![]() Isolation of KSHV genomic DNA and quantification of its copy number. 293T cells were cultured in Dulbecco's modified Eagle's medium with 10% fetal bovine serum and antibiotics. KSHV-positive PEL cells (BCBL1, BC-1, and BC-3) and both Epstein-Barr virus (EBV)- and KSHV-positive cells (JSC-1) were grown in RPMI medium (Gibco BRL) containing 10% heat-inactivated fetal bovine serum and the antibiotics penicillin and streptomycin (50 U/ml). ![]() We also show that the histone deacetylase inhibitor sodium butyrate (NaB) leads to a rapid loss of Rad21 and other cohesin subunits from chromatin, suggesting that cohesins play a regulatory role in the transcriptional repression of the KSHV immediate early lytic gene cluster. We also show that this region contains a bivalent histone modification pattern that is disrupted upon Rad21 depletion. We show that CTCF and cohesins bind to the region upstream of the divergent promoter for ORF50 and ORF45. ![]() We found that depletion of core cohesin subunits, especially Rad21, causes reactivation of latent KSHV immediate early gene expression and viral DNA replication.
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