scholarly journals Nanoscale FasL Organization on DNA Origami to Decipher Apoptosis Signal Activation in Cells

Small ◽  
2021 ◽  
pp. 2101678
Author(s):  
Ricarda M. L. Berger ◽  
Johann M. Weck ◽  
Simon M. Kempe ◽  
Oliver Hill ◽  
Tim Liedl ◽  
...  
Keyword(s):  
Dna Origami ◽  
Signal Activation ◽  
In Cells

scholarly journals Nanoscale Organization of FasL on DNA Origami as a Versatile Platform to Tune Apoptosis Signaling in Cells

2020 ◽  
Author(s):  
Ricarda M. L. Berger ◽  
Johann M. Weck ◽  
Simon M. Kempe ◽  
Tim Liedl ◽  
Joachim O. Rädler ◽  
...  
Keyword(s):  
Cell Biology ◽  
Fas Ligand ◽  
Dna Origami ◽  
Time To Death ◽  
Naturally Occurring ◽  
Soluble Fasl ◽  
Death Kinetics ◽  
Molecular Number ◽  
Signaling Efficiency ◽  
In Cells

AbstractNanoscale probes with fine-tunable properties are of key interest in cell biology and nanomedicine to elucidate and eventually control signaling processes in cells. A critical, still challenging issue is to conjugate these probes with molecules in a number- and spatially-controlled manner. Here, DNA origami-based nanoagents as nanometer precise scaffolds presenting Fas ligand (FasL) in well-defined arrangements to cells are reported. These nanoagents activate receptor molecules in the plasma membrane initiating apoptosis signaling in cells. Signaling for apoptosis depends sensitively on FasL geometry: fastest time-to-death kinetics are obtained for FasL nanoagents representing predicted structure models of hexagonal receptor ordering with 10 nm inter-molecular spacing. Slower kinetics are observed for one to two FasL on DNA origami or FasL coupled with higher flexibility. Nanoagents with FasL arranged in hexagons with small (5 nm) and large (30 nm) spacing impede signal transduction. Moreover, for predicted hexagonal FasL nanoagents, signaling efficiency is faster and 100× higher compared to naturally occurring soluble FasL. Incubation of the FasL-origami nanoagent in solution exhibited an EC50 value of only 90 pM. These studies present DNA origami as versatile signaling platforms to probe the significance of molecular number and nanoscale ordering for signal initiation in cells.

scholarly journals Brucella melitensis UGPase inhibits the activation of NF-κB by modulating the ubiquitination of NEMO

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Yucheng Zhou ◽  
Zhaoyang Bu ◽  
Jing Qian ◽  
Yuening Cheng ◽  
Lianjiang Qiao ◽  
...  
Keyword(s):  
Capsular Polysaccharide ◽  
Brucella Melitensis ◽  
Scientific Basis ◽  
Luciferase Assay ◽  
Wild Type ◽  
Proteomics Technology ◽  
Infected Cells ◽  
Related Proteins ◽  
Signal Activation ◽  
In Cells

Abstract Background UTP-glucose-1-phosphoryl transferase (UGPase) catalyzes the synthesis of UDP-glucose, which is essential for generating the glycogen needed for the synthesis of bacterial lipopolysaccharide (LPS) and capsular polysaccharide, which play important roles in bacterial virulence. However, the molecular function of UGPase in Brucella is still unknown. Results In this study, the ubiquitination modification of host immune-related protein in cells infected with UGPase-deleted or wild-type Brucella was analyzed using ubiquitination proteomics technology. The ubiquitination modification level and type of NF-κB Essential Modulator (NEMO or Ikbkg), a molecule necessary for NF-κB signal activation, was evaluated using Coimmunoprecipitation, Western blot, and dual-Luciferase Assay. We found 80 ubiquitin proteins were upregulated and 203 ubiquitin proteins were downregulated in cells infected with B. melitensis 16 M compared with those of B. melitensis UGPase-deleted strain (16 M-UGPase−). Moreover, the ubiquitin-modified proteins were mostly enriched in the categories of regulation of kinase/NF-κB signaling and response to a bacterium, suggesting Brucella UGPase inhibits ubiquitin modification of related proteins in the host NF-κB signaling pathway. Further analysis showed that the ubiquitination levels of NEMO K63 (K63-Ub) and Met1 (Met1-Ub) were significantly increased in the 16 M-UGPase−-infected cells compared with that of the 16 M-infected cells, further confirming that the ubiquitination levels of NF-κB signaling-related proteins were regulated by the bacterial UGPase. Besides, the expression level of IκBα was decreased, but the level of p-P65 was significantly increased in the 16 M-UGPase−-infected cells compared with that of the 16 M- and mock-infected cells, demonstrating that B. melitensis UGPase can significantly inhibit the degradation of IκBα and the phosphorylation of p65, and thus suppressing the NF-κB pathway. Conclusions The results of this study showed that Brucella melitensis UGPase inhibits the activation of NF-κB by modulating the ubiquitination of NEMO, which will provide a new scientific basis for the study of immune mechanisms induced by Brucella.

