Investigating YAP1 Promoter Regulation

For chromatin immunoprecipitation (ChIP), the promoter region of YAP1 was defined within 2000 base pairs upstream of the transcription start site and the sequence was obtained from the UCSC Table Browser [46 (link)]. Based on TRANSFAC (version 8.3), two putative binding sites of c-Jun were predicted at the region [47 (link), 48 (link)]. Crosslinking, lysis, and chromatin shearing were performed as previously described [25 (link)]. Targeted fragments (300–500 base pairs) were pulled down by Magnetic Dynabeads Protein G (1004, Life Tech), with c-Jun antibody or Normal IgG antibody. Primers for binding site sequences are listed in Supplementary Table S5. For co-immunoprecipitation, 500 µg cell lysate was used for immunoprecipitation with 20 µl magnetic beads, bonding with YAP1 antibody or normal IgG antibodyA light chain-specific secondary antibody (ab99632, Abcam) were used to avoid the signal of the heavy chain (~50 kDa).