Assessing the Phagosome Proteome by Quantitative Mass Spectrometry.
Phagocytosis is the process that engulfs particles in vesicles called phagosomes that are trafficked through a series of maturation steps, culminating in the destruction of the internalized cargo. Because phagosomes are in direct contact with the particle and undergo constant fusion and fission events with other organelles, characterization of the phagosomal proteome is a powerful tool to understand mechanisms controlling innate immunity as well as vesicle trafficking. The ability to isolate highly pure phagosomes through the use of latex beads led to an extensive use of proteomics to study phagosomes under different stimuli. Thousands of different proteins have been identified and quantified, revealing new properties and shedding new light on the dynamics and composition of maturing phagosomes and innate immunity mechanisms. In this chapter, we describe how quantitative-based proteomic methods such as label-free, dimethyl labeling or Tandem Mass Tag (TMT) labeling can be applied for the characterization of protein composition and translocation during maturation of phagosomes in macrophages.
MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dundee, DD1 5EH, UK.
Julien
J
Peltier
MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dundee, DD1 5EH, UK.
Anetta
A
Härtlova
MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dundee, DD1 5EH, UK. m.trost@dundee.ac.uk.
Matthias
M
Trost
D000818:Animals; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D010588:Phagosomes / Q000378:metabolism*; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics; D011786:Quality Control; D013194:Staining and Labeling; D013223:Statistics as Topic
D020543:Proteome
Dimethyl; Isotope labeling; Label free; Macrophages; Mass spectrometry; Maturation of phagosomes; Phagosome dynamics; Quantitative proteomics; TMT