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Regulation of pancreatic cancer cell migration and invasion by RhoC GTPase and caveolin-1.

Lin M, DiVito MM, Merajver SD, Boyanapalli M, van Golen KL - Mol. Cancer (2005)

Bottom Line: Furthermore, inhibition of RhoC or p38 activity in HPAF-II cells leads to partial restoration of cav-1 expression.Cav-1 expression inhibits RhoC GTPase activation and subsequent activation of the p38 MAPK pathway in primary PC cells thus restricting migration and invasion.In contrast, loss of cav-1 expression leads to RhoC-mediated migration and invasion in metastatic PC cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Hematology-Oncology, Department of Internal Medicine, The University of Michigan Comprehensive Cancer Center, Ann Arbor, Michigan 48109, USA. liming@yahoo.com

ABSTRACT

Background: In the current study we investigated the role of caveolin-1 (cav-1) in pancreatic adenocarcinoma (PC) cell migration and invasion; initial steps in metastasis. Cav-1 is the major structural protein in caveolae; small Omega-shaped invaginations within the plasma membrane. Caveolae are involved in signal transduction, wherein cav-1 acts as a scaffolding protein to organize multiple molecular complexes regulating a variety of cellular events. Recent evidence suggests a role for cav-1 in promoting cancer cell migration, invasion and metastasis; however, the molecular mechanisms have not been described. The small monomeric GTPases are among several molecules which associate with cav-1. Classically, the Rho GTPases control actin cytoskeletal reorganization during cell migration and invasion. RhoC GTPase is overexpressed in aggressive cancers that metastasize and is the predominant GTPase in PC. Like several GTPases, RhoC contains a putative cav-1 binding motif.

Results: Analysis of 10 PC cell lines revealed high levels of cav-1 expression in lines derived from primary tumors and low expression in those derived from metastases. Comparison of the BxPC-3 (derived from a primary tumor) and HPAF-II (derived from a metastasis) demonstrates a reciprocal relationship between cav-1 expression and p42/p44 Erk activation with PC cell migration, invasion, RhoC GTPase and p38 MAPK activation. Furthermore, inhibition of RhoC or p38 activity in HPAF-II cells leads to partial restoration of cav-1 expression.

Conclusion: Cav-1 expression inhibits RhoC GTPase activation and subsequent activation of the p38 MAPK pathway in primary PC cells thus restricting migration and invasion. In contrast, loss of cav-1 expression leads to RhoC-mediated migration and invasion in metastatic PC cells.

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Effect of disruption of caveolae and mis-localization of caveolin-1 after treatment of BxPC-3 cells with MβCD. A. Results of a colloidal gold random motility assay performed after a 1 h pre-treatment of BxPC-3 cells with 5 mM MβCD in growth medium. Phagokinetic tracks were measured 16 h after stimulation. MβCD treated BxPC-3 cells were significantly more migratory than untreated cells (* p = 0.0001) B. Total (light grey) and active GTP-bound (dark grey) levels of RhoC were measured in MβCD treated BxPC-3 cells 3 h after treatment. A statistical difference in RhoC activation was reached with a p value of 0.0001. C. Immunoblot analysis of basal levels of active (light grey) and total (dark grey) p42/p44 Erk and p38 MAPK in MβCD treated BxPC-3 cells 3 h after treatment.
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Figure 6: Effect of disruption of caveolae and mis-localization of caveolin-1 after treatment of BxPC-3 cells with MβCD. A. Results of a colloidal gold random motility assay performed after a 1 h pre-treatment of BxPC-3 cells with 5 mM MβCD in growth medium. Phagokinetic tracks were measured 16 h after stimulation. MβCD treated BxPC-3 cells were significantly more migratory than untreated cells (* p = 0.0001) B. Total (light grey) and active GTP-bound (dark grey) levels of RhoC were measured in MβCD treated BxPC-3 cells 3 h after treatment. A statistical difference in RhoC activation was reached with a p value of 0.0001. C. Immunoblot analysis of basal levels of active (light grey) and total (dark grey) p42/p44 Erk and p38 MAPK in MβCD treated BxPC-3 cells 3 h after treatment.

Mentions: Lastly, we treated the BxPC-3 cell line with methyl-β-cyclodextrin (MβCD), which sequesters cholesterol, disrupts caveolae and mis-localizes cav-1 in the cell [43]. As shown in Figure 6A, MβCD treatment significantly increased the area of BxPC-3 migration (p = 0.0001). Increased cellular migration was accompanied by a significant increase in active GTP-bound RhoC (Figure 6B; p = 0.0001). Consistent with previous observations, Figure 6C demonstrates that levels of phospho-p42/p44 Erk in the MβCD treated cells decreased while levels of phospho-p38 MAPK increased. The changes in MAPK proteins approached but did not achieve statistical significance. These data further suggest that activation of RhoC GTPase is regulated by cav-1 and that RhoC signals through the p38 arm of the MAPK pathway to induce cellular migration.


