FIG 7.

An acidic amino acid cluster is required for virion release. (A) Construction of BBLF1 mutant expression vectors. Through site-directed mutagenesis, alanine substitution mutations were generated at the targeted amino acids. G2A and C8A mutations were generated to obtain BBLF1 myristoylation (Myr) and BBLF1 palmitoylation (Pal) mutants, respectively. The BBLF1 acidic amino acid motifs, NDE (amino acids 28 to 31), SDE (amino acids 58 to 65), and both NDE and SDE, were also substituted with alanine. (B) IB of HEK293T cells transfected with mutant vectors showing the expression of BBLF1 mutants. (C) IB of cells transfected with BBLF1 mutants, with or without the proteasome inhibitor MG132. (D) IB of reactivated HEK293-dBBLF1 cells transcomplemented with mutant vectors showing the expression of BZLF1, BBLF1 mutants, and tubulin. Cell-free (E) and cell-associated (F) infectious viral titer of reactivated HEK293-dBBLF1 cells transcomplemented with the constructed mutant vectors. Results are means ± SD of three independent biological replicates.