scholarly journals Disruption of the Golgi Apparatus and Contribution of the Endoplasmic Reticulum to the SARS-CoV-2 Replication Complex

Viruses ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 1798
Author(s):  
Ted Hackstadt ◽  
Abhilash I. Chiramel ◽  
Forrest H. Hoyt ◽  
Brandi N. Williamson ◽  
Cheryl A. Dooley ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Rna Synthesis ◽  
Light And Electron Microscopy ◽  
Brefeldin A ◽  
Viral Proteins ◽  
Protein Markers ◽  
Replication Complex ◽  
Golgi Fragmentation

A variety of immunolabeling procedures for both light and electron microscopy were used to examine the cellular origins of the host membranes supporting the SARS-CoV-2 replication complex. The endoplasmic reticulum has long been implicated as a source of membrane for the coronavirus replication organelle. Using dsRNA as a marker for sites of viral RNA synthesis, we provide additional evidence supporting ER as a prominent source of membrane. In addition, we observed a rapid fragmentation of the Golgi apparatus which is visible by 6 h and complete by 12 h post-infection. Golgi derived lipid appears to be incorporated into the replication organelle although protein markers are dispersed throughout the infected cell. The mechanism of Golgi disruption is undefined, but chemical disruption of the Golgi apparatus by brefeldin A is inhibitory to viral replication. A search for an individual SARS-CoV-2 protein responsible for this activity identified at least five viral proteins, M, S, E, Orf6, and nsp3, that induced Golgi fragmentation when expressed in eukaryotic cells. Each of these proteins, as well as nsp4, also caused visible changes to ER structure as shown by correlative light and electron microscopy (CLEM). Collectively, these results imply that specific disruption of the Golgi apparatus is a critical component of coronavirus replication.

Development of the lateral palatal brush in larvae of Aedes aegypti (L.) (Diptera: Culicidae)

1990 ◽  
Vol 68 (7) ◽  
pp. 1454-1467 ◽  
Author(s):  
K. M. Fry ◽  
S. B. McIver
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Golgi Apparatus ◽  
Aedes Aegypti ◽  
Rough Endoplasmic Reticulum ◽  
Light And Electron Microscopy ◽  
Fourth Instar ◽  
Muscle Fibres ◽  
Final Length

Light and electron microscopy were used to observe development of the lateral palatal brush in Aedes aegypti (L.) larvae. Development was sampled at 4-h intervals from second- to third-instar ecdyses. Immediately after second-instar ecdysis, the epidermis apolyses from newly deposited cuticle in the lateral palatal pennicular area to form an extensive extracellular cavity into which the fourth-instar lateral palatal brush filaments grow as cytoplasmic extensions. On reaching their final length, the filaments deposit cuticulin, inner epicuticle, and procuticle sequentially on their outer surfaces. The lateral palatal crossbars, on which the lateral palatal brush filaments insert, form after filament development is complete. At the beginning of development, the organelles involved in plasma membrane and cuticle production are located at the base and middle of the cells. As the filament rudiments grow, most rough endoplasmic reticulum, mitochondria, and Golgi apparatus move to the apex of the epidermal cells and into the filament rudiments. After formation of the lateral palatal brush filaments and lateral palatal crossbars, extensive organelle breakdown occurs. Lateral palatal brush formation is unusual in that no digestion and resorption of old endocuticle occurs prior to deposition of new cuticle. No mucopolysaccharide secretion by the lateral palatal brush epidermis was observed, nor were muscle fibres observed to attach to the lateral palatal crossbars, as has been suggested by other workers.

scholarly journals SYNTHESIS, MIGRATION, AND RELEASE OF PRECURSOR COLLAGEN BY ODONTOBLASTS AS VISUALIZED BY RADIOAUTOGRAPHY AFTER [3H]PROLINE ADMINISTRATION

1974 ◽  
Vol 60 (1) ◽  
pp. 92-127 ◽  
Author(s):  
Melvyn Weinstock ◽  
C. P. Leblond
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Phosphotungstic Acid ◽  
Secretory Granules ◽  
Low Ph ◽  
Collagen Fibrils ◽  
Dentin Collagen ◽  
Odontoblast Process ◽  
High Radioactivity

