Abstract
BackgroundRecent advances have revealed a significant contribution of chemokines and their receptors in tumor growth, survival after chemotherapy, and organ-specific metastasis. The CXC chemokine receptor-7 (CXCR7) is the latest chemokine receptor implicated in cancer. Although over expressed in breast cancer cell lines and tumor tissues, its mechanism of action in breast cancer (BrCa) growth and metastasis is unclear. Studies in other cancers have implicated CXCR7 in cell proliferation, anti-apoptotic activity and cell-cell adhesion. The present study was initiated to examine the pattern of CXCR7 expression and its role in regulation of growth signaling in breast cancer.MethodsThe contribution of CXCR7 in BrCa cell proliferation was investigated in representative cell lines using real time quantitative PCR (q-PCR), proliferation assays, immunohistochemistry and immunoblotting. Phenotypic changes were examined after CXCR7 specific cDNA and siRNA transfection and expression levels were monitored by q-PCR. Further, the association of CXCR7 with epidermal growth factor receptor (EGFR) and modulation of its activity were investigated by western blotting, immunofluorescence, and in-situ proximity ligation assays in human BrCa cells and tissues.ResultsCXCR7 was expressed in both, estrogen receptor (ER) positive and negative BrCa cell lines. CXCR7 was also expressed unevenly in normal breast tissues and to a much higher extent in ER + cancer tissues. Depletion of CXCR7 in MCF7 BrCa cells by RNA interference decreased proliferation and caused cell cycle arrest. Further, proximity ligation assay (PLA) revealed colocalization of CXCR7 with EGFR in cancer tissues and cancer cell lines. CXCR7 depletion reduced levels of phospho-EGFR at Tyrosine1110 after EGF-stimulation and also reduced phosphorylation of ERK1/2, indicating a potentially direct impact on mitogenic signaling in MCF7 cells. Using siRNA to knockdown β-arrestin2 in cells with EGFR over expression we were able to nearly deplete the CXCR7-EGFR colocalization events, suggesting that β-arrestin2 acts as a scaffold to enhance CXCR7 dependent activation of EGFR after EGF stimulation.ConclusionsThese results demonstrate coupling of CXCR7 with EGFR to regulate proliferation of BrCa cells and suggest an important ligand-independent role of CXCR7 in BrCa growth. Thus, the CXCR7-EGFR axis is a promising target for breast cancer therapy.Electronic supplementary materialThe online version of this article (doi:10.1186/1476-4598-13-198) contains supplementary material, which is available to authorized users.
Highlights
Recent advances have revealed a significant contribution of chemokines and their receptors in tumor growth, survival after chemotherapy, and organ-specific metastasis
CXC chemokine receptor-7 (CXCR7) is expressed across breast cancer cell lines We performed quantitative PCR (q-PCR) to evaluate the expression level of CXCR7 in breast cancer cell lines
No epidermal growth factor receptor (EGFR) was detected in T47D and SKBR3 cells and CXCR4 was expressed across all breast cancer (BrCa) lines tested
Summary
Recent advances have revealed a significant contribution of chemokines and their receptors in tumor growth, survival after chemotherapy, and organ-specific metastasis. The present study was initiated to examine the pattern of CXCR7 expression and its role in regulation of growth signaling in breast cancer. Β-arrestins normally dock onto the phosphorylated cytoplasmic tail of an activated receptor, preventing further activation or downstream signaling, because they block the G proteins from docking onto the receptor. The arrestin scaffolds may serve as adapter molecules to assemble multi-protein complexes leading to receptor internalization, recycling back to the plasma membrane, and downstream signaling events, including ERK1/2 (extracellular signalregulated kinases) activation [9,10,11]. Arrestins may shuttle between the cell nucleus and cytoplasm [12]. This process is not fully elucidated for CXCR7
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