Mechanisms of procollagen and HSP47 sorting during ER-to-Golgi trafficking
TL;DR: It is suggested that procollagen and HSP47 sorting occurs at ERES beforeprocollagen is exported from the ER in Golgi-bound transport intermediates, providing new insights into mechanisms of Procollagen trafficking.
About: This article is published in Matrix Biology.The article was published on 2020-06-17 and is currently open access. It has received 21 citations till now. The article focuses on the topics: Procollagen peptidase & ER retention.
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TL;DR: In this article, a 3D view of early secretory compartments in mammalian cells with isotropic resolution and precise protein localization using whole-cell, focused ion beam scanning electron microscopy with cryo-structured illumination microscopy and live-cell synchronized cargo release approaches is provided.
100 citations
TL;DR: ER-phagy is dysfunctional in specific human diseases and its regulators are subverted by pathogens, highlighting its crucial role for cell and organism life.
Abstract: ER-phagy (reticulo-phagy) defines the degradation of portions of the endoplasmic reticulum (ER) within lysosomes or vacuoles. It is part of the self-digestion (i.e., auto-phagic) programs recycling cytoplasmic material and organelles, which rapidly mobilize metabolites in cells confronted with nutrient shortage. Moreover, selective clearance of ER subdomains participates to the control of ER size and activity during ER stress, the re-establishment of ER homeostasis after ER stress resolution and the removal of ER parts, in which aberrant and potentially cytotoxic material has been segregated. ER-phagy relies on the individual and/or concerted activation of the ER-phagy receptors, ER peripheral or integral membrane proteins that share the presence of LC3/Atg8-binding motifs in their cytosolic domains. ER-phagy involves the physical separation of portions of the ER from the bulk ER network, and their delivery to the endolysosomal/vacuolar catabolic district. This last step is accomplished by a variety of mechanisms including macro-ER-phagy (in which ER fragments are sequestered by double-membrane autophagosomes that eventually fuse with lysosomes/vacuoles), micro-ER-phagy (in which ER fragments are directly engulfed by endosomes/lysosomes/vacuoles), or direct fusion of ER-derived vesicles with lysosomes/vacuoles. ER-phagy is dysfunctional in specific human diseases and its regulators are subverted by pathogens, highlighting its crucial role for cell and organism life.
30 citations
TL;DR: A review of collagen regulation in osteogenesis imperfecta can be found in this paper, which highlights the significance of transport disruptions in the OI mechanism and aims to motivate exploration of skeletal fragility in OI from the perspective of these pathways.
Abstract: Osteogenesis Imperfecta (OI) comprises a heterogeneous group of patients who share bone fragility and deformities as the main characteristics, albeit with different degrees of severity. Phenotypic variation also exists in other connective tissue aspects of the disease, complicating disease classification and disease course prediction. Although collagen type I defects are long established as the primary cause of the bone pathology, we are still far from comprehending the complete mechanism. In the last years, the advent of next generation sequencing has triggered the discovery of many new genetic causes for OI, helping to draw its molecular landscape. It has become clear that, in addition to collagen type I genes, OI can be caused by multiple proteins connected to different parts of collagen biosynthesis. The production of collagen entails a complex process, starting from the production of the collagen Iα1 and collagen Iα2 chains in the endoplasmic reticulum, during and after which procollagen is subjected to a plethora of posttranslational modifications by chaperones. After reaching the Golgi organelle, procollagen is destined to the extracellular matrix where it forms collagen fibrils. Recently discovered mutations in components of the retrograde transport of chaperones highlight its emerging role as critical contributor of OI development. This review offers an overview of collagen regulation in the context of recent gene discoveries, emphasizing the significance of transport disruptions in the OI mechanism. We aim to motivate exploration of skeletal fragility in OI from the perspective of these pathways to identify regulatory points which can hint to therapeutic targets.
27 citations
TL;DR: In this paper, the authors used transgenic tagging in Drosophila flies and focused ion beam scanning electron microscopy (FIB-SEM) to characterize ERES-Golgi units in collagen-producing fat body, imaginal discs, and imaginal disc overexpressing ERES determinant Tango1.
14 citations
TL;DR: In this paper, the effect of the chemical chaperone 4-phenylbutyrate (4-PBA) on cell homeostasis, collagen trafficking, matrix production and mineralization was investigated in primary osteoblasts from two murine models of moderate osteogenesis imperfecta.
13 citations
References
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TL;DR: Results indicate that clonal osteogenic cell line MC3T3-E1 cells have the capacity to differentiate into osteoblasts and osteocytes and to form calcified bone tissue in vitro.
