FIG 7.

LANA mutants with deletions including amino acids 194 to 273 are deficient for episome maintenance after TR DNA transfection, as assessed by Gardella gel analysis. BJAB cells, BJAB cells stably expressing LANA, or BJAB cells stably expressing each deletion mutant were transfected with p8TR. Forty-eight hours after transfection, the cells were seeded into 96-well plates and selected for drug resistance encoded on the p8TR plasmid. Twenty-one days after plating, the cells were expanded from individual wells and analyzed by Gardella gel analysis for the presence of episomes. (A and B) Gardella gels containing LANA or LANA mutants, as indicated. Lanes with p8TR plasmid or with BCBL-1 cells (a KSHV-infected primary effusion lymphoma cell line) are also indicated. O, gel origin; E, BCBL-1 episomes; L, BCBL-1 linear forms (from lytic replication); ccc, p8TR cccDNA. The number of days of drug selection are indicated below each gel. (C and D) Western blot analyses of LANA, LANA mutants, or tubulin. The lanes correspond to those shown in the Gardella gels. Brightness and contrast were uniformly adjusted in each panel using Adobe Photoshop.