scholarly journals armadillo, bazooka, and stardust are critical for early stages in formation of the zonula adherens and maintenance of the polarized blastoderm epithelium in Drosophila.

1996 ◽  
Vol 134 (1) ◽  
pp. 149-163 ◽  
Author(s):  
H A Müller ◽  
E Wieschaus
Keyword(s):  
Plasma Membranes ◽  
Basolateral Membrane ◽  
Cell Sheet ◽  
Cell Monolayer ◽  
Test System ◽  
Sem Analysis ◽  
Drosophila Embryo ◽  
Early Stages ◽  
A Cell ◽  
The Stability

Cellularization of the Drosophila embryo results in the formation of a cell monolayer with many characteristics of a polarized epithelium. We have used antibodies specific to cellular junctions and nascent plasma membranes to study the formation of the zonula adherens (ZA) in relation to the establishment of basolateral membrane polarity. The same approach was then used as a test system to identify X-linked zygotically active genes required for ZA formation. We show that ZA formation begins during cellularization and that the basolateral membrane domain is established at mid-gastrulation. By creating deficiencies for defined regions of the X chromosome, we have identified genes that are required for the formation of the ZA and the generation of basolateral membrane polarity. We show that embryos mutant for both stardust (sdt) and bazooka (baz) fail to form a ZA. In addition to the failure to establish the ZA, the formation of the monolayered epithelium is disrupted after cellularization, resulting in formation of a multilayered cell sheet by mid-gastrulation. SEM analysis of mutant embryos revealed a conversion of cells exhibiting epithelial characteristics into cells exhibiting mesenchymal characteristics. To investigate how mutations that affect an integral component of the ZA itself influence ZA formation, we examined embryos with reduced maternal and zygotic supply of wild-type Arm protein. These embryos, like embryos mutant for both sdt and baz, exhibit an early disruption of ZA formation. These results suggest that early stages in the assembly of the ZA are critical for the stability of the polarized blastoderm epithelium.

Using FRET to Measure the Time it Takes for a Cell to Destroy a Virus

2019 ◽  
Author(s):  
Candace E. Benjamin ◽  
Zhuo Chen ◽  
Olivia Brohlin ◽  
Hamilton Lee ◽  
Stefanie Boyd ◽  
...  
Keyword(s):  
Cellular Uptake ◽  
Rhodamine B ◽  
Virus Like Particle ◽  
A Cell ◽  
Viral Nanoparticles ◽  
The Stability ◽  
Open Question ◽  
Viral Surface ◽  
Fret Pair

<div><div><div><p>The emergence of viral nanotechnology over the preceding two decades has created a number of intellectually captivating possible translational applications; however, the in vitro fate of the viral nanoparticles in cells remains an open question. Herein, we investigate the stability and lifetime of virus-like particle (VLP) Qβ - a representative and popular VLP for several applications - following cellular uptake. By exploiting the available functional handles on the viral surface, we have orthogonally installed the known FRET pair, FITC and Rhodamine B, to gain insight of the particle’s behavior in vitro. Based on these data, we believe VLPs undergo aggregation in addition to the anticipated proteolysis within a few hours of cellular uptake.</p></div></div></div>

Analysis of the Dynamics of the Electric Capacitance of a Cell Monolayer at the Late Stages of Caco-2 cell Differentiation

2019 ◽  
Vol 35 (6) ◽  
pp. 87-90
Author(s):  
S.V. Nikulin ◽  
V.A. Petrov ◽  
D.A. Sakharov
Keyword(s):  
Impedance Spectroscopy ◽  
Cell Monolayer ◽  
Microfluidic Chips ◽  
Russian Science ◽  
Electric Capacitance ◽  
A Cell ◽  
Static Conditions ◽  
Late Stages

The real-time monitoring of electric capacitance (impedance spectroscopy) allowed obtaining evidence that structures which look like intestinal villi can be formed during the cultivation under static conditions as well as during the cultivation in microfluidic chips. It was shown in this work via transcriptome analysis that the Hh signaling pathway is involved in the formation of villus-like structures in vitro, which was previously shown for their formation in vivo. impedance spectroscopy, intestine, villi, electric capacitance, Hh The study was funded by the Russian Science Foundation (Project 16-19-10597).

Polycystin expression is temporally and spatially regulated during renal development

1997 ◽  
Vol 272 (5) ◽  
pp. F602-F609 ◽  
Author(s):  
J. Van Adelsberg ◽  
S. Chamberlain ◽  
V. D'Agati
Keyword(s):  
Plasma Membranes ◽  
Predicted Amino Acid Sequence ◽  
Renal Development ◽  
Polycystic Kidney ◽  
Fetal Kidney ◽  
Spatial Regulation ◽  
Adult Kidney ◽  
A Cell ◽  
Autosomal Dominant Polycystic Kidney ◽  
Membrane Spanning

Mutations in PKD1 cause autosomal dominant polycystic kidney disease (ADPKD), a common genetic disease in which cysts form from kidney tubules. The predicted product of this gene is a novel protein with cell-adhesive and membrane-spanning domains. To test the hypothesis that polycystin, the product of the PKD1 gene, is a cell adhesion molecule, we raised antibodies against peptides derived from the unduplicated, membrane-spanning portion of the predicted amino acid sequence. These antibodies recognized membrane-associated polypeptides of 485 and 245 kDa in human fetal kidney homogenates. Expression was greater in fetal than adult kidney by both Western blot analysis and immunofluorescence. In fetal kidney, polycystin was localized to the plasma membranes of ureteric bud and comma and S-shaped bodies. However, in more mature tubules in fetal kidney, in adult kidney, and in polycystic kidney, the majority of polycystin staining was intracellular. The temporal and spatial regulation of polycystin expression during renal development lead us to speculate that polycystin may play a role in nephrogenesis.

