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. Author manuscript; available in PMC: 2018 Oct 14.
Published in final edited form as: Nat Cell Biol. 2015 Aug 10;17(9):1218–1227. doi: 10.1038/ncb3216

Figure 4. Co-occupancy of YAP/TAZ/TEAD and AP-1 at the same genomic loci.

Figure 4

(a) Density maps of YAP, TAZ, TEAD4 and JUN ChIP-seq reads at YAP/TAZ/TEAD-bound loci. See Supplementary Figures 4a-b for specificity controls of JUN antibody.

(b) Percentage of YAP/TAZ/TEAD4 peaks (n=5522) overlapping with JUN ChIP-seq peaks.

(c) Representative examples of YAP/TAZ/TEAD-bound enhancers co-occupied by JUN in the genome of MDA-MB-231 cells.

(d) Co-presence of YAP and JUN at the same genomic regions. Results are fold enrichment relative to FLAG IP in control (empty vector-transduced) cells (ChIP 1) or relative to IgG negative control (ChIP 2). Data from one representative experiment are shown; experiments were repeated twice with similar results.

(e) In situ PLA detection of endogenous YAP/AP-1 and TEAD1/AP-1 interactions in MDA-MB-231 cells. Nuclei were counterstained with DAPI (blue). The detected dimers are represented by fluorescent dots (red). The specificity of the interactions is revealed by the reduced number of dots detected after depletion of either of the partners with siRNAs. See Supplementary Figure 5a for different combinations of antibodies and Supplementary Table 5 for details about antibodies. Magnification is the same for all pictures.

(f) AP-1 proteins co-precipitate with FLAG-TEAD1 in protein lysates of MDA-MB-231 cells. Input and IP were run on different gels.

(g) TEAD1 co-precipitates with FOSL1 at endogenous protein levels in MDA-MB-231 cells. IP was performed with FOSL1 (N-17) antibodies. All samples were run on the same gel. JUN is a positive control.

(h) TEAD1 co-precipitates with JUND at endogenous protein levels in MDA-MB-231 cells. FOSL1 is a positive control. All samples were run on the same gel.

(i) CTGF and ANKRD1 promoters (either wild-type, or carrying mutations in AP-1 or TEAD binding sites) were cloned upstream of the luciferase coding sequence and their activity was measured in MDA-MB-231 cells. Data are normalized to wild-type promoter sequences. Data are presented as mean+SD of n=4 biological replicates from 2 independent experiments. Binding of TEAD and AP-1 proteins to their respective binding sites was verified by DNA pull down; mutations abolished binding (Supplementary Fig. 5j).

(j) Model of the complex formed by YAP/TAZ/TEAD and AP-1 on DNA.

See Supplementary Figure 7 for uncropped Western blots, and Methods for reproducibility of experiments.