Evidence for two endocytic transport pathways in plant cells (original) (raw)
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Visualising endocytosis in plants: past, present, and future
Journal of Microscopy, 2020
Chris Hawes had a lively fascination for the immensely complex organization of the endomembrane system, including the process of endocytosis. This is the method by which eukaryotic cells internalise membrane proteins, lipids, carbohydrates, and cell wall enzymes from the cell surface through membrane bound vesicles. Endocytosis occurs progressively, starting with early membrane deformation, scission, and finally the release of the vesicle into the cytoplasm. Next to secretion, endocytosis allows the cell to control the proteome composition of its inner and outer surface membrane and as such, its communication with the outside world. Whereas endocytosis was initially considered theoretically impossible in plants due to their high turgor pressure, it is now established as essential for plant life. Furthermore, endocytosis remains a highly active field of research, both in yeast, animal, and plant model systems. Over the past three decades, the tools and techniques used to visualise, quantify, and characterise endocytosis have resulted in an increasingly higher spatiotemporal understanding of this process. Here we provide a brief history of plant endocytosis research from the time when Chris Hawes was investigating the process, to the current state-of-the-This article is protected by copyright. All rights reserved. 2 art in the field. We will end this chapter with a discussion on some promising future developments for plant endocytosis research. This article is protected by copyright. All rights reserved. 3 A brief history of endocytosis in plants Chris Hawes had an intense fascination for understanding the intracellular organisation of a cell. Specifically, Chris was intrigued by the immensely complex organization of the endomembrane system, which covers the anterograde trafficking pathways from the endoplasmic reticulum to the Golgi apparatus, to the trans-Golgi network and to the plasma membrane, as well as the retrograde transport route, which recycles material from the Golgi apparatus back to the endoplasmic reticulum. Moreover, he also studied the endocytic pathway, which involves the delivery of material from the cell surface via endosomes and multi-vesicular bodies to the lytic vacuole. He approached his endomembrane research questions largely through imaging, and was very much interested in endocytosis, a process on which he co-authored a book 1. Endocytosis is the process by which eukaryotic cells internalise extracellular molecules and parts of their plasma membrane through the formation of endocytic vesicles. This allows cells to regulate the turnover of plasma membrane proteins (cargos), including cell wall enzymes, receptors, and transporters. Endocytosis is progressive, starting with membrane bending leading to the formation of an invagination, to scission, which releases the internalized vesicle into the cytoplasm (Figure 1a).
2018
Cellular vesicles are membrane-enclosed organelles that transport material inside and outside the cell. Plant-derived vesicles are receiving increasing attention due to their potential as nanovectors for the delivery of biologically active substances. We aimed to expand our understanding about the heterogeneity and the protein biocargo of citrus fruit juice sac cell-derived vesicles. Micro-and nanosized vesicle fractions were isolated from four citrus species, C. sinensis, C. limon, C. paradisi and C. aurantium, characterized using physicochemical methods and protein cargos were compared using label-free quantitative shotgun proteomics. In each sample approximately 600-800 proteins were identified. Orthologues of most of the top-ranking proteins have previously been reported in extracellular vesicles of mammalian origin. Patellin-3-like, clathrin heavy chain, heat shock proteins, 14-3-3 protein, glyceraldehyde-3-phosphate dehydrogenase and fructose-bisphosphate aldolase 6 were highly expressed in all citrus vesicle fractions. The presence of membrane channel aquaporin on the other hand characteristic of the nanovesicle fractions. Bioinformatics revealed more than hundred protein orthologues potentially implicated in vesicular trafficking. In particular, CCV, COPI and COPII coat proteins indicates the presence of highly heterogeneous populations of intracellular transport vesicles. Moreover, the different hydrolases and oxidoreductases transported within the citrus fruitderived vesicles can be responsible for the various biological activities possessed by the preparations.
The Plant Journal, 2005
The mechanism by which plasma membrane proteins are transported to vacuoles for degradation has not been well characterized in plants. To clarify how plasma membrane proteins are degraded, we monitored the endocytotic pathway in tobacco suspension-cultured BY-2 cells with a fluorescent endocytosis marker, FM4-64. Because of the efficient and rapid delivery of endosomes to the vacuoles, endosomes were scarcely detectable. Interestingly, we found that E-64d, an inhibitor of papain family proteases, caused the accumulation of a large number of endosomes in the cells under the sucrose-starved condition. This result indicates that E-64d attenuates the fusion of endosomes with vacuoles. We identified two papain homologues, which are localized in the endosomes, with a biotinylated inhibitor. We designated them as endosomelocalized papains (ENPs). Immunofluorescent analysis revealed that vacuolar sorting receptor, a marker of prevacuolar compartment (PVC), was localized in the endosomes. This result and their acidic nature show that the endosomes correspond to PVC. These results suggest that ENPs facilitate the final step in the vacuolar trafficking pathway under the sucrose-starved condition. We further examined the effects of E-64d on two transgenic Arabidopsis plants that constitutively express a fusion protein composed of green fluorescent protein (GFP) and a plasma membrane protein (GFP-PIP2a or GFP-LTI6b). GFP fluorescence was observed on the plasma membrane of root cells in these transgenic plants. Treatment with E-64d induced the accumulation of GFP-fluorescent endosomes and inhibited the degradation of these fusion proteins. No GFP fluorescence was observed in vacuoles in E-64d-treated transgenic plants. Taken together, these results suggest that endosomal proteases are required for the fusion of endosomes with vacuoles at the final step in the endocytotic pathway for degradation of plasma membrane proteins in plants.
