SUMMARYThe embryonic pyruvate kinase M2 (PKM2) isoform is highly expressed in human cancer. In contrast to the established role of PKM2 in aerobic glycolysis or the Warburg effect 1,2,3 , its nonmetabolic functions remain elusive. Here we demonstrate that EGFR activation induces translocation of PKM2, but not PKM1, into the nucleus, where K433 of PKM2 binds to c-Srcphosphorylated Y333 of β-catenin. This interaction is required for both proteins to be recruited to the CCND1 promoter, leading to HDAC3 removal from the promoter, histone H3 acetylation, and cyclin D1 expression. PKM2-dependent β-catenin transactivation is instrumental in EGFRpromoted tumor cell proliferation and brain tumor development. In addition, positive correlations have been identified among c-Src activity, β-catenin Y333 phosphorylation, and PKM2 nuclear accumulation in human glioblastoma specimens. Furthermore, levels of β-catenin phosphorylation and nuclear PKM2 have been correlated with grades of glioma malignancy and prognosis. These findings reveal that EGF induces β-catenin transactivation via a mechanism distinct from that induced by Wnt/wingless 4 and highlight the essential nonmetabolic functions of PKM2 in EGFRpromoted β-catenin transactivation, cell proliferation, and tumorigenesis.Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:http://www.nature.com/authors/editorial_policies/license.html#terms Correspondence: zhiminlu@mdanderson.org. Contributions This study was conceived by Z.L. Z.L and W.Y. designed the study; W.Y., Y.X., H.J., Y.Z., and J.L. performed experiments; K.A. provided pathology assistance; W.H. and X.G. provided reagents and conceptual advice; Z.L. wrote the paper with comments from all authors. Since both EGFR activation and PKM2 expression are instrumental in tumorigenesis 5,6,7 , we examined whether EGFR activation regulates PKM2 functions in a subcellular compartment-dependent manner. Immunofluorescence analysis showed that EGF treatment resulted in the nuclear accumulation of PKM2 in U87/EGFR human glioblastoma (GBM) cells (Fig. 1a). In addition, expression of the constitutively active EGFRvIII mutant in U87 cells had a higher amount of nuclear PKM2 than did EGF-untreated U87/EGFR cells ( Supplementary Fig. 2a). The finding that EGF induces nuclear translocation of PKM2 was further supported by cell fractionation analysis of DU145 prostate cancer cells, MDA-MB-231 breast cancer cells, and U87/EGFR cells ( Supplementary Fig. 2b). In addition, PKM1 failed to translocate into the nucleus upon EGF stimulation ( Supplementary Fig. 2c), indicating that EGF specifically regulates the subcellular distribution of PKM2 in multiple types of cancer cells. HHS Public AccessTo examine whether PKM2 directly regulates gene transcription and cell proliferation, we expressed PKM2 shRNA in U87/EGFR cells ( Supplementary Fig. 3a). PKM2 depletion largely reduced both basal and EGF-induced tumor cell prolife...

SUMMARY Pyruvate kinase M2 (PKM2) is upregulated in multiple cancer types and contributes to the Warburg effect by unclarified mechanisms. Here we demonstrate that EGFR-activated ERK2 binds directly to PKM2 I429/L431 via the ERK2 docking groove and phosphorylates PKM2 Ser37 but not PKM1. Phosphorylated PKM2 Ser37 recruits PIN1 for cis-trans isomerization of PKM2, which leads to PKM2 binding to importin α5 and nuclear translocation. Nuclear PKM2, acting as a coactivator of β-catenin, induces c-Myc expression, resulting in the upregulation of GLUT1, LDHA, and, in a positive feedback loop, PTB-dependent PKM2 expression. Replacement of wild type PKM2 with a nuclear translocation-deficient mutant (S37A) blocks the EGFR-promoted Warburg effect and brain tumor development. In addition, levels of PKM2 S37 phosphorylation correlate with EGFR and ERK1/2 activity in human glioblastoma specimens. Our findings highlight the importance of nuclear functions of PKM2 in the Warburg effect and tumorigenesis.

Increased transcriptional activity of ␤-catenin resulting from Wnt/Wingless-dependent or -independent signaling has been detected in many types of human cancer, but the underlying mechanism of Wnt-independent regulation is poorly understood. We have demonstrated that AKT, which is activated downstream from epidermal growth factor receptor signaling, phosphorylates ␤-catenin at Ser 552 in vitro and in vivo. AKTmediated phosphorylation of ␤-catenin causes its disassociation from cell-cell contacts and accumulation in both the cytosol and the nucleus and enhances its interaction with 14-3-3 via a binding motif containing Ser 552 . Phosphorylation of ␤-catenin by AKT increases its transcriptional activity and promotes tumor cell invasion, indicating that AKT-dependent regulation of ␤-catenin plays a critical role in tumor invasion and development.