Cytoplasmic Tubules in Cells Infected with Laryngotracheitis Virus

1969 ◽  
Vol 27 ◽  
pp. 376-377
Author(s):  
A. M. Watrach
Keyword(s):  
Tissue Culture ◽  
Small Proportion ◽  
Chicken Embryo ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Morphologic Characteristics ◽  
Infectious Laryngotracheitis ◽  
Laryngotracheitis Virus ◽  
In Cells

During a study of the development of infectious laryngotracheitis (LT) virus in tissue culture cells, unusual tubular formations were found in the cytoplasm of a small proportion of the affected cells. It is the purpose of this report to describe the morphologic characteristics of the tubules and to discuss their possible association with the development of virus.The source and maintenance of the strain of LT virus have been described. Prior to this study, the virus was passed several times in chicken embryo kidney (CEK) tissue culture cells.

Effect of Some Metabolic Inhibitors on Vinblastine-Induced Ribosomal-Granular Material Complexes

1971 ◽  
Vol 29 ◽  
pp. 548-549
Author(s):  
Awtar Krishan ◽  
Dora Hsu
Keyword(s):  
Granular Material ◽  
Rna Synthesis ◽  
Culture Medium ◽  
Actinomycin D ◽  
Metabolic Inhibitors ◽  
Protein And Rna Synthesis ◽  
Apparent Reduction ◽  
Plant Alkaloids ◽  
In Cells

Cells exposed to antitumor plant alkaloids, vinblastine and vincristine sulfate have large proteinacious crystals and complexes of ribosomes, helical polyribosomes and electron-dense granular material (ribosomal complexes) in their cytoplasm, Binding of H3-colchicine by the in vivo crystals shows that they contain microtubular proteins. Association of ribosomal complexes with the crystals suggests that these structures may be interrelated.In the present study cultured human leukemic lymphoblasts (CCRF-CEM), were incubated with protein and RNA-synthesis inhibitors, p. fluorophenylalanine, puromycin, cycloheximide or actinomycin-D before the addition of crystal-inducing doses of vinblastine to the culture medium. None of these compounds could completely prevent the formation of the ribosomal complexes or the crystals. However, in cells pre-incubated with puromycin, cycloheximide, or actinomycin-D, a reduction in the number and size of the ribosomal complexes was seen. Large helical polyribosomes were absent in the ribosomal complexes of cells treated with puromycin, while in cells exposed to cycloheximide, there was an apparent reduction in the number of ribosomes associated with the ribosomal complexes (Fig. 2).

Detection of Nucleic Acids in Cells or Tissue Sections Using In situ Polymerase Chain Reaction (PCR)

1996 ◽  
Vol 54 ◽  
pp. 16-17
Author(s):  
J. R. Hully ◽  
K. R. Luehrsen ◽  
K. Aoyagi ◽  
C. Shoemaker ◽  
R. Abramson
Keyword(s):  
Nucleic Acids ◽  
Viral Infections ◽  
Anatomical Location ◽  
Rt Pcr ◽  
Reverse Transcription Pcr ◽  
Cellular Source ◽  
Gene Transcripts ◽  
Initial Description ◽  
In Cells

