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Molecular and Cellular Pathobiology

Steroid Receptor Coactivator-1 Upregulates Integrin α5 Expression to Promote Breast Cancer Cell Adhesion and Migration

Li Qin, Xian Chen, Yelin Wu, Zhen Feng, Tao He, Li Wang, Lan Liao and Jianming Xu
DOI: 10.1158/0008-5472.CAN-10-3453 Published March 2011

Abstract

Metastatic breast cancer remains a lethal disease with poorly understood molecular mechanisms. Steroid receptor coactivator-1 (SRC-1 or NCOA1) is overexpressed in a subset of breast cancers with poor prognosis. It potentiates gene expression by serving as a coactivator for nuclear receptors and other transcription factors. We previously reported that SRC-1 promotes breast cancer metastasis without affecting primary mammary tumor formation. Herein, we found that SRC-1 deficiency in mouse and human breast cancer cells substantially reduced cell adhesion and migration capabilities on fibronectin and significantly extended the time of focal adhesion disassembly and reassembly. In agreement with this phenotype, SRC-1 expression positively correlated with integrin α5 (ITGA5) expression in estrogen receptor–negative breast tumors whereas SRC-1 deficiency decreased ITGA5 expression. Furthermore, ITGA5 reduction in SRC-1–deficient/insufficient breast cancer cells or knockdown of ITGA5 in SRC-1–expressing breast cancer cells was associated with a disturbed integrin-mediated signaling. Critical downstream changes included reduced phosphorylation and/or dampened activation of focal adhesion kinase, paxillin, Rac1, and Erk1/2 during cell adhesion. Finally, we found that SRC-1 enhanced ITGA5 promoter activity through an AP-1 (activator protein)–binding site proximal to the transcriptional initiation site; both SRC-1 and c-Jun were recruited to this promoter region in breast cancer cells. These results show that SRC-1 can promote breast cancer metastasis by directly enhancing ITGA5 expression and thus promoting ITGA5-mediated cell adhesion and migration. Therefore, targeting ITGA5 in SRC-1–positive breast cancers may result in inhibition of SRC-1–promoted breast cancer metastasis. Cancer Res; 71(5); 1742–51. ©2011 AACR.

Introduction

Steroid receptor coactivator-1 (SRC-1) boosts gene expression by serving as a transcriptional coactivator for nuclear hormone receptors and other transcription factors such as estrogen receptor α (ERα), progesterone receptor (PR), PEA3, activator protein (AP-1), HIF-1, and Ets-2 (1–6). Its expression in human breast cancer positively correlates with HER2 expression, endocrine therapy resistance, and poor prognosis (5, 7, 8). Knockout of SRC-1 in the MMTV-polyoma middle T antigen (PyMT) mammary tumor-prone mice dramatically suppresses lung metastasis without affecting primary tumor formation (9). These studies indicate that SRC-1 strongly promotes breast cancer metastasis.

SRC-1 upregulates the expression of several key regulators for breast cancer progression. In particular, SRC-1 deficiency in mouse mammary tumors reverses HER2 overexpression and reduces Akt activity (9). Knockout of SRC-1 in these tumors suppresses the expression of colony-stimulating factor (CSF-1; ref. 9), a chemoattractant that recruits macrophages to the tumor site. In turn, the macrophages secret epidermal growth factor (EGF) to stimulate tumor cell motility. SRC-1 also serves as a coactivator of PEA3 to enhance Twist1 expression in breast cancer cells. Elevated levels of Twist1 promote breast tumor cell epithelial mesenchymal transition (EMT), invasion, and metastasis by recruiting the NuRD protein complex to repress E-cadherin expression (2, 10). Furthermore, SRC-1 works with Ets-2 to induce c-Myc expression and with HOXC11 to induce calcium-binding protein S100beta expression, both of which are positively associated with acquired resistance to endocrine therapy (5, 7).

Recently, we discovered that the number of mammary tumor cells in the blood of SRC-1 wild-type (WT);PyMT mice is significantly higher than that in the blood of SRC-1–knockout (KO);PyMT mice, suggesting a contribution of SRC-1 to breast cancer cell migration and invasion from the primary tumor to the blood vessels (9). Local migration and invasion of tumor cells are early events partially induced by the tumor microenvironment in metastasis. Resident fibroblasts not only secret TGFβ to induce tumor cell EMT but also produce abundant collagen and fibronectin extracellular matrix (ECM) proteins to provide anchorages for tumor cell adhesion and migration (11–13). Integrins consist of 18 α- and 8 β-glycoprotein subunits, which form 24 distinct heterodimeric transmembrane receptors. These receptors bind to ECM proteins such as fibronectin to transport signals bidirectionally across the cell membrane, allowing cells to respond to environmental changes (14). Multiple integrins, including αvβ3, αvβ5, α5β1, α6β4, α4β1, and αvβ6, are detected in cancer cells and their expression levels are associated with tumorigenesis and cancer progression (15). In breast cancer, integrin β4 amplifies HER2 signaling to potentiate mammary tumorigenesis (16). Activation of integrin αvβ3 supports breast cancer cell adhesion to the vascular wall and promotes metastasis (17), whereas knockout of integrin β1 inhibits mammary tumorigenesis in mice (18). In addition, integrins also regulate tumor cell survival, growth, and metastasis in an anchorage-independent manner (15).

The mesenchymal integrins α5 (ITGA5) and β1 form heterodimers to mediate cell adhesion to fibronectin (15). Knockout of ITGA5 in mice results in embryonic lethality (19). In human hepatocarcinoma cells, ITGA5 promotes cell adhesion and migration on fibronectin through activating focal adhesion kinase (FAK; ref. 20). In transformed mammary epithelial cells, ITGA5 expression is increased along with the EMT process (21). These findings indicate that ITGA5 expression correlates with cancer progression and plays an important role to enhance cancer cell adhesion to and migration on fibronectin.

In this study, we found that SRC-1 works with AP-1 to potentiate ITGA5 expression. The increased ITGA5, in turn, significantly accelerates breast cancer cell adhesion and migration on fibronectin. The identification of ITGA5 as a target gene of SRC-1 and AP-1 in breast cancer cells uncovered a new molecular pathway: SRC-1 regulates ITGA5 expression to promote breast cancer metastasis.

Materials and Methods

Cell adhesion and migration assays

The primary and stable SRC-1 WT; PyMT (WT) and KO;PyMT (KO) mouse mammary tumor cell lines were generated as described previously (22). Adhesion assay was performed on fibronectin or laminin coated plates as described previously (23). Individual cell migration was tracked for 18 hours in 96-well plate precoated with fluorescent beads, and track areas were analyzed using NIH image software as described previously (2, 22).

