Limits...
EBP50 inhibits EGF-induced breast cancer cell proliferation by blocking EGFR phosphorylation.

Yao W, Feng D, Bian W, Yang L, Li Y, Yang Z, Xiong Y, Zheng J, Zhai R, He J - Amino Acids (2012)

Bottom Line: Knockdown of EBP50 in EBP50-overexpressed MDA-MB-231 cells abrogated the inhibitory effect of EBP50 on EGF-stimulated ERK1/2 phosphorylation and restoration of EBP50 expression in EBP50-knockdown MCF-7 cells rescued the inhibition of EBP50 on EGF-stimulated ERK1/2 phosphorylation, further confirming that the activation of EGF-induced downstream molecules could be specifically inhibited by EBP50 expression.Meanwhile, total expression levels of EGFR were unaffected during EGF stimulation.These results provide further insight into the molecular mechanism by which EBP50 regulates the development and progression of breast cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Capital Medical University, Beijing 100069, People's Republic of China.

ABSTRACT
Ezrin-radixin-moesin-binding phosphoprotein-50 (EBP50) suppresses breast cancer cell proliferation, potentially through its regulatory effect on epidermal growth factor receptor (EGFR) signaling, although the mechanism by which this occurs remains unknown. Thus in our studies, we aimed to determine the effect of EBP50 expression on EGF-induced cell proliferation and activation of EGFR signaling in the breast cancer cell lines, MDA-MB-231 and MCF-7. In MDA-MB-231 cells, which express low levels of EBP50, EBP50 overexpression inhibited EGF-induced cell proliferation, ERK1/2 and AKT phosphorylation. In MCF-7 cells, which express high levels of EBP50, EBP50 knockdown promoted EGF-induced cell proliferation, ERK1/2 and AKT phosphorylation. Knockdown of EBP50 in EBP50-overexpressed MDA-MB-231 cells abrogated the inhibitory effect of EBP50 on EGF-stimulated ERK1/2 phosphorylation and restoration of EBP50 expression in EBP50-knockdown MCF-7 cells rescued the inhibition of EBP50 on EGF-stimulated ERK1/2 phosphorylation, further confirming that the activation of EGF-induced downstream molecules could be specifically inhibited by EBP50 expression. Since EGFR signaling was triggered by EGF ligands via EGFR phosphorylation, we further detected the phosphorylation status of EGFR in the presence or absence of EBP50 expression. Overexpression of EBP50 in MDA-MB-231 cells inhibited EGF-stimulated EGFR phosphorylation, whereas knockdown of EBP50 in MCF-7 cells enhanced EGF-stimulated EGFR phosphorylation. Meanwhile, total expression levels of EGFR were unaffected during EGF stimulation. Taken together, our data shows that EBP50 can suppress EGF-induced proliferation of breast cancer cells by inhibiting EGFR phosphorylation and blocking EGFR downstream signaling in breast cancer cells. These results provide further insight into the molecular mechanism by which EBP50 regulates the development and progression of breast cancer.

Show MeSH

Related in: MedlinePlus

Establishment of breast cancer cells in which EBP50 expression was stably overexpressed or knocked down. a EBP50 was stably overexpressed in MDA-MB-231 breast cancer cells. HA-231 cells stably transfected with pBK-CMV-HA vector presented similar levels of EBP50 as that in its parental cells, and EBP50 was robustly expressed in EBP-231 cells stably transfected with pBK-CMV-HA-EBP50 constructs. b The expression of EBP50 was stably knocked down in MCF-7 cells. Luc-RNAi/MCF-7 cells stably transfected with Luciferase shRNA control plasmid presented the same level of EBP50 as that in parental cells, and the expression level of EBP50 in EBP-RNAi/MCF-7 cells was about 33 % as that in parental cells
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3472071&req=5

Fig1: Establishment of breast cancer cells in which EBP50 expression was stably overexpressed or knocked down. a EBP50 was stably overexpressed in MDA-MB-231 breast cancer cells. HA-231 cells stably transfected with pBK-CMV-HA vector presented similar levels of EBP50 as that in its parental cells, and EBP50 was robustly expressed in EBP-231 cells stably transfected with pBK-CMV-HA-EBP50 constructs. b The expression of EBP50 was stably knocked down in MCF-7 cells. Luc-RNAi/MCF-7 cells stably transfected with Luciferase shRNA control plasmid presented the same level of EBP50 as that in parental cells, and the expression level of EBP50 in EBP-RNAi/MCF-7 cells was about 33 % as that in parental cells

Mentions: The EBP50 stable transfection pool of cells, namely MDA-MB-231-HA-EBP50 (EBP-231) or its control MDA-MB-231-HA (HA-231), were generated by transfection with the neo-pBK-CMV-HA-EBP50 or neo-pBK-CMV-HA vector, respectively. Protein expression in these stable cells was verified by western blot analysis as shown in Fig. 1a. In HA-231 cells, transfection of the control vector had no effect on EBP50 expression, and similar levels of EBP50 were expressed in the parental cells. HA-tagged EBP50 protein expression was not detected in control cells (data not shown). In EBP-231 cells, exogenous HA-tagged EBP50 was robustly overexpressed.Fig. 1


EBP50 inhibits EGF-induced breast cancer cell proliferation by blocking EGFR phosphorylation.

