FIG. 7.

CTCF-RAD21 interaction represses KSHV transcription. (A) Schematic of Flag-tagged RAD21, CTCF, and CTCF-RAD21 (C-R) fusion protein constructs. FLAG-tagged constructs were transfected into 293T cells, and their expression was determined by Western blotting, with antibody to Flag. (B) Luciferase assays with pGL3-K14 or pGL3-73 luciferase constructs in the presence of ectopic expression of FLAG-CTCF, FLAG-RAD21, or a combination of the two, as indicated. (C) Same assays as in panel B, except FLAG-CTCF-RAD21 fusion protein was included. (D) BCBL1 cells were transfected with FLAG-RAD21, FLAG-CTCF, or FLAG-CTCF-RAD21, along with mGFP expression vector. GFP-positive BCBL1 cells were sorted, and KSHV gene transcription for K14, ORF74, ORF73, or ORF72 was quantified by RT-qPCR relative to cotransfected mGFP.