[Effects of K-ras gene mutation on colon cancer cell line Caco-2 metastasis by regulating E-cadherin/beta-catenin/p120 protein complex formation and RhoA protein activity].

Abstract

OBJECTIVE:

To explore the effects of K-ras gene mutation on colon cancer cell line Caco-2 metastasis by regulating E-cadherin/beta-catenin/p120 protein complex formation and RhoA protein activity.

METHODS:

K-ras wild-type colon cancer cell line Caco-2 was transiently transfected by phr-GFP vector (control group), transfected by mutant K-ras gene phr-K-ras (Val12) vector (transfection group), transfected by mutant K-ras gene phr-K-ras (Val12) vector and treated by specific MAPK pathway inhibitor PD98059 (MAPK inhibition group), or transfected by mutant K-ras gene phr-K-ras (Val12) vector and treated by specific PI-3K pathway inhibitor LY294002 (PI-3K inhibition group), respectively. Cell migration was tested by Transwell experiment. E-cadherin and beta-catenin protein expression and intracellular location were detected by cell immunofluorescence method. Intracellular p120 protein expression was detected by Western blot. beta-catenin protein level which combined with E-cadherin was detected by immunoprecipitation. RhoA activity was analyzed by Pull-down assay.

RESULTS:

The Caco-2 cell migration rate was (19.8 +/- 5.6) % in transfection group, which was significantly higher than that in control group [(14.0 +/- 4.2) %] (P = 0.001) and in MAPK inhibition group [(15.8 +/- 1.2) %] (P = 0.044), but was not significantly different from that in PI-3K inhibition group [(17.5 +/- 2.8) %] (P = 0.095). Immunofluorescence method showed that the E-cadherin and beta-catenin stain located in the cell membrane decreased in transfection group. Western blot showed that the total intracellular p120 protein decreased in transfection group and PI-3K inhibition group. Immunoprecipitation data showed that beta-catenin protein level combined with E-cadherin decreased in transfection group and PI-3K group. Pull-down test showed that RhoA protein activity was up-regulated in transfection group.

CONCLUSION:

K-ras gene mutation stimulates the migration of colon cancer cell Caco-2, which may be achieved by decreasing the E-cadherin/beta-catenin/p120 protein complex formation via MAPK pathway and increasing the RhoA protein activity.

• Authors: Jing-nan Li, Xiao Li, Jia-ming Qian, Xin-qing Lu, Hong Yang
• Journal: Zhongguo yi xue ke xue yuan xue bao. Acta Academiae Medicinae Sinicae (Zhongguo Yi Xue Ke Xue Yuan Xue Bao) Vol. 32 Issue 1 Pg. 46-50 (Feb 2010) ISSN: 1000-503X [Print] China
• PMID: 20236587 (Publication Type: English Abstract, Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Cadherins
• Catenins
• Multiprotein Complexes
• beta Catenin
• rhoA GTP-Binding Protein
• Delta Catenin

Topics

• Caco-2 Cells
• Cadherins (metabolism)
• Catenins (metabolism)
• Cell Movement
• Colonic Neoplasms (metabolism, pathology)
• Genes, ras (genetics)
• Humans
• Multiprotein Complexes (metabolism)
• Mutation
• Neoplasm Metastasis
• Transfection
• beta Catenin (metabolism)
• rhoA GTP-Binding Protein (metabolism)
• Delta Catenin