The elaboration of dentin collagen precursors by the odontoblasts in the incisor teeth of 30–40-g rats was investigated by electron microscopy, histochemistry, and radioautography after intravenous injection of tritium-labeled proline. At 2 min after injection, when the labeling of blood proline was high, radioactivity was restricted to the rough endoplasmic reticulum, indicating that it is the site of synthesis of the polypeptide precursors of collagen, the pro-alpha chains. At 10 min, when the labeling of blood proline had already declined, radioactivity was observed in spherical portions of Golgi saccules containing entangled threads, and, at 20 min, radioactivity appeared in cylindrical portions containing aggregates of parallel threads. The parallel threads measured 280–350 nm in length and stained with the low pH-phosphotungstic acid technique for carbohydrate and with the silver methenamine technique for aldehydes (as did extracellular collagen fibrils). The passage of label from spherical to cylindrical Golgi portions is associated with the reorganization of entangled into parallel threads, which is interpreted as the packing of procollagen molecules. Between 20 and 30 min, prosecretory and secretory granules respectively became labeled. These results indicate that the cylindrical portions of Golgi saccules transform into prosecretory and subsequently into secretory granules. Within these granules, the parallel threads, believed to be procollagen molecules, are transported to the odontoblast process. At 90 min and 4 h after injection, label was present in predentin, indicating that the labeled content of secretory granules had been released into predentin. This occurred by exocytosis as evidenced by the presence of secretory granules in fusion with the plasmalemma of the odontoblast process. It is proposed that pro-alpha chains give rise to procollagen molecules which assemble into parallel aggregates in the Golgi apparatus. Procollagen molecules are then transported within secretory granules to the odontoblast process and released by exocytosis. In predentin procollagen molecules would give rise to tropocollagen molecules, which would then polymerize into collagen fibrils.

Phase-contrast and electron-microscopic observations on a membranous complex in primary spermatocytes of the mouse

1975 ◽  
Vol 18 (1) ◽  
pp. 1-17
Author(s):  
A. Pleshkewych ◽  
L. Levine
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Phase Contrast ◽  
Cultured Cells ◽  
Light And Electron Microscopy ◽  
Cytoplasmic Inclusion ◽  
Electron Microscopic ◽  
Secondary Spermatocyte ◽  
Living Mouse ◽  
Circular Contour

A prominent cytoplasmic inclusion present in living mouse primary spermatocytes has been observed by both light and electron microscopy. It began to form at prometaphase and continued to increase in thickness and length as the cells developed. By metaphase it was a distinct sausage-shaped boundary that enclosed a portion of the cytoplasm between the spindle and the cell membrane. At the end of metaphase, the inclusion reached its maximum length. At telophase, it was divided between the daughter secondaries. The inclusion persisted as a circular contour in the interphase secondary spermatocyte. Electron microscopy of the same cultured cells that were previously observed with light microscopy revealed that the inclusion was a distinctive formation of membranes. It consisted of agranular cisternae and vesicles, and was therefore a membranous complex. Many of the smaller vesicles in the membranous complex resembled those found in the spindle. The cisternae in the membranous complex were identical to the cisternal endoplasmic reticulum of interphase primary spermatocytes. Nevertheless, the organization of vesicles and cisternae into the membranous complex was unique for the primaries in division stages, since such an organization was not present in their interphase stages.

Anatomy of the Unpollinated and Pollinated Watermelon Stigma

1982 ◽  
Vol 54 (1) ◽  
pp. 341-355
Author(s):  
M. SEDGLEY
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Light And Electron Microscopy ◽  
Freeze Fracture ◽  
Pollen Grain ◽  
Acidic Polysaccharides ◽  
Before And After ◽  
Em Autoradiography ◽  
Pollen Grain Wall ◽  
Stigma Secretion

The structure of the watermelon stigma before and after pollination was studied using light and electron microscopy, freeze-fracture and autoradiography. The wall thickenings of the papilla transfer cells contained callose and their presence prior to pollination was confirmed using EM-autoradiography, freeze-fracture and fixation. No further callose thickenings were produced following pollination. Pollination resulted in a rapid increase in aqueous stigma secretion and localized disruption of the cuticle, which appeared to remain on the surface of the secretion. Autolysis of the papilla cells, which had commenced prior to pollination, was accelerated and appeared to take place via cup-shaped vacuoles developed from distended endoplasmic reticulum. The reaction was localized to the papilla cells adjacent to the pollen tube only. Both pollen-grain wall and stigma secretion contained proteins, carbohydrates, acidic polysaccharides, lipids and phenolics.

