Cargo carriers from the Golgi to the cell surface.
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ABSTRACT:
SUBMITTER: Pfeffer SR
PROVIDER: S-EPMC3474924 | biostudies-other | 2012 Oct
REPOSITORIES: biostudies-other
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Project description:The plant hormone indole-acetic acid (auxin) is essential for many aspects of plant development. Auxin-mediated growth regulation typically involves the establishment of an auxin concentration gradient mediated by polarly localized auxin transporters. The localization of auxin carriers and their amount at the plasma membrane are controlled by membrane trafficking processes such as secretion, endocytosis, and recycling. In contrast to endocytosis or recycling, how the secretory pathway mediates the localization of auxin carriers is not well understood. In this study we have used the differential cell elongation process during apical hook development to elucidate the mechanisms underlying the post-Golgi trafficking of auxin carriers in Arabidopsis. We show that differential cell elongation during apical hook development is defective in Arabidopsis mutant echidna (ech). ECH protein is required for the trans-Golgi network (TGN)-mediated trafficking of the auxin influx carrier AUX1 to the plasma membrane. In contrast, ech mutation only marginally perturbs the trafficking of the highly related auxin influx carrier LIKE-AUX1-3 or the auxin efflux carrier PIN-FORMED-3, both also involved in hook development. Electron tomography reveals that the trafficking defects in ech mutant are associated with the perturbation of secretory vesicle genesis from the TGN. Our results identify differential mechanisms for the post-Golgi trafficking of de novo-synthesized auxin carriers to plasma membrane from the TGN and reveal how trafficking of auxin influx carriers mediates the control of differential cell elongation in apical hook development.
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Project description:Hepatocytes, the main epithelial cells of the liver, organize their polarized membrane domains differently from ductal epithelia. They also differ in their biosynthetic delivery of single-membrane-spanning and glycophosphatidylinositol-anchored proteins to the apical domain. While ductal epithelia target apical proteins to varying degrees from the trans-Golgi network (TGN) to the apical surface directly, hepatocytes target them first to the basolateral domain, from where they undergo basolateral-to-apical transcytosis. How TGN-to-surface transport differs in both scenarios is unknown. Here, we report that the basolateral detour of a hepatocyte apical protein is due, in part, to low RhoA activity at the TGN, which prevents its segregation from basolateral transport carriers. Activating Rho in hepatocytic cells, which switches their polarity from hepatocytic to ductal, also led to apical-basolateral cargo segregation at the TGN as is typical for ductal cells, affirming a central role for Rho-signaling in different aspects of the hepatocytic polarity phenotype. Nevertheless, Rho-induced cargo segregation was not sufficient to target the apical protein directly; thus, failure to recruit apical targeting machinery also contributes to its indirect itinerary.
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Project description:To describe trafficking of secretory cargo within the Golgi apparatus, the cisternal maturation model predicts that Golgi cisternae change their properties from cis to trans while cargo remains in the cisternae. Cisternal change has been demonstrated in living yeast Saccharomyces cerevisiae; however, the behavior of cargo has yet to be examined directly. In this study, we conducted simultaneous three-color and four-dimensional visualization of secretory transmembrane cargo together with early and late Golgi resident proteins. We show that cargo stays in a Golgi cisterna during maturation from cis-Golgi to trans-Golgi and further to the trans-Golgi network (TGN), which involves dynamic mixing and segregation of two zones of the earlier and later Golgi resident proteins. The location of cargo changes from the early to the late zone within the cisterna during the progression of maturation. In addition, cargo shows an interesting behavior during the maturation to the TGN. After most cargo has reached the TGN zone, a small amount of cargo frequently reappears in the earlier zone.
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Project description:The trans-Golgi network (TGN) is one of the main, if not the main, sorting stations in the process of intracellular protein trafficking. It is therefore of central importance to understand how the key players in the TGN-based sorting and delivery process, the post-Golgi carriers (PGCs), form and function. Over the last few years, modern morphological approaches have generated new insights into the questions of PGC biogenesis, structure and dynamics. Here, we present a view by which the "lifecycle" of a PGC consists of several distinct stages: the formation of TGN tubular export domains (where different cargoes are segregated from each other and from the Golgi enzymes); the docking of these tubular domains onto molecular motors and their extrusion towards the cell periphery along microtubules; the fission of the forming PGC from the donor membrane; and the delivery of the newly formed PGC to its specific acceptor organelle. It is now important to add the many molecular machineries that have been described as operating at the TGN to this "morphofunctional map" of the TGN export process.
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Project description:The Golgi is composed of a stack of cis, medial, trans cisternae that are biochemically distinct. The stable compartments model postulates that permanent cisternae communicate through bi-directional vesicles, while the cisternal maturation model postulates that transient cisternae biochemically mature to ensure anterograde transport. Testing either model has been constrained by the diffraction limit of light microscopy, as the cisternae are only 10-20 nm thick and closely stacked in mammalian cells. We previously described the unstacking of Golgi by the ectopic adhesion of Golgi cisternae to mitochondria. Here, we show that cargo processing and transport continue-even when individual Golgi cisternae are separated and "land-locked" between mitochondria. With the increased spatial separation of cisternae, we show using three-dimensional live imaging that cis-Golgi and trans-Golgi remain stable in their composition and size. Hence, we provide new evidence in support of the stable compartments model in mammalian cells.The different composition of Golgi cisternae gave rise to two different models for intra-Golgi traffic: one where stable cisternae communicate via vesicles and another one where cisternae biochemically mature to ensure anterograde transport. Here, the authors provide evidence in support of the stable compartments model.
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Project description:Transmembrane receptor clustering is a ubiquitous phenomenon in pro- and eukaryotic cells to physically sense receptor/ligand interactions and subsequently translate an exogenous signal into a cellular response. Despite that receptor cluster formation has been described for a wide variety of receptors, ranging from chemotactic receptors in bacteria to growth factor and neurotransmitter receptors in mammalian cells, a mechanistic understanding of the underlying molecular processes is still puzzling. In an attempt to fill this gap we followed a combined experimental and theoretical approach by dissecting and modulating cargo binding, internalization and cellular response mediated by KDEL receptors (KDELRs) at the mammalian cell surface after interaction with a model cargo/ligand. Using a fluorescent variant of ricin toxin A chain as KDELR-ligand (eGFP-RTA(H/KDEL)), we demonstrate that cargo binding induces dose-dependent receptor cluster formation at and subsequent internalization from the membrane which is associated and counteracted by anterograde and microtubule-assisted receptor transport to preferred docking sites at the plasma membrane. By means of analytical arguments and extensive numerical simulations we show that cargo-synchronized receptor transport from and to the membrane is causative for KDELR/cargo cluster formation at the mammalian cell surface.
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