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. 2016 Jun 3;36(3):e00343. doi: 10.1042/BSR20160123

Figure 4. Simulation of HEY2 phosphorylation at residue Ser-67 inhibits its ability to enhance p53 transcriptional activity.

Figure 4

(A) U2OS cells were transfected with 100 ng of PIG3-LUC and 200 ng of expression vectors for HEY1, HEY2 or HEYL (left panel) or expression vectors for HEY2 or HEY2-S67D (right panel). After transfection, cells were incubated 24 h. Subsequently cell lysates were assayed using a dual luciferase reporter system. Normalized values are expressed relative to the activity of the reporter in the absence of HEY1. The results shown represent the averages of results of three independent experiments assayed in duplicate + S.D. (B) Simulation of HEY2 phosphorylation at residue Ser-67 increases protein stability. U2OS cells were transfected with expression vectors for Flag-tagged HEY2 or HEY2-S67D. Twenty-four hours after transfection cells were treated with cycloheximide (CHX, 10 μg/ml) and HEY2 protein levels were analysed by western blotting at 0, 3 and 6 h after CHX addition.