Role for ezrin in breast cancer cell chemotaxis to CCL5.
This study was undertaken to observe the effects and the possible mechanism of membrane-cytoskeleton linker ezrin on the chemokine CC chemokine ligand 5 (CCL5)-induced invasive ability in human breast carcinoma MCF-7 cells. RNA interference (RNAi) using ezrin small hairpin RNAs (ezrin shRNA) was used to analyze the role of ezrin in the regulation of this CCL5-induced malignant behavior of MCF-7 cells. The effects of recombinant human CCL5 (rhCCL5) on the cell's invasive ability were detected by transwell assay. Western blotting was performed to examine the expression of the total and the phosphorylated ezrin at protein level. The CCL5-induced changes in the organization of the actin cytoskeleton in the transfected cells were determined using confocal microscopy. Flow cytometry was used to detect the cell cycle. MTT method was used to detect the proliferation of the cells. We found that the MCF-7 cells responded chemotactically to CCL5 in a dose- and time-dependent manner. After RNAi treatment, the proliferation was inhibited and the cell proportion in G2-M phase decreased. The CCL5-induced cell motility and invasiveness were obviously inhibited by the silencing of ezrin. In addition, the CCL5 induced a significant up-regulation in the total and the phosphorylated ezrin expression in MCF-7 cells, whereas in the presence of ezrin silencing, the CCL5 induced only a slight increase in the total and the phosphorylated ezrin expression. CCL5 was shown to induce changes in the organization of the actin cytoskeleton and the level of F-actin in MCF-7 cells, and the silencing of ezrin could inhibit these changes. Collectively, our data further show that CCL5 induced invasiveness in MCF-7 cells. These data indicate a potential role for ezrin in the processes of the CCL5-induced breast cancer cell migration, invasiveness and metastasis. It is suggested that ezrin may act as downstream effector of CCL5 and a new anti-invasive therapeutic target for human breast cancer.
Li
Juanjuan
J
Department of Breast and Thyroid Gland Surgery of Renmin Hospital of Wuhan University, The First College of Clinical Medicine of Wuhan University, Wuhan, Hubei 430060, P.R. China.
Tu
Yi
Y
Wen
Jing
J
Yao
Feng
F
Wei
Wen
W
Sun
Shengrong
S
D015153:Blotting, Western; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics*; D049109:Cell Proliferation; D002469:Cell Separation; D018946:Chemokine CCL5 / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D003599:Cytoskeleton / Q000187:drug effects; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D018613:Microscopy, Confocal; D009361:Neoplasm Invasiveness / Q000235:genetics* / Q000473:pathology; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000099149:Ezrin
D018946:Chemokine CCL5; D003598:Cytoskeletal Proteins; D000099149:Ezrin; C483024:CCL5 protein, human