Formation of chlamydospores in Gilbertella persicaria

1981 ◽  
Vol 59 (5) ◽  
pp. 908-928 ◽  
Author(s):  
Martha J. Powell ◽  
Charles E. Bracker ◽  
David J. Sternshein
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Acid Phosphatase ◽  
Light And Electron Microscopy ◽  
Multivesicular Bodies ◽  
Marker Enzyme ◽  
Transparent Layer ◽  
Thiamine Pyrophosphatase ◽  
Gilbertella Persicaria

The cytological events involved in the transformation of vegetative hyphae of the zygomycete Gilbertella persicaria (Eddy) Hesseltine into chlamydospores were studied with light and electron microscopy. Thirty hours after sporangiospores were inoculated into YPG broth, swellings appeared along the aseptate hyphae. Later, septa, traversed by plasmodesmata, delimited each end of the hyphal swellings and compartmentalized these hyphal regions as they differentiated into chlamydospores. Nonswollen regions adjacent to chlamydospores remained as isthmuses. Two additional wall layers appeared within the vegetative wall of the developing chlamydospores. An alveolate, electron-dense wall formed first, and then an electron-transparent layer containing concentrically oriented fibers formed between this layer and the plasma membrane. Rather than a mere condensation of cytoplasm, development and maturation of the multinucleate chlamydospores involved extensive cytoplasmic changes such as an increase in reserve products, lipid and glycogen, an increase and then disappearance of vacuoles, and the breakdown of many mitochondria. Underlying the plasma membrane during chlamydospore wall formation were endoplasmic reticulum, multivesicular bodies, vesicles with fibrillar contents, vesicles with electron-transparent contents, and cisternal rings containing the Golgi apparatus marker enzyme, thiamine pyrophosphatase. Acid phosphatase activity was localized cytochemically in a cisterna which enclosed mitochondria and in vacuoles which contained membrane fragments. Tightly packed membrane whorls and single membrane bounded sacs with finely granular matrices surrounding vacuoles were unique during chlamydospore development. Microbodies were rare in the mature chlamydospore, but endoplasmic reticulum was closely associated with lipid globules. As chlamydospores developed, the cytoplasm in the isthmus became highly vacuolated, lipid globules were closely associated with vacuoles, mitochondria were broken down in vacuoles, unusual membrane configurations appeared, and eventually the membranes degenerated. Unlike chlamydospores, walls of the isthmus did not thicken, but irregularly shaped appositions containing numerous channels formed at intervals on the inside of these walls. The pattern of cytoplasmic transformations during chlamydospore development is similar to events leading to the formation of zygospores and sporangiospores.

scholarly journals Remodeling the Endoplasmic Reticulum by Poliovirus Infection and by Individual Viral Proteins: an Autophagy-Like Origin for Virus-Induced Vesicles

2000 ◽  
Vol 74 (19) ◽  
pp. 8953-8965 ◽  
Author(s):  
David A. Suhy ◽  
Thomas H. Giddings ◽  
Karla Kirkegaard
Keyword(s):  
Endoplasmic Reticulum ◽  
Mammalian Cells ◽  
Buoyant Density ◽  
Rna Replication ◽  
Viral Proteins ◽  
Replication Complex ◽  
Poliovirus Infection ◽  
Endosomal Membranes ◽  
Infected Cells ◽  
Positive Strand Rna

