scholarly journals Taking apart the endocytic machinery

2008 ◽  
Vol 180 (6) ◽  
pp. 1059-1060 ◽  
Author(s):  
Marko Kaksonen
Keyword(s):  
Immunoelectron Microscopy ◽  
Vesicle Budding ◽  
Cell Biol ◽  
Endocytic Machinery ◽  
Molecular Machinery ◽  
Quantitative Immunoelectron Microscopy ◽  
Endocytic Vesicles ◽  
Endocytic Proteins

The formation of clathrin-coated endocytic vesicles is driven by a complex and highly dynamic molecular machinery. In this issue, Idrissi et al. (Idrissi, F.-Z., H. Grötsch, I.M. Fernández-Golbano, C. Presciatto-Baschong, H. Riezman, and M.-I. Geli. 2008. J. Cell Biol. 180:1219–1232) reveal some of the secrets of this machinery by analyzing the localizations of nine endocytic proteins during vesicle budding in yeast using quantitative immunoelectron microscopy.

Molecular machinery mediating vesicle budding, docking and fusion

1996 ◽  
Vol 52 (12) ◽  
pp. 1021-1025 ◽  
Author(s):  
T. H. Söllner ◽  
J. E. Rothman
Keyword(s):  
Vesicle Budding ◽  
Molecular Machinery

Parvalbumin in the monkey striate cortex: a quantitative immunoelectron-microscopy study

1991 ◽  
Vol 554 (1-2) ◽  
pp. 237-243 ◽  
Author(s):  
I. Blümcke ◽  
P.R. Hof ◽  
J.H. Morrison ◽  
M.R. Celio
Keyword(s):  
Immunoelectron Microscopy ◽  
Striate Cortex ◽  
Microscopy Study ◽  
Quantitative Immunoelectron Microscopy

scholarly journals Two distinct, calcium-mediated, signal transduction pathways can trigger deflagellation in Chlamydomonas reinhardtii

1994 ◽  
Vol 124 (5) ◽  
pp. 807-815 ◽  
Author(s):  
LM Quarmby ◽  
HC Hartzell
Keyword(s):  
Signal Transduction ◽  
Chlamydomonas Reinhardtii ◽  
Living Cells ◽  
Signalling Pathways ◽  
Signal Transduction Pathways ◽  
Channel Blockers ◽  
Wasp Venom ◽  
Intracellular Pool ◽  
Cell Biol ◽  
Molecular Machinery

The molecular machinery of deflagellation can be activated in detergent permeabilized Chlamydomonas reinhardtii by the addition of Ca2+ (Sanders, M. A., and J. L. Salisbury, 1989. J. Cell Biol. 108:1751-1760). This suggests that stimuli which induce deflagellation in living cells cause an increase in the intracellular concentration of Ca2+, but this has never been demonstrated. In this paper we report that the wasp venom peptide, mastoparan, and the permeant organic acid, benzoate, activate two different signalling pathways to trigger deflagellation. We have characterized each pathway with respect to: (a) the requirement for extracellular Ca2+; (b) sensitivity to Ca2+ channel blockers; and (c) 45Ca influx. We also report that a new mutant strain of C. reinhardtii, adf-1, is specifically defective in the acid-activated signalling pathway. Both signalling pathways appear normal in another mutant, fa-1, that is defective in the machinery of deflagellation (Lewin, R. and C. Burrascano. 1983. Experientia. 39:1397-1398; Sanders, M. A., and J. L. Salisbury. 1989. J. Cell Biol. 108:1751-1760). We conclude that mastoparan induces the release of an intracellular pool of Ca2+ whereas acid induces an influx of extracellular Ca2+ to activate the machinery of deflagellation.

scholarly journals Differential Requirements for COPI Coats in Formation of Replication Complexes among Three Genera of Picornaviridae

2002 ◽  
Vol 76 (21) ◽  
pp. 11113-11122 ◽  
Author(s):  
Elena V. Gazina ◽  
Jason M. Mackenzie ◽  
Rebecca J. Gorrell ◽  
David A. Anderson
Keyword(s):  
Endoplasmic Reticulum ◽  
Immunoelectron Microscopy ◽  
Brefeldin A ◽  
Rna Replication ◽  
Encephalomyocarditis Virus ◽  
Differential Sensitivity ◽  
Nonstructural Proteins ◽  
Vesicle Budding ◽  
Golgi Transport ◽  
Infected Cells

ABSTRACT Picornavirus RNA replication requires the formation of replication complexes (RCs) consisting of virus-induced vesicles associated with viral nonstructural proteins and RNA. Brefeldin A (BFA) has been shown to strongly inhibit RNA replication of poliovirus but not of encephalomyocarditis virus (EMCV). Here, we demonstrate that the replication of parechovirus 1 (ParV1) is partly resistant to BFA, whereas echovirus 11 (EV11) replication is strongly inhibited. Since BFA inhibits COPI-dependent steps in endoplasmic reticulum (ER)-Golgi transport, we tested a hypothesis that different picornaviruses may have differential requirements for COPI in the formation of their RCs. Using immunofluorescence and cryo-immunoelectron microscopy we examined the association of a COPI component, β-COP, with the RCs of EMCV, ParV1, and EV11. EMCV RCs did not contain β-COP. In contrast, β-COP appeared to be specifically distributed to the RCs of EV11. In ParV1-infected cells β-COP was largely dispersed throughout the cytoplasm, with some being present in the RCs. These results suggest that there are differences in the involvement of COPI in the formation of the RCs of various picornaviruses, corresponding to their differential sensitivity to BFA. EMCV RCs are likely to be formed immediately after vesicle budding from the ER, prior to COPI association with membranes. ParV1 RCs are formed from COPI-containing membranes but COPI is unlikely to be directly involved in their formation, whereas formation of EV11 RCs appears to be dependent on COPI association with membranes.

