Calcium/calmodulin-dependent protein kinase kinase 2 mediates pleiotropic effects of epidermal growth factor in cancer cells.

Engagement of epidermal growth factor (EGF) with its receptor (EGFR) produces a broad range of cancer phenotypes. The overriding aim of this study was to understand EGFR signaling and its regulation by the Ca2+/calmodulin (CaM) dependent protein kinase kinase 2 (CaMKK2) in cancer cells. In ovarian cancer cells and other cancer cell types, EGF-induced activation of oncogenic Akt is mediated by both the canonical PI3K-PDK1 pathway and by CaMKK2. Akt activation induced by EGF occurs by both calcium-dependent and calcium-independent mechanisms. In contrast to the canonical pathway, CaMKK2 neither binds to, nor is regulated by phosphoinositides but is activated by Ca2+/CaM. Akt activation at its primary activation site, T308 occurs by direct phosphorylation by CaMKK2, but activation at its secondary site (S473), is through an indirect mechanism requiring mTORC2. In cells in which another CaMKK2 target, 5'AMP-dependent protein kinase (AMPK) was deleted, Akt activation and calcium-dependency of activation were still observed. CaMKK2 accumulates in the nucleus in response to EGF and regulates transcription of phosphofructokinase platelet (PFKP) a glycolytic regulator. CaMKK2 is required for optimal PFK activity. CaMKK2 regulates transcription of plasminogen activator, urokinase (PLAU) a metastasis regulator. The EGFR inhibitor gefitinib synergizes with CaMKK2 inhibition in the regulation of cell survival and increases the dose-reduction index. CRISPR/Cas9 knockout of CaMKK2 leads to compensatory PTEN downregulation and upregulation of Akt activation. CaMKK2-mediation of EGFR action may enable cancer cells to use intracellular calcium elevation as a signal for growth and survival.