Human Coronavirus 229E Binds to CD13 in Rafts and Enters the Cell through Caveolae

ABSTRACT CD13, a receptor for human coronavirus 229E (HCoV-229E), was identified as a major component of the Triton X-100-resistant membrane microdomain in human fibroblasts. The incubation of living fibroblasts with an anti-CD13 antibody on ice gave punctate labeling that was evenly distributed on the cell surface, but raising the temperature to 37°C before fixation caused aggregation of the labeling. The aggregated labeling of CD13 colocalized with caveolin-1 in most cells. The HCoV-229E virus particle showed a binding and redistribution pattern that was similar to that caused by the anti-CD13 antibody: the virus bound to the cell evenly when incubated on ice but became colocalized with caveolin-1 at 37°C; importantly, the virus also caused sequestration of CD13 to the caveolin-1-positive area. Electron microscopy confirmed that HCoV-229E was localized near or at the orifice of caveolae after incubation at 37°C. The depletion of plasmalemmal cholesterol with methyl β-cyclodextrin significantly reduced the HCoV-229E redistribution and subsequent infection. A caveolin-1 knockdown by RNA interference also reduced the HCoV-229E infection considerably. The results indicate that HCoV-229E first binds to CD13 in the Triton X-100-resistant microdomain, then clusters CD13 by cross-linking, and thereby reaches the caveolar region before entering cells.

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Leukosialin (CD43, sialophorin) redistribution in uropods of polarized neutrophils is induced by CD43 cross-linking by antibodies, by colchicine or by chemotactic peptides

Journal of Cell Science ◽

10.1242/jcs.110.13.1465 ◽

1997 ◽

Vol 110 (13) ◽

pp. 1465-1475

Author(s):

S. Seveau ◽

S. Lopez ◽

P. Lesavre ◽

J. Guichard ◽

E.M. Cramer ◽

...

Keyword(s):

Electron Microscopy ◽

Cell Motility ◽

Small Proportion ◽

Cytochalasin B ◽

Cell Polarization ◽

Cross Linking ◽

Chemotactic Peptides ◽

Butanedione Monoxime ◽

Major Surface ◽

Triton X

We investigated a possible association of leukosialin (CD43), the major surface sialoglycoprotein of leukocytes, with neutrophil cytoskeleton. We first analysed the solubility of CD43 in Triton X-100 and observed that CD43 of resting neutrophils was mostly soluble. The small proportion of CD43 molecules, which ‘spontaneously’ precipitated in Triton, appeared associated with F-actin, as demonstrated by the fact that this insolubility did not occur when cells were incubated with cytochalasin B or when F-actin was depolymerized with DNase I in the Triton precipitate. Cell stimulation with anti-CD43 mAb (MEM59) enhanced this CD43-cytoskeleton association. By immunofluorescence as well as by electron microscopy, we observed a redistribution of CD43 on the neutrophil membrane, initially in patches followed by caps, during anti-CD43 cross-linking at 37 degrees C. This capping did not occur at 4 degrees C and was inhibited by cytochalasin B and by a myosin disrupting drug butanedione monoxime, thus providing evidence that the actomyosin contracile sytem is involved in the capping and further suggesting an association of CD43 with the cytoskeleton. Some of the capped cells exhibited a front-tail polarization with CD43 caps located in the uropod at the rear of the cell. Surprisingly, colchicine and the chemotactic factor fNLPNTL which induce neutrophil polarization associated with cell motility, also resulted in a clustering of CD43 in the uropod, independently of a cross-linking of the molecule by mAbs. An intracellular redistribution of F-actin, mainly at the leading front and of myosin in the tail, was observed during CD43 clustering induced by colchicine and in cells polarized by anti-CD43 mAbs cross-linking. We conclude that neutrophil CD43 interacts with the cytoskeleton, either directly or indirectly, to redistribute in the cell uropod under antibodies stimulation or during cell polarization by colchicine, thus highly suggesting that CD43 may be involved in cell polarization.

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Isoforms of caveolin-1 and caveolar structure

Journal of Cell Science ◽

10.1242/jcs.113.19.3509 ◽

2000 ◽

Vol 113 (19) ◽

pp. 3509-3517 ◽

Cited By ~ 9

Author(s):

T. Fujimoto ◽

H. Kogo ◽

R. Nomura ◽

T. Une

Keyword(s):

