Human activin type IIB receptor (ActRIIB) is a member of the TGF-β receptor superfamily. It mediates the actions of several ligands including activin A, myostatin, and GDF-11 to regulate cell growth, proliferation and differentiation. Dys-regulation of activin signal transduction such as reduced ActRIIB expression is associated with the development of high grade breast cancer in patients. This project aims to understand the mechanism by which ActRIIB regulates cell growth, proliferation and differentiation of breast cancer cells. First, we examined a number of human breast cancer cell lines and found that the level of expression of ActRIIB varies with different degree malignancy – its expression level is lower in the more malignant cancer cells. Since MCF-7, one of breast cancer cell line expresses a relatively higher level of ActRIIB, it is therefore chosen for our study to investigate the effects of knock-down of ActRIIB. Second, by using the siRNA approach, we observed that suppression of ActRIIB expression inhibits cell proliferation, and induces decreased telomerase activity and reduction in E-cadherin expression, suggesting that ActRIIB governs the proliferation, adhesion and migration of MCF-7 cells. The treatment of MCF-7 breast cancer cells with siRNA-ActRIIB at the concentration of 66nM resulted in massive lipid droplet accumulations. The activity of telomerase was constantly suppressed from as early as day 1 up to day 6 after the siRNA treatment, the inhibition of telomerase activity is consistent with that detected in the stable transfected cells with ActRIIB dominant negative expression. The cell cycle analysis by FACS further demonstrated that the knocking-down of ActRIIB receptor arrested cell cycle at G0/G1, accompanied by the inhibition of cell proliferation.
To further substantiate our findings, we obtained a stable ActRIIB gene knockdown cell line (named 76) by shRNA through lentivirus. In consistent to siRNA result, the cells demonstrated increased invasive ability as showed by matrigel invasive assay. The ActRIIB stable silencing cells present the elongated fibroblast- like morphology. Meanwhile, Immunofluorescent staining indicated loss of E-cadherin and gain of vimentin. It is most likely that the occurrence of EMT (epithelial-to-mesenchymal transition) in the 76 cells. Now we are using RNA sequencing to uncover the gene expression profiling in the 76 cells. Our study of the signalling mechanism of ActRIIB in breast cancer cells will shed light on the mechanism governing cancer cell proliferation and migration.