FOXO Proteins Regulate Tumor Necrosis Factor-related Apoptosis Inducing Ligand Expression

Abstract

Mutations in PTEN occur in 60 -80% of prostate cancers and lead to a constitutive activation of the phosphatidylinositol 3-kinase pathway and a resultant loss of activity of the FOXO family of forkhead transcription factors FKHRL1 and FKHR. To provide insight into the role of PTEN mutations in prostate cancer, we used microarrays to identify genes regulated by FKHRL1 and FKHR in LAPC4 prostate carcinoma cells. These studies revealed that adenoviral overexpression of FKHRL1 and FKHR in the LAPC4 prostate cancer cell line resulted in apoptosis and induced the expression of many genes that affect cellular proliferation or survival. The expression of one of these FOXO-regulated genes, TRAIL, a pro-apoptotic member of the tumor necrosis factor family, was decreased in human metastatic prostate tumors. The altered expression of TRAIL in these tumors correlated directly with decreased PTEN expression and the resultant loss of FKHRL1 and FKHR activity. Analysis of the effects of FOXO proteins on the TRAIL promoter localized the FKHRL1 responsive element of the TRAIL promoter to nucleotides ؊138 to ؊121 and demonstrated that TRAIL is a direct target of FKHRL1. These findings suggest that the decreased activity of FKHRL1 and FKHR in prostate cancers resulting from loss of PTEN leads to a decrease in TRAIL expression that may contribute to increased survival of the tumor cells.The forkhead transcription factors are DNA-binding proteins characterized by the presence of a conserved 110-amino acid winged helix DNA binding domain (1). They play important roles in embryogenesis, tumorigenesis, and maintenance of differentiation status. In Caenorhabditis elegans the forkhead transcription factor DAF-16 is under control of the insulin receptor/PI 3-kinase 1 pathway (2). The human orthologs of DAF-16 include FKHRL1, FKHR, and AFX and belong to the FOXO subfamily of forkhead transcription factors. The PI 3-kinase pathway, via activation of its downstream kinase Akt, phosphorylates each of the FOXO proteins at three different Ser/Thr residues (3). These phosphorylated FOXO proteins interact with 14-3-3 proteins and are subsequently sequestered in the cytoplasm where they are inactive. Inhibition of the PI 3-kinase pathway by PTEN overexpression or pharmacologic means leads to dephosphorylation and nuclear translocation of active FKHRL1, FKHR, and AFX, which in turn leads to cell cycle arrest and apoptosis (4). Conversely, loss of PTEN activity results in increased Akt activity leading to inhibition of FOXO protein activity through their phosphorylation and cytoplasmic sequestration. In addition, the expression of dominant negative FKHRL1 results in the inhibition of apoptosis, demonstrating that FOXO transcriptional activity controls cellular proliferation and apoptosis downstream of PTEN (5).The PTEN gene was initially identified by its frequent loss in glioblastomas (6), but subsequent studies have shown that PTEN is commonly mutated in prostate cancer (7-12). In prostate tumors, the loss of PTEN occurs late in the tumorigenic pro...

Forkhead protein FKHR and its phosphorylated form p-FKHR in human prostate cancer

“…Activation of these forkhead transcription proteins induces apoptosis through regulation of proapoptotic proteins including Fas ligand, TRAIL, and Bim in cancer cell lines [19,22,23]. Therefore, it seems possible that the higher the FKHR levels, the higher the tumor-suppressing effect FKHR would produce.…”

Deacetylation of FOXO3 by SIRT1 or SIRT2 leads to Skp2-mediated FOXO3 ubiquitination and degradation

“…Skp2 overexpression reduced FOXO3-mediated transcription, whereas Skp2 knockdown increased FOXO3 transactivation (Figures 3d and e). Furthermore, we found that Skp2 knockdown resulted in an increased expression of a FOXO3 target gene, tumor necrosis factor-related apoptosis inducing ligand (TRAIL) (Modur et al, 2002;Fu et al, 2009) (Figure 3f). Repression of Skp2 expression also enhanced cell death (Figure 3g).…”

NAMPT overexpression in prostate cancer and its contribution to tumor cell survival and stress response

“…Consistent with decreased CAT and SOD gene expression, the level of reactive oxygen species was significantly increased in PC3 cells upon NAMPT inhibition (Figure 8b). FOXO3a is a direct transcriptional activator of CAT and SOD promoters (Kops et al, 2002;Nemoto and Finkel, 2002), and is highly expressed in prostate cancer cell lines (Modur et al, 2002). Whereas nuclear localization of FOXO3a is seen in normal prostate cells, increasing cytoplasmic accumulation of FOXO3a occurs in more advanced prostate cancer (Shukla et al, 2009).…”