An In Vitro System to Analyze Generation and Degradation of Phagosomal Phosphatidylinositol Phosphates.
FYVE domain
Lipid phosphatases
Phagocytes
Phagocytosis
Phagosome
Phosphatidylinositol phosphates
Phosphoinositide detection
Phosphoinositide kinases
Phosphoinositide metabolism
Journal
Methods in molecular biology (Clifton, N.J.)
ISSN: 1940-6029
Titre abrégé: Methods Mol Biol
Pays: United States
ID NLM: 9214969
Informations de publication
Date de publication:
2023
2023
Historique:
medline:
28
6
2023
pubmed:
27
6
2023
entrez:
26
6
2023
Statut:
ppublish
Résumé
Phagosomes are formed when phagocytic cells take up large particles, and they develop into phagolysosomes where the particles are degraded. The transformation of nascent phagosomes into phagolysosomes is a complex multi-step process, and the precise timing of these steps depends at least in part on phosphatidylinositol phosphates (PIPs). Some such-called "intracellular pathogens" are not delivered to microbicidal phagolysosomes and manipulate the PIP composition of the phagosomes they reside in. Studying the dynamic changes of the PIP composition of inert-particle phagosomes will help to understand why the pathogens' manipulations reprogram phagosome maturation.We here describe a method to detect and to follow generation and degradation of PIPs on purified phagosomes. To this end, phagosomes formed around inert latex beads are purified from J774E macrophages and incubated in vitro with PIP-binding protein domains or PIP-binding antibodies. Binding of such PIP sensors to phagosomes indicates presence of the cognate PIP and is quantified by immunofluorescence microscopy. When phagosomes are incubated with PIP sensors and ATP at a physiological temperature, the generation and degradation of PIPs can be followed, and PIP-metabolizing enzymes can be identified using specific inhibitory agents.
Identifiants
pubmed: 37365474
doi: 10.1007/978-1-0716-3338-0_18
doi:
Substances chimiques
Phosphatidylinositol Phosphates
0
Antibodies
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
261-274Informations de copyright
© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.
Références
Fountain A, Inpanathan S, Alves P et al (2021) Phagosome maturation in macrophages: eat, digest, adapt, and repeat. Adv Biol Regul 82:100832. https://doi.org/10.1016/j.jbior.2021.100832
doi: 10.1016/j.jbior.2021.100832
pubmed: 34717137
Haas A (2007) The phagosome: compartment with a license to kill. Traffic 8:311–330. https://doi.org/10.1111/j.1600-0854.2006.00531.x
doi: 10.1111/j.1600-0854.2006.00531.x
pubmed: 17274798
Desjardins M, Huber LA, Parton RG et al (1994) Biogenesis of phagolysosomes proceeds through a sequential series of interactions with the endocytic apparatus. J Cell Biol 124:677–688. https://doi.org/10.1083/jcb.124.5.677
doi: 10.1083/jcb.124.5.677
pubmed: 8120091
Lancaster CE, Fountain A, Dayam RM et al (2021) Phagosome resolution regenerates lysosomes and maintains the degradative capacity in phagocytes. J Cell Biol 220. https://doi.org/10.1083/jcb.202005072
Levin-Konigsberg R, Montaño-Rendón F, Keren-Kaplan T et al (2019) Phagolysosome resolution requires contacts with the endoplasmic reticulum and phosphatidylinositol-4-phosphate signalling. Nat Cell Biol 21:1234–1247. https://doi.org/10.1038/s41556-019-0394-2
doi: 10.1038/s41556-019-0394-2
pubmed: 31570833
pmcid: 8340083
Pizarro-Cerdá J, Cossart P (2004) Subversion of phosphoinositide metabolism by intracellular bacterial pathogens. Nat Cell Biol 6:1026–1033. https://doi.org/10.1038/ncb1104-1026
doi: 10.1038/ncb1104-1026
pubmed: 15516995
Weber SS, Ragaz C, Hilbi H (2009) Pathogen trafficking pathways and host phosphoinositide metabolism. Mol Microbiol 71:1341–1352. https://doi.org/10.1111/j.1365-2958.2009.06608.x
doi: 10.1111/j.1365-2958.2009.06608.x
pubmed: 19208094