Western blot analysis of human breast tumors

A total of 24 human breast cancer specimens were collected from surgically removed tumor tissues at Luzhou Medical College Affiliated Hospital in 2009. All patients were Asian women and aged 33 to 65 years. No patient survival data were available at this stage. A portion of the specimen was used for clinical diagnosis of tumor pathology and immunohistochemistry for ERα, PR, and HER2. The remaining tumor tissues were immediately frozen in liquid nitrogen and stored at −80°C. Tumor tissue lysates were prepared after homogenizing the tissues in a lysis buffer containing the protease inhibitor cocktail. The tissue lysates with 50 μg protein were analyzed by Western blotting, using antibodies against SRC-1, ITGA5, and β-actin. Band intensities were determined by densitometry and normalized to the β-actin band intensity.

Other methods

Immunostaining, knockdown, and expression of SRC-1, focal adhesion disassembly assay, Western blotting, quantitative RT-PCR (qPCR), cell transfection, luciferase assay, and chromatin immunoprecipitation (ChIP) assay were performed as descried in the Supplementary Methods.

Results

SRC-1 deficiency reduces mammary tumor cell adhesion and migration on fibronectin-coated plate

To define the role of SRC-1 in breast cancer cell adhesion to ECM, we compared the adhesion capabilities of 2 SRC-1 KO (KO1 and KO2) to 2 WT (WT1 and WT2) mouse mammary tumor cell lines. As expected, SRC-1 protein was detected in both WT cell lines but was absent in both KO cell lines (Fig. 1A). After seeded in an fibronectin-coated plate, about 90% of WT1 and 75% of WT2 cells adhered and spread within 1 hour, and these percentages increased to nearly 100% by 2.5 hours. On the contrary, only about 20% of KO1 and 36% of KO2 cells adhered and spread on fibronectin-coated plate by 1 hour, increasing to only 60% and 70%, respectively, by 2.5 hours (Fig. 1A; data not shown). After culturing overnight, both SRC-1 WT and KO cell lines appeared to adhere and spread well on the fibronectin-coated plate (Fig. 1A). Next, we repeated these assays by using 5 WT and 5 SRC-1 KO primary cell preparations isolated from individual mammary tumors in WT;PyMT and KO;PyMT mice. About 60% to 80% of WT primary cells adhered to fibronectin-coated plate in 1 hour, whereas only 40% to 50% of KO primary cells adhered under the same conditions (Fig. 1B). These results show that SRC-1 is required for effective adhesion of mammary tumor cells to fibronectin.

Figure 1.
Figure 1.

SRC-1 deficiency reduces breast cancer cell adhesion and migration on fibronectin-coated plates. A, phase-contrast images of SRC-1 WT1 and KO1 mammary tumor cell lines after culturing on fibronectin-coated plates for time periods indicated (left), and percentages of adhered KO1, KO2, WT1, and WT2 cells at 1 hour culture time (right). *, P < 0.05 by t test. Western blot results showed the presence of SRC-1 protein in WT1 and WT2 cells and the absence in KO1 and KO2 cells. B, percentages of adhered SRC-1 KO;PyMT (n = 5) and WT; PyMT (n = 5) primary mammary tumor cells after culturing on fibronectin-coated plates for 1 hour. C, left, migration track areas of SRC-1 KO1, KO2, WT1, and WT2 cell lines on fibronectin-coated plates. Average migration area was calculated from the track areas of 50 cells. Middle, knockdown of SRC-1 mRNA in WT1 and WT2 cells by using siRNAs; nontargeting siRNA was used as a control. Right, SRC-1 knockdown reduced the average migration areas of WT1 and WT2 cells. D, left, knockdown of SRC-1 mRNA in MDA-MB-231 cells by using 3 shRNA constructs; nontargeting shRNA was used as a control. Right, MDA-MB-231 cells with SRC-1 knockdown showed reduced cell migration as measured by tracing the migration areas with fluorescent beads.

Next, the migration of individual SRC-1 WT and KO cells was tracked on fibronectin-coated plates for 18 hours. The average track areas swept by the 2 WT cell lines were 60,000 and 90,000 pixels, respectively, whereas those of the SRC-1 KO cell lines were only 25,000 and 20,000 pixels (Fig. 1C). Knockdown of SRC-1 by using siRNAs in both mouse WT cell lines effectively reduced their migration areas by 50% (Fig. 1C). Similarly, knockdown of SRC-1 mRNA in MDA-MB-231 human breast cancer cells by using 3 different shRNAs also significantly reduced cell migration on fibronectin-coated plate by 50% (Fig. 1D). These results indicate that SRC-1 expression facilitates the migration of mammary tumor cells along fibronectin and SRC-1 deficiency attenuates the tumor cell migration.

SRC-1 deficiency attenuates disassembly and reassembly of focal adhesion complexes

Focal adhesions connect the cell cytoskeleton and the ECM through integrins. In motile cells, the rate of focal adhesion assembly at the leading edge and disassembly at the trailing edge determines the speed of cell movement (24). Immunostaining of vinculin, an adaptor protein located in focal adhesion complexes (FAC), showed that focal adhesions are distributed in a polarized pattern in most WT cells, suggesting an active migration status of these cells. However, the FAC distribution in most (∼70%) of SRC-1 KO cells was nonpolarized and, instead, distributed evenly over the cell membrane. This suggested most SRC-1 KO cells remained in an inactive migration status (Fig. 2A). Accordingly, F-actin filaments in SRC-1 KO cells were much more abundant than in WT cells (Fig. 2A).

Figure 2.
Figure 2.

The disassembly and reassembly of FACs and ITGA5 expression in SRC-1 KO and WT mammary tumor cells. A, different distribution patterns of FACs and F-actin in SRC-1 KO1 and WT1 cells as revealed by vinculin immunostaining (green) and philloidin staining (red). Cells were cultured on fibronectin-coated plate for 1 hour. Arrows indicate FACs. B, FAC disassembly and reassembly in SRC-1 KO1 and WT1 cells were monitored at the time points indicated by vinculin immunostaining. C, FAC disassembly in SRC-1 siRNA–transfected WT1 cells was slower than that in control siRNA–transfected WT1 cells. D, measurements of ITGB1, ITGB3, ITGAv, and ITGA5 mRNA levels by qPCR and ITGA5 protein levels by Western blot in SRC-1 KO1, KO2, WT1, and WT2 cells. *, P < 0.05 by t test.