2,4-Dichlorophenol and MCPA in a V79 Test

1988 ◽  
Vol 15 (3) ◽  
pp. 245-250
Author(s):  
Geirid Fiskesjö
Keyword(s):  
Chinese Hamster ◽  
Chinese Hamster Cell ◽  
Test System ◽  
Similar Degree ◽  
Human Beings ◽  
Industrial Chemicals ◽  
Allium Test ◽  
Chinese Hamster Cell Line ◽  
A Cell ◽  
Mutagenic Effects

Two industrial chemicals, 2,4-dichlorophenol and 4-chloro-2-methylphenoxyacetic acid (MCPA), which have no toxic effects on the Chinese hamster cell line V79 alone, were tested for toxicity and mutagenicity in a cell-mediated test, where mixed-function oxidase (MFO) enzymes are active in the metabolism of xenobiotics. For 2,4-dichlorophenol, a dose-dependent toxicity as well as a slight mutagenicity could be shown when oxygenation enzymes were present. A similar degree of toxicity in a plant test system (the Allium test) indicates a similar risk of damage from exposure to dichlorophenol treatments in both these systems. MCPA did not induce any toxic or mutagenic effects at the concentrations tested. These results were not in agreement with previous results in plant material, where MCPA was clearly toxic at relatively low doses. However, since chlorophenols have been found in plants sprayed with phenoxyacetic acids, further investigations should be performed concerning potential risk to human beings.

scholarly journals A cell-free assay for the insertion of a viral glycoprotein into the plasma membrane.

1986 ◽  
Vol 103 (5) ◽  
pp. 1829-1835 ◽  
Author(s):  
P G Woodman ◽  
J M Edwardson
Keyword(s):  
Influenza Virus ◽  
Plasma Membrane ◽  
Trichloroacetic Acid ◽  
Plasma Membranes ◽  
Semliki Forest Virus ◽  
Fusion Reaction ◽  
Viral Glycoprotein ◽  
Acetylneuraminic Acid ◽  
Donor And Acceptor ◽  
A Cell

A cell-free assay has been developed for the delivery of influenza virus neuraminidase to the plasma membrane. Two types of postnuclear supernatant, which acted as donor and acceptor of the enzyme, were prepared from baby hamster kidney cells. Donor preparations were obtained from cells infected with influenza virus and containing neuraminidase en route to the plasma membrane. Acceptor preparations were obtained from cells containing, bound to their plasma membranes, Semliki Forest virus with envelope glycoproteins bearing [3H]N-acetylneuraminic acid. Fusion between vesicles from these two preparations permits access of the enzyme to its substrate, which results in the release of free [3H]N-acetylneuraminic acid. This release was detected through the transfer of radioactivity from a trichloroacetic acid-insoluble to a trichloroacetic acid-soluble fraction. An ATP-dependent component of release was found, which appears to be a consequence of vesicle fusion. This component was enhanced when the donor was prepared from cells in which the enzyme had been concentrated in a compartment between the Golgi complex and the plasma membrane, which indicates that a specific exocytic fusion event has been reconstituted. The extent of fusion is greatly reduced by pre-treatment of donor and acceptor preparations with trypsin, which points to the involvement of proteins in the fusion reaction.

Syndesmos, a protein that interacts with the cytoplasmic domain of syndecan-4, mediates cell spreading and actin cytoskeletal organization

2000 ◽  
Vol 113 (2) ◽  
pp. 315-324 ◽  
Author(s):  
P.C. Baciu ◽  
S. Saoncella ◽  
S.H. Lee ◽  
F. Denhez ◽  
D. Leuthardt ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Focal Adhesions ◽  
Cell Spreading ◽  
Cytoplasmic Domain ◽  
Cellular Protein ◽  
Actin Stress Fiber ◽  
Independent Manner ◽  
Intracellular Proteins ◽  
Central Regions ◽  
A Cell

Syndecan-4 is a cell surface heparan sulfate proteoglycan which, in cooperation with integrins, transduces signals for the assembly of focal adhesions and actin stress fibers in cells plated on fibronectin. The regulation of these cellular events is proposed to occur, in part, through the interaction of the cytoplasmic domains of these transmembrane receptors with intracellular proteins. To identify potential intracellular proteins that interact with the cytoplasmic domain of syndecan-4, we carried out a yeast two-hybrid screen in which the cytoplasmic domain of syndecan-4 was used as bait. As a result of this screen, we have identified a novel cellular protein that interacts with the cytoplasmic domain of syndecan-4 but not with those of the other three syndecan family members. The interaction involves both the membrane proximal and variable central regions of the cytoplasmic domain. We have named this cDNA and encoded protein syndesmos. Syndesmos is ubiquitously expressed and can be myristylated. Consistent with its myristylation and syndecan-4 association, syndesmos colocalizes with syndecan-4 in the ventral plasma membranes of cells plated on fibronectin. When overexpressed in NIH 3T3 cells, syndesmos enhances cell spreading, actin stress fiber and focal contact formation in a serum-independent manner.