scholarly journals Delivery of ligands from sorting endosomes to late endosomes occurs by maturation of sorting endosomes

1992 ◽  
Vol 117 (2) ◽  
pp. 301-310 ◽  
Author(s):  
KW Dunn ◽  
FR Maxfield
Keyword(s):  
Cho Cells ◽  
Digital Image Analysis ◽  
Compartment Model ◽  
Half Time ◽  
Chase Period ◽  
Cell Biol ◽  
Late Endosomes ◽  
Pass Through ◽  
Endocytic Vesicles ◽  
To Receive

After endocytosis, lysosomally targeted ligands pass through a series of endosomal compartments. The endocytic apparatus that accomplishes this passage may be considered to take one of two forms: (a) a system in which lysosomally targeted ligands pass through preexisting, long-lived early sorting endosomes and are then selectively transported to long-lived late endosomes in carrier vesicles, or (b) a system in which lysosomally targeted ligands are delivered to early sorting endosomes which themselves mature into late endosomes. We have previously shown that sorting endosomes in CHO cells fuse with newly formed endocytic vesicles (Dunn, K. W., T. E. McGraw, and F. R. Maxfield. 1989. J. Cell Biol. 109:3303-3314) and that previously endocytosed ligands lose their accessibility to fusion with a half-time of approximately 8 min (Salzman, N. H., and F. R. Maxfield. 1989. J. Cell Biol. 109:2097-2104). Here we have studied the properties of individual endosomes by digital image analysis to distinguish between the two mechanisms for entry of ligands into late endosomes. We incubated TRVb-1 cells (derived from CHO cells) with diO-LDL followed, after a variable chase, by diI-LDL, and measured the diO content of diI-containing endosomes. As the chase period was lengthened, an increasing percentage of the endosomes containing diO-LDL from the initial incubation had no detectable diI-LDL from the second incubation, but those endosomes that contained both probes showed no decrease in the amount of diO-LDL per endosomes. These results indicate that (a) a pulse of fluorescent LDL is retained by individual sorting endosomes, and (b) with time sorting endosomes lose the ability to fuse with primary endocytic vesicles. These data are inconsistent with a preexisting compartment model which predicts that the concentration of ligand in sorting endosomes will decline during a chase interval, but that the ability of the stable sorting endosome to receive newly endocytosed ligands will remain high. These data are consistent with a maturation mechanism in which the sorting endosome retains and accumulates lysosomally directed ligands until it loses its ability to fuse with newly formed endocytic vesicles and matures into a late endosome. We also find that, as expected according to the maturation model, new sorting endosomes are increasingly labeled during the chase period indicating that new sorting endosomes are continuously formed to replace those that have matured into late endosomes.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Synaptic Vesicles Form by Budding from Tubular Extensions of Sorting Endosomes in PC12 Cells

1999 ◽  
Vol 10 (12) ◽  
pp. 4163-4176 ◽  
Author(s):  
Heidi de Wit ◽  
Yael Lichtenstein ◽  
Hans J. Geuze ◽  
Regis B. Kelly ◽  
Peter van der Sluijs ◽  
...  
Keyword(s):  
Pc12 Cells ◽  
Transferrin Receptor ◽  
T Antigen ◽  
Marker Proteins ◽  
Late Endosomes ◽  
Early Endosomes ◽  
Quantitative Immunoelectron Microscopy ◽  
Recycling Pathway ◽  
Endocytic Vesicles

The putative role of sorting early endosomes (EEs) in synaptic-like microvesicle (SLMV) formation in the neuroendocrine PC12 cell line was investigated by quantitative immunoelectron microscopy. By BSA-gold internalization kinetics, four distinct endosomal subcompartments were distinguished: primary endocytic vesicles, EEs, late endosomes, and lysosomes. As in other cells, EEs consisted of vacuolar and tubulovesicular subdomains. The SLMV marker proteins synaptophysin and vesicle-associated membrane protein 2 (VAMP-2) localized to both the EE vacuoles and associated tubulovesicles. Quantitative analysis showed that the transferrin receptor and SLMV proteins colocalized to a significantly higher degree in primary endocytic vesicles then in EE-associated tubulovesicles. By incubating PC12 cells expressing T antigen-tagged VAMP (VAMP-TAg) with antibodies against the luminal TAg, the recycling pathway of SLMV proteins was directly visualized. At 15°C, internalized VAMP-TAg accumulated in the vacuolar domain of EEs. Upon rewarming to 37°C, the labeling shifted to the tubular part of EEs and to newly formed SLMVs. Our data delineate a pathway in which SLMV proteins together with transferrin receptor are delivered to EEs, where they are sorted into SLMVs and recycling vesicles, respectively.

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