A LONG-TERM 10G-HYPERGRAVITY EXPOSURE PROMOTES CELL-CELL CONTACTS AND REDUCES ADHESIVENESS TO A SUBSTRATE, MIGRATION, AND INVASIVENESS OF MCF-7HUMAN BREAST CANCER CELLS.

BACKGROUND: G-force is a fundamental force controlling human cells. Cancer is one of the 4 major health challenges in the Space missions. Cancer in Space project evaluates the reaction of human cancer cells to the conditions of the space flights, including an exposure to high g-forces. AIM: Explore an impact of 10 g force on the oncogenic properties of human breast adenocarcinoma cells MCF-7. MATERIALS AND METHODS: Cells were exposed to 10 g force for 10 days, as part of a 6-week simulation of conditions of a space flight. Then the cells were cultured for one week under normal culture conditions, before performing tests. Cell proliferation, cell viability, cell-cell contact inhibition, migration, and invasiveness were measured. Immunoblotting was used to evaluate expression of proteins. RESULTS: Proliferation, cell-cell interaction and formation of 3D structures, migration, and invasiveness of cells exposed to 10 g were compared to parental cells cultured at 1 g condition. 10 g exposed cells showed a higher propensity for cell-cell contact inhibitions and lower for 3-dimensional growth in dense culture. This correlated with the decrease of proliferation in a dense culture as compared to the parental cells. The decrease of migration, adherence to a surface, and invasiveness was observed for cells subjected to the hypergravity, as compared to the parental MCF-7 cells. Enhanced expression of E-cadherin and phosphorylated pY576-FAK were observed in 10 g exposed cells but no impact on the expression of Erk, pErk, FAK and p53 was detected. CONCLUSION: The prolonged exposure of MCF-7 cells to 10 g force targets cell-cell and cell-substrate interactions.

Proteomics of transforming growth factor ß1 (TGF ß1) signaling in 184A1 human breast epithelial cells suggests the involvement of casein kinase 2α in TGF ß1-dependent p53 phosphorylation at Ser392.

AIM: Transforming growth factor ß1 (TGF ß1) is a potent regulator of breast tumorigenesis. It inhibits proliferation of carcinoma cells, but the strength of its inhibitory action varies for cells from benigh, non-metastatic or metastatic tumors. The aim of this work was to generate a proteome profile of TGF ß1 action on non-tumorigenic human breast epithelial cells 184A1, and validate predicted involvement of casein kinase 2α (CK2α), p53 and structure-specific recognition protein-1 (SSRP1). MATERIALS AND METHODS: Two-dimensional gel electrophoresis and mass spectrometry were used to identify TGF ß1-regulated proteins in 184A1 human breast immortalized non-tumorigenic cells. 184A1 cells may serve as a model of benign breast neoplasia. These cells were obtained from normal mammary tissue, were immortalized but are not malignant, and were obtained from the American Type Culture Collection. The systemic analysis was performed by using the Cytoscape tool. Transfection of cells with CK2α construct and small interfering RNAs to CK2α and SSRP1 were used to assess an impact of CK2α and SSRP1 on phosphorylation of the p53 and cell proliferation. RESULTS: Proliferation of 184A1 cells was transiently inhibited by TGF ß1. We identified 100 and 47 unique proteins which changed their expression and/or 35S-incorporation, respectively, upon treatment with TGF ß1 for 2 h, 8 h or 24 h. Cell proliferation, death, migration, and metabolism were among the biological regulatory processes retrieved by the network analysis as affected by the identified proteins. The network analysis suggested that TGF ß1 may affect the phosphorylation of p53 at Ser392 by engaging CK2α. This was confirmed by the immunoblotting and cell proliferation assays. CONCLUSION: We report here the list of 147 TGF ß1-regulated proteins in immortalized non-tumorigenic human breast epithelial cells, and show involvement of CK2α in the regulation of p53 Ser392 phosphorylation.