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Angiomotin: an angiostatin binding protein that regulates endothelial cell migration and tube formation.

Troyanovsky B, Levchenko T, Månsson G, Matvijenko O, Holmgren L - J. Cell Biol. (2001)

Bottom Line: Transfected angiomotin as well as endogenous angiomotin protein were localized to the leading edge of migrating endothelial cells.Expression of angiomotin in endothelial cells resulted in increased cell migration, suggesting a stimulatory role of angiomotin in cell motility.These findings indicate that angiostatin inhibits cell migration by interfering with angiomotin activity in endothelial cells.

View Article: PubMed Central - PubMed

Affiliation: Center for Genomics Research and Microbiology and Tumor Biology Center, Karolinska Institutet, S-171 76 Stockholm, Sweden.

ABSTRACT
Angiostatin, a circulating inhibitor of angiogenesis, was identified by its ability to maintain dormancy of established metastases in vivo. In vitro, angiostatin inhibits endothelial cell migration, proliferation, and tube formation, and induces apoptosis in a cell type-specific manner. We have used a construct encoding the kringle domains 1--4 of angiostatin to screen a placenta yeast two-hybrid cDNA library for angiostatin-binding peptides. Here we report the identification of angiomotin, a novel protein that mediates angiostatin inhibition of migration and tube formation of endothelial cells. In vivo, angiomotin is expressed in the endothelial cells of capillaries as well as larger vessels of the human placenta. Upon expression of angiomotin in HeLa cells, angiomotin bound and internalized fluorescein-labeled angiostatin. Transfected angiomotin as well as endogenous angiomotin protein were localized to the leading edge of migrating endothelial cells. Expression of angiomotin in endothelial cells resulted in increased cell migration, suggesting a stimulatory role of angiomotin in cell motility. However, treatment with angiostatin inhibited migration and tube formation in angiomotin-expressing cells but not in control cells. These findings indicate that angiostatin inhibits cell migration by interfering with angiomotin activity in endothelial cells.

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Angiostatin induces FAK activity in vitro in cells transfected with angiomotin. (A) Angiomotin-negative EaHy926 or (B) MAE cells were transfected with angiomotin or empty vector. Cells were stimulated with angiostatin at the indicated time points. FAK was immunoprecipitated and subjected to in vitro kinase analysis as described in Materials and Methods. Arrows indicate the localization of p125 FAK.
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Figure 6: Angiostatin induces FAK activity in vitro in cells transfected with angiomotin. (A) Angiomotin-negative EaHy926 or (B) MAE cells were transfected with angiomotin or empty vector. Cells were stimulated with angiostatin at the indicated time points. FAK was immunoprecipitated and subjected to in vitro kinase analysis as described in Materials and Methods. Arrows indicate the localization of p125 FAK.

Mentions: Previous studies have demonstrated that angiostatin induces FAK activity in in vitro assays (Claesson-Welsh et al. 1998). This is, to our knowledge, the only reported signaling pathway that is affected by angiostatin. To assess the effect of angiomotin on FAK activity, we transfected angiomotin into the human cell line EaHy926 (derived from a fusion hybrid between HUVECs and a carcinoma cell line) or into the immortalized MAE cell line. Expression of endogenous angiomotin was not detectable by RT-PCR analysis in neither of these two cell lines (data not shown). Addition of angiostatin to angiomotin-transfected cells increased the in vitro kinase activity of FAK within 1 h (Fig. 6A and Fig. B). By contrast, no effect of angiostatin was detected in cells transfected with the empty vector.


Angiomotin: an angiostatin binding protein that regulates endothelial cell migration and tube formation.

Troyanovsky B, Levchenko T, Månsson G, Matvijenko O, Holmgren L - J. Cell Biol. (2001)

Angiostatin induces FAK activity in vitro in cells transfected with angiomotin. (A) Angiomotin-negative EaHy926 or (B) MAE cells were transfected with angiomotin or empty vector. Cells were stimulated with angiostatin at the indicated time points. FAK was immunoprecipitated and subjected to in vitro kinase analysis as described in Materials and Methods. Arrows indicate the localization of p125 FAK.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199208&req=5

Figure 6: Angiostatin induces FAK activity in vitro in cells transfected with angiomotin. (A) Angiomotin-negative EaHy926 or (B) MAE cells were transfected with angiomotin or empty vector. Cells were stimulated with angiostatin at the indicated time points. FAK was immunoprecipitated and subjected to in vitro kinase analysis as described in Materials and Methods. Arrows indicate the localization of p125 FAK.
Mentions: Previous studies have demonstrated that angiostatin induces FAK activity in in vitro assays (Claesson-Welsh et al. 1998). This is, to our knowledge, the only reported signaling pathway that is affected by angiostatin. To assess the effect of angiomotin on FAK activity, we transfected angiomotin into the human cell line EaHy926 (derived from a fusion hybrid between HUVECs and a carcinoma cell line) or into the immortalized MAE cell line. Expression of endogenous angiomotin was not detectable by RT-PCR analysis in neither of these two cell lines (data not shown). Addition of angiostatin to angiomotin-transfected cells increased the in vitro kinase activity of FAK within 1 h (Fig. 6A and Fig. B). By contrast, no effect of angiostatin was detected in cells transfected with the empty vector.

Bottom Line: Transfected angiomotin as well as endogenous angiomotin protein were localized to the leading edge of migrating endothelial cells.Expression of angiomotin in endothelial cells resulted in increased cell migration, suggesting a stimulatory role of angiomotin in cell motility.These findings indicate that angiostatin inhibits cell migration by interfering with angiomotin activity in endothelial cells.

View Article: PubMed Central - PubMed

Affiliation: Center for Genomics Research and Microbiology and Tumor Biology Center, Karolinska Institutet, S-171 76 Stockholm, Sweden.

ABSTRACT
Angiostatin, a circulating inhibitor of angiogenesis, was identified by its ability to maintain dormancy of established metastases in vivo. In vitro, angiostatin inhibits endothelial cell migration, proliferation, and tube formation, and induces apoptosis in a cell type-specific manner. We have used a construct encoding the kringle domains 1--4 of angiostatin to screen a placenta yeast two-hybrid cDNA library for angiostatin-binding peptides. Here we report the identification of angiomotin, a novel protein that mediates angiostatin inhibition of migration and tube formation of endothelial cells. In vivo, angiomotin is expressed in the endothelial cells of capillaries as well as larger vessels of the human placenta. Upon expression of angiomotin in HeLa cells, angiomotin bound and internalized fluorescein-labeled angiostatin. Transfected angiomotin as well as endogenous angiomotin protein were localized to the leading edge of migrating endothelial cells. Expression of angiomotin in endothelial cells resulted in increased cell migration, suggesting a stimulatory role of angiomotin in cell motility. However, treatment with angiostatin inhibited migration and tube formation in angiomotin-expressing cells but not in control cells. These findings indicate that angiostatin inhibits cell migration by interfering with angiomotin activity in endothelial cells.

Show MeSH
Related in: MedlinePlus