Objective: Forkhead box O1 (FoxO1) plays a crucial role in the development of many tumors. Cyclin D kinase (CDK) 1 could influence the nuclear export and activity of FoxO1 through phosphorylation of serine (S)249. However, the effects of S249 phosphorylation in the development of glioma remain unclear. The aim of the present study is to assess the function of FoxO1:S249V mutant, which was converted S249 phosphorylation site into valine (V) residues in the glioma development.
Methods: FoxO1-knockdown U251 glioma cells (U251-KD cells) were established by infection of retrovirus particles with FoxO1 siRNA and FoxO1 restored cells (FoxO1:S249V) were obtained by re-introduction of FoxO1:S249V cDNA. We detected mRNA expression by real-time PCR, and cell cycle arrest and apoptosis by flow cytometric assay, and cell proliferation by BrdU assay and CCK-8 assay. The protective effects of FoxO1:S249V were detected by the xenograft tumor formation assay.
Results: The FoxO1 mRNA expression was significantly decreased in the glioma specimens (n = 24). The U251-KD cells showed downregulation of p27 and Bim, while the phosphorylation of CDK1 was upregulated. FoxO1:S249V cells inhibited the phosphorylation of S249, and induced G2/M cell cycle arrest, following reduced cell growth and increased apoptosis. Moreover, FoxO1:S249V expression effectively inhibits the glioma growth.
Conclusion: Our findings suggest that the forced FoxO1:S249V suppressed the cell growth through G2/M cell cycle arrests and increased apoptosis in glioma.
Keywords: FoxO1; apoptosis; cell cycle; cell proliferation; glioblastoma.