Yao W, Feng D, Bian W, Yang L, Li Y, Yang Z, Xiong Y, Zheng J, Zhai R, He J - Amino Acids (2012)

Establishment of breast cancer cells in which EBP50 expression was stably overexpressed or knocked down. a EBP50 was stably overexpressed in MDA-MB-231 breast cancer cells. HA-231 cells stably transfected with pBK-CMV-HA vector presented similar levels of EBP50 as that in its parental cells, and EBP50 was robustly expressed in EBP-231 cells stably transfected with pBK-CMV-HA-EBP50 constructs. b The expression of EBP50 was stably knocked down in MCF-7 cells. Luc-RNAi/MCF-7 cells stably transfected with Luciferase shRNA control plasmid presented the same level of EBP50 as that in parental cells, and the expression level of EBP50 in EBP-RNAi/MCF-7 cells was about 33 % as that in parental cells
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3472071&req=5

Fig1: Establishment of breast cancer cells in which EBP50 expression was stably overexpressed or knocked down. a EBP50 was stably overexpressed in MDA-MB-231 breast cancer cells. HA-231 cells stably transfected with pBK-CMV-HA vector presented similar levels of EBP50 as that in its parental cells, and EBP50 was robustly expressed in EBP-231 cells stably transfected with pBK-CMV-HA-EBP50 constructs. b The expression of EBP50 was stably knocked down in MCF-7 cells. Luc-RNAi/MCF-7 cells stably transfected with Luciferase shRNA control plasmid presented the same level of EBP50 as that in parental cells, and the expression level of EBP50 in EBP-RNAi/MCF-7 cells was about 33 % as that in parental cells
Mentions: The EBP50 stable transfection pool of cells, namely MDA-MB-231-HA-EBP50 (EBP-231) or its control MDA-MB-231-HA (HA-231), were generated by transfection with the neo-pBK-CMV-HA-EBP50 or neo-pBK-CMV-HA vector, respectively. Protein expression in these stable cells was verified by western blot analysis as shown in Fig. 1a. In HA-231 cells, transfection of the control vector had no effect on EBP50 expression, and similar levels of EBP50 were expressed in the parental cells. HA-tagged EBP50 protein expression was not detected in control cells (data not shown). In EBP-231 cells, exogenous HA-tagged EBP50 was robustly overexpressed.Fig. 1

Bottom Line: Knockdown of EBP50 in EBP50-overexpressed MDA-MB-231 cells abrogated the inhibitory effect of EBP50 on EGF-stimulated ERK1/2 phosphorylation and restoration of EBP50 expression in EBP50-knockdown MCF-7 cells rescued the inhibition of EBP50 on EGF-stimulated ERK1/2 phosphorylation, further confirming that the activation of EGF-induced downstream molecules could be specifically inhibited by EBP50 expression.Meanwhile, total expression levels of EGFR were unaffected during EGF stimulation.These results provide further insight into the molecular mechanism by which EBP50 regulates the development and progression of breast cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Capital Medical University, Beijing 100069, People's Republic of China.

ABSTRACT
Ezrin-radixin-moesin-binding phosphoprotein-50 (EBP50) suppresses breast cancer cell proliferation, potentially through its regulatory effect on epidermal growth factor receptor (EGFR) signaling, although the mechanism by which this occurs remains unknown. Thus in our studies, we aimed to determine the effect of EBP50 expression on EGF-induced cell proliferation and activation of EGFR signaling in the breast cancer cell lines, MDA-MB-231 and MCF-7. In MDA-MB-231 cells, which express low levels of EBP50, EBP50 overexpression inhibited EGF-induced cell proliferation, ERK1/2 and AKT phosphorylation. In MCF-7 cells, which express high levels of EBP50, EBP50 knockdown promoted EGF-induced cell proliferation, ERK1/2 and AKT phosphorylation. Knockdown of EBP50 in EBP50-overexpressed MDA-MB-231 cells abrogated the inhibitory effect of EBP50 on EGF-stimulated ERK1/2 phosphorylation and restoration of EBP50 expression in EBP50-knockdown MCF-7 cells rescued the inhibition of EBP50 on EGF-stimulated ERK1/2 phosphorylation, further confirming that the activation of EGF-induced downstream molecules could be specifically inhibited by EBP50 expression. Since EGFR signaling was triggered by EGF ligands via EGFR phosphorylation, we further detected the phosphorylation status of EGFR in the presence or absence of EBP50 expression. Overexpression of EBP50 in MDA-MB-231 cells inhibited EGF-stimulated EGFR phosphorylation, whereas knockdown of EBP50 in MCF-7 cells enhanced EGF-stimulated EGFR phosphorylation. Meanwhile, total expression levels of EGFR were unaffected during EGF stimulation. Taken together, our data shows that EBP50 can suppress EGF-induced proliferation of breast cancer cells by inhibiting EGFR phosphorylation and blocking EGFR downstream signaling in breast cancer cells. These results provide further insight into the molecular mechanism by which EBP50 regulates the development and progression of breast cancer.

Show MeSH
Related in: MedlinePlus