Figure 2

HDACi-induced NKG2D ligand regulation did not involve the DDR. (a) HEK-293 cells were treated with 100 nM LBH589 for 16 h, washed and incubated with primary human NK cells of healthy donors in indicated ratios for 3 h. Mean and s.e. of four independent experiments. (b) FACS analysis of HEK-293 cells after 1 h of preincubation with 2 μM actinomycin D or 10 μM cycloheximide, and treatment with LBH589 (100 nM) for 6 h. (c) Real-time PCR for HEK-293 cells treated with 10 μM Ara-C or 100 nM LBH589 for the indicated periods relative to GAPDH. (d) HEK-293 cells were treated with the ATM/ATR inhibitor CGK733 (5 μM) together with the damage inducer Ara-C (10 μM) or the HDACis LBH589 (100 nM) and trichostatin A (TSA; 250 nM) for 16 h to analyze the surface expression of MICA/B, ULBP1, ULBP2 and ULBP3. (e) MDA-231 and 911 cells were incubated with ATM/ATR inhibitor CGK733 (5 μM) or ATM inhibitor KU55933 (10 μM) and TSA (250 nM) for 16 h and subsequently studied for MICA/B expression by FACS. (f) Intracellular FACS staining of HEK-293 cells for DNA damage marker phospho-γH2AX (S139 phosphorylated) after incubation with 10 μM Ara-C or 100 nM LBH589 for 1 h.