Figure 2.

Role of TEAD in YAP-driven transcriptional activity. (A) Localization of endogenous YAP/TEAD1 complexes by in situ PLA in HOS cells. The red signal was obtained using Alexa555-labeled hybridization oligo nucleotides targeting amplified in situ PLA products. DAPI (blue) staining was used for nuclear visualization (left panel). Bars indicate means ± SD of three independent experiments (* p < 0.05, ** p < 0.01, right panel). (B HEK293 were transiently co-transfected with the YAP-S94A, YAP-S127A, TEAD1 or empty vector as indicated. 48 h after transfection lysates were subjected to immunoprecipitation (IP) with anti-Flag antibody followed by Western blotting (WB) by anti-Flag and anti-HA antibodies as indicated. (C) HOS cells were co-transfected with the TEAD-specific construct (TEAD)8-lux with or without empty, YAPS94A and YAPS127A expression vectors. Bars indicate means ± SD of four independent experiments, each performed in triplicate (** p < 0.01). (D) YAP mRNA steady-state levels were quantified by RT-q-PCR analysis in mock-, YAPS94A- and YAPS127A-transfected K-HOS cells. Bars indicate means ± SD of four independent experiments, each performed in duplicate (* p < 0.05, left panel). YAP production was detected by Western blot analysis in mock-, YAPS94A- and YAPS127A-transfected K-HOS cells. Results shown are representative of two independent experiments (right panel). (E) Localization of YAP/TEAD1 complexes by in situ PLA experiments in mock-, YAPS94A- and YAPS127A-transfected K-HOS cells. The red signal was obtained using Alexa555-labeled hybridization oligo nucleotides targeting amplified in situ PLA products. DAPI (blue) staining was used for nuclear visualization (left panel). Bars indicate means ± S.D. of three independent experiments (** p < 0.01, right panel). (F) Mock-, YAPS94A- and YAPS127A-transfected cells were transiently transfected with the TEAD-specific construct (TEAD)8-lux. Bars indicate means ± SD of four independent experiments, each performed in duplicate (* p < 0.05).