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Concept: Endomembrane system

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The Golgi apparatus has attracted intense attentions due to its fascinating morphology and vital role as the pivot of cellular secretory pathway since its discovery. However, its complex structure at the molecular level remains elusive due to limited approaches. In this study, the structure of Golgi apparatus, including the Golgi stack, cisternal structure, relevant tubules and vesicles, were directly visualized by high-resolution atomic force microscope. We imaged both sides of Golgi apparatus membranes and revealed that the outer leaflet of Golgi membranes is relatively smooth while the inner membrane leaflet is rough and covered by dense proteins. With the treatment of methyl-β-cyclodextrin and Triton X-100, we confirmed the existence of lipid rafts in Golgi apparatus membrane, which are mostly in the size of 20 nm -200 nm and appear irregular in shape. Our results may be of significance to reveal the structure-function relationship of the Golgi complex and pave the way for visualizing the endomembrane system in mammalian cells at the molecular level.

Concepts: Cell, Cell membrane, Golgi apparatus, Organelle, Endoplasmic reticulum, Protein targeting, Endomembrane system, Secretory pathway

3

As the most abundant biopolymer on Earth, cellulose is a key structural component of the plant cell wall. Cellulose is produced at the plasma membrane by cellulose synthase (CesA) complexes (CSCs), which are assembled in the endomembrane system and trafficked to the plasma membrane. While several proteins that affect CesA activity have been identified, components that regulate CSC assembly and trafficking remain unknown. Here we show that STELLO1 and 2 are Golgi-localized proteins that can interact with CesAs and control cellulose quantity. In the absence of STELLO function, the spatial distribution within the Golgi, secretion and activity of the CSCs are impaired indicating a central role of the STELLO proteins in CSC assembly. Point mutations in the predicted catalytic domains of the STELLO proteins indicate that they are glycosyltransferases facing the Golgi lumen. Hence, we have uncovered proteins that regulate CSC assembly in the plant Golgi apparatus.

Concepts: Cell, Bacteria, Cell membrane, Golgi apparatus, Endoplasmic reticulum, Cell wall, Cellulose, Endomembrane system

1

The endomembrane system consists of the secretory and endocytic pathways, which communicate by transport to and from the trans-Golgi network (TGN). In mammalian cells, the endocytic pathway includes early, late, and recycling endosomes. In budding yeast, different types of endosomes have been described, but the organization of the endocytic pathway has remained unclear. We performed a spatial and temporal analysis of yeast endosomal markers and endocytic cargoes. Our results indicate that the yeast TGN also serves as an early and recycling endosome. In addition, as previously described, yeast contains a late or prevacuolar endosome (PVE). Endocytic cargoes localize to the TGN shortly after internalization, and manipulations that perturb export from the TGN can slow the passage of endocytic cargoes to the PVE. Yeast apparently lacks a distinct early endosome. Thus, yeast has a simple endocytic pathway that may reflect the ancestral organization of the endomembrane system.

Concepts: Golgi apparatus, Hypha, Yeast, Endosome, Endomembrane system, Receptor-mediated endocytosis, Exosome, Mannose 6-phosphate receptor

0

The identity of organelles in the endomembrane system of any eukaryotic cell critically depends on the correctly localized Rab GTPase, which binds effectors and thus promotes membrane remodeling or fusion. However, it is still unresolved which factors are required and therefore define the localization of the correct fusion machinery. Using SNARE-decorated proteoliposomes that cannot fuse on their own, we now demonstrate that full fusion activity can be achieved by just four soluble factors: a soluble SNARE (V am7), a guanine nucleotide exchange factor (GEF, Mon1-Ccz1), a Rab- GDP dissociation inhibitor (GDI) complex (prenylated Ypt7:GDI), and a Rab effector complex (HOPS). Our findings reveal that the GEF Mon1-Ccz1 is necessary and sufficient for stabilizing prenylated Ypt7 on membranes. HOPS binding to Ypt7-GTP then drives SNARE-mediated fusion, which is fully GTP- dependent. We conclude that an entire fusion cascade can be controlled by a GEF.

Concepts: DNA, Cell, Eukaryote, Golgi apparatus, Organelle, Guanine nucleotide exchange factor, Necessary and sufficient condition, Endomembrane system

0

AP-4 is a member of the heterotetrameric adaptor protein (AP) complex family involved in protein sorting in the endomembrane system of eukaryotic cells. Interest in AP-4 has recently risen with the discovery that mutations in any of its four subunits cause a form of hereditary spastic paraplegia (HSP) with intellectual disability. The critical sorting events mediated by AP-4 and the pathogenesis of AP-4 deficiency, however, remain poorly understood. Here we report the identification of ATG9A, the only multispanning membrane component of the core autophagy machinery, as a specific AP-4 cargo. AP-4 promotes signal-mediated export of ATG9A from the trans-Golgi network to the peripheral cytoplasm, contributing to lipidation of the autophagy protein LC3B and maturation of preautophagosomal structures. These findings implicate AP-4 as a regulator of autophagy and altered autophagy as a possible defect in AP-4-deficient HSP.