To define the role of SRC-1 in focal adhesion turnover, we examined FAC disassembly and reassembly in WT and SRC-1 KO cells by immunostaining vinculin. Cells were treated with nocodazole to stimulate FAC formation (Fig. 2B). After removing nocodazole, most WT cells showed FAC disassembly in 1 hour and newly formed FACs in 2 hours. However, most SRC-1 KO cells took as long as 2 hours for FAC disassembly and 4 hours for FAC reassembly (Fig. 2B). Furthermore, FACs in WT cells transfected with nontargeting siRNAs disassembled in 1 hour and reassembled in 2 hours whereas FACs in the same cells transfected with SRC-1 siRNAs disassembled in about 2 hours and reassembled in about 4 hours (Fig. 2C). These results suggest that SRC-1 is required for faster FAC turnover in the mammary tumor cells.

Integrins are essential components of FACs. They connect ECM with their extracellular domains to cytoskeleton through interaction of their intracellular domains with multiple proteins such as talin, paxillin, and FAK (14). They also transfer signals across the cell membrane from both sides (14). Since SRC-1 deficiency reduced cell adhesion to fibronectin, we measured the expression levels of integrins β1, β3, αv, and α5, which form αvβ3 and α5β1 heterodimers necessary for binding fibronectin. We found no consensus changes in mRNA levels of integrins β1, αv, and β3 in WT and SRC-1 KO cells (Fig. 2D). However, we found that both mRNA and protein levels of integrin α5 in the 2 SRC-1 KO cell lines were significantly lower than those in the 2 WT cell lines (Fig. 2D). These results suggest that the decreased adhesion capability of SRC-1 KO cells may be related to the lower ITGA5 expression level in these cells.

SRC-1 expression positively correlates with ITGA5 expression

To explore the possibility that SRC-1 may regulate ITGA5 expression, we ectopically expressed SRC-1 in SRC-1 KO cell lines. We found that SRC-1 restoration in these cells promoted ITGA5 expression (Fig. 3A). Conversely, knockdown of SRC-1 in WT cell lines dramatically reduced ITGA5 mRNA and protein levels (Fig. 3A; data not shown). Knockdown of SRC-1 in MDA-MB-231 human breast cancer cells also reduced ITGA5 expression (Fig. 3B). These results indicate that SRC-1 expression levels positively correlate with ITGA5 expression levels in mammary tumor cell lines.

Figure 3.
Figure 3.

SRC-1 expression positively correlates with ITGA5 expression. A, adenovirus-mediated SRC-1 expression in SRC-1 KO1 and KO2 cells enhanced ITGA5 expression. Adenovirus-mediated GFP expression served as a control (left). Conversely, siRNA-mediated knockdown of SRC-1 decreased ITGA5 mRNA and protein levels in SRC-1 WT1 and WT2 cells. The nontargeting siRNA served as a control (middle and right). *, P < 0.05 by t test. B, the shRNA-mediated stable knockdown of SRC-1 in MDA-MB-231 cells reduced ITGA5 expression. C, immunohistochemical staining (brown color) of ITGA5 in SRC-1 WT;PyMT and KO;PyMT tumors isolated from 8- and 14-week-old mice. Slides were counterstained with hematoxylin. D1, Western blot analysis of SRC-1 and ITGA5 proteins in ERα+; PR+ (n = 11), ERα+; PR− (n = 5) and ERα−; PR− (n = 8) human breast tumors was performed as shown in Supplementary Figure 1. β-Actin was assayed as a loading control. Band intensities were measured by densitometry. Relative ITGA5 and SRC-1 levels were normalized to β-actin and presented here as bar graphs. *, P < 0.05, and **, P < 0.01 by F test. D2, SRC-1 mRNA levels correlate with ITGA5 mRNA levels in human ductal breast carcinoma. The microarray data were collected by M. Bittner and downloaded from Oncomine Database. The SRC-1 and ITGA5 expression data shown in Supplementary Figure 2 were plotted and statistically analyzed by Pearson's correlation. The P value indicates that the expression levels of SRC-1 significantly correlate with the expression levels of ITGA5 in the human ductal breast carcinomas.

Next, we examined ITGA5 expression levels in SRC-1 WT;PyMT, and KO; PyMT mouse mammary tumors. In all 4 samples of each tumor type, ITGA5 immunoreactivity was detected on the membrane and in the cytoplasm of tumor cells. However, both the immunostaining signals of ITGA5 and the number of ITGA5-positive tumor cells in WT; PyMT tumors were much higher than those in KO; PyMT tumors isolated from 8- and 14-week-old mice (Fig. 3C; data not shown). The average ITGA5 immunoreactivity detected in WT; PyMT tumors (n = 12) was about 2-fold higher than that detected in KO;PyMT tumors (n = 13). About 40% to 60% of WT; PyMT tumor cells were ITGA5 positive, whereas only 20% to 30% of KO; PyMT tumor cells were ITGA5 positive. These results suggest that ITGA5 expression is also positively associated with SRC-1 expression in mouse mammary tumors.

Finally, we examined the association between SRC-1 and ITGA5 expression levels in human breast tumors by Western blot analysis. The levels of both SRC-1 and ITGA5 proteins were low in ERα- and PR-positive tumors and in ERα-positive and PR-negative tumors. However, the average levels of both SRC-1 and ITGA5 proteins were significantly elevated in ERα- and PR-negative tumors (Fig. 3D1 and Supplementary Fig. S1). In agreement with the protein levels, the expression levels of SRC-1 and ITGA5 mRNAs were also positively correlated in 264 human ductal breast carcinomas according to the data provided to Oncomine Database by M. Bittner (Fig. 3D2 and Supplementary Fig. S2).

SRC-1 potentiates ITGA5-mediated signaling and tumor cell migration

Integrins bind to ECM and integrin clustering activates downstream signaling cascades involving phosphorylation or activation of FAK, Src, paxillin, Rac1, and Erk1/2 (14). To examine the role of SRC-1 in the integrin signaling pathway, we incubated SRC-1 WT and KO cells on fibronectin-coated plates for 0.5 or 1 hour and assessed adhesion-induced FAK activation by measuring pY397-FAK, an autophosphorylation site for Src association (14). Western blot analysis revealed that total FAK levels were similar in SRC-1 WT and KO cell lines whereas the pY397-FAK levels in the 2 SRC-1 KO cell lines were significantly lower than those in the 2 WT cell lines at both time points examined. Knockdown of SRC-1 or ITGA5 in WT cells also reduced the pY397-FAK levels at both or one of the time points examined (Fig. 4A). Similarly, knockdown of SRC-1 in MDA-MB-231 human breast cancer cells consistently decreased the pY397-FAK levels (Fig. 4B).

Figure 4.
Figure 4.

