Desmosomal localization of beta-catenin in the skin of plakoglobin null-mutant mice

Development ◽  
1999 ◽  
Vol 126 (2) ◽  
pp. 371-381 ◽  
Author(s):  
C. Bierkamp ◽  
H. Schwarz ◽  
O. Huber ◽  
R. Kemler
Keyword(s):  
Heart Defects ◽  
Microscopic Analysis ◽  
Beta Catenin ◽  
Null Mutant ◽  
Electron Microscopic ◽  
Developmental Defects ◽  
Keratin Filaments ◽  
Embryonic Skin ◽  
Null Mutant Mice ◽  
And Function

Plakoglobin, a protein belonging to the Armadillo-repeat gene family, is the only component that adherens junctions and desmosomes have in common. Plakoglobin null-mutant mouse embryos die because of severe heart defects and may exhibit an additional skin phenotype, depending on the genetic background. Lack of plakoglobin affects the number and structure of desmosomes, resulting in visible defects when cells are subjected to increasing mechanical stress, e.g. when embryonic blood starts circulating or during skin differentiation. By analysing plakoglobin-negative embryonic skin differentiation in more detail, we show here that, in the absence of plakoglobin, its closest homologue, beta-catenin, becomes localized to desmosomes and associated with desmoglein. This substitution may account for the relatively late appearance of the developmental defects seen in plakoglobin null-mutant embryos. beta-catenin cannot, however, fully compensate a lack of plakoglobin. In the absence of plakoglobin, there was reduced cell-cell adhesion, resulting in large intercellular spaces between keratinocytes, subcorneal acantholysis and necrosis in the granular layer of the skin. Electron microscopic analysis documented a reduced number of desmosomes, and those present lacked the inner dense plaque and had fewer keratin filaments anchored. Our analysis underlines the central role of plakoglobin for desmosomal assembly and function during embryogenesis.

Microscopic radiography: A combined technique for improved analytical microscopic analysis and interpretration

1996 ◽  
Vol 54 ◽  
pp. 248-249
Author(s):  
A. Angel ◽  
R.C. Moretz
Keyword(s):  
Microscopic Analysis ◽  
Device Structure ◽  
Electron Microscopic ◽  
Drug Products ◽  
Microscopic Evaluation ◽  
Final Interpretation ◽  
And Function ◽  
Non Destructive ◽  
Combined Technique ◽  
Selection Of

Microscopic analysis of pharmaceutical devices and products relies primarily on destructive sample preparative methods, including sectioning, sawing, grinding and embedding. Reconstruction of images obtained from the prepared samples is often necessary to interpret the results. The preparative methods can introduce artifacts or distortion, which can affect the analysis, and the inability to visualize the intact object can also affect the interpretation. Radiography has been used to assist in the selection of preparative methods for microscopy, determine optimal orientation during preparation and analysis and to aid in the visualization and integration of the microscopic results in the final interpretation. The application of x-ray radiography to the examination of intact devices or manufactured drug products in conjunction with standard light and scanning electron microscopic evaluation presents a novel non-destructive technique to assess device structure and function as well as locate potential inclusions in manufactured drug products.

Electron microscopic analysis of the microtubular structure of HEp-2 cells

1990 ◽  
Vol 48 (3) ◽  
pp. 470-471
Author(s):  
Miklós S.Z. Kellermayer ◽  
Tamás Henics ◽  
György Szücs ◽  
Gerald H. Pollack
Keyword(s):  
Microscopic Analysis ◽  
Unknown Origin ◽  
Tubular Structure ◽  
Minimal Essential Medium ◽  
Electron Microscopic ◽  
Brij 58 ◽  
Cacodylate Buffer ◽  
Microscopic Studies ◽  
And Function ◽  
Hank’S Solution

The HEp-2 cell line was first established by Moore, et al. and today it serves as a widely used subject for a large variety of microbiological and cell-biological experiments. A tubular structure of unknown origin and function has been described first in virally infected, and later in uninfected HEp-2 cells. However, this tubular structure has not been further analyzed. We performed transmission (TEM) and whole cell mount electron microscopic studies of monolayer HEp-2 cells to morphologically describe the structure and relate it to other organelles of the cell.For embedding, HEp-2 cells were grown on glass coverslips in Eagle's minimal essential medium. 24 hours after passage, three types of procedures were carried out: (1) The cells were fixed with 2.5% glutaraldehyde in 0.14 M Na-cacodylate buffer. (2) The cells were treated for 10 min. with Hank's solution containing 0.2% Brij-58 and 2.5% glutaraldehyde. Fixation was completed with 2.5% glutaraldehyde in 0.14 M Na-cacodylate. (3) Cells were treated with 0.2% Brij-58 in Hank’s solution for 10 min. Subsequently, they were fixed with 2.5% glutaraldehyde in 0.14 M Na-cacodylate.

