Abstract
BackgroundTGF-β acts as an antiproliferative factor in normal epithelial cells and at early stages of oncogenesis. However, later in tumor development TGF-β can become tumor promoting through mechanisms including the induction of epithelial-to-mesenchymal transition (EMT), a process that is thought to contribute to tumor progression, invasion and metastasis. To identify EMT-related breast cancer therapeutic targets and biomarkers, we have used two proteomic approaches to find proteins that change in abundance upon the induction of EMT by TGF-β in two mouse mammary epithelial cell lines, NMuMG and BRI-JM01.ResultsPreliminary experiments based on two-dimensional electrophoresis of a hydrophobic cell fraction identified only 5 differentially expressed proteins from BRI-JM01 cells. Since 3 of these proteins were glycoproteins, we next used the lectin, wheat germ agglutinin (WGA), to enrich for glycoproteins, followed by relative quantification of tryptic peptides using a label-free LC-MS based method. Using these approaches, we identified several proteins that are modulated during the EMT process, including cell adhesion molecules (several members of the Integrin family, Fibronectin, Activated leukocyte cell adhesion molecule, and Neural cell adhesion molecule 1) and regulators of cellular signaling (Tumor-associated calcium signal transducer 2, Basigin).ConclusionInterestingly, despite the fact that TGF-β induces similar EMT phenotypes in NMuMG and BRI-JM01 cells, the proteomic results for the two cell lines showed only minimal overlap. These differences likely result in part from the conservative cut-off values used to define differentially-expressed proteins in these experiments. Alternatively, it is possible that the two cell lines may use different mechanisms to achieve an EMT transition.
Highlights
TGF-β acts as an antiproliferative factor in normal epithelial cells and at early stages of oncogenesis
Phenotypic effects of TGF- treatment on BRI-JM01 and Namru murine mammary gland (NMuMG) cells Both the BRI-JM01 and NMuMG cells adopt a spindleshape fibroblast-like morphology after 24 hrs in the presence of TGF-β1, as shown by phase contrast microscopy (Figure 1)
The epithelial-to-mesenchymal transition (EMT) process is thought to be directly linked to the invasive potential of a cancer cell
Summary
TGF-β acts as an antiproliferative factor in normal epithelial cells and at early stages of oncogenesis. Later in tumor development TGF-β can become tumor promoting through mechanisms including the induction of epithelial-to-mesenchymal transition (EMT), a process that is thought to contribute to tumor progression, invasion and metastasis. NMuMG is a nontransformed cell line that is a well-established EMT model system, whereas the BRI-JM01 cell line was recently presented as an alternative for studying TGF-β-induced EMT [7,8]. In both cell lines, treatment with TGF-β induces similar changes in phenotype, including the loss or relocalization of epithelial markers (e.g. ZO-1 and E-cadherin), the rearrangement of actin and vimentin filaments, and an increase in motility
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