Effects of ITGA5 reduction in SRC-1–deficient cells on integrin signaling pathways and cell migration. A, Western blot analysis of pY397-FAK, total FAK (t-FAK), p-paxillin, t-paxillin, active Rac1 (a-RAC1), and total Rac1 (t-RAC1) in SRC-1 WT1, WT2, KO1, and KO2 cells and WT1 cells transfected with siRNAs against SRC-1 (siSRC-1) or ITGA5 (siITGA5) mRNA or with nontargeting siRNA (control). Cells were allowed to adhere to fibronectin-coated plates for 30 or 60 minutes prior to the analysis. The relative band intensities (indicated) were normalized to total protein band intensities. B, Western blot analysis of pY397-FAK, p-paxillin, and β-actin in MDA-MB-231 cells with stable expression of nontargeting shRNA (Ctrl) or 1 of the 3 shRNAs targeting different regions of the SRC-1 mRNA. C, Western blot analysis of pY925-FAK, t-FAK, p-Erk1/2, and t-Erk1/2 in SRC-1 WT1, WT2, KO1, and KO2 cells after cultured on the fibronectin-coated plates for 30 or 60 minutes. Relative band intensities are indicated. D, WT1 and WT2 cells were transfected with either control siRNA or siRNAs targeting ITGA5 mRNA. ITGA5 mRNA levels in these cells were measured by qPCR (left). These cells were subjected to migration assays on fibronectin- and fluorescence bead–coated plate (middle). The track areas of individual cell migration were traced and averaged from about 50 cells for each group. *, P < 0.05 by t test.

It is known that a decreased pY397-FAK level should be associated with a reduced assembly of FAK and Src complex and the formation of this tyrosine kinase complex is responsible for phosphorylating Y118-paxillin and several other phosphorylation sites of FAK including pY925 for binding GRB2 and activating Erk1/2 (14). In agreement with a lower assembly/activity of the FAK/Src complex, we found that pY118-paxillin, active Rac1, pY925-FAK and p-Erk1/2 levels in SRC-1 KO cells were significantly reduced when compared with WT cells at both 30- and 60-minute adhesion time points (Fig. 4A and C). Accordingly, knockdown of SRC-1 or ITGA5 in SRC-1 WT tumor cells or knockdown of SRC-1 in MDA-MB-231 human breast cancer cells also reduced the levels of p-paxillin (Fig. 4A and B). Furthermore, p-Erk1/2 levels in SRC-1 KO cell lines were much lower than that in WT cells, which was consistent with the reduced levels of pY925-FAK for GRB2 recruitment (Fig. 4C). Taken together, these results suggest that SRC-1 deficiency and ITGA5 insufficiency disturbed integrin-mediated signaling, leading to reduced activation of FAK, Src, Rac1, and Erk1/2, the important players of cell adhesion and migration.

To assess the requirement of ITGA5 in cell migration, ITGA5 mRNA was knocked down in the 2 SRC-1 WT tumor cell lines and these cells were subjected to migration assays on a fluorescent bead–coated plate. The WT cells transfected with ITGA5 siRNAs reduced 60% to 70% of cell motility on the fibronectin-coated plate when compared with WT cells transfected with nontargeting siRNA (Fig. 4D). These results suggest that the elevated ITGA5 level resulting from SRC-1 overexpression in the mouse and human mammary tumors plays an important role in promoting breast cancer cell migration.

SRC-1 promotes AP-1–mediated ITGA5 expression

To examine whether SRC-1 can enhance transcriptional activity of the ITGA5 gene, we constructed 3 luciferase reporter constructs containing different fragments (F1, F2, and F3) of the ITGA5 promoter (Fig. 5A). Expression of SRC-1 in HeLa cells significantly activated all 3 reporters in an SRC-1 dose-dependent manner, suggesting that SRC-1 may coactivate certain transcription factors associated with all 3 fragments of the ITGA5 promoter (Fig. 5B). Sequence analysis identified potential binding sites for several transcription factors known to use SRC-1 as a coactivator, including PEA3, NF-κB, C/EBPα/β, and AP-1 (refs. 2, 3, 25–27; Fig. 5A). Expression of PEA3 or NF-κB, either alone or combined with SRC-1, had no significant effects on the activities of all 3 promoter-reporters. Expression of C/EBPα alone also showed no obvious activation of these promoter-reporters, whereas coexpression of SRC-1 slightly increased the activities of F1-Luc (luciferase) and F3-Luc promoter-reporters. Expression of C/EBPβ alone activated F2-Luc and F3-Luc promoter-reporters, but coexpression of SRC-1 only slightly increased the activities of these promoter-reporters in cells transfected with high concentrations of SRC-1 plasmids (Fig. 5C). These results suggest that PEA3, NF-κB, and C/EBPα/β are not the major transcription factors that work with SRC-1 to enhance ITGA5 promoter activity.

Figure 5.
Figure 5.

SRC-1 regulates ITGA5 promoter activity. A, the pGL3 plasmid constructs containing different ITGA5 promoter fragments (F1, F2, or F3), which are linked to the luciferase (Luc) reporter sequence. The computer-mapped binding sites for AP-1, NF-κB, PEA3, and C/EBP are indicated. The AP-1 site at bp −9 is deleted in pGL3-MF1-Luc, pGL3-MF2-Luc, and pGL3-MF3-Luc constructs. B, expression of SRC-1 in HeLa cells enhances pGL3-F1-Luc, pGL3-F2-Luc, and pGL3-F3-Luc activities in a dose-dependent manner. C, effects of SRC-1 coexpression with PEA3, NF-κB, C/EBPα, or C/EBPβ on the activities of pGL3-F1-Luc, pGL3-F2-Luc, and pGL3-F3-Luc in HeLa cells. D, coexpression of SRC-1 and c-Jun in HeLa cells significantly enhanced the activities of pGL3-F1-Luc, pGL3-F2-Luc, and pGL3-F3-Luc but not those of the mutants pGL3-MF1-Luc, pGL3-MF2-Luc, and pGL3-MF3-Luc.

Figure 6.
Figure 6.

SRC-1 and AP-1 are associated with a proximal region of the ITGA5 promoter. A, ChIP assays were designed to detect regions a, b, c, d, and e of the ITGA5 promoter with specific PCR primer pairs as indicated. Predicted AP-1 binding sites are indicated. B and C, ChIP assays were performed with MDA-MB-231 and MDA-MB-435 cells and antibodies against SRC-1 (B) and c-Jun (C). DNA extracted from cross-linked and sonicated cell lysate was used as input control. IgG served as a control for antibody specificity in ChIP assays. No template (−template) PCR reaction served as a negative control for PCR. PCR-amplified fragments a, b, c, d, and e are indicated. P, the primer bands; ns, a nonspecific PCR product.