Electron Microscopy ◽

Hepg2 Cells ◽

Immunofluorescence Microscopy ◽

Human Fibroblasts ◽

Freeze Fracture ◽

Molecular Composition ◽

Immunogold Electron Microscopy ◽

Caveolin 1 ◽

Different Depths ◽

The Relationship

The relationship between caveolin-1 isoforms alpha and beta and caveolar ultrastructure was studied. By immunofluorescence microscopy of human fibroblasts, caveolae were observed as dots positive for caveolin-1, but many dots labeled by an antibody recognizing both isoforms (anti-alphabeta) were not labeled by another antibody specific for the alpha isoform (anti-alpha). Immunogold electron microscopy of freeze-fracture replicas revealed caveolae of different depths, and indicated that anti-alpha labeled deep caveolae preferentially over shallow ones, whereas anti-alphabeta labeled both forms with an equivalent frequency and intensity. The presence of the beta isoform in deep caveolae was confirmed by labeling epitope-tagged beta-caveolin. When made to be expressed in HepG2 cells lacking endogenous caveolins, the alpha isoform formed caveolar depressions efficiently, but the beta isoform hardly did so. Caveolae were also formed in cells expressing the two isoforms, but their frequency was variable among cells of the same clone. Coexpression of caveolin-1 and caveolin-2 caused more efficient formation of deep caveolae than caveolin-1 alone. The result indicates that the two isoforms of caveolin-1 have a different potential for forming caveolae structure, and more importantly, that deep and shallow caveolae may be diversified in their molecular composition.

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Embedding Procedure for Water Swollen Samples

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010006951x ◽

1970 ◽

Vol 28 ◽

pp. 504-505

Author(s):

Ann M. Thomas ◽

Virginia Shemeley

Keyword(s):

Electron Microscopy ◽

Transmission Electron Microscopy ◽

Ion Exchange ◽

Structural Changes ◽

Cross Linking ◽

Ion Exchange Resins ◽

Transmission Electron ◽

First Pass ◽

Exchange Resins

Those samples which swell rapidly when exposed to water are, at best, difficult to section for transmission electron microscopy. Some materials literally burst out of the embedding block with the first pass by the knife, and even the most rapid cutting cycle produces sections of limited value. Many ion exchange resins swell in water; some undergo irreversible structural changes when dried. We developed our embedding procedure to handle this type of sample, but it should be applicable to many materials that present similar sectioning difficulties.The purpose of our embedding procedure is to build up a cross-linking network throughout the sample, while it is in a water swollen state. Our procedure was suggested to us by the work of Rosenberg, where he mentioned the formation of a tridimensional structure by the polymerization of the GMA biproduct, triglycol dimethacrylate.

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The Current Situation in Chemical Stabilization of Biological Materials

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100093912 ◽

1972 ◽

Vol 30 ◽

pp. 132-133

Author(s):

John H. Luft

Keyword(s):

Electron Microscopy ◽

Electron Microscope ◽

Osmium Tetroxide ◽

Molecular Level ◽

Cross Linking ◽

Chemical Stabilization ◽

Specimen Preparation ◽

Sufficient Condition ◽

Radio Telescopes

With information processing devices such as radio telescopes, microscopes or hi-fi systems, the quality of the output often is limited by distortion or noise introduced at the input stage of the device. This analogy can be extended usefully to specimen preparation for the electron microscope; fixation, which initiates the processing sequence, is the single most important step and, unfortunately, is the least well understood. Although there is an abundance of fixation mixtures recommended in the light microscopy literature, osmium tetroxide and glutaraldehyde are favored for electron microscopy. These fixatives react vigorously with proteins at the molecular level. There is clear evidence for the cross-linking of proteins both by osmium tetroxide and glutaraldehyde and cross-linking may be a necessary if not sufficient condition to define fixatives as a class.

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CRYO-Electron Microscopy of the Acto-Myosin-ATP Complex

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100179993 ◽

1990 ◽

Vol 48 (1) ◽

pp. 248-249

Author(s):

John Trinickt ◽

Howard White

Keyword(s):

Electron Microscopy ◽

Atp Hydrolysis ◽

Myosin Head ◽

Bound State ◽

Cross Linking ◽

Weakly Bound ◽

Cryo Electron Microscopy ◽

Myosin Subfragment 1 ◽

Attached State ◽

High Fraction

The primary force of muscle contraction is thought to involve a change in the myosin head whilst attached to actin, the energy coming from ATP hydrolysis. This change in attached state could either be a conformational change in the head or an alteration in the binding angle made with actin. A considerable amount is known about one bound state, the so-called strongly attached state, which occurs in the presence of ADP or in the absence of nucleotide. In this state, which probably corresponds to the last attached state of the force-producing cycle, the angle between the long axis myosin head and the actin filament is roughly 45°. Details of other attached states before and during power production have been difficult to obtain because, even at very high protein concentration, the complex is almost completely dissociated by ATP. Electron micrographs of the complex in the presence of ATP have therefore been obtained only after chemically cross-linking myosin subfragment-1 (S1) to actin filaments to prevent dissociation. But it is unclear then whether the variability in attachment angle observed is due merely to the cross-link acting as a hinge.We have recently found low ionic-strength conditions under which, without resorting to cross-linking, a high fraction of S1 is bound to actin during steady state ATP hydrolysis. The structure of this complex is being studied by cryo-electron microscopy of hydrated specimens. Most advantages of frozen specimens over ambient temperature methods such as negative staining have already been documented. These include improved preservation and fixation rates and the ability to observe protein directly rather than a surrounding stain envelope. In the present experiments, hydrated specimens have the additional benefit that it is feasible to use protein concentrations roughly two orders of magnitude higher than in conventional specimens, thereby reducing dissociation of weakly bound complexes.

