Assessment of development and chemotaxis in dictyostelium discoideum mutants

Yulia Artemenko, Kristen F. Swaney, Peter N. Devreotes

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

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 LimE Δcoil or PIP3-binding PH domains to the plasma membrane in response to cAMP stimulation.

Original languageEnglish (US)
Title of host publicationCell Migration
Subtitle of host publicationDevelopmental Methods and Protocols
EditorsClaire M. Wells, Maddy Parsons
Pages287-309
Number of pages23
DOIs
StatePublished - Aug 3 2011

Publication series

NameMethods in Molecular Biology
Volume769
ISSN (Print)1064-3745

Keywords

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

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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  • Cite this

    Artemenko, Y., Swaney, K. F., & Devreotes, P. N. (2011). Assessment of development and chemotaxis in dictyostelium discoideum mutants. In C. M. Wells, & M. Parsons (Eds.), Cell Migration: Developmental Methods and Protocols (pp. 287-309). (Methods in Molecular Biology; Vol. 769). https://doi.org/10.1007/978-1-61779-207-6_20