Quantitative and Dynamic Assessment of the Contribution of the ER to Phagosome Formation

Abstract

Phagosomes were traditionally thought to originate from an invagination and scission of the plasma membrane to form a distinct intracellular vacuole. An alternative model implicating the endoplasmic reticulum (ER) as a major component of nascent and maturing phagosomes was recently proposed (Gagnon et al., 2002). To reconcile these seemingly disparate hypotheses, we used a combination of biochemical, fluorescence imaging, and electron microscopy techniques to quantitatively and dynamically assess the contribution of the plasmalemma and of the ER to phagosome formation and maturation. We could not verify even a transient physical continuity between the ER and the plasma membrane, nor were we able to detect a significant contribution of the ER to forming or maturing phagosomes in either macrophages or dendritic cells. Instead, our data indicate that the plasma membrane is the main constituent of nascent and newly formed phagosomes, which are progressively remodeled by fusion with endosomal and eventually lysosomal compartments as phagosomes mature into acidic, degradative organelles.

A new role for the P2X7 receptor: a scavenger receptor for bacteria and apoptotic cells in the absence of serum and extracellular ATP

“…Confocal microscopy has shown that internal membranes within the cell fuse with plasma membrane during the course of particle ingestion, and that recycling endosomes are the primary source of membrane for enlargement of phagocytic cup [22]. Flow cytometric assessment of particle engulfment has to some extent replaced microscopic observation [23] particularly as the kinetics of uptake of fluorescent targets by phagocytes can be followed by instruments capable of time-resolved measurements in a stirred cuvette at 37°C [24].…”

Vesicle-Associated Membrane Protein-8/Endobrevin Negatively Regulates Phagocytosis of Bacteria in Dendritic Cells

“…Other studies suggested the existence of endoplasmic reticulum (ER)-mediated phagocytosis (8 -10), whereby the ER membrane was targeted to the phagocytic site to aid in phagosome formation. However, by using a combination of biochemical, fluorescence imaging and electron microscopy techniques, Grinstein and colleagues (11) did not obtain evidence to prove the direct fusion between the ER and the plasma membrane. ER-mediated phagocytosis remains a subject of controversy.…”

Caractéristiques dynamiques et fonctionnelles de la membrane du phagosome

“…Mais la majeure partie de l'attention portée à ce modèle était due à sa capacité d'expliquer le mécanisme mal compris de la présentation croisée des antigènes [4,5]. La participation du RE dans la formation du phagosome a été récemment ré-examinée en utilisant diverses approches biophysiques et biochimiques [6]. Une quantification des composants du phagosome présents au cours de sa formation par microscopie de fluorescence ou électronique, et par une approche biochimique intégrative, n'a indiqué aucune contribution significative du RE.…”