Cell Migration

Volume 769 of the series Methods in Molecular Biology pp 287-309


Assessment of Development and Chemotaxis in Dictyostelium discoideum Mutants

  • Yulia Artemenko
  • , Kristen F. Swaney
  • , Peter N. DevreotesAffiliated withDepartment of Cell Biology, Johns Hopkins University School of Medicine Email author 

* Final gross prices may vary according to local VAT.

Get Access


Studies using the social amoeba Dictyostelium discoideum have greatly contributed to the current understanding of the signaling network that underlies chemotaxis. Since directed migration is essential for normal D. discoideum multicellular development, mutants with chemotactic impairments are likely to have abnormal developmental morphologies. We have used multicellular development as a readout in a screen of mutants to identify new potential regulators of chemotaxis. In this chapter, we describe how mutants generated by restriction enzyme-mediated integration (REMI) are analyzed, from assessment of development to detailed characterization of 3′,5′-cyclic adenosine monophosphate (cAMP)-induced responses. Two complementary approaches, plating cells either clonally on a bacterial lawn or as a population on non-nutrient agar, are used to evaluate multicellular development. Once mutants with aberrant developmental phenotypes are identified, their chemotaxis toward cAMP is assessed by both small population and micropipette assays. Furthermore, mutants are tested for defects in both general and specific signaling pathways by examining the recruitment of actin-binding LimEΔcoil or PIP3-binding PH domains to the plasma membrane in response to cAMP stimulation.

Key words

Multicellular development Differentiation Under buffer assay cAR1 Small population assay Micropipette assay CMFDA labeling PIP3 dynamics Actin