Abstract
Many types of human tumour cells overexpress the dual-specificity phosphatase Cdc25A. Cdc25A dephosphorylates cyclin-dependent kinase and regulates the cell cycle, but other substrates of Cdc25A and their relevant cellular functions have yet to be identified. We demonstrate here that EGFR activation results in c-Src-mediated Cdc25A phosphorylation at Y59, which interacts with nuclear pyruvate kinase M2 (PKM2). Cdc25A dephosphorylates PKM2 at S37, and promotes PKM2-dependent β-catenin transactivation and c-Myc-upregulated expression of the glycolytic genes GLUT1, PKM2 and LDHA, and of CDC25A; thus, Cdc25A upregulates itself in a positive feedback loop. Cdc25A-mediated PKM2 dephosphorylation promotes the Warburg effect, cell proliferation and brain tumorigenesis. In addition, we identify positive correlations among Cdc25A Y59 phosphorylation, Cdc25A and PKM2 in human glioblastoma specimens. Furthermore, levels of Cdc25A Y59 phosphorylation correlate with grades of glioma malignancy and prognosis. These findings reveal an instrumental function of Cdc25A in controlling cell metabolism, which is essential for EGFR-promoted tumorigenesis.
Highlights
Many types of human tumour cells overexpress the dual-specificity phosphatase Cdc25A
Alternate splicing of PKM pre-mRNA by heterogeneous nuclear ribonucleoproteins A1/2 and polypyrimidine-tract binding protein splicing factors leads to pyruvate kinase M2 (PKM2) generation by the inclusion of exon 10 and the exclusion of exon 9, which is specific for PKM1
We found that nuclear PKM2 binds to c-Src phosphorylated Cdc25A at Y59, leading to Cdc25A-dependent PKM2 dephosphorylation, which is instrumental for PKM2 to interact with and activate b-catenin. b-catenin-mediated c-Myc expression subsequently upregulates expression of Cdc25A and glycolytic genes, which promotes the Warburg effect and cell proliferation
Summary
Nuclear PKM2 pS37 is dephosphorylated by Cdc25A. Epidermal growth factor receptor (EGFR) activation induces ERK-mediated PKM2 S37 phosphorylation in the cytosol, which results in nuclear translocation of about 10% cytosolic PKM2 (ref. 28). Overexpression of Flag-tagged wild-type (WT) Cdc25A, but not that of a catalytically inactive Cdc25A C431S mutant, dephosphorylated PKM2 at pS37 upon EGF treatment for 3 h in U87/EGFR cells (Fig. 1d) and U251 cells (Supplementary Fig. 1b). To identify the signal pathway or the protein kinase that regulates the interaction between Cdc25A and PKM2, we pretreated U87/EGFR cells with the following inhibitors: the phosphoinositide 3-kinase inhibitor LY294002, c-Jun N-terminal kinases (JNK) inhibitor SP600125, a NF-kB inhibitor, a JAK2 inhibitor and Src inhibitor SU6656, which blocked EGF-induced phosphorylation of AKT and c-Jun, TNF-ainduced and NF-kB-dependent IkBa promoter activation, and EGF-induced phosphorylation of Stat[3] and c-Src, respectively (Supplementary Fig. 2b–f). Analysis of the Cdc25A amino acid sequence using PPSP (http://ppsp.biocuckoo.org) and DISPHOS a EGF 0 1 3 6 (h)
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