Rapid escape of the dot/icm mutants of Legionella pneumophila into the cytosol of mammalian and protozoan cells

Maëlle Molmeret, Marina Santic', Rexford Asare, Reynold A. Carabeo, Yousef Abu Kwaik

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

The Legionella pneumophila-containing phagosome evades endocytic fusion and intercepts endoplasmic reticulum (ER)-to-Golgi vesicle traffic, which is believed to be mediated by the Dot/Icm type IV secretion system. Although phagosomes harboring dot/icm mutants are thought to mature through the endosomal-lysosomal pathway, colocalization studies with lysosomal markers have reported contradictory results. In addition, phagosomes harboring the dot/icm mutants do not interact with endocytosed materials, which is inconsistent with maturation of the phagosomes in the endosomal-lysosomal pathway. Using multiple strategies, we show that the dot/icm mutants defective in the Dot/Icm structural apparatus are unable to maintain the integrity of their phagosomes and escape into the cytoplasm within minutes of entry into various mammalian and protozoan cells in a process independent of the type II secretion system. In contrast, mutants defective in cytoplasmic chaperones of Dot/Icm effectors and rpoS, letA/S, and letE regulatory mutants are all localized within intact phagosomes. Importantly, non-dot/icm L. pneumophila mutants whose phagosomes acquire late endosomal-lysosomal markers are all located within intact phagosomes. Using high-resolution electron microscopy, we show that phagosomes harboring the dot/icm transporter mutants do not fuse to lysosomes but are free in the cytoplasm. Inhibition of ER-to-Golgi vesicle traffic by brefeldin A does not affect the integrity of the phagosomes harboring the parental strain of L. pneumophila. We conclude that the Dot/Icm transporter is involved in maintaining the integrity of the L. pneumophila phagosome, independent of interception of ER-to-Golgi vesicle traffic, which is a novel function of type IV secretion systems.
Original languageEnglish
Pages (from-to)3290-3304
Number of pages15
JournalInfection and Immunity
Volume75
Issue number7
Early online date16 Apr 2007
DOIs
Publication statusPublished - Jul 2007

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Legionella pneumophila
Phagosomes
Cytosol
Endoplasmic Reticulum
Cytoplasm
Brefeldin A
Endocytosis
Lysosomes
Electron Microscopy

Keywords

  • Macrophages
  • Microscopy, Confocal
  • Animals
  • Legionella pneumophila
  • Carrier Proteins
  • Humans
  • Phagosomes
  • Membrane Proteins
  • Acanthamoeba
  • Bacterial Proteins
  • Epithelial Cells
  • Cytosol
  • Time Factors
  • Mutation
  • U937 Cells

Cite this

Rapid escape of the dot/icm mutants of Legionella pneumophila into the cytosol of mammalian and protozoan cells. / Molmeret, Maëlle; Santic', Marina; Asare, Rexford; Carabeo, Reynold A. ; Abu Kwaik, Yousef.

In: Infection and Immunity, Vol. 75, No. 7, 07.2007, p. 3290-3304.

Research output: Contribution to journalArticle

Molmeret, Maëlle ; Santic', Marina ; Asare, Rexford ; Carabeo, Reynold A. ; Abu Kwaik, Yousef. / Rapid escape of the dot/icm mutants of Legionella pneumophila into the cytosol of mammalian and protozoan cells. In: Infection and Immunity. 2007 ; Vol. 75, No. 7. pp. 3290-3304.
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abstract = "The Legionella pneumophila-containing phagosome evades endocytic fusion and intercepts endoplasmic reticulum (ER)-to-Golgi vesicle traffic, which is believed to be mediated by the Dot/Icm type IV secretion system. Although phagosomes harboring dot/icm mutants are thought to mature through the endosomal-lysosomal pathway, colocalization studies with lysosomal markers have reported contradictory results. In addition, phagosomes harboring the dot/icm mutants do not interact with endocytosed materials, which is inconsistent with maturation of the phagosomes in the endosomal-lysosomal pathway. Using multiple strategies, we show that the dot/icm mutants defective in the Dot/Icm structural apparatus are unable to maintain the integrity of their phagosomes and escape into the cytoplasm within minutes of entry into various mammalian and protozoan cells in a process independent of the type II secretion system. In contrast, mutants defective in cytoplasmic chaperones of Dot/Icm effectors and rpoS, letA/S, and letE regulatory mutants are all localized within intact phagosomes. Importantly, non-dot/icm L. pneumophila mutants whose phagosomes acquire late endosomal-lysosomal markers are all located within intact phagosomes. Using high-resolution electron microscopy, we show that phagosomes harboring the dot/icm transporter mutants do not fuse to lysosomes but are free in the cytoplasm. Inhibition of ER-to-Golgi vesicle traffic by brefeldin A does not affect the integrity of the phagosomes harboring the parental strain of L. pneumophila. We conclude that the Dot/Icm transporter is involved in maintaining the integrity of the L. pneumophila phagosome, independent of interception of ER-to-Golgi vesicle traffic, which is a novel function of type IV secretion systems.",
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T1 - Rapid escape of the dot/icm mutants of Legionella pneumophila into the cytosol of mammalian and protozoan cells

AU - Molmeret, Maëlle

AU - Santic', Marina

AU - Asare, Rexford

AU - Carabeo, Reynold A.