scholarly journals Methylation at nucleotide C62 in spliceosomal RNA U6 alters mRNA splicing which is important for embryonic development

2019 ◽  
Author(s):  
Allison Ogren ◽  
Nataliya Kibiryeva ◽  
Jennifer Marshall ◽  
James E. O’Brien ◽  
Douglas C. Bittel
Keyword(s):  
Alternative Splicing ◽  
Congenital Heart Defects ◽  
Congenital Heart ◽  
Heart Defects ◽  
Mrna Splicing ◽  
Developmental Defects ◽  
Small Noncoding Rnas ◽  
Spliceosomal Rnas ◽  
Mrna Transcripts ◽  
And Function

AbstractUnderstanding the regulation of development can help elucidate the pathogenesis behind many developmental defects found in humans and other vertebrates. Evidence has shown that alternative splicing of messenger RNA (mRNA) plays a role in developmental regulation, but our knowledge of the underlying mechanisms that regulate alternative splicing are inadequate. Notably, a subset of small noncoding RNAs known as scaRNAs (small cajal body associated RNAs) contribute to spliceosome maturation and function through covalently modifying spliceosomal RNAs by either methylating or pseudouridylating specific nucleotides, but the developmental significance of these modifications is not well understood. Our focus is on one such scaRNA, known as SNORD94 or U94, that methylates one specific cytosine (C62) on spliceosomal RNA U6, thus potentially altering spliceosome function during embryogenesis. We previously showed that mRNA splicing is significantly different in myocardium from infants with congenital heart defects (CHD) compared to controls. Furthermore, we showed that modifying expression of scaRNAs alters mRNA splicing in human cells, and zebrafish embryos. Here we present evidence that SNORD94 levels directly influence levels of methylation at C62 in U6, which we have previously shown is associated with altered splicing and congenital heart defects. The potential importance of scaRNAs as a developmentally important regulatory mechanism controlling alternative splicing of mRNA is unappreciated and needs more research.Author summarySplicing of mRNA transcripts by removal of introns and some non-critical exons is a crucial part of mRNA processing, gene expression, and cell function, and regulation of this process is still under investigation. Alternative splicing of mRNA transcripts of genes is tissue and time specific throughout life, although this process occurs everywhere in the body according to local tissue needs and signals. The spliceosome, the large ribonucleoprotein complex that carries out splicing, is biochemically modified by small noncoding RNAs, which is important for its structure and function. Here we show that the amount of 2’-O-ribose methylation at nucleotide C62 in spliceosomal RNA U6 is dependent on the level of the scaRNA SNORD94. We hypothesize that alternative splicing is dependent, at least in part, on biochemical modification to the spliceosomal RNAs. Furthermore, when scaRNA directed modifications are dysregulated, the result causes inappropriate alternative splicing that may contribute to developmental defects such as congenital heart defects. To our knowledge, this is the first demonstration that 2’-O-ribose methylation is indeed dependent on scaRNA levels in human cells and tissues.

scholarly journals Development, Maintenance, and Function of the Adrenal Gland in Early Postnatal Proopiomelanocortin-Null Mutant Mice

Endocrinology ◽  
2005 ◽  
Vol 146 (6) ◽  
pp. 2555-2562 ◽  
Author(s):  
Jason Karpac ◽  
Dirk Ostwald ◽  
Stephanie Bui ◽  
Peggy Hunnewell ◽  
Malini Shankar ◽  
...  
Keyword(s):  
Adrenal Glands ◽  
Mutant Mice ◽  
Null Mutant ◽  
Wild Type ◽  
Normal Morphology ◽  
Adrenal Hormones ◽  
Adrenal Cells ◽  
Mouse Mutants ◽  
Null Mutant Mice ◽  
And Function

Abstract Adult mouse mutants homozygous for an engineered proopiomelanocortin (POMC)-null allele lack macroscopically distinct adrenal glands and circulating adrenal hormones. To understand the basis for this adrenal defect, we compared the development of adrenal primordia in POMC-null mice and littermate controls. POMC-null mutant mice are born with adrenal glands that are morphologically indistinguishable from those of their wild-type littermates. However, in mutants adrenal cells fail to proliferate postnatally and adrenals atrophy until they have disappeared macroscopically in the adult. While present, mutant adrenals are differentiated as evidenced by the presence of enzymes for the final steps in the synthesis of corticosterone, aldosterone, and catecholamines. However, in contrast to adrenals of wild-type littermates, adrenals of POMC-null mutants do not produce corticosterone, not even in response to acute stimulation with exogenous ACTH. They do produce aldosterone; however, it is produced at reduced levels correlating with adrenal size. Transplantation of POMC-null mutant adrenals to adrenalectomized wild-type littermates results in adrenals with normal morphology and production of both corticosterone and aldosterone. These findings demonstrate that POMC peptides are not required for prenatal adrenal development and that POMC peptides in addition to ACTH are required for postnatal proliferation and maintenance of adrenal structures capable of producing both glucocorticoids and mineralocorticoids.