Interestingly, although expression of c-Jun alone did not increase the activity of the 3 ITGA5 promoter-reporters, coexpression of c-Jun and SRC-1 robustly activated these constructs in a SRC-1 dose-dependent manner, suggesting that the transcriptional activation function of c-Jun and SRC-1 is associated with AP-1 binding site(s) located in the F3 common region of the ITGA5 promoter (Fig. 5A and D). Furthermore, deletion of the single AP-1 binding site at bp −9 position significantly reduced the basal activity of all 3 ITGA5 promoter-reporters in cells transfected with or without c-Jun plasmids and completely abolished SRC-1–enhanced activities of these ITGA5 promoter regions (Fig. 5D).

Since SRC-1 interacted with c-Jun (3) to activate the ITGA5 promoter (Fig. 5D), we performed ChIP assays to examine whether SRC-1 and c-Jun are recruited to the ITGA5 promoter. ChIP assays showed that the endogenous SRC-1 protein is associated with ITGA5 promoter regions d (bp −420 to −220) and e (bp −115 to +73) in both MDA-MB-231 and MDA-MB-435 cancer cells (Fig. 6A and B). Region e contains the functional AP-1 site (Fig. 5A). In contrast, SRC-1 was not associated with ITGA5 promoter regions a, b, and c (Fig. 6A and B). In agreement with the presence of the functional AP-1 binding site in region e of the ITGA5 promoter, c-Jun was found strongly associated with region e in both MDA-MB-231 and MDA-MB-435 cells but only weakly associated with region d in MDA-MB-435 cells (Fig. 6A and C). These results suggest that both c-Jun and SRC-1 are recruited to region e of the ITGA5 promoter. The association of SRC-1 with region d might be due to either SRC-1 interaction with another transcription factor or a cross PCR reaction from an extended template of region e which is associated with both SRC-1 and c-Jun.

Discussion

The 3 transcriptional coactivators in the SRC family are gene expression amplifiers regulated by multiple signaling pathways (28). These coactivators are expressed in normal cells at limiting concentrations so that their changes in concentration and/or activity can effectively modulate gene expression (28). These coactivators are commonly overexpressed in cancers, acting to promote carcinogenesis and/or metastasis. Specifically, SRC-3 (AIB1) is amplified and overexpressed in a subset of breast cancers and its overexpression is associated with HER2 expression and resistance to endocrine therapy (29, 30). In mouse breast cancer models, knockout of SRC-3 suppresses oncogene and carcinogen-induced carcinogenesis and metastasis whereas SRC-3 overexpression is sufficient to induce high frequency of mammary tumors (22, 31–33). Recent studies also showed that SRC-3 is required for epidermal growth factor receptor (EGFR) and HER2 phosphorylation and activation in breast cancer cells (33). SRC-3Delta4, a splicing isoform of SRC-3, can also serve as a signaling adaptor that links EGFR and FAK and promotes EGF-induced phosphorylation of FAK and c-Src to enhance cell migration (34). SRC-2 (NCOA2) is identified as an overexpressed oncogene in prostate cancer (35). SRC-1 is overproduced in a subset of breast cancers and its expression is positively associated with HER2 expression, tamoxifen resistance, and poor prognosis (6, 8, 28). In mouse models, knockout of SRC-1 does not affect primary mammary tumor formation but effectively suppresses metastasis (9). These findings highlight the crucial roles of the SRC family coactivators in cancer initiation, progression, and metastasis.

Because SRCs work as transcriptional coregulators for many nuclear receptors and other transcription factors, it has been difficult to identify key SRC-associated transcription factors and their target genes responsible for promoting tumorigenesis and metastasis. Although recent studies have made progress in understanding the mechanisms responsible for SRC-1 to promote breast cancer metastasis (2, 5, 9), it is just the beginning to identify key genes and gene networks regulated by SRC-1.

In this study, we have found that SRC-1 deficiency slows breast cancer cell adhesion and migration on fibronectin, which correlates well with the attenuated disassembly and reassembly of FACs in SRC-1 KO and knockdown cells. Furthermore, we identified ITGA5 as a new SRC-1 target gene on the basis of multiple lines of evidence. First, SRC-1 expression levels are associated with ITGA5 expression levels in both mouse and human breast cancer cell lines and primary tumors. Knockout or knockdown of SRC-1 reduces ITGA5 expression, whereas SRC-1 expression enhances ITGA5 expression. Second, SRC-1 is a known coactivator of AP-1 (3), and we found that both AP-1 and SRC-1 associate with the ITGA5 promoter to enhance ITGA5 promoter activity. Although experiment to knock down c-Jun was not performed to further validate the functional contribution of c-Jun to ITGA5 expression in this study, previous studies have shown that AP-1 strongly enhances ITGA5 expression (36, 37). Finally, the reduced ITGA5 expression caused by SRC-1 deficiency or insufficiency in breast cancer cells partially impairs the function of the fibronectin-integrin-FAK cell migration pathway. In this pathway, the heterodimers of α5β1 integrins bind fibronectin to induce phosphorylation of pY397-FAK and formation of the active FAK/Src complex, followed by further phosphorylation of FAK and phosphorylation/activation of downstream signaling components including paxillin, RAC1, and Erk1/2 (14, 38–40). In agreement with an important role of SRC-1–mediated ITGA5 expression in the fibronectin and integrin interaction–initiated signaling and cell migration, the FAK phosphorylation at Y397, paxillin phosphorylation, and activation of Rac-1 and Erk1/2 during cell adhesion process are significantly reduced in SRC-1–knockout and SRC-1- or ITGA5-knockdown breast cancer cells. Taken together, these findings show that SRC-1–mediated ITGA5 expression is partially responsible for SRC-1–promoted adhesion, migration, and metastasis of breast cancer cells.

