LANA-binding to the lytic KSHV promoters is required for the EZH2 recruitment during de novo infection.

(A) Immunoblot analysis of SLK cells infected with lentiviruses expressing 3xFLAG-tagged WT or mutant LANA using anti-FLAG antibody. (B) ChIP assays for the recruitment of LANA (using αLANA antibody) and EZH2 on viral promoters in RTA KO and LANA/RTA dKO KSHV-infected SLK cells as well as in LANA/RTA dKO KSHV-infected SLK cells expressing WT or mutant LANA. (C) Total lysates of 293T cells expressing either 3xF-LANA or 3xF-GMRRL mutant LANA were subjected to FLAG immunoprecipitation followed by immunoblotting with anti-FLAG and anti-EZH2 antibodies. Input lysates were included as controls. (D) ChIP assays for LANA and EZH2 occupancy on the KSHV promoters during de novo LANA/RTA dKO KSHV infection of WT- or GMRRL mutant LANA-expressing SLK cells. (E) Model showing LANA-mediated recruitment PRCs onto the KSHV genome during de novo infection.