ABSTRACT All positive-strand RNA viruses of eukaryotes studied assemble RNA replication complexes on the surfaces of cytoplasmic membranes. Infection of mammalian cells with poliovirus and other picornaviruses results in the accumulation of dramatically rearranged and vesiculated membranes. Poliovirus-induced membranes did not cofractionate with endoplasmic reticulum (ER), lysosomes, mitochondria, or the majority of Golgi-derived or endosomal membranes in buoyant density gradients, although changes in ionic strength affected ER and virus-induced vesicles, but not other cellular organelles, similarly. When expressed in isolation, two viral proteins of the poliovirus RNA replication complex, 3A and 2C, cofractionated with ER membranes. However, in cells that expressed 2BC, a proteolytic precursor of the 2B and 2C proteins, membranes identical in buoyant density to those observed during poliovirus infection were formed. When coexpressed with 2BC, viral protein 3A was quantitatively incorporated into these fractions, and the membranes formed were ultrastructurally similar to those in poliovirus-infected cells. These data argue that poliovirus-induced vesicles derive from the ER by the action of viral proteins 2BC and 3A by a mechanism that excludes resident host proteins. The double-membraned morphology, cytosolic content, and apparent ER origin of poliovirus-induced membranes are all consistent with an autophagic origin for these membranes.

scholarly journals Light and dark cells in the lumbar sensory ganglia of pre and post-hatching domestic fowl

1988 ◽  
Vol 46 (1) ◽  
pp. 3-5
Author(s):  
Claudio A. Ferraz de Carvalho ◽  
Ciro F. da Silva
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Satellite Cell ◽  
Satellite Cells ◽  
Domestic Fowl ◽  
Sensory Ganglia ◽  
Dark Cells ◽  
Junctional Complexes ◽  
Functional Features

Clear and dark satellite cell classes were identified by electron microscopy in the lumbar sensory ganglia of domestic fowl in 8 pre and 4 post-hatching stages of development. Some cytologic differences found between the two classes relating to the rough-endoplasmic reticulum, ribosomes, Golgi apparatus and junctional complexes suggest the existence of distinct functional features for both types of satellite cells.

scholarly journals Capacity of the Golgi Apparatus for Biogenesis from the Endoplasmic Reticulum

2003 ◽  
Vol 14 (12) ◽  
pp. 5011-5018 ◽  
Author(s):  
Sapna Puri ◽  
Adam D. Linstedt
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Self Assembly ◽  
De Novo ◽  
Brefeldin A ◽  
The Other ◽  
Matrix Proteins ◽  
Er Export ◽  
Golgi Matrix

It is unclear whether the mammalian Golgi apparatus can form de novo from the ER or whether it requires a preassembled Golgi matrix. As a test, we assayed Golgi reassembly after forced redistribution of Golgi matrix proteins into the ER. Two conditions were used. In one, ER redistribution was achieved using a combination of brefeldin A (BFA) to cause Golgi collapse and H89 to block ER export. Unlike brefeldin A alone, which leaves matrix proteins in relatively large remnant structures outside the ER, the addition of H89 to BFA-treated cells caused ER accumulation of all Golgi markers tested. In the other, clofibrate treatment induced ER redistribution of matrix and nonmatrix proteins. Significantly, Golgi reassembly after either treatment was robust, implying that the Golgi has the capacity to form de novo from the ER. Furthermore, matrix proteins reemerged from the ER with faster ER exit rates. This, together with the sensitivity of BFA remnants to ER export blockade, suggests that presence of matrix proteins in BFA remnants is due to cycling via the ER and preferential ER export rather than their stable assembly in a matrix outside the ER. In summary, the Golgi apparatus appears capable of efficient self-assembly.

scholarly journals LOCALIZATION OF PEROXIDASE ACTIVITY IN MICROBODIES OF FETAL MOUSE LIVER

1969 ◽  
Vol 17 (7) ◽  
pp. 454-466 ◽  
Author(s):  
EDWARD ESSNER
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Acid Phosphatase ◽  
Peroxidase Activity ◽  
Mouse Liver ◽  
Fetal Liver ◽  
Light And Electron Microscopy ◽  
Small Population ◽  
Fetal Mouse ◽  
Fetal Mouse Liver

The peroxidase activity of microbodies in fetal mouse liver was studied by light and electron microscopy. Two types of microbodies were present; a small population of bodies that lacked a nucleoid, predominant on the 16th day of gestation, and a larger population of nucleoid-bearing microbodies, predominant on the 19th day, in association with the rough endoplasmic reticulum from which they probably originate. Both types of bodies were visualized when incubated for peroxidase activity but were negative (19th day) for acid phosphatase activity. The findings suggest that the anucleoid- and nucleoid-bearing organelles together constitute the microbody population of the fetal liver.

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