Abstract
The Wnt-induced planar cell polarity (PCP) signaling pathway is essential for polarized cell migration and morphogenesis. Dishevelled (Dvl) and its binding protein Daam1 mediate RhoA activation in this pathway. WGEF, a member of the Rho-guanine nucleotide exchange factor (Rho-GEF) family, was shown to play a role in Wnt-induced RhoA activation in Xenopus embryos. However, it has remained unknown which member(s) of a Rho-GEF family are involved in Wnt/Dvl-induced RhoA activation in mammalian cells. Here we identified p114-RhoGEF and Lfc (also called GEF-H1) as the Rho-GEFs responsible for Wnt-3a-induced RhoA activation in N1E-115 mouse neuroblastoma cells. We screened for Rho-GEF-silencing short-hairpin RNAs (shRNAs) that are capable of suppressing Dvl-induced neurite retraction in N1E-115 cells and found that p114-RhoGEF and Lfc shRNAs, but not WGEF shRNA, suppressed Dvl- and Wnt-3a-induced neurite retraction. p114-RhoGEF and Lfc shRNAs also inhibited Dvl- and Wnt-3a-induced RhoA activation, and p114-RhoGEF and Lfc proteins were capable of binding to Dvl and Daam1. Additionally, the Dvl-binding domains of p114-RhoGEF and Lfc inhibited Dvl-induced neurite retraction. Our results suggest that p114-RhoGEF and Lfc are critically involved in Wnt-3a- and Dvl-induced RhoA activation and neurite retraction in N1E-115 cells.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adaptor Proteins, Signal Transducing / metabolism*
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Animals
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Cell Line, Tumor
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Dishevelled Proteins
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Enzyme Activation / drug effects
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Gene Expression Profiling
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Gene Expression Regulation, Neoplastic / drug effects
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Gene Knockdown Techniques
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Guanine Nucleotide Exchange Factors / metabolism*
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Humans
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Lysophospholipids / pharmacology
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Mice
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Microfilament Proteins / metabolism
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Neurites / enzymology*
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Neuroblastoma / enzymology*
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Neuroblastoma / genetics
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Neuroblastoma / pathology
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Phosphoproteins / metabolism*
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Protein Binding / drug effects
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Protein Interaction Mapping
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Proto-Oncogene Proteins / metabolism*
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RNA, Small Interfering / metabolism
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Rho Guanine Nucleotide Exchange Factors
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Wnt Proteins / metabolism*
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Wnt3 Protein
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Wnt3A Protein
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Xenopus Proteins
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rho GTP-Binding Proteins / metabolism
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rhoA GTP-Binding Protein / metabolism*
Substances
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Adaptor Proteins, Signal Transducing
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Arhgef2 protein, mouse
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DVL1 protein, Xenopus
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Dishevelled Proteins
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GEF-H1 protein, Xenopus
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Guanine Nucleotide Exchange Factors
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Lysophospholipids
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Microfilament Proteins
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Phosphoproteins
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Proto-Oncogene Proteins
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RNA, Small Interfering
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Rho Guanine Nucleotide Exchange Factors
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WNT3A protein, Xenopus
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WNT3A protein, human
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Wnt Proteins
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Wnt3 Protein
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Wnt3A Protein
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Wnt3a protein, mouse
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Xenopus Proteins
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Daam1 protein, mouse
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rho GTP-Binding Proteins
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rhoA GTP-Binding Protein
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lysophosphatidic acid