Metabolic reprogramming in transformed mouse cortical astrocytes: A proteomic study Azeddine Bentaib a , Pascal De Tullio b , Hervé Chneiweiss c , 1 , Emmanuel Hermans d , Marie-Pierre Junier c , 1 , Pierre Leprince a, ⁎ a GIGA-Neuroscience, University of Liège, Liège, Belgium b Pharmaceutical chemistry, Department of Pharmacy, University of Liège, Liège, Belgium c Glial Plasticity and Cerebral Tumors, UMR8246 CNRS/U1130 Inserm/ UMCR18, Université Pierre et Marie Curie, Paris, France d Institute of Neurosciences, Group of Neuropharmacology, Université Catholique de Louvain, Brussels, Belgium ARTICLE INFO ABSTRACT Article history: Received 6 April 2014 Accepted 22 September 2014 Metabolic reprogramming is thought to play a key role in sustaining the survival and proliferation of cancer cells. These changes facilitate for example the uptake and release of nutrients required for nucleotide, protein and lipid synthesis necessary for macromolecule assembly and tumor growth. We applied a 2D-DIGE (two-dimensional differential in-gel electrophoresis) quantitative proteomic analysis to characterize the proteomes of mouse astrocytes that underwent in vitro cancerous transformation, and of their normal counterparts. Metabolic reprogramming effects on enzymatic and structural protein expression as well as associated metabolites abundance were quantified. Using enzymatic activity measurements and zymography, we documented and confirmed several changes in abundance and activity of various isoenzymes likely to participate in metabolic reprogramming. We found that after transformation, the cells increase their expression of glycolytic enzymes, thus augmenting their ability to use aerobic glycolysis (Warburg effect). An increased capacity to dispose of reducing equivalents through lactate production was also documented. Major effects on carbohydrates, amino acids and nucleotides metabolic enzymes were also observed. Conversely, the trans- formed cells reduced their enzymatic capacity for reactions of tricarboxylic acid oxidation, for neurotransmitter (glutamate) metabolism, for oxidative stress defense and their expression of astroglial markers. Biological significance The use of a global approach based on a 2D DIGE analysis allows obtaining a comprehensive view of the metabolic reprogramming undergone by astrocytes upon cancerous transfor- mation. Indeed, except for a few enzymes such as pyruvate carboxylase and glutaminase that were not detected in our initial analysis, pertinent information on the abundance of most enzymes belonging to pathways relevant to metabolic reprogramming was directly obtained. In this in vitro model, transformation causes major losses of astrocyte-specific proteins and functions and the acquisition of metabolic adaptations that favor intermedi- ate metabolites production for increased macromolecule biosynthesis. Thus our approach Keywords: 2D-DIGE Cancerous transformation Warburg effect Glycolysis Glia Glioblastoma JOURNAL OF PROTEOMICS 113 (2015) 292 – 314 ⁎ Corresponding author at: Giga-Neuroscience, University of Liège. CHU B36, 1 Av de l'hopital, B4000 Liège, Belgium. E-mail address: pleprince@ulg.ac.be (P. Leprince). 1 These two authors contributed equally to this work. http://dx.doi.org/10.1016/j.jprot.2014.09.019 1874-3919/© 2014 Elsevier B.V. All rights reserved. Available online at www.sciencedirect.com ScienceDirect www.elsevier.com/locate/jprot