EBNA1 partitions Epstein-Barr virus plasmids in yeast cells by attaching to human EBNA1-binding protein 2 on mitotic chromosomes.
Epstein-Barr virus (EBV) episomal genomes are stably maintained in human cells and are partitioned during cell division by mitotic chromosome attachment. Partitioning is mediated by the viral EBNA1 protein, which binds both the EBV segregation element (FR) and a mitotic chromosomal component. We previously showed that the segregation of EBV-based plasmids can be reconstituted in Saccharomyces cerevisiae and is absolutely dependent on EBNA1, the EBV FR sequence, and the human EBNA1-binding protein 2 (EBP2). We have now used this yeast system to elucidate the functional contribution of human EBP2 to EBNA1-mediated plasmid partitioning. Human EBP2 was found to attach to yeast mitotic chromosomes in a cell cycle-dependent manner and cause EBNA1 to associate with the mitotic chromosomes. The domain of human EBP2 that binds both yeast and human chromosomes was mapped and shown to be functionally distinct from the EBNA1-binding domain. The functionality and localization of human EBP2 mutants and fusion proteins indicated that the attachment of EBNA1 to mitotic chromosomes is crucial for EBV plasmid segregation in S. cerevisiae, as it is in humans, and that this is the contribution of human EBP2. The results also indicate that plasmid segregation in S. cerevisiae can occur through chromosome attachment.
Kapoor
Priya
P
Department of Medical Genetics and Microbiology, University of Toronto, Toronto, Canada M5S 1A8.
Frappier
Lori
L
D002352:Carrier Proteins / Q000378:metabolism*; D020090:Chromosome Segregation; D015825:Chromosomes, Fungal / Q000378:metabolism*; D019309:Epstein-Barr Virus Nuclear Antigens / Q000378:metabolism*; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D008938:Mitosis; D009687:Nuclear Proteins / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics*; D016601:RNA-Binding Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D020798:Two-Hybrid System Techniques
D002352:Carrier Proteins; C432687:EBNA1BP2 protein, human; C499008:EBP2 protein, S cerevisiae; D019309:Epstein-Barr Virus Nuclear Antigens; D009687:Nuclear Proteins; D016601:RNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; C417029:EBV-encoded nuclear antigen 1