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. 2016 Mar 3;6:24–31. doi: 10.1016/j.bbrep.2016.02.015

Fig. 2.

Fig. 2.

YAP transactivation, PDZ-binding, and TEAD-binding domains are required for TEAD-mediated transcriptional activity. (A) Schematic illustration of the pGT4Tluc luciferase TEAD reporter. HEK293T cells were transfected with pGT4Tluc (Control), FLAG-tagged wild-type (WT), ΔTAD, ΔPDZ or ΔTEAD mYAP constructs (B), and pCI-HA-TEAD-2 (HA-TEAD2) as indicated. (C) After 24 h cells were harvested and luciferase activity was determined. Firefly luciferase activity was normalised to Renilla luciferase activity. The average luciferase activities were normalised to WT mYAP without HA-TEAD2, which was set to 100%. Data is presented as mean+SEM from three independent experiments. *p<0.05, **p<0.01, ***p<0.001. (D) Cell lysates from (C) were separated by SDS-PAGE, transferred to membrane and immunoblotted for FLAG and β-Actin as indicated. Size markers are shown in kilodaltons. (E) HEK293T cells were transfected with pGT4Tluc (Control), mYAP or hYAP1-2α constructs, and pCI-HA-TEAD-2 (HA-TEAD2) as indicated. After 24 h cells were harvested and processed as in (B). Cell lysates were immunoblotted for YAP and β-Actin as indicated. Size markers are shown in kilodaltons. Vertical line indicates the samples were blotted on separate gels.