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Alpha2beta1 integrin regulates lineage commitment in multipotent human colorectal cancer cells.

Kirkland SC, Ying H - J. Biol. Chem. (2008)

Bottom Line: Function-blocking antibodies to alpha2 integrin also blocked both HRA-19 endocrine lineage commitment and enterocytic differentiation by Caco-2 human colon cancer cells; both effects being abrogated by the MEK inhibitor, PD98059, suggesting a role for ERK signaling in alpha2-mediated regulation of colorectal cancer cell differentiation.To further explore the role of alpha2 integrin in multilineage differentiation, we established multipotent cells expressing high levels of wild-type alpha2 integrin or a non-signaling chimeric alpha2 integrin.Overexpression of wild-type alpha2 integrin in HRA-19 cells significantly enhanced endocrine and mucous lineage commitment, while cells expressing the non-signaling chimeric alpha2 integrin had negligible ability for either endocrine or mucous lineage commitment.

View Article: PubMed Central - PubMed

Affiliation: Department of Histopathology, Faculty of Medicine, Imperial College London, London W12 ONN, United Kingdom. s.kirkland@imperial.ac.uk

ABSTRACT
The human colorectal epithelium is maintained by multipotent stem cells that give rise to absorptive, mucous, and endocrine lineages. Recent evidence suggests that human colorectal cancers are likewise maintained by a minority population of so-called cancer stem cells. We have previously established a human colorectal cancer cell line with multipotent characteristics (HRA-19) and developed a serum-free medium that induces endocrine, mucous and absorptive lineage commitment by HRA-19 cells in vitro. In this study, we investigate the role of the beta1 integrin family of cell surface extracellular matrix receptors in multilineage differentiation by these multipotent human colorectal cancer cells. We show that endocrine and mucous lineage commitment is blocked in the presence of function-blocking antibodies to beta1 integrin. Function-blocking antibodies to alpha2 integrin also blocked both HRA-19 endocrine lineage commitment and enterocytic differentiation by Caco-2 human colon cancer cells; both effects being abrogated by the MEK inhibitor, PD98059, suggesting a role for ERK signaling in alpha2-mediated regulation of colorectal cancer cell differentiation. To further explore the role of alpha2 integrin in multilineage differentiation, we established multipotent cells expressing high levels of wild-type alpha2 integrin or a non-signaling chimeric alpha2 integrin. Overexpression of wild-type alpha2 integrin in HRA-19 cells significantly enhanced endocrine and mucous lineage commitment, while cells expressing the non-signaling chimeric alpha2 integrin had negligible ability for either endocrine or mucous lineage commitment. This study indicates that the collagen receptor alpha2beta1 integrin is a regulator of cell fate in human multipotent colorectal cancer cells.

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Integrin expression in HRA-19, human colorectal cancer cells. Western blot analysis of β1(A) and α2(B) integrin expression in lysates of HRA-19 cells. C, surface expression of α2β1 integrin in HRA-19 cells, demonstrated by biotinylation, lysis, and immunoprecipitation with antibodies to β1, α2, and α2β1 integrin. Biotinylated proteins were detected with streptavidinHRP.
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fig3: Integrin expression in HRA-19, human colorectal cancer cells. Western blot analysis of β1(A) and α2(B) integrin expression in lysates of HRA-19 cells. C, surface expression of α2β1 integrin in HRA-19 cells, demonstrated by biotinylation, lysis, and immunoprecipitation with antibodies to β1, α2, and α2β1 integrin. Biotinylated proteins were detected with streptavidinHRP.

Mentions: HRA-19 Cells Express α2β1 Integrin—Immunoblotting was used to analyze integrin expression in HRA-19 cells. Lysates contained two β1 integrin bands representing the immature (smaller band) and the mature glycosylated forms (Fig. 3A) (36). α2 integrin expression was also demonstrated (Fig. 3B). α2β1 integrin was demonstrated at the cell surface by biotinylation of live cells and immunoprecipitation with mAb to β1 integrin (Fig. 3C). Only the fully glycosylated β1 integrin band is seen at the cell surface along with a β1 integrin-associated protein which co-migrates with α2 integrin (Fig. 3C). Immunoprecipitation with antibodies to α2 integrin and α2β1 integrin complex also revealed two biotinylated protein bands corresponding in molecular weight to the α2 and β1 integrin (Fig. 3C).