Posor Y, Jang W, Haucke V (2022) Phosphoinositides as membrane organizers. Nat Rev Mol Cell Biol:1–20. https://doi.org/10.1038/s41580-022-00490-x
Balla A, Balla T (2006) Phosphatidylinositol 4-kinases: old enzymes with emerging functions. Trends Cell Biol 16:351–361. https://doi.org/10.1016/j.tcb.2006.05.003
doi: 10.1016/j.tcb.2006.05.003
pubmed: 16793271
de Matteis MA, Godi A (2004) PI-loting membrane traffic. Nat Cell Biol 6:487–492. https://doi.org/10.1038/ncb0604-487
doi: 10.1038/ncb0604-487
pubmed: 15170460
Rusten TE, Stenmark H (2006) Analyzing phosphoinositides and their interacting proteins. Nat Methods 3:251–258. https://doi.org/10.1038/nmeth867
doi: 10.1038/nmeth867
pubmed: 16554828
Kutateladze TG (2010) Translation of the phosphoinositide code by PI effectors. Nat Chem Biol 6:507–513. https://doi.org/10.1038/nchembio.390
doi: 10.1038/nchembio.390
pubmed: 20559318
pmcid: 3182472
Levin R, Grinstein S, Schlam D (2015) Phosphoinositides in phagocytosis and macropinocytosis. Biochim Biophys Acta 1851:805–823. https://doi.org/10.1016/j.bbalip.2014.09.005
doi: 10.1016/j.bbalip.2014.09.005
pubmed: 25238964
Vieira OV, Harrison RE, Scott CC et al (2004) Acquisition of Hrs, an essential component of phagosomal maturation, is impaired by mycobacteria. Mol Cell Biol 24:4593–4604. https://doi.org/10.1128/MCB.24.10.4593-4604.2004
doi: 10.1128/MCB.24.10.4593-4604.2004
pubmed: 15121875
pmcid: 400451
Jeschke A, Zehethofer N, Lindner B et al (2015) Phosphatidylinositol 4-phosphate and phosphatidylinositol 3-phosphate regulate phagolysosome biogenesis. Proc Natl Acad Sci U S A 112:4636–4641. https://doi.org/10.1073/pnas.1423456112
doi: 10.1073/pnas.1423456112
pubmed: 25825728
pmcid: 4403170
Defacque H, Bos E, Garvalov B et al (2002) Phosphoinositides regulate membrane-dependent actin assembly by latex bead phagosomes. Mol Biol Cell 13:1190–1202. https://doi.org/10.1091/mbc.01-06-0314
doi: 10.1091/mbc.01-06-0314
pubmed: 11950931
pmcid: 102261
Gillooly DJ, Morrow IC, Lindsay M et al (2000) Localization of phosphatidylinositol 3-phosphate in yeast and mammalian cells. EMBO J 19:4577–4588. https://doi.org/10.1093/emboj/19.17.4577
doi: 10.1093/emboj/19.17.4577
pubmed: 10970851
pmcid: 302054
Dowler S, Currie RA, Campbell DG et al (2000) Identification of pleckstrin-homology-domain-containing proteins with novel phosphoinositide-binding specificities. Biochem J 351:19–31. https://doi.org/10.1042/0264-6021:3510019
doi: 10.1042/0264-6021:3510019
pubmed: 11001876
pmcid: 1221362
Fiani ML, Beitz J, Turvy D et al (1998) Regulation of mannose receptor synthesis and turnover in mouse J774 macrophages. J Leukoc Biol 64:85–91. https://doi.org/10.1002/jlb.64.1.85
doi: 10.1002/jlb.64.1.85
pubmed: 9665280
Siddhanta U, McIlroy J, Shah A et al (1998) Distinct roles for the p110alpha and hVPS34 phosphatidylinositol 3′-kinases in vesicular trafficking, regulation of the actin cytoskeleton, and mitogenesis. J Cell Biol 143:1647–1659. https://doi.org/10.1083/jcb.143.6.1647
doi: 10.1083/jcb.143.6.1647
pubmed: 9852157
pmcid: 2132989
Ronan B, Flamand O, Vescovi L et al (2014) A highly potent and selective Vps34 inhibitor alters vesicle trafficking and autophagy. Nat Chem Biol 10:1013–1019. https://doi.org/10.1038/nchembio.1681
doi: 10.1038/nchembio.1681
pubmed: 25326666
Bago R, Malik N, Munson MJ et al (2014) Characterization of VPS34-IN1, a selective inhibitor of Vps34, reveals that the phosphatidylinositol 3-phosphate-binding SGK3 protein kinase is a downstream target of class III phosphoinositide 3-kinase. Biochem J 463:413–427. https://doi.org/10.1042/BJ20140889
doi: 10.1042/BJ20140889
pubmed: 25177796
Endemann GC, Graziani A, Cantley LC (1991) A monoclonal antibody distinguishes two types of phosphatidylinositol 4-kinase. Biochem J 273(Pt 1):63–66. https://doi.org/10.1042/bj2730063
doi: 10.1042/bj2730063
pmcid: 1149879