Overexpression of PP2A inhibitory subunits promotes expression and activation of oncogenic signatures

Abstract

The phosphatase enzyme PP2A regulates various signalling pathways, including the mTOR pathway, and is responsible for control of cell growth, proliferation, and apoptosis. In this investigation, the effect of overexpressing the PP2A inhibitory subunits CIP2A, SET and IGBP1 was studied in selected breast cancer cell lines, MCF-7 and MDA-MB-453. Polysome bound RNA studies revealed that recruitment of c-MYC transcript to the polysomes increases significantly upon overexpression of CIP2A, SET and IGBP1 in both MCF-7 and MDA-MB-453. DigiWest technology was then used to measure total protein and phospho-proteins using lysates from MDA-MB-453 cells overexpressing the PP2A inhibitory subunits and compared to the originator cells. A list of proteins that are upregulated upon overexpression (protein signature) as well as a phospho-profile was generated. The protein signature showed upregulation of beta-catenin, MCL1, c-MYC and RICTOR. The shift towards expression of ERK2 upon overexpression of CIP2A, SET and IGBP1 offers an opportunity to further investigate the role of the PP2A inhibitory subunits in Epithelial and Mesenchymal Transition (EMT)-induction. The phospho-profile showed an upregulation of p4E-BP1(phosphoThr37/Thr46) suggesting the release and activation of eIF4E; upregulation of PKC alpha (phosphoThr497/Thr638/Thr641); and dephosphorylation of p70 S6 kinase (phosphoThr389). Correlation of the protein signature with the expression of the PP2A inhibitory subunits was performed using the publicly available patient dataset, namely the Breast Invasive Carcinoma (TCGA), generating an oncogenic signature, also including c-MYC and beta catenin. The results show that the expression of beta-catenin protein is regulated via various components of the PP2A complex and confirm AURKA as the surrogate marker of low PP2A activity.