Your browser doesn't support javascript.
loading
Phagosome-lysosome fusion is a calcium-independent event in macrophages.
Zimmerli, S; Majeed, M; Gustavsson, M; Stendahl, O; Sanan, D A; Ernst, J D.
Afiliação
  • Zimmerli S; Division of Infectious Diseases, San Francisco General Hospital, University of California at San Francisco 94143-0868, USA.
J Cell Biol ; 132(1-2): 49-61, 1996 Jan.
Article em En | MEDLINE | ID: mdl-8567729
Phagosome-lysosome membrane fusion is a highly regulated event that is essential for intracellular killing of microorganisms. Functionally, it represents a form of polarized regulated secretion, which is classically dependent on increases in intracellular ionized calcium ([Ca2+]i). Indeed, increases in [Ca2+]i are essential for phagosome-granule (lysosome) fusion in neutrophils and for lysosomal fusion events that mediate host cell invasion by Trypanosoma cruzi trypomastigotes. Since several intracellular pathogens survive in macrophage phagosomes that do not fuse with lysosomes, we examined the regulation of phagosome-lysosome fusion in macrophages. Macrophages (M phi) were treated with 12.5 microM bis-(2-amino-S-methylphenoxy) ethane-N,N,N',N',-tetraacetic acid tetraacetoxymethyl ester (MAPT/AM), a cell-permeant calcium chelator which reduced resting cytoplasmic [Ca2+]; from 80 nM to < or = 20 nM and completely blocked increases in [Ca2+]i in response to multiple stimuli, even in the presence of extracellular calcium. Subsequently, M phi phagocytosed serum-opsonized zymosan, staphylococci, or Mycobacterium bovis. Microbes were enumerated by 4',6-diamidino-2-phenylindole, dihydrochloride (DAPI) staining, and phagosome-lysosome fusion was scored using both lysosome-associated membrane protein (LAMP-1) as a membrane marker and rhodamine dextran as a content marker for lysosomes. Confirmation of phagosome-lysosome fusion by electron microscopy validated the fluorescence microscopy findings. We found that phagosome-lysosome fusion in M phi occurs noramlly at very low [Ca2+]i (< or = 20 nM). Kinetic analysis showed that in M phi none of the steps leading from particle binding to eventual phagosome-lysosome fusion are regulated by [Ca2+]i in a rate-limiting way. Furthermore, confocal microscopy revealed no difference in the intensity of LAMP-1 immunofluorescence in phagolysosome membranes in calcium-buffered vs. control macrophages. We conclude that neither membrane recognition nor fusion events in the phagosomal pathway in macrophages are dependent on or regulated by calcium.
Assuntos

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Fagocitose / Fagossomos / Cálcio / Lisossomos / Macrófagos / Fusão de Membrana Limite: Animals / Humans Idioma: En Ano de publicação: 1996 Tipo de documento: Article

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Fagocitose / Fagossomos / Cálcio / Lisossomos / Macrófagos / Fusão de Membrana Limite: Animals / Humans Idioma: En Ano de publicação: 1996 Tipo de documento: Article