Changes of cell cycle distribution and nuclear structure in NK cells by EBV infection

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Although considerable part of natural killer (NK) cell neoplasms possess EBV genome, there has been no direct evidence that EBV infects human NK cells in vitro. In this study, we demonstrated EBV entry into the NK cells using enhanced green fluorescence (EGFP)-containing recombinant EBV. After 48 hr, about 30% of NK cell lines and more than 40% of normal peripheral blood NK cells were positive for EGFP signal. In situ hybridization for EBNAs and BHLFs showed that latent and lytic infections coexisted at the early phase of EBV infection. NK cells in latent but not lytic EBV infection tended to show a bizzare and giant shape. Cdc2, cyclin B, and Cdc25C, which are important for G2/M transition, increased in protein levels and partially migrated to the nucleus though their activation was inhibited. Flow cytometric analysis showed mainly G2 arrest in EBV-infected NK cells. Although nuclear distribution of PML, SC35, and HP1 was unchanged, cyto- and nuclear-skeletons made of actin, tubulin and lamin has apparently rearranged and abnormalities of centrosome numbers were detected. We established eight EBV-carrying clones, which showed both latency types I and II. Both of them are recognized in EBV-associated NK-cell neoplasms. The cell size became larger and the frequency of micronuclei formation, a hallmark of genomic instability, increased in EBV-carrying NKL clones. Furthermore, these clones were resistant to various chemotherapeutic agents and cell dilution compared to the parent NK cell lines.