In the literature, the role of integrin α5β1 in cancer is somewhat controversial. One early study showed that elevated levels of integrin α5β1 suppressed the transformed phenotype of CHO cells (41). Some studies also reported an inhibitory effect of integrins α5β1 on tumorigenesis and metastasis (12, 42). However, multiple lines of evidence suggest that integrin α5β1 expression positively correlates with cancer progression and metastasis. First, integrin α5β1 expression is upregulated in malignant breast cancer cells and its upregulation correlates with poor prognosis (43). Second, SDF1-activated CXCR4 upregulates integrin α5β1 expression and enhances prostate tumor cell adhesion, invasion, and metastasis (44). Third, a mutant p53 has been shown to drive cell invasion and metastatic behavior via activating EGFR/integrins α5β1 signaling (45). Fourth, fibronectin and ITGA5 are required for switching cell–cell adhesion to cell–ECM adhesion and this switch is required for tumor cells to invade into the stromal tissue (46). Fifth, E-cadherin expression in ovarian and breast cancers negatively correlates with ITGA5 expression, suggesting an important role of ITGA5 in EMT (47, 48). Finally, blocking the function of integrin α5β1 by an antibody (volociximab) or a non–RGD-based peptide inhibitor (ATN-161) significantly inhibits breast cancer growth and metastasis (49, 50). In this study, we have shown that SRC-1 upregulates ITGA5 expression to promote breast cancer cell adhesion and migration on fibronectin, facilitating local invasion of these tumor cells. On the basis of the important contributions of ITGA5 in cancer cell migration, invasion, and metastasis described earlier, it is reasonable to conclude that ITGA5 functions as one of the key target genes of SRC-1 to mediate SRC-1–promoted breast cancer cell migration and metastasis.

Since SRC-1 serves as a coactivator for multiple transcription factors such as AP-1, PEA3, Ets-2, and HOXC11 to upregulate the expression of multiple target genes such as ITGA5, Twist1, CSF-1, c-Myc, and S100beta in promotion of breast cancer metastasis (2, 5, 7, 9), targeting SRC-1 could be a potential strategy to interfere multiple pathways involved in cancer metastasis.

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.

Grant Support

This work is supported by NIH grants (CA112403, CA119689, and DK58242) and an American Cancer Society scholar award (RSG-05-082-01-TBE).

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Acknowledgments

We thank Hongwu Chen for providing SRC-1 expression adenovirus, Brian York, Junjiang Fu and Jun Hong for experimental assistance, and Jean Tien for critical reading of the manuscript.

Footnotes

  • Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/).

  • Received September 20, 2010.
  • Revision received December 13, 2010.
  • Accepted December 17, 2010.