AU - Abu Kwaik, Yousef

PY - 2007/7

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N2 - The Legionella pneumophila-containing phagosome evades endocytic fusion and intercepts endoplasmic reticulum (ER)-to-Golgi vesicle traffic, which is believed to be mediated by the Dot/Icm type IV secretion system. Although phagosomes harboring dot/icm mutants are thought to mature through the endosomal-lysosomal pathway, colocalization studies with lysosomal markers have reported contradictory results. In addition, phagosomes harboring the dot/icm mutants do not interact with endocytosed materials, which is inconsistent with maturation of the phagosomes in the endosomal-lysosomal pathway. Using multiple strategies, we show that the dot/icm mutants defective in the Dot/Icm structural apparatus are unable to maintain the integrity of their phagosomes and escape into the cytoplasm within minutes of entry into various mammalian and protozoan cells in a process independent of the type II secretion system. In contrast, mutants defective in cytoplasmic chaperones of Dot/Icm effectors and rpoS, letA/S, and letE regulatory mutants are all localized within intact phagosomes. Importantly, non-dot/icm L. pneumophila mutants whose phagosomes acquire late endosomal-lysosomal markers are all located within intact phagosomes. Using high-resolution electron microscopy, we show that phagosomes harboring the dot/icm transporter mutants do not fuse to lysosomes but are free in the cytoplasm. Inhibition of ER-to-Golgi vesicle traffic by brefeldin A does not affect the integrity of the phagosomes harboring the parental strain of L. pneumophila. We conclude that the Dot/Icm transporter is involved in maintaining the integrity of the L. pneumophila phagosome, independent of interception of ER-to-Golgi vesicle traffic, which is a novel function of type IV secretion systems.

AB - The Legionella pneumophila-containing phagosome evades endocytic fusion and intercepts endoplasmic reticulum (ER)-to-Golgi vesicle traffic, which is believed to be mediated by the Dot/Icm type IV secretion system. Although phagosomes harboring dot/icm mutants are thought to mature through the endosomal-lysosomal pathway, colocalization studies with lysosomal markers have reported contradictory results. In addition, phagosomes harboring the dot/icm mutants do not interact with endocytosed materials, which is inconsistent with maturation of the phagosomes in the endosomal-lysosomal pathway. Using multiple strategies, we show that the dot/icm mutants defective in the Dot/Icm structural apparatus are unable to maintain the integrity of their phagosomes and escape into the cytoplasm within minutes of entry into various mammalian and protozoan cells in a process independent of the type II secretion system. In contrast, mutants defective in cytoplasmic chaperones of Dot/Icm effectors and rpoS, letA/S, and letE regulatory mutants are all localized within intact phagosomes. Importantly, non-dot/icm L. pneumophila mutants whose phagosomes acquire late endosomal-lysosomal markers are all located within intact phagosomes. Using high-resolution electron microscopy, we show that phagosomes harboring the dot/icm transporter mutants do not fuse to lysosomes but are free in the cytoplasm. Inhibition of ER-to-Golgi vesicle traffic by brefeldin A does not affect the integrity of the phagosomes harboring the parental strain of L. pneumophila. We conclude that the Dot/Icm transporter is involved in maintaining the integrity of the L. pneumophila phagosome, independent of interception of ER-to-Golgi vesicle traffic, which is a novel function of type IV secretion systems.

KW - Macrophages

KW - Microscopy, Confocal

KW - Animals

KW - Legionella pneumophila

KW - Carrier Proteins

KW - Humans

KW - Phagosomes

KW - Membrane Proteins

KW - Acanthamoeba

KW - Bacterial Proteins

KW - Epithelial Cells

KW - Cytosol

KW - Time Factors

KW - Mutation

KW - U937 Cells

U2 - 10.1128/IAI.00292-07

DO - 10.1128/IAI.00292-07

M3 - Article

VL - 75

SP - 3290

EP - 3304

JO - Infection and Immunity

JF - Infection and Immunity

SN - 0019-9567

IS - 7

ER -