Mechanisms underlying p53 regulation of PIK3CA transcription in ovarian surface epithelium and in ovarian cancer

Abstract

Inactivation of the transcription factor and tumor suppressor p53, and overexpression or mutational activation of PIK3CA, which encodes the p110α catalytic subunit of phosphatidylinositol-3-kinase (PI3K), are two of the most common deleterious genomic changes in cancer, including in ovarian carcinomas. We investigated molecular mechanisms underlying interactions between these two mediators and their possible roles in ovarian tumorigenesis. We identified two alternate PIK3CA promoters and showed direct binding of and transcriptional inhibition by p53 to one of these promoters. Conditional suppression of functional p53 increased p110α transcripts, protein levels and PI3K activity in immortalized, non-tumorigenic ovarian surface epithelial (OSE) cells, the precursors of ovarian carcinoma. Conversely, overexpression of p53 by adenoviral infection and activation of p53 by γ-irradiation both diminished p110α protein levels in normal OSE and ovarian cancer cells. The demonstration that p53 binds directly to the PIK3CA promoter and inhibits its activity identifies a novel mechanism whereby these two mediators regulate cellular functions, and whereby inactivation of p53 and subsequent upregulation of PIK3CA might contribute to the pathophysiology of ovarian cancer.

The transcriptional induction of PIK3CA in tumor cells is dependent on the oncoprotein Y-box binding protein-1

“…YB-1 binds to the PIK3CA promoter at three validated sites The PIK3CA promoter region was characterized by us for the very first time (Astanehe et al, 2008), enabling the study of PIK3CA transcriptional regulation. The 5 0 UTR of PIK3CA is more than 50 000 bp upstream of the translational start site that lies within exon 2 (Figure 1a).…”

“…This 5 0 UTR consists of two alternate first exons (1a and 1b) that splice differentially with exon 2. Thereby, there are two alternate promoter regions 1a and 1b (Astanehe et al, 2008). Evaluation of the PIK3CA promoter for YB-1 binding identified six putative sites ( Figure 1a).…”

“…EMSAs were carried out using the Lightshift Chemiluminescent EMSA kit (Pierce Biotechnology) as previously described . Oligonucleotides used are listed in Supplementary Table S3. PIK3CA luciferase assay PIK3CA promoter 1a subcloned into a pGL3-basic vector (Promega, Madison, WI, USA) was previously described by us (Astanehe et al, 2008). For the overexpression studies, cells were co-transfected with the YB-1 expression vector and both of PIK3CA 1a-pGL3 and pRL-TK.…”

“…We recently characterized the 5 0 -untranslated region (5 0 UTR) of PIK3CA and published the first report on the transcriptional regulation of this important oncogene (Astanehe et al, 2008). We demonstrated that PIK3CA transcription is repressed by the tumor suppressor protein p53 in ovarian surface epithelial cells, and provided evidence to show that loss of p53 in ovarian cancer is one mechanism that leads to elevated p110a levels (Astanehe et al, 2008).…”

“…We recently characterized the 5 0 -untranslated region (5 0 UTR) of PIK3CA and published the first report on the transcriptional regulation of this important oncogene (Astanehe et al, 2008). We demonstrated that PIK3CA transcription is repressed by the tumor suppressor protein p53 in ovarian surface epithelial cells, and provided evidence to show that loss of p53 in ovarian cancer is one mechanism that leads to elevated p110a levels (Astanehe et al, 2008). In addition, it has recently been demonstrated that the forkhead transcription factor, FOXO3a, induces PIK3CA by directly binding to its promoter rendering chronic myelogenous leukemia cells resistant to doxorubicin (Hui et al, 2008).…”