Journal: Journal of cell science

Article Title: Promoter occupancy of MLL1 histone methyltransferase seems to specify the proliferative and apoptotic functions of E2F1 in a tumour microenvironment.

doi: 10.1242/jcs.126235

Figure Lengend Snippet: Fig. 8. Mutually exclusive promoter occupancy of E2F1 co-activator/repressor complexes in hepatoma cells. (A) Liver tissue lysates from 3- and 6-month-old C57B6 control (C) and X15-myc transgenic (T) mice were immunoprecipitated with anti-E2F1 and immunoblotted for CBP, GCN5 and Brg1. One-tenth of the tissue lysate was used to probe the expression of GCN5 and HBx. Actin was used as loading control. (B–E) HepG2.2.15 cells were transfected with either scrambled (scr) siRNA or CBP-specific siRNA. Cells were serum starved 24 hours post-silencing and subjected to serum stimulation (SS) either in the absence or presence of cisplatin (SS + cisplatin) and processed for ChIP-qPCR analysis. Enrichment of CBP and Brg1 occupancy over IgG control was measured on either MCM4 (B) or Apaf-1 gene promoters (D). Similarly, the same samples were used to measure the enrichment of MLL1 and E2F1 occupancy over IgG control on either MCM4 (C) or Apaf-1 gene promoters (E). Data are means 6 s.d. of three independent experiments; *P,0.05 and #P,0.01.

Article Snippet: For RNA interference studies, 2.0 mg of HBx shRNA (X-D) was transfected using Fugene 6 (Invitrogen). siRNA against E2F1, MLL1 and CBP (all from Santa Cruz Biotechnology) were used for transfection according to manufacturer’s instructions.

Techniques: Control, Transgenic Assay, Immunoprecipitation, Expressing, Transfection, ChIP-qPCR