Abstract
The phagosome is a redox-active organelle. Numerous reductive and oxidative systems play both direct and indirect roles in phagosomal function. With the advent of newer methodologies to study these redox events in live cells, the details of how redox conditions change within the maturing phagosome, how they are regulated, and how they influence other phagosomal functions can be investigated. In this chapter, we detail phagosome-specific, fluorescence-based assays that measure disulfide reduction and the production of reactive oxygen species in live phagocytes such as macrophages and dendritic cells, in real time.
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Yates RM (2013) Redox considerations in the phagosome: current concepts, controversies, and future challenges. Antioxid Redox Signal 18:628–629
Collins DS, Unanue ER, Harding CV (1991) Reduction of disulfide bonds within lysosomes is a key step in antigen processing. J Immunol 147:4054–4059
Phan UT, Arunachalam B, Cresswell P (2000) Gamma-interferon-inducible lysosomal thiol reductase (GILT). Maturation, activity, and mechanism of action. J Biol Chem 275:25907–25914
Balce DR, Allan ER, McKenna N et al (2014) Gamma-interferon-inducible lysosomal thiol reductase (GILT) maintains phagosomal proteolysis in alternatively activated macrophages. J Biol Chem 289:31891–31904
Allan ER, Tailor P, Balce DR et al (2014) NADPH oxidase modifies patterns of MHC class II-restricted epitopic repertoires through redox control of antigen processing. J Immunol 192:4989–5001
Balce DR, Yates RM (2013) Redox-sensitive probes for the measurement of redox chemistries within phagosomes of macrophages and dendritic cells. Redox Biol 1:467–474
Yates RM, Russell DG (2008) Real-time spectrofluorometric assays for the lumenal environment of the maturing phagosome. Methods Mol Biol 445:311–325
Balce DR, Li B, Allan ER et al (2011) Alternative activation of macrophages by IL-4 enhances the proteolytic capacity of their phagosomes through synergistic mechanisms. Blood 118:4199–4208
Balce DR, Greene CJ, Tailor P et al (2015) Endogenous and exogenous pathways maintain the reductive capacity of the phagosome. J Leukocyte Biol 100(1):17–26
VanderVen BC, Yates RM, Russell DG (2009) Intraphagosomal measurement of the magnitude and duration of the oxidative burst. Traffic 10:372–378
Rybicka JM, Balce DR, Khan MF et al (2010) NADPH oxidase activity controls phagosomal proteolysis in macrophages through modulation of the lumenal redox environment of phagosomes. Proc Natl Acad Sci U S A 107:10496–10501
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Balce, D.R., Yates, R.M. (2017). Fluorometric Approaches to Measuring Reductive and Oxidative Events in Phagosomes. In: Botelho, R. (eds) Phagocytosis and Phagosomes. Methods in Molecular Biology, vol 1519. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6581-6_14
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DOI: https://doi.org/10.1007/978-1-4939-6581-6_14
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Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-6579-3
Online ISBN: 978-1-4939-6581-6
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