The development of PCR technology has greatly accelerated medical research at the genetic and molecular levels. Until recently, the inherent sensitivity of this technique has been limited to isolated preparations of nucleic acids which lack or at best have limited morphological information. With the obvious exception of cell lines, traditional PCR or reverse transcription-PCR (RT-PCR) cannot identify the cellular source of the amplified product. In contrast, in situ hybridization (ISH) by definition, defines the anatomical location of a gene and/or it’s product. However, this technique lacks the sensitivity of PCR and cannot routinely detect less than 10 to 20 copies per cell. Consequently, the localization of rare transcripts, latent viral infections, foreign or altered genes cannot be identified by this technique. In situ PCR or in situ RT-PCR is a combination of the two techniques, exploiting the sensitivity of PCR and the anatomical definition provided by ISH. Since it’s initial description considerable advances have been made in the application of in situ PCR, improvements in protocols, and the development of hardware dedicated to in situ PCR using conventional microscope slides. Our understanding of the importance of viral latency or viral burden in regards to HIV, HPV, and KSHV infections has benefited from this technique, enabling detection of single viral copies in cells or tissue otherwise thought to be normal. Clearly, this technique will be useful tool in pathobiology especially carcinogenesis, gene therapy and manipulations, the study of rare gene transcripts, and forensics.

Absence of temporal ordering of apoptotic features in heat-shock treated leukemia and lymphoma cell lines

1996 ◽  
Vol 54 ◽  
pp. 758-759
Author(s):  
D. W. Fairbain ◽  
M.D. Standing ◽  
K.L. O'Neill
Keyword(s):  
Heat Shock ◽  
Cell Lines ◽  
Chromatin Condensation ◽  
Membrane Integrity ◽  
Resistant Cell ◽  
Membrane Blebbing ◽  
Late Loss ◽  
Induced Apoptosis ◽  
Dying Cells ◽  
In Cells

Apoptosis is a genetically defined response to physiological stimuli that results in cellular suicide. Features common to apoptotic cells include chromatin condensation, oligonucleosomal DNA fragmentation, membrane blebbing, nuclear destruction, and late loss of ability to exclude vital dyes. These characteristics contrast markedly from pathological necrosis, in which membrane integrity loss is demonstrated early, and other features of apoptosis, which allow a non-inflammatory removal of dead and dying cells, are absent. Using heat shock-induced apoptosis as a model for examining stress response in cells, we undertook to categorize a variety of human leukemias and lymphomas with regard to their response to heat shock. We were also interested in determining whether a common temporal order was followed in cells dying by apoptosis. In addition, based on our previous results, we investigated whether increasing heat load resulted in increased apoptosis, with particular interest in relatively resistant cell lines, or whether the mode of death changed from apoptosis to necrosis.

Umatilla Virus Association with Fibrils and Tubules in Cultured Cells

1980 ◽  
Vol 38 ◽  
pp. 476-477
Author(s):  
Neil M. Foster ◽  
Ruth D. Breckon
Keyword(s):  
Bluetongue Virus ◽  
Linear Array ◽  
Cultured Cells ◽  
Intimate Association ◽  
Infected Cells ◽  
Dark Staining ◽  
In Cells

Macrotubules have been described1 in cells infected with Umatilla virus (UMAV), an orbivirus for which bluetongue virus (BTV) is the protype. Macrotubules, often in linear array, were observed in the cytoplasm and in intimate association with viroplasms of infected cells. Macrotubules had outside and inside diameters of 20 and 15 nm and many had dark-staining centers with diameters similar to the interiors of the tubules. UMAV was 60 nm and the RNA core was 30 nm in diameter. This report describes the association of UMAV with macrotubules and two types of microtubules.

Photo-tethered molecular beacons for superior light-induction

2021 ◽  
Author(s):  
Robin Klimek ◽  
Mantian Wang ◽  
Vivien R. McKenney ◽  
Erin M. Schuman ◽  
Alexander Heckel
Keyword(s):  
Molecular Beacons ◽  
Light Induction ◽  
In Cells

Photolabile circularization of molecular beacons via backbone phosphates leads to superior probes to study spatiotemporal aspects of RNA in cells.

Use of the comet assay to measure DNA damage in cells exposed to photosensitizers and gamma radiation

1999 ◽  
Vol 96 (1) ◽  
pp. 143-146 ◽  
Author(s):  
J.-P. Pouget ◽  
J.-L. Ravanat ◽  
T. Douki ◽  
M.-J. Richard ◽  
J. Cadet
Keyword(s):  
Dna Damage ◽  
Comet Assay ◽  
Gamma Radiation ◽  
In Cells
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