title: Monitoring time-dependent maturation changes in purified phagosomes from Dictyostelium discoideum.
abstract: The amoeba Dictyostelium discoideum is an established model to study phagocytosis. The sequence of events leading to the internalization and degradation of a particle is conserved in D. discoideum compared to metazoan cells. As its small haploid genome has been sequenced, it is now amenable to genome-wide analysis including organelle proteomics. Therefore, we adapted to Dictyostelium the classical protocol to purify phagosomes formed by ingestion of latex beads particles. The pulse-chase protocol detailed here gives easy access to pure, intact, and synchronized phagosomes from representative stages of the entire process of phagosome maturation. Recently, this protocol was used to generate individual temporal profiles of proteins and lipids during phagosome maturation generating a proteomic fingerprint of six maturation stages (1). In addition, immunolabeling of phagosomes on a coverslip was developed to visualize and quantitate antigen distribution at the level of individual phagosomes.
authors.affiliation: Department of Biochemistry, University of Geneva, Geneva, Switzerland.
authors.forename: Régis
authors.initials: R
authors.lastname: Dieckmann
authors.forename: Navin
authors.initials: N
authors.lastname: Gopaldass
authors.forename: Caroline
authors.initials: C
authors.lastname: Escalera
authors.forename: Thierry
authors.initials: T
authors.lastname: Soldati
mesh_terms: D000818:Animals; D004023:Dictyostelium / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D010587:Phagocytosis / Q000502:physiology; D010588:Phagosomes / Q000378:metabolism*; D013997:Time Factors