Proteomic characterization of peroxisome proliferator-activated receptor-γ (PPARγ) overexpressing or silenced colorectal cancer cells unveils a novel protein network associated with an aggressive phenotype

Peroxisome proliferator-activated receptor-gamma (PPAR gamma) is a transcription factor of the nuclear hormone receptor superfamily implicated in a wide range of processes, including tumorigenesis. Its role in colorectal cancer (CRC) is still debated; most reports support that PPAR gamma reduced expression is associated with poor prognosis. We employed 2 -Dimensional Differential InGel Electrophoresis (2-D DIGE) followed by Liquid Chromatography (LC)-tandem Mass Spectrometry (MS/MS) to identify differentially expressed proteins and the molecular pathways underlying PPARy expression in CRC progression. We identified several differentially expressed proteins in HT29 and HCT116 CRC cells and derived clones either silenced or overexpressing PPAR gamma, respectively. In Ingenuity Pathway Analysis (IPA) they showed reciprocal relation with PPAR gamma and a strong relationship with networks linked to cell death, growth and survival. Interestingly, five of the identified proteins, ezrin (EZR), isoform C of prelamin-A/C (LMNA), alpha-enolase (ENOA), prohibitin (PHB) and RuvB-like 2 (RUVBL2) were shared by the two cell models with opposite expression levels, suggesting a possible regulation by PPAR gamma. mRNA and western blot analysis were undertaken to obtain a technical validation and confirm the expression trend observed by 2-D DIGE data. We associated EZR upregulation with increased cell surface localization in PPAR gamma-overexpressing cells by flow cytometry and immunofluorescence staining. We also correlated EZR and PPAR gamma expression in our series of CRC specimens and the expression profiling of all five proteins levels in the publicly available colon cancer genomic data from Oncomine and Cancer Genome Atlas (TCGA) colon adenocarcinoma (COAD) datasets. In summary, we identified a panel of proteins correlated with PPAR gamma expression that could be associated with CRC unveiling new pathways to be investigated for the selection of novel potential prognostic/predictive biomarkers and/or therapeutic targets. (C) 2016 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.