Rho Isoform-specific Interaction with IQGAP1 Promotes Breast Cancer Cell Proliferation and Migration
Copyright policy )Rho Isoform-specific Interaction with IQGAP1 Promotes Breast Cancer Cell Proliferation and Migration
We performed a proteomics screen for Rho isoform-specific binding proteins to clarify the tumor-promoting effects of RhoA and C that contrast with the tumor-suppressive effects of RhoB. We found that the IQ-motif-containing GTPase-activating protein IQGAP1 interacts directly with GTP-bound, prenylated RhoA and RhoC, but not with RhoB. Co-immunoprecipitation of IQGAP1 with endogenous RhoA/C was enhanced when RhoA/C were activated by epidermal growth factor (EGF) or transfection of a constitutively active guanine nucleotide exchange factor (GEF). Overexpression of IQGAP1 increased GTP-loading of RhoA/C, while siRNA-mediated depletion of IQGAP1 prevented endogenous RhoA/C activation by growth factors. IQGAP1 knockdown also reduced the amount of GTP bound to GTPase-deficient RhoA/C mutants, suggesting that IQGAP enhances Rho activation by GEF(s) or stabilizes Rho-GTP. IQGAP1 depletion in MDA-MB-231 breast cancer cells blocked EGF- and RhoA-induced stimulation of DNA synthesis. Infecting cells with adenovirus encoding constitutively active RhoA(L63) and measuring absolute amounts of RhoA-GTP in infected cells demonstrated that the lack of RhoA(L63)-induced DNA synthesis in IQGAP1-depleted cells was not due to reduced GTP-bound RhoA. These data suggested that IQGAP1 functions downstream of RhoA. Overexpression of IQGAP1 in MDA-MB-231 cells increased DNA synthesis irrespective of siRNA-mediated RhoA knockdown. Breast cancer cell motility was increased by expressing a constitutively-active RhoC(V14) mutant or overexpressing IQGAP1. EGF- or RhoC-induced migration required IQGAP1, but IQGAP1-stimulated migration independently of RhoC, placing IQGAP1 downstream of RhoC. We conclude that IQGAP1 acts both upstream of RhoA/C, regulating their activation state, and downstream of RhoA/C, mediating their effects on breast cancer cell proliferation and migration, respectively.
- University of California, San Diego United States
- University of California, San Diego United States
Proteomics, rho GTP-Binding Proteins, Blotting, Western, Protein Prenylation, Breast Neoplasms, Binding, Competitive, HEK293 Cells, Cell Movement, ras GTPase-Activating Proteins, rhoC GTP-Binding Protein, Cell Line, Tumor, Humans, Immunoprecipitation, Female, RNA Interference, Guanosine Triphosphate, rhoA GTP-Binding Protein, rhoB GTP-Binding Protein, Cell Proliferation, Protein Binding
Proteomics, rho GTP-Binding Proteins, Blotting, Western, Protein Prenylation, Breast Neoplasms, Binding, Competitive, HEK293 Cells, Cell Movement, ras GTPase-Activating Proteins, rhoC GTP-Binding Protein, Cell Line, Tumor, Humans, Immunoprecipitation, Female, RNA Interference, Guanosine Triphosphate, rhoA GTP-Binding Protein, rhoB GTP-Binding Protein, Cell Proliferation, Protein Binding
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