scholarly journals Expression and Externalization of Annexin 1 in the Adrenal Gland: Structure and Function of the Adrenal Gland in Annexin 1-Null Mutant Mice

Endocrinology ◽  
2007 ◽  
Vol 148 (3) ◽  
pp. 1030-1038 ◽  
Author(s):  
Evelyn Davies ◽  
Selma Omer ◽  
Julia C. Buckingham ◽  
John F. Morris ◽  
Helen C. Christian
Keyword(s):  
Adrenal Gland ◽  
Structure And Function ◽  
Mutant Mice ◽  
Null Mutant ◽  
Annexin 1 ◽  
Null Mutant Mice ◽  
And Function

scholarly journals Immunochemical characterization of three components of the hemidesmosome and their expression in cultured epithelial cells.

1989 ◽  
Vol 109 (6) ◽  
pp. 3377-3390 ◽  
Author(s):  
D H Klatte ◽  
M A Kurpakus ◽  
K A Grelling ◽  
J C Jones
Keyword(s):  
Epithelial Cells ◽  
Microscopic Analysis ◽  
Serum Samples ◽  
Electron Microscopic ◽  
Protein Preparation ◽  
Protein Mixture ◽  
Mouse Keratinocytes ◽  
And Function ◽  
Structure Assembly

Treatment of bovine tongue mucosa with 1 M KCl induced a split in the lamina densa of the basement membrane zone (BMZ). The epithelium was then separated from the underlying connective tissue. Electron microscopic analysis of the stripped epithelium revealed that hemidesmosomes and their associated intermediate filaments (IF) remain along the basal surface of the epithelium. This surface was solubilized in an SDS/urea-containing buffer. Characterization of components of this protein mixture was undertaken using human autoantibodies from bullous pemphigoid (BP) patients that have been shown to recognize hemidesmosomal plaque elements (Mutasim, D. F., Y. Takahashi, R. S. Labib, G. J. Anhalt, H. P. Patel, and L. A. Diaz. 1985. J. Invest. Dermatol. 84:47-53) and by production of mAbs. Affinity-purified autoantibodies directed against 180- and 240-kD polypeptides present in the protein preparation generated strong immunofluorescence staining patterns along the BMZ of bovine tongue mucosa. Furthermore, immunogold localization revealed that these two polypeptides are associated with the hemidesmosomal plaque. A mAb preparation directed against a 125-kD polypeptide present in this same protein mixture lamina lucida side of the hemidesmosome. Autoantibodies in BP serum samples, affinity-purified 180-kD autoantibodies and the mAb preparation generated a punctate stain along the substratum attached surface of epithelial cells maintained on glass substrata for approximately 1 wk. The spots appeared to be associated with bundles of IF in cultured mouse keratinocytes. These monospecific antibody probes should prove invaluable for the study of hemidesmosome structure, assembly, and function.

scholarly journals 016.Role of cited genes in placental morphogenesis: studies in null mutant mice

2004 ◽  
Vol 16 (9) ◽  
pp. 16
Author(s):  
S. L. Dunwoodie ◽  
S. L. Withington ◽  
D. B. Sparrow ◽  
A. N. Scott ◽  
J. I. Preis ◽  
...  
Keyword(s):  
Hypoxia Inducible Factor ◽  
Giant Cells ◽  
Maternal Blood ◽  
Postnatal Period ◽  
Null Mutant ◽  
Placental Development ◽  
Null Mutant Mice ◽  
And Function ◽  
Trophoblast Giant Cells ◽  
Null Mutants

