ABSTRACT
Rho GDP dissociation inhibitor 2 (RhoGDI2) expression correlates with tumor growth, metastasis, and chemoresistance in gastric cancer. Here, we show that RhoGDI2 functions in the epithelial-mesenchymal transition (EMT), which is responsible for invasiveness during tumor progression. This tumorigenic activity is associated with repression of E-cadherin by RhoGDI2 via upregulation of Snail. Overexpression of RhoGDI2 induced phenotypic changes consistent with EMT in gastric cancer cells, including abnormal epithelial cell morphology, fibroblast-like properties, and reduced intercellular adhesion. RhoGDI2 overexpression also resulted in decreased expression of the epithelial markers E-cadherin and ß-catenin and increased expression of the mesenchymal markers vimentin and fibronectin. Importantly, RhoGDI2 overexpression also stimulated the expression of Snail, a repressor of E-cadherin and inducer of EMT, but not other family members such as Slug or Twist. RNA interference-mediated knockdown of Snail expression suppressed RhoGDI2-induced EMT and invasion, confirming that the effect was Snail-specific. These results indicate that RhoGDI2 plays a critical role in tumor progression in gastric cancer through induction of EMT. Targeting RhoGDI2 may thus be a useful strategy to inhibit gastric cancer cell invasion and metastasis.
Subject(s)
Cadherins/metabolism , Cell Movement , Cell Proliferation , Epithelial-Mesenchymal Transition , Stomach Neoplasms/pathology , Transcription Factors/metabolism , rho Guanine Nucleotide Dissociation Inhibitor beta/metabolism , Antineoplastic Agents/pharmacology , Apoptosis , Blotting, Western , Cadherins/genetics , Drug Resistance, Neoplasm , Fluorescent Antibody Technique , Humans , RNA, Messenger/genetics , RNA, Small Interfering/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Snail Family Transcription Factors , Stomach Neoplasms/drug therapy , Stomach Neoplasms/metabolism , Transcription Factors/antagonists & inhibitors , Transcription Factors/genetics , Tumor Cells, Cultured , rho Guanine Nucleotide Dissociation Inhibitor beta/geneticsABSTRACT
Rho GDP dissociation inhibitor 2 (RhoGDI2) was initially identified as a regulator of the Rho family of GTPases. Our recent works suggest that RhoGDI2 promotes tumor growth and malignant progression, as well as enhances chemoresistance in gastric cancer. Here, we delineate the mechanism by which RhoGDI2 promotes gastric cancer cell invasion and chemoresistance using two-dimensional gel electrophoresis (2-DE) on proteins derived from a RhoGDI2-overexpressing SNU-484 human gastric cancer cell line and control cells. Differentially expressed proteins were identified using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF-MS). In total, 47 differential protein spots were identified; 33 were upregulated, and 14 were downregulated by RhoGDI2 overexpression. Upregulation of SAE1, Cathepsin D, Cofilin1, CIAPIN1, and PAK2 proteins was validated by Western blot analysis. Loss-of-function analysis using small interference RNA (siRNA) directed against candidate genes reveals the need for CIAPIN1 and PAK2 in RhoGDI2-induced cancer cell invasion and Cathepsin D and PAK2 in RhoGDI2-mediated chemoresistance in gastric cancer cells. These data extend our understanding of the genes that act downstream of RhoGDI2 during the progression of gastric cancer and the acquisition of chemoresistance.
Subject(s)
Guanine Nucleotide Dissociation Inhibitors/metabolism , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathology , Cell Line, Tumor , Drug Resistance, Neoplasm , Electrophoresis, Gel, Two-Dimensional , Humans , Intracellular Signaling Peptides and Proteins/analysis , Intracellular Signaling Peptides and Proteins/metabolism , Neoplasm Metastasis , Proteome/analysis , Proteome/metabolism , Proteomics/methods , Reproducibility of Results , Stomach Neoplasms/drug therapy , Up-Regulation , p21-Activated Kinases/analysis , p21-Activated Kinases/metabolism , rho Guanine Nucleotide Dissociation Inhibitor beta , rho-Specific Guanine Nucleotide Dissociation InhibitorsABSTRACT
Rho GDP dissociation inhibitor 2 (RhoGDI2) is a regulator of the Rho family GTPases. Recent work from our laboratory suggests that RhoGDI2 expression potentially enhances resistance to cisplatin as well as promotes tumor growth and malignant progression in gastric cancer. In this study, we demonstrate that phospholipase C-gamma (PLCγ) is required for RhoGDI2-mediated cisplatin resistance and cancer cell invasion in gastric cancer. The levels of phosphorylated PLCγ are markedly enhanced in RhoGDI2-overexpressing SNU-484 cells and, by contrast, repressed in RhoGDI2-depleted MKN-28 cells. Depletion of PLCγ expression or inhibition of its activity not only significantly increases cisplatin-induced apoptosis but also suppresses the invasive ability of RhoGDI2-overexpressing SNU-484 cells. Taken together, our results suggest that PLCγ plays a key role in RhoGDI2-mediated cisplatin resistance and cell invasion in gastric cancer cells.
