids contain the sequences required for interaction with major capsid protein. These studies have identified the multifunctional role of the gammaherpesvirus small capsid proteins.
Henson
Brandon W
BW
Viral Oncology Program, The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University, 353 CRB1 Viral Oncology, 1650 New Orleans St., Baltimore, MD 21117, USA.
Perkins
Edward M
EM
Cothran
Jonathan E
JE
Desai
Prashant
P
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002213:Capsid / Q000378:metabolism*; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002499:Centrifugation, Density Gradient; D004252:DNA Mutational Analysis; D004854:Herpesvirus 4, Human / Q000502:physiology*; D046529:Microscopy, Electron, Transmission; D011485:Protein Binding; D025941:Protein Interaction Mapping; D018411:Spodoptera; D019065:Virus Assembly
D036022:Capsid Proteins