Abstract 2881: Posttranslational inactivation of PTEN promoted by loss of Notch1 signaling in T-ALL
Authors: Hales Eric C. , Gedman Amanda Larson , Ge Yubin , Taub Jeffrey W. , Matherly Larry H. 
Published: June 2012

Abstract
T-cell acute lymphoblastic leukemias (T-ALL) are frequently associated with mutations in Notch 1, many of which lead to increased susceptibilities to cleavage by gamma secretase and generation of ICN1, promoting enhanced Notch1 signaling. Increased Notch1 activity in T-ALL was reported to indirectly repress transcription of PTEN, the major tumor suppressor involved in antagonizing PIP3-dependent cell signaling and in maintaining tight control over the pro-survival AKT pathway. While use of gamma secretase inhibitors (GSIs) prevents Notch1-activating cleavage and restores PTEN protein levels, a link between GSI treatments and PTEN posttranslational inactivation was also implied (Haematologica 95: 674-678, 2010). Phosphorylation by casein kinase II (CK2) of the S380/T382/3 cluster in the carboxyl-terminal region of PTEN is an efficient means of regulating PTEN activity, and of maintaining tight control over cell viability. PTEN phosphorylation impacts its cytosolic versus membrane localization, and PTEN protein stability and levels. However, the relative impact of Notch1 and GSIs on PTEN levels resulting from effects on PTEN transcription versus potential PTEN posttranslational effects has not been established.
To begin to address this important question, we used conditional ectopic PTEN expression under control of a tetracycline-inducible promoter in PTEN-null Jurkat cells (designated Jkt/tet-onPten) to assess whether Notch1 can regulate PTEN posttranslationally. Cells were treated with doxycycline to induce PTEN, then with compound E (CompE), a potent GSI. CompE treatment increased phosphorylation of PTEN and AKT (S473 and T308), leading to elevated phosphorylation of GSK3αβ and S6RP downstream, consistent with inactivation of PTEN. This was accompanied by a moderate increase in cell proliferation. Analogous results were seen in Jkt/tet-onPten cells in which Notch1 was knocked down with a shRNA lentiviral vector (Jkt/tet-onPtenKD cells). Knockdown of Notch1 was accompanied by increased cytosolic localization of PTEN, consistent with its phosphorylation and inactivation. Treatment of Jkt/tet-onPtenKD cells with a CK2 inhibitor (tetrabromobenzimidazole) reduced PTEN phosphorylation and restored AKT activation, establishing that the effects of Notch1 on PTEN phosphorylation were via CK2. Our results suggest a mechanism whereby reduced Notch1 signaling in T-ALL cells increases PTEN posttranslational inactivation and AKT signaling, through an effect on CK2-mediated phosphorylation of PTEN. Although GSI treatment should transcriptionally increase PTEN levels, our data suggest that GSI treatment could potentially result in a more aggressive T-ALL due to increased PTEN inactivation and increased AKT signaling. Thus, combined use of a GSI with a downstream inhibitor to CK2, AKT or mTOR could provide significant benefit in treating T-ALL.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2881. doi:10.1158/1538-7445.AM2011-2881