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 publicationMethods in Molecular Biology
Pages287-309
Number of pages23
Volume769
DOIs
StatePublished - 2011
Externally publishedYes

Publication series

NameMethods in Molecular Biology
Volume769
ISSN (Print)10643745

Fingerprint

Dictyostelium
Chemotaxis
Cyclic AMP
Amoeba
Population
Agar
Actins
Cell Membrane
Phenotype
Enzymes

Keywords

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

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

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

Assessment of development and chemotaxis in dictyostelium discoideum mutants. / Artemenko, Yulia; Swaney, Kristen F.; Devreotes, Peter N.

Methods in Molecular Biology. Vol. 769 2011. p. 287-309 (Methods in Molecular Biology; Vol. 769).

Research output: Chapter in Book/Report/Conference proceedingChapter

Artemenko, Y, Swaney, KF & Devreotes, PN 2011, Assessment of development and chemotaxis in dictyostelium discoideum mutants. in Methods in Molecular Biology. vol. 769, Methods in Molecular Biology, vol. 769, pp. 287-309. https://doi.org/10.1007/978-1-61779-207-6_20
Artemenko Y, Swaney KF, Devreotes PN. Assessment of development and chemotaxis in dictyostelium discoideum mutants. In Methods in Molecular Biology. Vol. 769. 2011. p. 287-309. (Methods in Molecular Biology). https://doi.org/10.1007/978-1-61779-207-6_20
Artemenko, Yulia ; Swaney, Kristen F. ; Devreotes, Peter N. / Assessment of development and chemotaxis in dictyostelium discoideum mutants. Methods in Molecular Biology. Vol. 769 2011. pp. 287-309 (Methods in Molecular Biology).
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