Vacuolar ATPase in phagosome-lysosome fusion

Sandra Kissing, Christina Hermsen, Urska Repnik, Cecilie Kåsi Nesset, Kristine Von Bargen, Gareth Griffiths, Atsuhiro Ichihara, Beth S. Lee, Michael Schwake, Jef De Brabander, Albert Haas, Paul Saftig

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Abstract

The vacuolar H<sup>+</sup>-ATPase (v-ATPase) complex is instrumental in establishing and maintaining acidification of some cellular compartments, thereby ensuring their functionality. Recently it has been proposed that the transmembrane V<inf>0</inf> sector of v-ATPase and its a-subunits promote membrane fusion in the endocytic and exocytic pathways independent of their acidification functions. Here, we tested if such a proton-pumping independent role of v-ATPase also applies to phagosome-lysosome fusion. Surprisingly, endo(lyso)somes in mouse embryonic fibroblasts lacking the V<inf>0</inf> a3 subunit of the v-ATPase acidified normally, and endosome and lysosome marker proteins were recruited to phagosomes with similar kinetics in the presence or absence of the a3 subunit. Further experiments used macrophages with a knockdown of v-ATPase accessory protein 2 (ATP6AP2) expression, resulting in a strongly reduced level of the V<inf>0</inf> sector of the v-ATPase. However, acidification appeared undisturbed, and fusion between latex bead-containing phagosomes and lysosomes, as analyzed by electron microscopy, was even slightly enhanced, as was killing of non-pathogenic bacteria by V<inf>0</inf> mutant macrophages. Pharmacologically neutralized lysosome pH did not affect maturation of phagosomes in mouse embryonic cells or macrophages. Finally, locking the two large parts of the v-ATPase complex together by the drug saliphenylhalamide A did not inhibit in vitro and in cellulo fusion of phagosomes with lysosomes. Hence, our data do not suggest a fusion-promoting role of the v-ATPase in the formation of phagolysosomes.

Original languageEnglish (US)
Pages (from-to)14166-14180
Number of pages15
JournalJournal of Biological Chemistry
Volume290
Issue number22
DOIs
StatePublished - May 29 2015

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ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Kissing, S., Hermsen, C., Repnik, U., Nesset, C. K., Von Bargen, K., Griffiths, G., Ichihara, A., Lee, B. S., Schwake, M., De Brabander, J., Haas, A., & Saftig, P. (2015). Vacuolar ATPase in phagosome-lysosome fusion. Journal of Biological Chemistry, 290(22), 14166-14180. https://doi.org/10.1074/jbc.M114.628891