References

  1. 1.↵
    1. Xu J,
    2. Qiu Y,
    3. DeMayo FJ,
    4. Tsai SY,
    5. Tsai MJ,
    6. O'Malley BW
    . Partial hormone resistance in mice with disruption of the steroid receptor coactivator-1 (SRC-1) gene. Science 1998;279:1922–5.
    OpenUrlAbstract/FREE Full Text
  2. 2.↵
    1. Qin L,
    2. Liu Z,
    3. Chen H,
    4. Xu J
    . The steroid receptor coactivator-1 regulates twist expression and promotes breast cancer metastasis. Cancer Res 2009;69:3819–27.
    OpenUrlAbstract/FREE Full Text
  3. 3.↵
    1. Lee SK,
    2. Kim HJ,
    3. Na SY,
    4. Kim TS,
    5. Choi HS,
    6. Im SY,
    7. et al.
    Steroid receptor coactivator-1 coactivates activating protein-1-mediated transactivations through interaction with the c-Jun and c-Fos subunits. J Biol Chem 1998;273:16651–4.
    OpenUrlAbstract/FREE Full Text
  4. 4.↵
    1. Carrero P,
    2. Okamoto K,
    3. Coumailleau P,
    4. O'Brien S,
    5. Tanaka H,
    6. Poellinger L
    . Redox-regulated recruitment of the transcriptional coactivators CREB-binding protein and SRC-1 to hypoxia-inducible factor 1alpha. Mol Cell Biol 2000;20:402–15.
    OpenUrlAbstract/FREE Full Text
  5. 5.↵
    1. Al-azawi D,
    2. Ilroy MM,
    3. Kelly G,
    4. Redmond AM,
    5. Bane FT,
    6. Cocchiglia S,
    7. et al.
    Ets-2 and p160 proteins collaborate to regulate c-Myc in endocrine resistant breast cancer. Oncogene 2008;27:3021–31.
    OpenUrlCrossRefPubMed
  6. 6.↵
    1. Myers E,
    2. Hill AD,
    3. Kelly G,
    4. McDermott EW,
    5. O'Higgins NJ,
    6. Buggy Y,
    7. et al.
    Associations and interactions between Ets-1 and Ets-2 and coregulatory proteins, SRC-1, AIB1, and NCoR in breast cancer. Clin Cancer Res 2005;11:2111–22.
    OpenUrlAbstract/FREE Full Text
  7. 7.↵
    1. McIlroy M,
    2. McCartan D,
    3. Early S,
    4. P OG,
    5. Pennington S,
    6. Hill AD,
    7. et al.
    Interaction of developmental transcription factor HOXC11 with steroid receptor coactivator SRC-1 mediates resistance to endocrine therapy in breast cancer [corrected]. Cancer Res 2010;70:1585–94.
    OpenUrlAbstract/FREE Full Text
  8. 8.↵
    1. Redmond AM,
    2. Bane FT,
    3. Stafford AT,
    4. McIlroy M,
    5. Dillon MF,
    6. Crotty TB,
    7. et al.
    Coassociation of estrogen receptor and p160 proteins predicts resistance to endocrine treatment; SRC-1 is an independent predictor of breast cancer recurrence. Clin Cancer Res 2009;15:2098–106.
    OpenUrlAbstract/FREE Full Text
  9. 9.↵
    1. Wang S,
    2. Yuan Y,
    3. Liao L,
    4. Kuang SQ,
    5. Tien JC,
    6. O'Malley BW,
    7. et al.
    Disruption of the SRC-1 gene in mice suppresses breast cancer metastasis without affecting primary tumor formation. Proc Natl Acad Sci U S A 2009;106:151–6.
    OpenUrlAbstract/FREE Full Text
  10. 10.↵
    1. Fu J,
    2. Qin L,
    3. He T,
    4. Qin J,
    5. Hong J,
    6. Wong J,
    7. et al.
    The TWIST/Mi2/NuRD protein complex and its essential role in cancer metastasis. Cell Res 2010 Aug 17. [Epub ahead of print].
  11. 11.↵
    1. Thiery JP
    . Epithelial-mesenchymal transitions in tumour progression. Nat Rev Cancer 2002;2:442–54.
    OpenUrlCrossRefPubMed
  12. 12.↵
    1. Larsen M,
    2. Artym VV,
    3. Green JA,
    4. Yamada KM
    . The matrix reorganized: extracellular matrix remodeling and integrin signaling. Curr Opin Cell Biol 2006;18:463–71.
    OpenUrlCrossRefPubMed
  13. 13.↵
    1. Bierie B,
    2. Moses HL
    . Tumour microenvironment: TGFbeta: the molecular Jekyll and Hyde of cancer. Nat Rev Cancer 2006;6:506–20.
    OpenUrlCrossRefPubMed
  14. 14.↵
    1. Luo BH,
    2. Carman CV,
    3. Springer TA
    . Structural basis of integrin regulation and signaling. Annu Rev Immunol 2007;25:619–47.
    OpenUrlCrossRefPubMed
  15. 15.↵
    1. Desgrosellier JS,
    2. Cheresh DA
    . Integrins in cancer: biological implications and therapeutic opportunities. Nat Rev Cancer 2010;10:9–22.
    OpenUrlCrossRefPubMed
  16. 16.↵
    1. Guo W,
    2. Pylayeva Y,
    3. Pepe A,
    4. Yoshioka T,
    5. Muller WJ,
    6. Inghirami G,
    7. et al.
    Beta 4 integrin amplifies ErbB2 signaling to promote mammary tumorigenesis. Cell 2006;126:489–502.
    OpenUrlCrossRefPubMed
  17. 17.↵
    1. Felding-Habermann B,
    2. O'Toole TE,
    3. Smith JW,
    4. Fransvea E,
    5. Ruggeri ZM,
    6. Ginsberg MH,
    7. et al.
    Integrin activation controls metastasis in human breast cancer. Proc Natl Acad Sci U S A 2001;98:1853–8.
    OpenUrlAbstract/FREE Full Text
  18. 18.↵
    1. White DE,
    2. Kurpios NA,
    3. Zuo D,
    4. Hassell JA,
    5. Blaess S,
    6. Mueller U,
    7. et al.
    Targeted disruption of beta1-integrin in a transgenic mouse model of human breast cancer reveals an essential role in mammary tumor induction. Cancer Cell 2004;6:159–70.
    OpenUrlCrossRefPubMed
  19. 19.↵
    1. Watt FM,
    2. Hodivala KJ
    . Cell adhesion. Fibronectin and integrin knockouts come unstuck. Curr Biol 1994;4:270–2.
    OpenUrlCrossRefPubMed
  20. 20.↵
    1. Wang QY,
    2. Zhang Y,
    3. Shen ZH,
    4. Chen HL
    . Alpha1,3 fucosyltransferase-VII up-regulates the mRNA of alpha5 integrin and its biological function. J Cell Biochem 2008;104:2078–90.
    OpenUrlCrossRefPubMed
  21. 21.↵
    1. Maschler S,
    2. Wirl G,
    3. Spring H,
    4. Bredow DV,
    5. Sordat I,
    6. Beug H,
    7. et al.
    Tumor cell invasiveness correlates with changes in integrin expression and localization. Oncogene 2005;24:2032–41.
    OpenUrlCrossRefPubMed
  22. 22.↵
    1. Qin L,
    2. Liao L,
    3. Redmond A,
    4. Young L,
    5. Yuan Y,
    6. Chen H,
    7. et al.
    The AIB1 oncogene promotes breast cancer metastasis by activation of PEA3-mediated matrix metalloproteinase 2 (MMP2) and MMP9 expression. Mol Cell Biol 2008;28:5937–50.
    OpenUrlAbstract/FREE Full Text
  23. 23.↵
    1. Qin L,
    2. Zhang M
    . Maspin regulates endothelial cell adhesion and migration through an integrin signaling pathway. J Biol Chem 2010;285:32360–9.
    OpenUrlAbstract/FREE Full Text
  24. 24.↵
    1. Ridley AJ,
    2. Schwartz MA,
    3. Burridge K,
    4. Firtel RA,
    5. Ginsberg MH,
    6. Borisy G,
    7. et al.
    Cell migration: integrating signals from front to back. Science 2003;302:1704–9.
    OpenUrlAbstract/FREE Full Text
  25. 25.↵
    1. Na SY,
    2. Lee SK,
    3. Han SJ,
    4. Choi HS,
    5. Im SY,
    6. Lee JW
    . Steroid receptor coactivator-1 interacts with the p50 subunit and coactivates nuclear factor kappaB-mediated transactivations. J Biol Chem 1998;273:10831–4.
    OpenUrlAbstract/FREE Full Text
  26. 26.↵
    1. Yin L,
    2. Wang Y,
    3. Dridi S,
    4. Vinson C,
    5. Hillgartner FB
    . Role of CCAAT/enhancer-binding protein, histone acetylation, and coactivator recruitment in the regulation of malic enzyme transcription by thyroid hormone. Mol Cell Endocrinol 2005;245:43–52.
    OpenUrlCrossRefPubMed
  27. 27.↵
    1. Dong J,
