Fig. 5

IRF4 is a repressor of MICA promoter activity in MM cells. a MICA cell surface expression was analyzed by flow cytometry on pEF.CMV.EGFP-IRF4-lentivirus (or pEF.CMV.EGFP control virus) infected SKO-007(J3) cells (72 h), by gating on GFP⁺ cells as indicated in the panel. After infection, cells were separated into different drug treatment groups, untreated or treated with JQ1 (0.5 μM). The ratio MFI between JQ1-treated on untreated cells is shown in a representative experiment. b SKO-007(J3) cells were cotransfected with 3 μg of the indicated luciferase reporter vector and pRL-TK as described above + 1 μg of an expression vector encoding a truncated form of the human IRF4, IRF4-DN, or an empty control vector pcDNA3. After 48 h, cells were harvested and protein extracts were prepared for the luciferase and Renilla assays. Results are expressed as relative luciferase activity normalized to protein concentration as well as to Renilla activity produced off the internal control plasmid and represent the mean value of four independent experiments (*P < 0.05). c SKO-007(J3) cells were transfected with 3 μg of the indicated luciferase reporter vector and pRL-TK as described above. After 48 h, cells were harvested and protein extracts were prepared for the luciferase and Renilla assays. Results are expressed as relative luciferase activity normalized to protein concentration as well as to Renilla activity produced off the internal control plasmid and represent the mean value of four independent experiments (*P < 0.05). d Schematic representation of the IRF4 response element deleted in the mutant promoter MICA/-270-DEL