Figure 5.

Role of phosphoinositide 3-kinase/mechanistic target of rapamycin in the regulation of translation of SOX2 protein and aldehyde dehydrogenase (ALDH)1A1 expression. A) Silencing AKT1 or EIF4E statistically significantly reduces SOX2 protein levels in 013C and 067C cells. B) Treatment of 013C and 067C cells with the EIF4E inhibitor 4EGI-1 decreased SOX2 protein. C) The EIF4E protein physically binds SOX2 mRNA as measured by RNA immunoprecipitation using EIF4E specific antibodies. Western blot analysis showing successful precipitation of EIF4E protein. D) Exogenous expression of SOX2 (013C, 067C cells) increases ALDH1A1 protein. E) ALDH1A1 levels statistically significantly increase within six days following exogenous expression of SOX2. F) SOX2 expression increased the ALDH⁺ population in both 013C and 067C cells. G and H) Silencing of SOX2 in overexpression cells decreases ALDH1A1 expression and protein levels and (I) the ALDH⁺ population. J) Exogenous expression of SOX2 activates an ALDH1A1-promoter as measured by a luciferase assay. Chromatin immunoprecipitation using SOX2 antibodies suggests SOX2 associates with the ALDH1A1 promoter in 013C cells. Graphed results are presented as mean ± SD of three or more independent experiments. Statistical significance was calculated by the two-tailed Student’s t test (*P < .05; †P < .01). ALDH = aldehyde dehydrogenase; mTOR = mechanistic target of rapamycin; PI3K = phosphoinositide 3-kinase.