Chemotactic Blebbing in Dictyostelium Cells.
Many researchers use the social amoeba Dictyostelium discoideum as a model organism to study various aspects of the eukaryotic cell chemotaxis. Traditionally, Dictyostelium chemotaxis is considered to be driven mainly by branched F-actin polymerization. However, recently it has become evident that Dictyostelium, as well as many other eukaryotic cells, can also employ intracellular hydrostatic pressure to generate force for migration. This process results in the projection of hemispherical plasma membrane protrusions, called blebs, that can be controlled by chemotactic signaling.Here we describe two methods to study chemotactic blebbing in Dictyostelium cells and to analyze the intensity of the blebbing response in various strains and under different conditions. The first of these methods-the cyclic-AMP shock assay-allows one to quantify the global blebbing response of cells to a uniform chemoattractant stimulation. The second one-the under-agarose migration assay-induces directional blebbing in cells moving in a gradient of chemoattractant. In this assay, the cells can be switched from a predominantly F-actin-driven mode of motility to a bleb-driven chemotaxis, allowing one to compare the efficiency of both modes and explore the molecular machinery controlling chemotactic blebbing.
Department of Biology, Stanford University, 337 Campus Drive, Stanford, CA, 94305, USA. evgeny@stanford.edu.
Evgeny
E
Zatulovskiy
MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus, Cambridge, CB2 0QH, UK.
Robert R
RR
Kay
D002478:Cells, Cultured; D002630:Chemotactic Factors / Q000378:metabolism; D002633:Chemotaxis; D000242:Cyclic AMP / Q000378:metabolism; D004023:Dictyostelium / Q000166:cytology* / Q000502:physiology*; D018613:Microscopy, Confocal; D059008:Time-Lapse Imaging
D002630:Chemotactic Factors; D000242:Cyclic AMP
3D migration; Agarose overlay; Bleb-driven motility; Blebbing; Chemotaxis; Cyclic-AMP shock; Dictyostelium