Gli1 h300

The following antibodies and dilution were used: Gli1 (# 2643 S Cell Signaling) 1:1000, Gli1 H300 Santa Cruz Biotechnology (sc-20687) 1:100, Gli2 (# AF3635 R&D) 1:1000, HDAC1 (ab31263-100 AbCam) 1:2000, HDAC2 (sc-7899X Santa Cruz) 1:2000, Vinculin (sc-7649 Santa Cruz) 1:1000, Actin (sc-1616 Santa Cruz) 1:1000, PARP (# 9542 S Cell Signaling) 1:1000, Tubulin (Santa Cruz Biotechnology sc-8035) 1:1000, Cleaved Caspase 3 (# 9661 S Cell Signaling) 1:1000, p21 (sc-817 Santa Cruz) 1:1000, p53 (#2524 Cell signaling) 1:1000, Bcl-2 (sc-492 Santa Cruz) 1:1000, Acetyl Histone H3 (# 06-599 Millipore) 1:1000, Ki67 (MA5-14520 Pierce) 1:100, Flag-M2 Peroxidase (A8592 Sigma) 1:5000, AcGli1 was described previously ref. 16 (link) and was used at 1:50 dilution for IHC and 1:500 for western blot.

ChIP was performed as described earlier.^{57 (link), 58 (link)} Immunoprecipitated DNA from HEK293T with Gli1 (H300) (cat.#sc-20687; Santa Cruz Biotechnology) or Maml1 (D3E9) or IgG (normal rabbit IgG) antibodies was eluted and analyzed by semiquantitative PCR, using a primer set encompassing two predicted binding sites for Gli zinc finger family (V$GLIF Matrix Family) (from −538 to −521; from −461 to −445, corresponding to dark circles in Figure 1f, low panel), on human Patched1 promoter (GXP_227868, from −891 to −87 relative to start codon). Human Patched1 promoter was identified using MatInspector (Software GmbH, Munich, Germany). The primer set used to specifically amplify Gli binding sites is the following: 5′-GAACCCAGCAGCCAGAGC-3′ and 5′-CGACCCCTTCACTGCAGAA-3′.