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Integrin α9β1 promotes malignant tumor growth and metastasis by potentiating epithelial-mesenchymal transition.

Gupta SK, Oommen S, Aubry MC, Williams BP, Vlahakis NE - Oncogene (2012)

Bottom Line: In addition, we found that α9β1 forms a tri-partite protein complex with β-catenin and E-cadherin, which dissociates following integrin activation and subsequent src and β-catenin phosphorylation.These in vitro results are biologically significant as α9β1-expressing cancer cells induce greater tumor growth and metastases in mice as compared to the cells without α9β1 expression or when integrin expression is suppressed.Furthermore, integrin α9β1 is expressed in primary human small cell lung cancer and patients having a high expression of α9β1 demonstrated significantly worse long-term survival compared with patients with low α9β1 expression.

View Article: PubMed Central - PubMed

Affiliation: Thoracic Disease Research Unit, Division of Pulmonary & Critical Care Medicine, Mayo Clinic, Rochester 55905, MN, USA.

ABSTRACT
The integrin α9β1 binds a number of extracellular matrix components to mediate cell adhesion, migration and tissue invasion. Although expressed in a variety of normal human cells including endothelium, it is also expressed in cancer cells. We have previously shown that α9β1 binds VEGF-A to facilitate angiogenesis, an important component of the tumor microenvironment. As α9β1 induces accelerated cancer cell migration, we wished to determine what role it played in cancer growth and metastasis. In this study, we show that α9β1 expression induces molecular changes consistent with epithelial-mesenchymal transition. In addition, we found that α9β1 forms a tri-partite protein complex with β-catenin and E-cadherin, which dissociates following integrin activation and subsequent src and β-catenin phosphorylation. These findings were consistent in cells in which: α9β1 was exogenously over-expressed, or when its expression was suppressed in cancer cells endogenously expressing α9β1. These in vitro results are biologically significant as α9β1-expressing cancer cells induce greater tumor growth and metastases in mice as compared to the cells without α9β1 expression or when integrin expression is suppressed. Furthermore, integrin α9β1 is expressed in primary human small cell lung cancer and patients having a high expression of α9β1 demonstrated significantly worse long-term survival compared with patients with low α9β1 expression. These findings highlight a novel mechanism of integrin α9β1 function in human cancer.

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α9β1-induced malignant tumor growth and metastasis is inhibited followingα9 knockdownA, Flow cytometry for expression of α9β1 in LLC-1 cells transduced with non-targeted (shRNA-Cont) or α9β1-targeted (shRNA-α9) shRNA. B, Cell adhesion C, Haptotaxis (using α9β1-specific ligand, TnfnRAA) and chemotaxis (using 10% FBS) and D, Cell invasion assays in shRNA-Cont (grey bars) or shRNA-α9 (black bars) LLC-1 grown on plastic (no matrix) or TnfnRAA or pFN as indicated. E, Immunoblots to detect EMT and signaling proteins as indicated in lysates from LLC-1 cells expressing shRNA-cont (grey) or shRNA-α9, and grown with (+) or without (−) TnfnRAA. F, Volume of mouse flank tumors from subcutaneous injection of LLC-1 cells expressing shRNA-Cont (grey) or shRNA-α9 (black) ** p<0.01. G, Representative photomicrographs of H&E stained lung (2 mice) showing metastatic lesions derived from LLC-1 cells expressing shRNA-cont or shRNA-α9.
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Figure 5: α9β1-induced malignant tumor growth and metastasis is inhibited followingα9 knockdownA, Flow cytometry for expression of α9β1 in LLC-1 cells transduced with non-targeted (shRNA-Cont) or α9β1-targeted (shRNA-α9) shRNA. B, Cell adhesion C, Haptotaxis (using α9β1-specific ligand, TnfnRAA) and chemotaxis (using 10% FBS) and D, Cell invasion assays in shRNA-Cont (grey bars) or shRNA-α9 (black bars) LLC-1 grown on plastic (no matrix) or TnfnRAA or pFN as indicated. E, Immunoblots to detect EMT and signaling proteins as indicated in lysates from LLC-1 cells expressing shRNA-cont (grey) or shRNA-α9, and grown with (+) or without (−) TnfnRAA. F, Volume of mouse flank tumors from subcutaneous injection of LLC-1 cells expressing shRNA-Cont (grey) or shRNA-α9 (black) ** p<0.01. G, Representative photomicrographs of H&E stained lung (2 mice) showing metastatic lesions derived from LLC-1 cells expressing shRNA-cont or shRNA-α9.

Mentions: Having shown that endogenously expressed α9β1 confers an EMT phenotype and more aggressive cancer attributes, we next assessed EMT when α9 expression was suppressed using α9-targeted lentiviral shRNA (Fig 5A). As expected α9 depleted LLC-1 demonstrated significantly decreased adhesion on TnfnRAA (Fig 5B) whereas cell adhesion to pFN, a non-α9β1 ligand, was not affected. Similarly, migration and invasion (Fig 5C, D) were significantly decreased in cells expressing α9-targeted shRNA suggesting an inhibitory effect on EMT.


