1.

IL-1β inhibits transcriptional activity of the human TβRII promoter via–47/–15 region. A: HACs were cultured for 5–6 days in 10% FCS-containing DMEM and transiently transfected by nucleofection with TβRII promoter constructs as described in Methods. Then, they were treated with IL-1β for 24 hrs, and luciferase activities were determined and expressed as RLU fold induction over control. Means of triplicate absolute values are given on each histogram. B: Schematic representation of the human TβRII promoter. Putative binding sites are represented: transcription initiation site; PRE1/2: Positive regulatory element 1/2; NRE: Negative regulatory element. C: HACs were transfected with double-strand oligonucleotides corresponding to the wild-type (WT) or mutated (Mut) −47/−15 sequence of the human TβRII promoter. Then, they were incubated in DMEM + 2% FCS in the presence or not of IL-1β (1 ng/ml) for 24 hrs. Thereafter, TβRII mRNA levels were analysed by real-time RT-PCR. The modulation of TβRII mRNA expression was expressed as per cent of controls, after normalization to GAPDH signal. D: HACs were transiently transfected by nucleofection with wild-type or mutated p219TbRII or p47TbRII vectors as described in Methods. Then, they were treated with IL-1β for 24 hrs, and luciferase activities were determined and expressed as RLU.