FIG 8.

The C-terminal region of BBLF1 can partially suppress virion release. (A) Amino acid sequences of mutants with a BBLF1-C-terminal domain (CTD) mutation obtained through gene editing. (B) Akata-EBV cells were reactivated through treatment with anti-human IgG for 3 days and then subjected to IB to assess the expression of BZLF1, BMRF1, BALF4, and BBLF1. (C, D) Cell-free (C) and cell-associated (D) virion titers of Akata-EBV cells at 3 days postreactivation. Cell-free and cell-associated virion samples were obtained as described in Fig. 3A. (E) EBV genome copies in DNase-treated culture supernatant. (F to H) Transcomplementation of the BBLF1-knockout virus. HEK293-dBBLF1 cells were transfected with BBLF1-HA expression vectors (WT or BBLF1-CTD mutant) during lytic reactivation and their cell-free (F), cell-associated (G), and total (H) progeny virions were quantified. (I) IB of transcomplemented dBBLF1 cells showing the expression of BZLF1, BBLF1-WT-HA, BBLF1-CTD-HA, and tubulin as a housekeeping gene. (J) A similar experiment to that shown in Fig. 8H was conducted, except that WT HEK293-EBV cells were used. (K) IB of transcomplemented WT cells showing the expression of BZLF1, BBLF1-WT-HA, BBLF1-CTD-HA, and tubulin as a housekeeping gene. The data are means ± SD of three independent biological replicates. Student's t test was performed to assess significance. No significant difference (ns), P > 0.05; *, P < 0.05; **, P < 0.01.