Alpha2beta1 integrin regulates lineage commitment in multipotent human colorectal cancer cells.

Kirkland SC, Ying H - J. Biol. Chem. (2008)

Integrin expression in HRA-19, human colorectal cancer cells. Western blot analysis of β1(A) and α2(B) integrin expression in lysates of HRA-19 cells. C, surface expression of α2β1 integrin in HRA-19 cells, demonstrated by biotinylation, lysis, and immunoprecipitation with antibodies to β1, α2, and α2β1 integrin. Biotinylated proteins were detected with streptavidinHRP.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2562061&req=5

fig3: Integrin expression in HRA-19, human colorectal cancer cells. Western blot analysis of β1(A) and α2(B) integrin expression in lysates of HRA-19 cells. C, surface expression of α2β1 integrin in HRA-19 cells, demonstrated by biotinylation, lysis, and immunoprecipitation with antibodies to β1, α2, and α2β1 integrin. Biotinylated proteins were detected with streptavidinHRP.
Mentions: HRA-19 Cells Express α2β1 Integrin—Immunoblotting was used to analyze integrin expression in HRA-19 cells. Lysates contained two β1 integrin bands representing the immature (smaller band) and the mature glycosylated forms (Fig. 3A) (36). α2 integrin expression was also demonstrated (Fig. 3B). α2β1 integrin was demonstrated at the cell surface by biotinylation of live cells and immunoprecipitation with mAb to β1 integrin (Fig. 3C). Only the fully glycosylated β1 integrin band is seen at the cell surface along with a β1 integrin-associated protein which co-migrates with α2 integrin (Fig. 3C). Immunoprecipitation with antibodies to α2 integrin and α2β1 integrin complex also revealed two biotinylated protein bands corresponding in molecular weight to the α2 and β1 integrin (Fig. 3C).

Bottom Line: Function-blocking antibodies to alpha2 integrin also blocked both HRA-19 endocrine lineage commitment and enterocytic differentiation by Caco-2 human colon cancer cells; both effects being abrogated by the MEK inhibitor, PD98059, suggesting a role for ERK signaling in alpha2-mediated regulation of colorectal cancer cell differentiation.To further explore the role of alpha2 integrin in multilineage differentiation, we established multipotent cells expressing high levels of wild-type alpha2 integrin or a non-signaling chimeric alpha2 integrin.Overexpression of wild-type alpha2 integrin in HRA-19 cells significantly enhanced endocrine and mucous lineage commitment, while cells expressing the non-signaling chimeric alpha2 integrin had negligible ability for either endocrine or mucous lineage commitment.

View Article: PubMed Central - PubMed

Affiliation: Department of Histopathology, Faculty of Medicine, Imperial College London, London W12 ONN, United Kingdom. s.kirkland@imperial.ac.uk

ABSTRACT
The human colorectal epithelium is maintained by multipotent stem cells that give rise to absorptive, mucous, and endocrine lineages. Recent evidence suggests that human colorectal cancers are likewise maintained by a minority population of so-called cancer stem cells. We have previously established a human colorectal cancer cell line with multipotent characteristics (HRA-19) and developed a serum-free medium that induces endocrine, mucous and absorptive lineage commitment by HRA-19 cells in vitro. In this study, we investigate the role of the beta1 integrin family of cell surface extracellular matrix receptors in multilineage differentiation by these multipotent human colorectal cancer cells. We show that endocrine and mucous lineage commitment is blocked in the presence of function-blocking antibodies to beta1 integrin. Function-blocking antibodies to alpha2 integrin also blocked both HRA-19 endocrine lineage commitment and enterocytic differentiation by Caco-2 human colon cancer cells; both effects being abrogated by the MEK inhibitor, PD98059, suggesting a role for ERK signaling in alpha2-mediated regulation of colorectal cancer cell differentiation. To further explore the role of alpha2 integrin in multilineage differentiation, we established multipotent cells expressing high levels of wild-type alpha2 integrin or a non-signaling chimeric alpha2 integrin. Overexpression of wild-type alpha2 integrin in HRA-19 cells significantly enhanced endocrine and mucous lineage commitment, while cells expressing the non-signaling chimeric alpha2 integrin had negligible ability for either endocrine or mucous lineage commitment. This study indicates that the collagen receptor alpha2beta1 integrin is a regulator of cell fate in human multipotent colorectal cancer cells.

Show MeSH
Related in: MedlinePlus