Abstract
Background: Tropomyosin 1 alpha chain (Tm1) is an actin-binding protein that regulates the endothelial cell response to oxidative stress following its phosphorylation at Serine 283 (S283). Tm1 is also a major tumor suppressor in breast cancer. In the present study, we investigated the role of phosphorylation of Tm1 in regulating its tumor suppressor properties. Methods: MDA MB231 breast cancer cells stably overexpressing wild type form of Tm1 or Tm1 mutants (S283A and S283E) were generated. Proliferation and cell viability were assayed by means of the enzymatic cleavage of the tetrazolium salt WST-1 to formazan dye by cellular mitochondrial dehydrogenases. Adhesion assays were performed at various periods of time on cells grown on plastic. Cell migration was evaluated by using the wound-healing assay and by measuring transendothelial migration of cancer cells. Malignant transformation in vitro was determined by using the anchorage-independent growth assay on soft agar. Results: We found that cells expressing the phosphomimetic form of Tm1 S283E/Tm1 are characterized by an increased adhesion to the substratum. Moreover, the migration of MDA-MB231/S283E/Tm1 cells in a wound closure assay is reduced compared to parental cells or those expressing the non-phosphorylatable form of Tm1 (S283A). Similarly, the transendothelial migration of MDA-MB231/S283E/Tm1 cells is also reduced as compared to the other cell lines. Moreover, we found that the cells expressing the S283A mutants form more colonies in soft agar that those expressing the S283E mutants. Conclusion: Phosphorylation of Tm1 at Ser283 contributes to its anti-tumor properties, and this effect results mainly from an increase in cell adhesion associated with a decrease in their migratory and invasive potentials.
Highlights
Breast cancer is the most common malignancy among women in Europe and North America
We reported previously that Tropomyosin 1 alpha chain (Tm1) is phosphorylated at Serine 283 (S283) in endothelial cells exposed to oxidative stress, which contributes to maintain the integrity of the endothelium [11] [12] [22]
We found that the migration potential of MDA MB231/S283E/Tm1 cells was reduced as compared to parental cells that do not express Tm1 or to the MDA 231/S283A/Tm1 cells
Summary
Breast cancer is the most common malignancy among women in Europe and North America. Despite its high incidence, early detection and modern treatments have significantly increased patient survival. Genome analyses indicate that there are only a few genes that are commonly mutated in breast cancer. These genes include the oncogens ErbB2, PI3KCA, MYC, and CCND1 as well as the tumor suppressors BRCA1/2 and p53 [2]. Results: We found that cells expressing the phosphomimetic form of Tm1 S283E/Tm1 are characterized by an increased adhesion to the substratum. The migration of MDA-MB231/S283E/Tm1 cells in a wound closure assay is reduced compared to parental cells or those expressing the non-phosphorylatable form of Tm1 (S283A). Conclusion: Phosphorylation of Tm1 at Ser283 contributes to its anti-tumor properties, and this effect results mainly from an increase in cell adhesion associated with a decrease in their migratory and invasive potentials
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