Figure 3 | Scientific Reports

Figure 3

From: Rab-GTPase binding effector protein 2 (RABEP2) is a primed substrate for Glycogen Synthase kinase-3 (GSK3)

Figure 3

RABEP2 is phosphorylated in intact cells by GSK3. (A) FLAG-RABEP2 WT, Ser200Ala, Ser204Ala or Ser200/204Ala mutants were expressed in HEK293 cells which were then incubated ± 10 μM CT99021 (GSK3 inhibitor, CT) for 1 h prior to cell lysis. Protein lysates were immunoblotted with the antibodies as indicated. (B) and (C) HEK293 or H9C2 cells were incubated ± 10 μM CT99021 (GSK3 inhibitor, CT) for 1 h prior to cell lysis. Endogenous RABEP2 was immunoprecipitated from cell lysates and immunoblotted with the indicated antibodies. A representative image of two separate experiments performed in at least duplicate are provided. Quantification was performed using the Licor Odyssey software and the ratio of phosphor to total RABEP2 is provided in the graphs to the right of each figure. In all parts of the figure the fluorescent images were cropped between 60–70 kDa (original images of complete gels are provided in Supplementary Information), and the separate red 700 nm or green 800 nm Licor Odyssey scan (from same gel) plus the merged image are provided. In C the greyscale image of the same gel is also provided for clarity.

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