Apical beta 1 integrin in polarized MDCK cells mediates tubulocyst formation in response to type I collagen overlay

1996 ◽  
Vol 109 (7) ◽  
pp. 1875-1889 ◽  
Author(s):  
A. Zuk ◽  
K.S. Matlin
Keyword(s):  
Plasma Membranes ◽  
Type I Collagen ◽  
Mdck Cells ◽  
Basolateral Membrane ◽  
Collagen Gel ◽  
Type I ◽  
Membrane Markers ◽  
Polarized Epithelia ◽  
Beta 1 Integrin ◽  
Type I Collagen Gel

A number of epithelia form tubulocysts in vitro when overlaid with type I collagen gel. Because collagen receptors are generally believed to be expressed on the basolateral domain, the mechanism by which collagen elicits this morphogenetic response from the apical surface is unclear. To investigate the role of beta 1 integrins, the major receptor family for collagen, in this process, we overlaid polarized monolayers of MDCK II cells grown on permeable supports with type I collagen gel and correlated integrin polarity with the polarity of other apical and basolateral membrane markers during tubulocyst formation. Polarized monolayers of one clone of MDCK II cells, referred to as Heidelberg MDCK, initially respond to collagen overlay by stratifying; within 48 hours, lumena develop between the cell layers giving rise to tubulocysts. Tight junctions remain intact during tubulocyst formation because transepithelial electrical resistance does not significantly change. Major alterations are observed, however, in the expression and localization of apical and basolateral membrane markers. beta 1 integrins are necessary for tubulocyst morphogenesis because a function-blocking antibody administered to the apical pole of the cells completely inhibits the formation of these structures. To determine how apical-cell collagen interactions elicit tubulocyst formation, we examined whether beta 1 integrins are mobilized to apical plasma membranes in response to collagen overlay. We found that in the absence of collagen, polarized monolayers of Heidelberg MDCK cells endogenously express on apical plasma membranes a small pool of the beta 1 family, including alpha 2 beta 1 and alpha 3 beta 1. Collagen overlay does not mobilize additional beta 1 integrins to apical domains. If beta 1 integrins are not already apically expressed, as in the C6 MDCK cell line (Schoenenberger et al. (1994) J. Cell Biol. 107, 527–541), beta 1 integrins are not directed apically and tubulocysts do not develop in response to collagen. Thus, interaction of beta 1 integrin pre-existing on apical plasma membranes of polarized epithelia with type I collagen gel is the mechanism by which apical application of collagen elicits the formation of tubulocysts. Depolarized integrins on apical plasma membranes of polarized epithelia may be relevant to the pathogenesis of disease and injury.

scholarly journals The Krebs Cycle Enzyme α-Ketoglutarate Decarboxylase Is an Essential Glycosomal Protein in Bloodstream African Trypanosomes

2014 ◽  
Vol 14 (3) ◽  
pp. 206-215 ◽  
Author(s):  
Steven Sykes ◽  
Anthony Szempruch ◽  
Stephen Hajduk
Keyword(s):  
Plasma Membranes ◽  
Fluorescent Protein ◽  
Krebs Cycle ◽  
Content Type ◽  
African Trypanosomes ◽  
Atp Production ◽  
Cycle Enzyme ◽  
A Cell ◽  
Direct Localization ◽  
Green Fluorescent

ABSTRACT α-Ketoglutarate decarboxylase (α-KDE1) is a Krebs cycle enzyme found in the mitochondrion of the procyclic form (PF) of Trypanosoma brucei . The bloodstream form (BF) of T. brucei lacks a functional Krebs cycle and relies exclusively on glycolysis for ATP production. Despite the lack of a functional Krebs cycle, α-KDE1 was expressed in BF T. brucei and RNA interference knockdown of α-KDE1 mRNA resulted in rapid growth arrest and killing. Cell death was preceded by progressive swelling of the flagellar pocket as a consequence of recruitment of both flagellar and plasma membranes into the pocket. BF T. brucei expressing an epitope-tagged copy of α-KDE1 showed localization to glycosomes and not the mitochondrion. We used a cell line transfected with a reporter construct containing the N-terminal sequence of α-KDE1 fused to green fluorescent protein to examine the requirements for glycosome targeting. We found that the N-terminal 18 amino acids of α-KDE1 contain overlapping mitochondrion- and peroxisome-targeting sequences and are sufficient to direct localization to the glycosome in BF T. brucei . These results suggest that α-KDE1 has a novel moonlighting function outside the mitochondrion in BF T. brucei .

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