Spatiotemporally different DNA repair systems participate in Epstein-Barr virus genome maturation.

Productive replication of Epstein-Barr virus occurs in discrete sites in nuclei, called replication compartments, where viral DNA replication proteins and host homologous recombinational repair (HRR) and mismatch repair (MMR) factors are recruited. Three-dimensional (3D) surface reconstruction imaging clarified the spatial arrangements of these factors within the replication compartments. Subnuclear domains, designated BMRF1 cores, which were highly enriched in viral polymerase processivity factor BMRF1 could be identified inside the replication compartments. Pulse-chase experiments revealed that newly synthesized viral genomes organized around the BMRF1 cores were transferred inward. HRR factors could be demonstrated mainly outside BMRF1 cores, where de novo synthesis of viral DNA was ongoing, whereas MMR factors were found predominantly inside. These results imply that de novo synthesis of viral DNA is coupled with HRR outside the cores, followed by MMR inside cores for quality control of replicated viral genomes. Thus, our approach unveiled a viral genome manufacturing plant.