Planta, 1987
Freeze-fracture electron microscopy of propane-jet-frozen samples has been employe~l to investigate vesicle-mediated secretion and ~em-~ brane recycling events in carrot (Daucus carota L.) and sycamore maple (Acer pseudoplatanus L.)~uspension-culture cells. Stabilization of the cell~by means of ultrarapid freezing has enabled us to preserve the cells in a turgid state and to visuMize new intermediate membrane configurations related to these events. Indeed, many of the observed membrane configurations, such as flattened mlembrane vesicles with slit-shaped membrane fusion sites and horseshoe-shaped membrane infoldi~gs, appear to result from the action of turgor forces on the plasma membrane. Individual cells exhibited great variations in numbers and types of n~embrahe configurations postulated to be related to secretion and membrane-recycling events. In the majority of cells, the different membrane profiles displayed a patchy distribution, and within each patch the membrane configurations tended t~ be of the same stage. This result indicates that s~cretory events are triggered in domains meas+ing from 0.1 to about 10 gm in diameter. Based on an extensive analysis of the different membrane configurations seen in our samples, we have formulated the following model of vesicle-mediated secretion in plant cells : Fusion of a secretory ve~icle with the plasma membrane leads to the formation of a single, narrow-necked pore that increasds in diameter up to about 60 nm. During discharge I the vesicle is flattened, forming a disc-shaped struqtured perpendicular to the plane of the plasma n)embrane. As the vesicle is flattened, the pore is ton
Membrane transport in the endocytic pathway: Animal versus plant cells
Protoplasma, 2000
The endocytic pathway is a well established process in animal cells, but it is not well understood in plant cells. At the morphological level, all the compartments involved in endocytosis in animal cells seem to have counterparts in plant cells, and the organization of the pathway appears to share some striking similarities. Several Rab homologues have been found in plant cells, including homologues of Rab5, Rab7, and Rabll, markers of endocytic compartments in animal cells. Coat proteins are also present in plant cells, including clathrin, adaptins, and ADP ribosylation factor proteins. However, endocytic compartments in plant cells also exhibit specific features both in organization and function. The molecular composition of these compartments remains to be established, and future work will be necessary to identify the key regulators of endocytic trafficking in plant cells.
In situ Localization and in vitro Induction of Plant COPI-Coated Vesicles
The Plant Cell, 2000
Coat protein (COP)-coated vesicles have been shown to mediate protein transport through early steps of the secretory pathway in yeast and mammalian cells. Here, we attempt to elucidate their role in vesicular trafficking of plant cells, using a combined biochemical and ultrastructural approach. Immunogold labeling of cryosections revealed that COPI proteins are localized to microvesicles surrounding or budding from the Golgi apparatus. COPI-coated buds primarily reside on the cis -face of the Golgi stack. In addition, COPI and Arf1p show predominant labeling of the cis -Golgi stack, gradually diminishing toward the trans -Golgi stack. In vitro COPI-coated vesicle induction experiments demonstrated that Arf1p as well as coatomer could be recruited from cauliflower cytosol onto mixed endoplasmic reticulum (ER)/ Golgi membranes. Binding of Arf1p and coatomer is inhibited by brefeldin A, underlining the specificity of the recruitment mechanism. In vitro vesicle budding was confirmed by identification of COPI-coated vesicles through immunogold negative staining in a fraction purified from isopycnic sucrose gradient centrifugation. Similar in vitro induction experiments with tobacco ER/Golgi membranes prepared from transgenic plants overproducing barley ␣ -amylase-HDEL yielded a COPI-coated vesicle fraction that contained ␣ -amylase as well as calreticulin.
Endosomes and Golgi vesicles in adsorptive and fluid phase endocytosis
The Journal of Cell Biology, 1984
We studied with morphometric methods the endocytosis by pheochromocytoma cells of a conjugate of wheat germ agglutinin with ferritin (WGA-Ft) and of horseradish peroxidase (HRP). Quantitative studies indicated that WGA-Ft was cleared slowly from cell surfaces and that it was not recycled to the surface. Cells labeled with WGA-Ft for 15 min at room temperature were washed and incubated in medium containing HRP for 15 or 30 min at 37 degrees C. The greatest proportion of labeled vesicles and tubules contained only WGA-Ft (83.4% at 15 min and 85.3% at 30 min). A very small fraction of labeled vesicles and tubules contained only HRP (0.2% at 15 min and 0.9% at 30 min). Vesicles and tubules at the Golgi apparatus were labeled almost exclusively with WGA-Ft (97% at 15 min and 30 min); the rest had both labels. Most labeled lysosomes contained both labels (80.1% at 15 min and 80.8% at 30 min). Of the remainder more contained WGA-Ft alone (20% at 15 min and 10.9% at 30 min), then HRP alone ...
Journal of plant physiology, 2018
Cell-derived vesicles are membrane-enclosed organelles that transport material inside and outside the cell. Plant-derived vesicles are receiving more and more attention due to their potential as nanovectors for the delivery of biologically active substances. Here, we studied the heterogeneity and protein biocargo in citrus fruit juice sac cell-derived vesicles populations. Micro- and nano-sized vesicle fractions were isolated from four citrus species, C. sinensis, C. limon, C. paradisi and C. aurantium, characterized using physicochemical methods and protein cargos were compared using label-free quantitative shotgun proteomics. In each sample approximately 600-800 proteins were identified. Orthologues of most of the top-ranking proteins have previously been reported in extracellular vesicles of mammalian origin. High expression levels of patellin-3-like, clathrin heavy chain, heat shock proteins, 14-3-3 protein, glyceraldehyde-3-phosphate dehydrogenase and fructose-bisphosphate aldo...