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Proteome analysis reveals novel proteins associated with proliferation and differentiation of the colorectal cancer cell line Caco-2

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Author: Stierum, R. · Gaspari, M. · Dommels, Y. · Ouatas, T. · Pluk, H. · Jespersen, S. · Vogels, J. · Verhoeckx, K. · Groten, J. · Ommen, B. van
Type:article
Date:2003
Source:Biochimica et Biophysica Acta - Proteins and Proteomics, 1-2, 1650, 73-91
Identifier: 237230
doi: doi:10.1016/S1570-9639(03)00204-8
Keywords: Health Nutrition · Analytical research · Caco-2 · Differentiation · Mass spectrometry · Proteome analysis · Two-dimensional gel electrophoresis · cofilin · creatine kinase · disulfide · enolase · fatty acid binding protein · glutathione transferase · heat shock protein · isomerase · liver protein · mitochondrial protein · nucleoside diphosphate kinase · nucleotide · proteome · tumor protein · alkaline phosphatase · article · brush border · carcinogenesis · cell culture · cell differentiation · cell growth · cell maturation · cell proliferation · cell strain CACO 2 · colon carcinogenesis · colorectal cancer · controlled study · cytoskeleton · genetic transcription · glycolysis · human · human cell · intestine cell · matrix assisted laser desorption ionization time of flight mass spectrometry · microvillus · nucleotide metabolism · pH · phenotype · priority journal · protein analysis · protein expression · protein folding · protein function · proteomics · signal transduction · two dimensional gel electrophoresis · cell division · cytology · metabolism · physiology · principal component analysis · Western blotting · Alkaline Phosphatase · Blotting, Western · Caco-2 Cells · Cell Differentiation · Cell Division · Humans · Principal Component Analysis · Proteome

Abstract

Here, we describe a proteomics approach to study protein expression changes in differentiating Caco-2 cells. Caco-2 is a colorectal carcinoma cell line, which upon differentiation loses its tumorigenic phenotype and displays characteristics of mature enterocytes, including brush borders with microvilli. Cells were grown in culture flasks and harvested at different stages of differentiation (days post-confluence: -3, 0, 3, 7, 10, 14, and 18). Two-dimensional gel electrophoresis was used to analyse proteome changes. Approximately 1400 protein spots were detected within the Caco-2 proteome, within the pH 4-7 range. Two-dimensional gel electrophoresis allowed for the detection of 18 proteins from which the levels of expression were found to be associated with differentiation. Of these proteins, 11 were identified by means of MALDI-TOF or NANO-ESI-MS/MS mass spectrometry and include liver fatty acid binding protein (FABL), three forms of α-enolase (ENOA), nucleoside diphosphate kinase A (NDKA), cofilin-1 (COF1), translationally controlled tumour protein (TCTP), mitochondrial 60-kDa heat shock protein (CH60), probable protein disulfide isomerase (ER60), creatine kinase B (KCRB), and glutathione S-transferase α (GTA1). Thus, proteomics revealed that the differentiation-related change in phenotype of Caco-2 involves changes in a variety of distinct biochemical pathways. Some of these proteins have not been shown before to be associated with Caco-2 differentiation (ER60; COF1; CH60; NDKA; TCTP and ENOA). Therefore, processes related to protein folding and disulfide bridge formation, cytoskeleton formation and maintenance, nucleotide metabolism, glycolysis as well as tumorigenesis-associated proteins may be involved in Caco-2 differentiation. Changes in the expression of CH60, TCTP, GTA1, NDKA, and FABL have also been reported to be associated with in vivo colon carcinogenesis. These findings illustrate that a combination of proteomics and cell culture is a useful approach to find markers for Caco-2 differentiation, which could contribute to the comprehension of the process of colon carcinogenesis. © 2003 Elsevier B.V. All rights reserved.