Regulation of pancreatic cancer cell migration and invasion by RhoC GTPase and caveolin-1.

Lin M, DiVito MM, Merajver SD, Boyanapalli M, van Golen KL - Mol. Cancer (2005)

Effect of disruption of caveolae and mis-localization of caveolin-1 after treatment of BxPC-3 cells with MβCD. A. Results of a colloidal gold random motility assay performed after a 1 h pre-treatment of BxPC-3 cells with 5 mM MβCD in growth medium. Phagokinetic tracks were measured 16 h after stimulation. MβCD treated BxPC-3 cells were significantly more migratory than untreated cells (* p = 0.0001) B. Total (light grey) and active GTP-bound (dark grey) levels of RhoC were measured in MβCD treated BxPC-3 cells 3 h after treatment. A statistical difference in RhoC activation was reached with a p value of 0.0001. C. Immunoblot analysis of basal levels of active (light grey) and total (dark grey) p42/p44 Erk and p38 MAPK in MβCD treated BxPC-3 cells 3 h after treatment.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 6: Effect of disruption of caveolae and mis-localization of caveolin-1 after treatment of BxPC-3 cells with MβCD. A. Results of a colloidal gold random motility assay performed after a 1 h pre-treatment of BxPC-3 cells with 5 mM MβCD in growth medium. Phagokinetic tracks were measured 16 h after stimulation. MβCD treated BxPC-3 cells were significantly more migratory than untreated cells (* p = 0.0001) B. Total (light grey) and active GTP-bound (dark grey) levels of RhoC were measured in MβCD treated BxPC-3 cells 3 h after treatment. A statistical difference in RhoC activation was reached with a p value of 0.0001. C. Immunoblot analysis of basal levels of active (light grey) and total (dark grey) p42/p44 Erk and p38 MAPK in MβCD treated BxPC-3 cells 3 h after treatment.
Mentions: Lastly, we treated the BxPC-3 cell line with methyl-β-cyclodextrin (MβCD), which sequesters cholesterol, disrupts caveolae and mis-localizes cav-1 in the cell [43]. As shown in Figure 6A, MβCD treatment significantly increased the area of BxPC-3 migration (p = 0.0001). Increased cellular migration was accompanied by a significant increase in active GTP-bound RhoC (Figure 6B; p = 0.0001). Consistent with previous observations, Figure 6C demonstrates that levels of phospho-p42/p44 Erk in the MβCD treated cells decreased while levels of phospho-p38 MAPK increased. The changes in MAPK proteins approached but did not achieve statistical significance. These data further suggest that activation of RhoC GTPase is regulated by cav-1 and that RhoC signals through the p38 arm of the MAPK pathway to induce cellular migration.

Bottom Line: Furthermore, inhibition of RhoC or p38 activity in HPAF-II cells leads to partial restoration of cav-1 expression.Cav-1 expression inhibits RhoC GTPase activation and subsequent activation of the p38 MAPK pathway in primary PC cells thus restricting migration and invasion.In contrast, loss of cav-1 expression leads to RhoC-mediated migration and invasion in metastatic PC cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Hematology-Oncology, Department of Internal Medicine, The University of Michigan Comprehensive Cancer Center, Ann Arbor, Michigan 48109, USA. liming@yahoo.com

ABSTRACT

Background: In the current study we investigated the role of caveolin-1 (cav-1) in pancreatic adenocarcinoma (PC) cell migration and invasion; initial steps in metastasis. Cav-1 is the major structural protein in caveolae; small Omega-shaped invaginations within the plasma membrane. Caveolae are involved in signal transduction, wherein cav-1 acts as a scaffolding protein to organize multiple molecular complexes regulating a variety of cellular events. Recent evidence suggests a role for cav-1 in promoting cancer cell migration, invasion and metastasis; however, the molecular mechanisms have not been described. The small monomeric GTPases are among several molecules which associate with cav-1. Classically, the Rho GTPases control actin cytoskeletal reorganization during cell migration and invasion. RhoC GTPase is overexpressed in aggressive cancers that metastasize and is the predominant GTPase in PC. Like several GTPases, RhoC contains a putative cav-1 binding motif.

Results: Analysis of 10 PC cell lines revealed high levels of cav-1 expression in lines derived from primary tumors and low expression in those derived from metastases. Comparison of the BxPC-3 (derived from a primary tumor) and HPAF-II (derived from a metastasis) demonstrates a reciprocal relationship between cav-1 expression and p42/p44 Erk activation with PC cell migration, invasion, RhoC GTPase and p38 MAPK activation. Furthermore, inhibition of RhoC or p38 activity in HPAF-II cells leads to partial restoration of cav-1 expression.

Conclusion: Cav-1 expression inhibits RhoC GTPase activation and subsequent activation of the p38 MAPK pathway in primary PC cells thus restricting migration and invasion. In contrast, loss of cav-1 expression leads to RhoC-mediated migration and invasion in metastatic PC cells.

Show MeSH
Related in: MedlinePlus