Figure 5.

GSK3β inhibits Neurog2-mediated promoter transactivation. A–C, Effects of GSK3β activity on Neurog2-dependent transactivation of Neurod1 (A), Dll1 (B), and Rnd2 (C) promoters in P19 embryonal carcinoma cells. D, Phosphorylation of Neurog2 at S231/234 by GSK3β in HEK293 cells, and abrogation by S231A/S234A point mutations. E, GSK3β-mediated reduction of Neurog2-mediated Neurod1 promoter transactivation persists when the S231/234 phosphoacceptor residues are mutated to alanines. F, Schematic of Neurog2 deletion constructs. TAD, Transactivation domain; b, basic domain; HLH, helix-loop-helix domain. G, Effect of GSK3β on Neurog2 deletion construct-mediated transactivation of the Neurod1 promoter. The p values are denoted as follows: *p < 0.05, **p < 0.001, ***p < 0.0001. H, Half-life analysis of mutated Neurog2 proteins over a 5 h timeframe. Protein ladder is marked at the left of each panel (red line, 60 kDa). Lanes are labeled with the hour each protein sample was removed from the degradation reaction. I, Quantification of protein half-lives based on autoradiography and densitometry. Error bars indicate SEM.