Integrin α9β1 promotes malignant tumor growth and metastasis by potentiating epithelial-mesenchymal transition.

Gupta SK, Oommen S, Aubry MC, Williams BP, Vlahakis NE - Oncogene (2012)

α9β1-induced malignant tumor growth and metastasis is inhibited followingα9 knockdownA, Flow cytometry for expression of α9β1 in LLC-1 cells transduced with non-targeted (shRNA-Cont) or α9β1-targeted (shRNA-α9) shRNA. B, Cell adhesion C, Haptotaxis (using α9β1-specific ligand, TnfnRAA) and chemotaxis (using 10% FBS) and D, Cell invasion assays in shRNA-Cont (grey bars) or shRNA-α9 (black bars) LLC-1 grown on plastic (no matrix) or TnfnRAA or pFN as indicated. E, Immunoblots to detect EMT and signaling proteins as indicated in lysates from LLC-1 cells expressing shRNA-cont (grey) or shRNA-α9, and grown with (+) or without (−) TnfnRAA. F, Volume of mouse flank tumors from subcutaneous injection of LLC-1 cells expressing shRNA-Cont (grey) or shRNA-α9 (black) ** p<0.01. G, Representative photomicrographs of H&E stained lung (2 mice) showing metastatic lesions derived from LLC-1 cells expressing shRNA-cont or shRNA-α9.
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Figure 5: α9β1-induced malignant tumor growth and metastasis is inhibited followingα9 knockdownA, Flow cytometry for expression of α9β1 in LLC-1 cells transduced with non-targeted (shRNA-Cont) or α9β1-targeted (shRNA-α9) shRNA. B, Cell adhesion C, Haptotaxis (using α9β1-specific ligand, TnfnRAA) and chemotaxis (using 10% FBS) and D, Cell invasion assays in shRNA-Cont (grey bars) or shRNA-α9 (black bars) LLC-1 grown on plastic (no matrix) or TnfnRAA or pFN as indicated. E, Immunoblots to detect EMT and signaling proteins as indicated in lysates from LLC-1 cells expressing shRNA-cont (grey) or shRNA-α9, and grown with (+) or without (−) TnfnRAA. F, Volume of mouse flank tumors from subcutaneous injection of LLC-1 cells expressing shRNA-Cont (grey) or shRNA-α9 (black) ** p<0.01. G, Representative photomicrographs of H&E stained lung (2 mice) showing metastatic lesions derived from LLC-1 cells expressing shRNA-cont or shRNA-α9.
Mentions: Having shown that endogenously expressed α9β1 confers an EMT phenotype and more aggressive cancer attributes, we next assessed EMT when α9 expression was suppressed using α9-targeted lentiviral shRNA (Fig 5A). As expected α9 depleted LLC-1 demonstrated significantly decreased adhesion on TnfnRAA (Fig 5B) whereas cell adhesion to pFN, a non-α9β1 ligand, was not affected. Similarly, migration and invasion (Fig 5C, D) were significantly decreased in cells expressing α9-targeted shRNA suggesting an inhibitory effect on EMT.

Bottom Line: In addition, we found that α9β1 forms a tri-partite protein complex with β-catenin and E-cadherin, which dissociates following integrin activation and subsequent src and β-catenin phosphorylation.These in vitro results are biologically significant as α9β1-expressing cancer cells induce greater tumor growth and metastases in mice as compared to the cells without α9β1 expression or when integrin expression is suppressed.Furthermore, integrin α9β1 is expressed in primary human small cell lung cancer and patients having a high expression of α9β1 demonstrated significantly worse long-term survival compared with patients with low α9β1 expression.

View Article: PubMed Central - PubMed

Affiliation: Thoracic Disease Research Unit, Division of Pulmonary & Critical Care Medicine, Mayo Clinic, Rochester 55905, MN, USA.

ABSTRACT
The integrin α9β1 binds a number of extracellular matrix components to mediate cell adhesion, migration and tissue invasion. Although expressed in a variety of normal human cells including endothelium, it is also expressed in cancer cells. We have previously shown that α9β1 binds VEGF-A to facilitate angiogenesis, an important component of the tumor microenvironment. As α9β1 induces accelerated cancer cell migration, we wished to determine what role it played in cancer growth and metastasis. In this study, we show that α9β1 expression induces molecular changes consistent with epithelial-mesenchymal transition. In addition, we found that α9β1 forms a tri-partite protein complex with β-catenin and E-cadherin, which dissociates following integrin activation and subsequent src and β-catenin phosphorylation. These findings were consistent in cells in which: α9β1 was exogenously over-expressed, or when its expression was suppressed in cancer cells endogenously expressing α9β1. These in vitro results are biologically significant as α9β1-expressing cancer cells induce greater tumor growth and metastases in mice as compared to the cells without α9β1 expression or when integrin expression is suppressed. Furthermore, integrin α9β1 is expressed in primary human small cell lung cancer and patients having a high expression of α9β1 demonstrated significantly worse long-term survival compared with patients with low α9β1 expression. These findings highlight a novel mechanism of integrin α9β1 function in human cancer.

Show MeSH
Related in: MedlinePlus