Concepts: DNA, Cell, Eukaryote, Golgi apparatus, Organelle, Endoplasmic reticulum, Cytoplasm, Endomembrane system

0

GTP-ases of the Rab family (about 70 in human) are key regulators of intracellular transport and membrane trafficking in eukaryotic cells. Remarkably, almost one third associate with membranes of the Golgi complex and TGN (trans-Golgi network). Through interactions with a variety of effectors that include molecular motors, tethering complexes, scaffolding proteins and lipid kinases, they play an important role in maintaining Golgi architecture.

Concepts: DNA, Cell, Cell membrane, Golgi apparatus, Organelle, Endoplasmic reticulum, Lysosome, Endomembrane system

0

Approximately one-third of all eukaryotic proteins are delivered to their destination by trafficking within the endomembrane system. Such cargo proteins are incorporated into forming membrane vesicles on donor compartments and delivered to acceptor compartments by vesicle fusion. How cargo proteins are sorted into forming vesicles is still largely unknown. Here we review the roles of small GTPases of the ARF/SAR1 family, their regulators designated ARF guanine-nucleotide exchange factors (ARF-GEFs) and ARF GTPase-activating proteins (ARF-GAPs) as well as coat protein complexes during membrane vesicle formation. Although conserved across eukaryotes, these four functional groups of proteins display plant-specific modifications in composition, structure and function.

Concepts: Cell, Archaea, Eukaryote, Cytosol, Endoplasmic reticulum, Prokaryote, Membrane biology, Endomembrane system

0

The small GTPase Arf4 and the Arf GTPase activating protein (GAP) ASAP1 cooperatively sequester sensory receptor cargo into transport carriers targeted to primary cilia, but the input that drives Arf4 activation in this process remains unknown. Here, we show that during the carrier biogenesis from the photoreceptor Golgi/trans-Golgi network (TGN) a functional complex is formed between Arf4, the Arf guanine nucleotide exchange factor (GEF) GBF1 and the light-sensing receptor, rhodopsin. Rhodopsin and Arf4 bind the regulatory N-terminal DCB-HUS domain of GBF1. The complex is sensitive to Golgicide A (GCA), a selective inhibitor of GBF1 that accordingly blocks rhodopsin delivery to the cilia, without disrupting the photoreceptor Golgi. The emergence of newly synthesized rhodopsin in the endomembrane system is essential for GBF1-Arf4 complex formation in vivo. Notably, GBF1 interacts with the Arf GAP ASAP1 in a GCA-resistant manner. Our findings implicate that converging signals on GBF1 from the influx of cargo into the Golgi/TGN and the feedback from Arf4, combined with an input from ASAP1, control Arf4 activation during sensory membrane trafficking to primary cilia.

Concepts: Proteins, Protein, Cell, Signal transduction, Golgi apparatus, Guanine nucleotide exchange factor, Endomembrane system, Sensory receptors

0

The vacuole is a prominent organelle that is essential for plant viability. The vacuole size, and its role in ion homeostasis, protein degradation and storage, place significant demands for trafficking of vacuolar cargo along the endomembrane system. Recent studies indicate that sorting of vacuolar cargo initiates at the ER and Golgi, but not the trans-Golgi network/early endosome, as previously thought. Furthermore, maturation of the trans-Golgi network into pre-vacuolar compartments seems to contribute to a major route for plant vacuolar traffic that works by bulk flow and ends with membrane fusion between the pre-vacuolar compartment and the tonoplast. Here we summarize recent evidence that indicates conserved and plant-specific mechanisms involved in sorting and trafficking of proteins to this major organelle.

Concepts: Cell, Cell membrane, Golgi apparatus, Organelle, Endoplasmic reticulum, Organelles, Vacuole, Endomembrane system

0

Nuclear pore complexes (NPCs) are the sole gateway between the cytoplasm and the nucleus serving both as stringent permeability barrier and active transporters between the two compartments of eukaryotic cells. Complete mechanistic understanding of how these two functions are implemented within one and the same transport machine has not been attained to date. Based on several lines of structural evidence, a hypothesis was proposed postulating that NPCs shares common evolutionary origin with other intracellular systems responsible for active management of endomembranes. In this review we attempt to summarize the evidence supporting this hypothesis. The structural data obtained so far is evaluated and supplemented with the analysis of the functional evidence. Based on this analysis, a model is proposed which integrates the knowledge from the field of NPC function with that obtained from other endomembrane management systems in an attempt to shed new light on the mechanism of the NPC active transport.

Concepts: DNA, Cell nucleus, Archaea, Eukaryote, Cytoplasm, Nuclear pore, Nuclear envelope, Endomembrane system