Differential proteomic analysis of a highly metastatic variant of human breast cancer cells using two-dimensional differential gel electrophoresis

Distant metastasis represents the major lethal cause of breast cancer. To understand the molecular mechanisms of breast cancer metastasis and identify markers with metastatic potential, we established a highly metastatic variant of parental MDA-MB-231 cells (MDA-MB-231HM). Using two-dimensional electrophoresis (2-DE), we performed a proteomic comparison of the two kinds of cells. As much as 51 protein spots were differentially expressed between the selected variant and its parental counterpart in at least 3 experiments. Ten unique proteins were identified using matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS), liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS), and database searching software. Among them, nine proteins were up-regulated in MDA-MB-231HM cells, including Macrophage-capping protein (CapG), Galectin-1, Chloride intracellular channel protein 1, Endoplasmic reticulum protein ERp29 precursor, Stathmin-1 (STMN1), Isoform 1 of uridine–cytidine kinase 2(UCK2), Rho GDP-dissociation inhibitor 2 (ARHGDIB), isocitrate dehydrogenase [NADP] cytoplasmic (IDH1), and N-myc downstream regulated gene 1 (NDRG1) protein. Only transgelin-2 was down-regulated. Differential expression was confirmed for three proteins including CapG, STMN1, and transgelin-2 by Western blotting analysis. Transgelin-2 was chosen for further verification by immunohistochemistry. The results suggested that 2-DE would be an efficient way to screen the proteins responsible for specific biological function. Furthermore, the findings imply that different proteins may be involved in the metastatic process in breast carcinomas.