Nodal Regulates Cyclin G2 Gene Expression via FoxO3a and SMAD in Human
Epithelial Ovarian Cancer Cells
Guodong Fu and Chun Peng
Department of Biology, York University, Toronto, ON
We have recently shown that Nodal, a member of the transforming growth factor-

(TGF-

) superfamily, acts through activin receptor-like kinase 7 (ALK7) to up-regulate cyclin G2 (CCNG2) mRNA levels. However, the molecular mechanisms underlying the Nodal/ALK7 up-regulation of CCNG2 gene are not fully understood. The aim of this study was to elucidate how CCNG2 gene expression is regulated at the transcriptional level by Nodal. FoxO3a, a transcription factor belonging to the forkhead transcription factor family, has been shown to regulate transcription of genes involved in cell cycle progression. Examination of CCNG2 promoter sequence revealed the presence of several potential FoxO binding sites. Using luciferase assays, we found that overexpression of FoxO3a increased, while knockdown of FoxO3a decreased, CCNG2 promoter activity. FoxO3a is phosphorylated and thereby inactivated by Akt, a downstream kinase of the PI3K pathway. Inhibition of PI3K pathway also resulted in an increase in CCNG2 transcription. Deletion and mutagenesis analysis, as well as ChIP assay identified a functional FoxO3a binding site located in the core promoter region of CCNG2 gene. Overexpression of Smad2 and Smad3, which are downstream signaling molecules of Nodal, enhanced FoxO3a-induced
CCNG2 transcription. Nodal inhibited Akt activation and FoxO3a inactivation but induced the association of FoxO3a with Smad and thereby CCNG2 promoter activity.
Knockdown of FoxO3a by siRNA reduced Nodal/ALK7 stimulated CCNG2 mRNA level and promoter activity. Taken together, these results suggest that the
Nodal/ALK7 pathway upregulates CCNG2 gene expression through activation of
FoxO3a and Smads.
(Supported by CIHR grants and a mid-career award from OWHC/CIHR to CP)