Abstract: We investigated the capacity of a clonal osteogenic cell line MC3T3-E1, established from newborn mouse calvaria and selected on the basis of high alkaline phosphatase (ALP) activity in the confluent state, to differentiate into osteoblasts and mineralize in vitro. The cells in the growing state showed a fibroblastic morphology and grew to form multiple layers. On day 21, clusters of cells exhibiting typical osteoblastic morphology were found in osmiophilic nodular regions. Such nodules increased in number and size with incubation time and became easily identifiable with the naked eye by day 40-50. In the central part of well-developed nodules, osteocytes were embedded in heavily mineralized bone matrix. Osteoblasts were arranged at the periphery of the bone spicules and were surrounded by lysosome-rich cells and a fibroblastic cell layer. Numerous matrix vesicles were scattered around the osteoblasts and young osteocytes. Matrix vesicles and plasma membranes of osteoblasts, young osteocytes, and lysosome-rich cells showed strong reaction to cytochemical stainings for ALP activity and calcium ions. Minerals were initially localized in the matrix vesicles and then deposited on well-banded collagen fibrils. Deposited minerals consisted exclusively of calcium and phosphorus, and some of the crystals had matured into hydroxyapatite crystals. These results indicate that MC3T3-E1 cells have the capacity to differentiate into osteoblasts and osteocytes and to form calcified bone tissue in vitro.
1,676 citations
TL;DR: Inspired by molecular modeling, the N,N-dimethylamino substituents in tetramethylrhodamine are replaced with four-membered azetidine rings, which doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging.
Abstract: Specific labeling of biomolecules with bright fluorophores is the keystone of fluorescence microscopy. Genetically encoded self-labeling tag proteins can be coupled to synthetic dyes inside living cells, resulting in brighter reporters than fluorescent proteins. Intracellular labeling using these techniques requires cell-permeable fluorescent ligands, however, limiting utility to a small number of classic fluorophores. Here we describe a simple structural modification that improves the brightness and photostability of dyes while preserving spectral properties and cell permeability. Inspired by molecular modeling, we replaced the N,N-dimethylamino substituents in tetramethylrhodamine with four-membered azetidine rings. This addition of two carbon atoms doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging. The novel substitution is generalizable, yielding a palette of chemical dyes with improved quantum efficiencies that spans the UV and visible range.
1,140 citations
TL;DR: Results indicate that these pre-Golgi carrier structures moving unidirectionally along microtubule tracks are responsible for transporting VSVG-GFP through the cytoplasm to the Golgi complex, which contrasts with the traditional focus on small vesicles as the primary vehicles for ER-to-golgi transport.
Abstract: Newly synthesized proteins that leave the endoplasmic reticulum (ER) are funnelled through the Golgi complex before being sorted for transport to their different final destinations. Traditional approaches have elucidated the biochemical requirements for such transport and have established a role for transport intermediates. New techniques for tagging proteins fluorescently have made it possible to follow the complete life history of single transport intermediates in living cells, including their formation, path and velocity en route to the Golgi complex. We have now visualized ER-to-Golgi transport using the viral glycoprotein ts045 VSVG tagged with green fluorescent protein (VSVG-GFP). Upon export from the ER, VSVG-GFP became concentrated in many differently shaped, rapidly forming pre-Golgi structures, which translocated inwards towards the Golgi complex along microtubules by using the microtubule minus-end-directed motor complex of dynein/dynactin. No loss of fluorescent material from pre-Golgi structures occurred during their translocation to the Golgi complex and they frequently stretched into tubular shapes. Together, our results indicate that these pre-Golgi carrier structures moving unidirectionally along microtubule tracks are responsible for transporting VSVG-GFP through the cytoplasm to the Golgi complex. This contrasts with the traditional focus on small vesicles as the primary vehicles for ER-to-Golgi transport.
1,117 citations
TL;DR: Analysis of the human genome reveals that approximately a third of all open reading frames code for proteins that enter the endoplasmic reticulum (ER), demonstrating the importance of this organelle for global protein maturation.
Abstract: Analysis of the human genome reveals that approximately a third of all open reading frames code for proteins that enter the endoplasmic reticulum (ER), demonstrating the importance of this organelle for global protein maturation. The path taken by a polypeptide through the secretory pathway starts with its translocation across or into the ER membrane. It then must fold and be modified correctly in the ER before being transported via the Golgi apparatus to the cell surface or another destination. Being physically segregated from the cytosol means that the ER lumen has a distinct folding environment. It contains much of the machinery for fulfilling the task of protein production, including complex pathways for folding, assembly, modification, quality control, and recycling. Importantly, the compartmentalization means that several modifications that do not occur in the cytosol, such as glycosylation and extensive disulfide bond formation, can occur to secreted proteins to enhance their stability before their exposure to the extracellular milieu. How these various machineries interact during the normal pathway of folding and protein secretion is the subject of this review.
595 citations
TL;DR: The results suggest a sequential mode of COPII and COPI action and indicate that the transport complexes are ER-to-Golgi transport intermediates from which COPI may be involved in recycling material to the ER.
500 citations