    2. Tsai-Morris CH,
    3. Dufau ML
    . A novel estradiol/estrogen receptor alpha-dependent transcriptional mechanism controls expression of the human prolactin receptor. J Biol Chem 2006;281:18825–36.
    OpenUrlAbstract/FREE Full Text
  28. 28.↵
    1. Xu J,
    2. Wu RC,
    3. O'Malley BW
    . Normal and cancer-related functions of the p160 steroid receptor co-activator (SRC) family. Nat Rev Cancer 2009;9:615–30.
    OpenUrlCrossRefPubMed
  29. 29.↵
    1. Anzick SL,
    2. Kononen J,
    3. Walker RL,
    4. Azorsa DO,
    5. Tanner MM,
    6. Guan XY,
    7. et al.
    AIB1, a steroid receptor coactivator amplified in breast and ovarian cancer. Science 1997;277:965–8.
    OpenUrlAbstract/FREE Full Text
  30. 30.↵
    1. Osborne CK,
    2. Bardou V,
    3. Hopp TA,
    4. Chamness GC,
    5. Hilsenbeck SG,
    6. Fuqua SA,
    7. et al.
    Role of the estrogen receptor coactivator AIB1 (SRC-3) and HER-2/neu in tamoxifen resistance in breast cancer. J Natl Cancer Inst 2003;95:353–61.
    OpenUrlAbstract/FREE Full Text
  31. 31.↵
    1. Kuang SQ,
    2. Liao L,
    3. Zhang H,
    4. Lee AV,
    5. O'Malley BW,
    6. Xu J
    . AIB1/SRC-3 deficiency affects insulin-like growth factor I signaling pathway and suppresses v-Ha-ras-induced breast cancer initiation and progression in mice. Cancer Res 2004;64:1875–85.
    OpenUrlAbstract/FREE Full Text
  32. 32.↵
    1. Kuang SQ,
    2. Liao L,
    3. Wang S,
    4. Medina D,
    5. O'Malley BW,
    6. Xu J
    . Mice lacking the amplified in breast cancer 1/steroid receptor coactivator-3 are resistant to chemical carcinogen-induced mammary tumorigenesis. Cancer Res 2005;65:7993–8002.
    OpenUrlAbstract/FREE Full Text
  33. 33.↵
    1. Fereshteh MP,
    2. Tilli MT,
    3. Kim SE,
    4. Xu J,
    5. O'Malley BW,
    6. Wellstein A,
    7. et al.
    The nuclear receptor coactivator amplified in breast cancer-1 is required for Neu (ErbB2/HER2) activation, signaling, and mammary tumorigenesis in mice. Cancer Res 2008;68:3697–706.
    OpenUrlAbstract/FREE Full Text
  34. 34.↵
    1. Long W,
    2. Yi P,
    3. Amazit L,
    4. LaMarca HL,
    5. Ashcroft F,
    6. Kumar R,
    7. et al.
    SRC-3Delta4 mediates the interaction of EGFR with FAK to promote cell migration. Mol Cell 37:321–32.
  35. 35.↵
    1. Taylor BS,
    2. Schultz N,
    3. Hieronymus H,
    4. Gopalan A,
    5. Xiao Y,
    6. Carver BS,
    7. et al.
    Integrative genomic profiling of human prostate cancer. Cancer Cell 2010;18:11–22.
    OpenUrlCrossRefPubMed
  36. 36.↵
    1. Corbi AL,
    2. Jensen UB,
    3. Watt FM
    . The alpha2 and alpha5 integrin genes: identification of transcription factors that regulate promoter activity in epidermal keratinocytes. FEBS Lett 2000;474:201–7.
    OpenUrlCrossRefPubMed
  37. 37.↵
    1. Han S,
    2. Roman J
    . COX-2 inhibitors suppress integrin alpha5 expression in human lung carcinoma cells through activation of Erk: involvement of Sp1 and AP-1 sites. Int J Cancer 2005;116:536–46.
    OpenUrlCrossRefPubMed
  38. 38.↵
    1. Cheresh DA,
    2. Leng J,
    3. Klemke RL
    . Regulation of cell contraction and membrane ruffling by distinct signals in migratory cells. J Cell Biol 1999;146:1107–16.
    OpenUrlAbstract/FREE Full Text
  39. 39.↵
    1. Cho SY,
    2. Klemke RL
    . Extracellular-regulated kinase activation and CAS/Crk coupling regulate cell migration and suppress apoptosis during invasion of the extracellular matrix. J Cell Biol 2000;149:223–36.
    OpenUrlAbstract/FREE Full Text
  40. 40.↵
    1. Mitra SK,
    2. Schlaepfer DD
    . Integrin-regulated FAK-Src signaling in normal and cancer cells. Curr Opin Cell Biol 2006;18:516–23.
    OpenUrlCrossRefPubMed
  41. 41.↵
    1. Giancotti FG,
    2. Ruoslahti E
    . Elevated levels of the alpha 5 beta 1 fibronectin receptor suppress the transformed phenotype of Chinese hamster ovary cells. Cell 1990;60:849–59.
    OpenUrlCrossRefPubMed
  42. 42.↵
    1. Fang Z,
    2. Yao W,
    3. Xiong Y,
    4. Zhang J,
    5. Liu L,
    6. Li J,
    7. et al.
    Functional elucidation and methylation-mediated downregulation of ITGA5 gene in breast cancer cell line MDA-MB-468. J Cell Biochem 2010;110:1130–41.
    OpenUrlCrossRefPubMed
  43. 43.↵
    1. Nam JM,
    2. Onodera Y,
    3. Bissell MJ,
    4. Park CC
    . Breast cancer cells in three-dimensional culture display an enhanced radioresponse after coordinate targeting of integrin alpha5beta1 and fibronectin. Cancer Res 70:5238–48.
  44. 44.↵
    1. Engl T,
    2. Relja B,
    3. Marian D,
    4. Blumenberg C,
    5. Muller I,
    6. Beecken WD,
    7. et al.
    CXCR4 chemokine receptor mediates prostate tumor cell adhesion through alpha5 and beta3 integrins. Neoplasia 2006;8:290–301.
    OpenUrlCrossRefPubMed
  45. 45.↵
    1. Muller PA,
    2. Caswell PT,
    3. Doyle B,
    4. Iwanicki MP,
    5. Tan EH,
    6. Karim S,
    7. et al.
    Mutant p53 drives invasion by promoting integrin recycling. Cell 2009;139:1327–41.
    OpenUrlCrossRefPubMed
  46. 46.↵
    1. Guo W,
    2. Giancotti FG
    . Integrin signalling during tumour progression. Nat Rev Mol Cell Biol 2004;5:816–26.
    OpenUrlCrossRefPubMed
  47. 47.↵
    1. Sawada K,
    2. Mitra AK,
    3. Radjabi AR,
    4. Bhaskar V,
    5. Kistner EO,
    6. Tretiakova M,
    7. et al.
    Loss of E-cadherin promotes ovarian cancer metastasis via alpha 5-integrin, which is a therapeutic target. Cancer Res 2008;68:2329–39.
    OpenUrlAbstract/FREE Full Text
  48. 48.↵
    1. Wu H,
    2. Liang YL,
    3. Li Z,
    4. Jin J,
    5. Zhang W,
    6. Duan L,
    7. et al.
    Positive expression of E-cadherin suppresses cell adhesion to fibronectin via reduction of alpha5beta1 integrin in human breast carcinoma cells. J Cancer Res Clin Oncol 2006;132:795–803.
    OpenUrlCrossRefPubMed
  49. 49.↵
    1. Khalili P,
    2. Arakelian A,
    3. Chen G,
    4. Plunkett ML,
    5. Beck I,
    6. Parry GC,
    7. et al.
    A non-RGD-based integrin binding peptide (ATN-161) blocks breast cancer growth and metastasis in vivo . Mol Cancer Ther 2006;5:2271–80.
    OpenUrlAbstract/FREE Full Text
  50. 50.↵
    1. Ricart AD,
    2. Tolcher AW,
    3. Liu G,
    4. Holen K,
    5. Schwartz G,
    6. Albertini M,
    7. et al.
    Volociximab, a chimeric monoclonal antibody that specifically binds alpha5beta1 integrin: a phase I, pharmacokinetic, and biological correlative study. Clin Cancer Res 2008;14:7924–9.
    OpenUrlAbstract/FREE Full Text
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Cancer Research: 71 (5)
March 2011
Volume 71, Issue 5

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Steroid Receptor Coactivator-1 Upregulates Integrin α5 Expression to Promote Breast Cancer Cell Adhesion and Migration
Li Qin, Xian Chen, Yelin Wu, Zhen Feng, Tao He, Li Wang, Lan Liao and Jianming Xu
Cancer Res March 1 2011 (71) (5) 1742-1751; DOI: 10.1158/0008-5472.CAN-10-3453
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