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Lessons Learned From Serial Isolations Of Human Coronavirus 229E From Environmental Surfaces Of A Classroom During Influenza Season

10.31988/scitrends.16688 ◽

2018 ◽

Author(s):

John Lednicky

Keyword(s):

Influenza Season ◽

Lessons Learned ◽

Human Coronavirus ◽

Environmental Surfaces ◽

Human Coronavirus 229E

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A protocol for adding knowledge to Wikidata: aligning resources on human coronaviruses

BMC Biology ◽

10.1186/s12915-020-00940-y ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Andra Waagmeester ◽

Egon L. Willighagen ◽

Andrew I. Su ◽

Martina Kutmon ◽

Jose Emilio Labra Gayo ◽

...

Keyword(s):

Common Ground ◽

Knowledge Bases ◽

Public Knowledge ◽

Medical Problems ◽

Human Coronavirus ◽

Data Schema ◽

Helping Others ◽

Human Coronaviruses ◽

The Right ◽

Human Coronavirus 229E

Abstract Background Pandemics, even more than other medical problems, require swift integration of knowledge. When caused by a new virus, understanding the underlying biology may help finding solutions. In a setting where there are a large number of loosely related projects and initiatives, we need common ground, also known as a “commons.” Wikidata, a public knowledge graph aligned with Wikipedia, is such a commons and uses unique identifiers to link knowledge in other knowledge bases. However, Wikidata may not always have the right schema for the urgent questions. In this paper, we address this problem by showing how a data schema required for the integration can be modeled with entity schemas represented by Shape Expressions. Results As a telling example, we describe the process of aligning resources on the genomes and proteomes of the SARS-CoV-2 virus and related viruses as well as how Shape Expressions can be defined for Wikidata to model the knowledge, helping others studying the SARS-CoV-2 pandemic. How this model can be used to make data between various resources interoperable is demonstrated by integrating data from NCBI (National Center for Biotechnology Information) Taxonomy, NCBI Genes, UniProt, and WikiPathways. Based on that model, a set of automated applications or bots were written for regular updates of these sources in Wikidata and added to a platform for automatically running these updates. Conclusions Although this workflow is developed and applied in the context of the COVID-19 pandemic, to demonstrate its broader applicability it was also applied to other human coronaviruses (MERS, SARS, human coronavirus NL63, human coronavirus 229E, human coronavirus HKU1, human coronavirus OC4).

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Examining the persistence of human Coronavirus 229E on fresh produce

Food Microbiology ◽

10.1016/j.fm.2021.103780 ◽

2021 ◽

pp. 103780

Author(s):

Madeleine Blondin-Brosseau ◽

Jennifer Harlow ◽

Tanushka Doctor ◽

Neda Nasheri

Keyword(s):

Fresh Produce ◽

Human Coronavirus ◽

Human Coronavirus 229E

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Effect of pH and temperature on the infectivity of human coronavirus 229E

Canadian Journal of Microbiology ◽

10.1139/m89-160 ◽

1989 ◽

Vol 35 (10) ◽

pp. 972-974 ◽

Cited By ~ 40

Author(s):

Alain Lamarre ◽

Pierre J. Talbot

Keyword(s):

Incubation Period ◽

Storage Conditions ◽

Viral Infectivity ◽

Human Coronavirus ◽

Virus Growth ◽

The Stability ◽

And Storage ◽

Influence Of Ph ◽

Human Coronavirus 229E

The stability of human coronavirus 229E infectivity was maximum at pH 6.0 when incubated at either 4 or 33 °C. However, the influence of pH was more pronounced at 33 °C. Viral infectivity was completely lost after a 14-day incubation period at 22, 33, or 37 °C but remained relatively constant at 4 °C for the same length of time. Finally, the infectious titer did not show any significant reduction when subjected to 25 cycles of thawing and freezing. These studies will contribute to optimize virus growth and storage conditions, which will facilitate the molecular characterization of this important pathogen.Key words: coronavirus, pH, temperature, infectivity, human coronavirus.

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