Journal: PLoS Pathogens
Article Title: Chloroquine triggers Epstein-Barr virus replication through phosphorylation of KAP1/TRIM28 in Burkitt lymphoma cells
Figure Lengend Snippet: Cellular KAP1 regulates EBV lytic cycle in Burkitt Lymphoma cells. A-C. HH514-16 Burkitt lymphoma (BL) cells were transfected with scrambled siRNA (non-targeting control; open bars in B, C) or siRNA to KAP1 (black bars in B, C); D-F. BL cells were transfected with empty vector (open bars in E, F) or pFLAG-CMV2-KAP1 (black bars in E, F). Transfected cells were treated with NaB 24 hours after transfection, and harvested at indicated times (A, D), 24 hours (B, E), or 48 hours (C, F) post-treatment to determine relative amounts of ZEBRA, KAP1 and β-actin by immunoblotting (A, D), relative levels of transcripts from EBV lytic genes BZLF1 , BMRF1 and BFRF3 by qRT-PCR after normalization to 18S rRNA using the ΔΔC T method (B, E), and relative levels of cell-associated EBV DNA by qPCR (C, F). Error bars: SEM of 3 experiments with 4 technical replicates. Western blots are representative of 2 experiments. Numbers below blots indicate relative amounts of protein after normalization to β-actin.
Article Snippet: BACmid p2089 was a gift from Professor Henri-Jacques Delecluse [ ]. siRNAs targeting human KAP1 and ATM transcripts and scrambled/non-targeting siRNA were purchased from Santa Cruz Biotechnology (sc-38550, sc-29761 and sc-37007) and Dharmacon (D-001206-13-05 and M-005046-01) and reconstituted with nuclease free water.
Techniques: Transfection, Plasmid Preparation, Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Western Blot