Cited1 and Cited2 interact with CBP and p300. CBP/p300 bind numerous proteins and evidence exists, for Cited2 at least, that Cited binding prevents the binding of other proteins to CBP/p300. Since CBP/p300 interact with many proteins, can acetylate protein and DNA, and act as a ubiquitin ligase, it is likely that Cited1 and Cited2 function at a number of sites during development. We have generated mice that carry a null mutant allele for each of these genes. Analysis of null mutant embryos demonstrates that both Cited1 and Cited2 are required for normal embryonic development and survival. Although both Cited1 and Cited2 are expressed in the developing embryo and placenta, it appears that abnormal placental development and function is the cause of embryonic death. The defect that develops in the placentas of Cited1 null mutants is not apparent until late in gestation (16.5dpc). Cited1 null mutants are smaller than controls at birth and die during the early postnatal period. The placentas of these mutants are disorganised, with spongiotrophoblasts projecting in to the labyrinthine layer. In addition, resin casts of the maternal blood spaces within these placentas revealed extremely enlarged blood sinuses. We are searching for factors that could result in the increased size of the maternal blood sinuses. Cited2 null placentas and embryos are significantly smaller than controls; mutants die 3/4 the way through gestation (15.5dpc). The null mutant placentas have proportionally fewer spongiotrophoblasts, trophoblast giant cells and invasive trophoblasts. In addition, resin casts of fetal vasculature of the placenta reveal that the capillary network is underdeveloped. Through the isolation of trophoblast stem (TS) cells we are exploring the possibility that TS cell proliferation and/or differentiation is impaired due to a lack of Cited2. We suspect that the development of the phenotype may relate to the Hypoxia Inducible Factor-1a (HIF1a) transcription factor as Cited2 expression is induced by HIF1 and it acts to negatively regulate its activity.

Preparatory Procedures for Electron Microscopic Analysis at the Molecular Level of Resolution

1978 ◽  
Vol 36 (3) ◽  
pp. 516-520
Author(s):  
F.J. Sjostrand
Keyword(s):  
Electron Microscope ◽  
Molecular Level ◽  
Microscopic Analysis ◽  
Resolving Power ◽  
Cellular Structures ◽  
Electron Microscopic ◽  
Systematic Analysis ◽  
Technical Developments ◽  
Thin Sectioning ◽  
Obvious Reason

In the 1940's and 1950's electron microscopy conferences were attended with everybody interested in learning about the latest technical developments for one very obvious reason. There was the electron microscope with its outstanding performance but nobody could make very much use of it because we were lacking proper techniques to prepare biological specimens. The development of the thin sectioning technique with its perfectioning in 1952 changed the situation and systematic analysis of the structure of cells could now be pursued. Since then electron microscopists have in general become satisfied with the level of resolution at which cellular structures can be analyzed when applying this technique. There has been little interest in trying to push the limit of resolution closer to that determined by the resolving power of the electron microscope.

The Effect of Oxidized Cholesterol on the Aorta of Rabbits- Scanning Electron Microscopic Observations

1982 ◽  
Vol 40 ◽  
pp. 340-341
Author(s):  
S. K. Pena ◽  
C. B. Taylor ◽  
J. Hill ◽  
J. Safarik
Keyword(s):  
Cholesterol Biosynthesis ◽  
Cholesterol Content ◽  
Arterial Injury ◽  
Electron Microscopic ◽  
Aortic Smooth Muscle ◽  
Oxidized Cholesterol ◽  
Initial Event ◽  
Membrane Structure And Function ◽  
Scanning Electron Microscopic ◽  
And Function

Introduction: Oxidized cholesterol derivatives have been demonstrated in various cell cultures to be very potent inhibitors of 3-hvdroxy-3- methylglutaryl Coenzyme A reductase which is a principle regulator of cholesterol biosynthesis in the cell. The cholesterol content in the cells exposed to oxidized cholesterol was found to be markedly decreased. In aortic smooth muscle cells, the potency of this effect was closely related to the cytotoxicity of each derivative. Furthermore, due to the similarity of their molecular structure to that of cholesterol, these oxidized cholesterol derivatives might insert themselves into the cell membrane, alter membrane structure and function and eventually cause cell death. Arterial injury has been shown to be the initial event of atherosclerosis.

The Scanning Electron Microscopic Observation of the Vestibular Sensory Epithelia

1969 ◽  
Vol 27 ◽  
pp. 40-41
Author(s):  
D.J. Lim ◽  
W.C. Lane
Keyword(s):  
Semicircular Canals ◽  
Electron Microscopic Observation ◽  
Gravitational Acceleration ◽  
Electron Microscopic ◽  
Sensory Epithelia ◽  
Scanning Electron Microscopic ◽  
Vestibular Sensory Epithelia ◽  
And Function ◽  
Sand Piles ◽  
Active Investigation

The morphology and function of the vestibular sensory organs has been extensively studied during the last decade with the advent of electron microscopy and electrophysiology. The opening of the space age also accelerated active investigation in this area, since this organ is responsible for the sensation of balance and of linear, angular and gravitational acceleration.The vestibular sense organs are formed by the saccule, utricle and three ampullae of the semicircular canals. The maculae (sacculi and utriculi) have otolithic membranes on the top of the sensory epithelia. The otolithic membrane is formed by a layer of thick gelatin and sand-piles of calcium carbonate crystals (Fig.l).

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