Abstract
Signaling proteins are tightly regulated spatially and temporally to perform multiple functions. For Cdc42 and other guanosine triphosphatases, the subcellular location of activation is a critical determinant of cell behavior. However, current approaches are limited in their ability to examine the dynamics of Cdc42 activity in living cells. We report the development of a biosensor capable of visualizing the changing activation of endogenous, unlabeled Cdc42 in living cells. With the use of a dye that reports protein interactions, the biosensor revealed localized activation in the trans-Golgi apparatus, microtubule-dependent Cdc42 activation at the cell periphery, and activation kinetics precisely coordinated with cell extension and retraction.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Actins / metabolism
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Algorithms
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Animals
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Biosensing Techniques*
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Cell Adhesion
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Cell Line
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Cell Membrane / metabolism*
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Cell Polarity
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Cell Surface Extensions / metabolism
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Cell Surface Extensions / ultrastructure
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Endothelial Cells / metabolism
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Endothelial Cells / ultrastructure
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Fibroblasts
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Fluorescence
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Fluorescent Dyes / chemistry
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Fluorescent Dyes / metabolism
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Green Fluorescent Proteins
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Humans
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Luminescent Proteins
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Mice
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Microtubules / metabolism
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Neutrophil Activation
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Neutrophils / metabolism*
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Proteins / chemistry
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Proteins / metabolism
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Pseudopodia / metabolism
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Pyrimidinones / metabolism
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Sensitivity and Specificity
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Wiskott-Aldrich Syndrome Protein
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cdc42 GTP-Binding Protein / metabolism*
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rho GTP-Binding Proteins / metabolism
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trans-Golgi Network / metabolism*
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trans-Golgi Network / ultrastructure
Substances
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Actins
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Fluorescent Dyes
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Luminescent Proteins
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Proteins
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Pyrimidinones
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WAS protein, human
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Was protein, mouse
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Wiskott-Aldrich Syndrome Protein
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Green Fluorescent Proteins
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merocyanine dye
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RHOQ protein, human
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Rhoq protein, mouse
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cdc42 GTP-Binding Protein
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rho GTP-Binding Proteins