Uniform cAMP stimulation of Dictyostelium cells induces localized patches of signal transduction and pseudopodia

M. Postma, J. Roelofs, J. Goedhart, T.W.J. Gadella, A.J.W.G. Visser, P.J.M. van Haastert

Research output: Contribution to journalArticleAcademicpeer-review

79 Citations (Scopus)

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 1 muM cAMP we observed two distinct phases of PHCrap-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 similar to20 s. A second translocation of PHCrac-GFP starts after similar to30 s and persists as long as cAMP remains present. Translocation during this second response occurs to small patches with radius of similar to4-5 mum, each covering similar to10% of the cell surface. Membrane patches of PHCrac-GFP are both temporally and spatially closely associated with pseudopodia, which are extended at similar to10 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.
Original languageEnglish
Pages (from-to)5019-5027
JournalMolecular Biology of the Cell
Volume14
DOIs
Publication statusPublished - 2003

Keywords

  • living cells
  • chemotaxis
  • discoideum
  • activation
  • adaptation
  • polarity
  • protein
  • relay

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