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Uniform cAMP Stimulation of Dictyostelium Cells Induces Localized Patches of Signal Transduction and Pseudopodia

By Marten Postma, Jeroen Roelofs, Joachim Goedhart, Theodorus W.J. Gadella, Antonie J.W.G. Visser and Peter J.M. Van Haastert

Abstract

The chemoattractant cAMP induces the translocation of cytosolic PHCrac-GFP to the plasma membrane. PHCrac-GFP is a green fluorescent protein fused to a PH domain that presumably binds to phosphatydylinositol polyphosphates in the membrane. We determined the relative concentration of PHCrac-GFP in the cytosol and at different places along the cell boundary. In cells stimulated homogeneously with 1μM cAMP we observed two distinct phases of PHCrac-GFP translocation. The first translocation is transient and occurs to nearly the entire boundary of the cell; the response is maximal at 6-8 s after stimulation and disappears after ∼20 s. A second translocation of PHCrac-GFP starts after ∼30 s and persists as long as cAMP remains present. Translocation during this second response occurs to small patches with radius of ∼4-5 μm, each covering ∼10% of the cell surface. Membrane patches of PHCrac-GFP are both temporally and spatially closely associated with pseudopodia, which are extended at ∼10 s from the area with a PHCrac-GFP patch. These signaling patches in pseudopodia of homogeneously stimulated cells resemble the single patch of PHCrac-GFP at the leading edge of a cell in a gradient of cAMP, suggesting that PHCrac-GFP is a spatial cue for pseudopod formation also in uniform cAMP

Topics: Articles
Publisher: The American Society for Cell Biology
Year: 2003
DOI identifier: 10.1091/mbc.E03-08-0566
OAI identifier: oai:pubmedcentral.nih.gov:284803
Provided by: PubMed Central
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