scholarly journals Ribonucleoprotein staining of centrioles and kinetochores in newt lung cell spindles.

1979 ◽  
Vol 80 (1) ◽  
pp. 1-9 ◽  
Author(s):  
C L Rieder
Keyword(s):  
Mitotic Spindle ◽  
Lung Epithelial Cells ◽  
Staining Reaction ◽  
Voltage Electron ◽  
High Voltage Electron Microscope ◽  
Lung Epithelial ◽  
Pericentriolar Material ◽  
Rnase Digestion ◽  
Spindle Microtubules ◽  
Positively Charged

The distribution of ribonucleoprotein (RNP) within the mitotic spindle of newt lung epithelial cells was studied with the high voltage electron microscope (HVEM) using Bernhard's uranyl-EDTA-lead staining of thick sections in conjunction with the ribonuclease digestion of fixed cells. The results indicate that aside from ribosomes, the major RNP-containing components of the spindle are the kinetochores and centrioles, both of which stain electron-opaque after EDTA treatment. In both cases, the electron-opaque material associated with these microtubule organizing centers (MTOC's) can be removed by RNAse digestion and cold perchloric acid (PCA) extraction under conditions which leave the spindle microtubules (Mts) centrioles, and kinetochores intact. The staining reaction is not abolished by cold PCA extraction alone or by substituting other positively charged proteins (i.e., cytochrome c or lysozyme) for RNAse. The RNP component of the kinetochore is closely associated with the bases of the kinetochore microtubules. The RNP component of the centriole can be seen to surround the microtubules of the triplet blades. No evidence was found to indicate the presence of RNP in the pericentriolar material. The possible function of both kinetochore and centriolar RNP is discussed.

The Xenopus protein kinase pEg2 associates with the centrosome in a cell cycle-dependent manner, binds to the spindle microtubules and is involved in bipolar mitotic spindle assembly

1998 ◽  
Vol 111 (5) ◽  
pp. 557-572 ◽  
Author(s):  
C. Roghi ◽  
R. Giet ◽  
R. Uzbekov ◽  
N. Morin ◽  
I. Chartrain ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Mitotic Spindle ◽  
Cultured Cells ◽  
Dependent Manner ◽  
Egg Extracts ◽  
Pericentriolar Material ◽  
Spindle Microtubules ◽  
Cycle Dependent

By differential screening of a Xenopus laevis egg cDNA library, we have isolated a 2,111 bp cDNA which corresponds to a maternal mRNA specifically deadenylated after fertilisation. This cDNA, called Eg2, encodes a 407 amino acid protein kinase. The pEg2 sequence shows significant identity with members of a new protein kinase sub-family which includes Aurora from Drosophila and Ipl1 (increase in ploidy-1) from budding yeast, enzymes involved in centrosome migration and chromosome segregation, respectively. A single 46 kDa polypeptide, which corresponds to the deduced molecular mass of pEg2, is immunodetected in Xenopus oocyte and egg extracts, as well as in lysates of Xenopus XL2 cultured cells. In XL2 cells, pEg2 is immunodetected only in S, G2 and M phases of the cell cycle, where it always localises to the centrosomal region of the cell. In addition, pEg2 ‘invades’ the microtubules at the poles of the mitotic spindle in metaphase and anaphase. Immunoelectron microscopy experiments show that pEg2 is located precisely around the pericentriolar material in prophase and on the spindle microtubules in anaphase. We also demonstrate that pEg2 binds directly to taxol stabilised microtubules in vitro. In addition, we show that the presence of microtubules during mitosis is not necessary for an association between pEg2 and the centrosome. Finally we show that a catalytically inactive pEg2 kinase stops the assembly of bipolar mitotic spindles in Xenopus egg extracts.

Analysis of SARS-CoV-2 ORF3a structure reveals chloride binding sites

2020 ◽  
Author(s):  
Valeria Marquez-Miranda ◽  
Maximiliano Rojas ◽  
Yorley Duarte ◽  
Ignacio Diaz-Franulic ◽  
Miguel Holmgren ◽  
...  
Keyword(s):  
Transmembrane Domain ◽  
Lung Epithelial Cells ◽  
Transmembrane Helices ◽  
Cellular Processes ◽  
Charged Amino Acids ◽  
Coordination Sites ◽  
Lung Epithelial ◽  
Dynamics Simulations ◽  
Positively Charged

AbstractSARS-CoV-2 ORF3a is believed to form ion channels, which may be involved in the modulation of virus release, and has been implicated in various cellular processes like the up-regulation of fibrinogen expression in lung epithelial cells, downregulation of type 1 interferon receptor, caspase-dependent apoptosis, and increasing IFNAR1 ubiquitination. ORF3a assemblies as homotetramers, which are stabilized by residue C133. A recent cryoEM structure of a homodimeric complex of ORF3a has been released. A lower-resolution cryoEM map of the tetramer suggests two dimers form it, arranged side by side. The dimer’s cryoEM structure revealed that each protomer contains three transmembrane helices arranged in a clockwise configuration forming a six helices transmembrane domain. This domain’s potential permeation pathway has six constrictions narrowing to about 1 Å in radius, suggesting the structure solved is in a closed or inactivated state. At the cytosol end, the permeation pathway encounters a large and polar cavity formed by multiple beta strands from both protomers, which opens to the cytosolic milieu. We modeled the tetramer following the arrangement suggested by the low-resolution tetramer cryoEM map. Molecular dynamics simulations of the tetramer embedded in a membrane and solvated with 0.5 M of KCl were performed. Our simulations show the cytosolic cavity is quickly populated by both K+ and Cl-, yet with different dynamics. K+ ions moved relatively free inside the cavity without forming proper coordination sites. In contrast, Cl- ions enter the cavity, and three of them can become stably coordinated near the intracellular entrance of the potential permeation pathway by an inter-subunit network of positively charged amino acids. Consequently, the central cavity’s electrostatic potential changed from being entirely positive at the beginning of the simulation to more electronegative at the end.

Electron-microscopic study of the spindle and chromosome movement in the yeast Saccharomyces cerevisiae

1976 ◽  
Vol 22 (2) ◽  
pp. 219-242 ◽  
Author(s):  
J.B. Peterson ◽  
H. Ris
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Spindle Pole ◽  
Linkage Groups ◽  
Electron Microscopic ◽  
Yeast Saccharomyces Cerevisiae ◽  
Genetic Studies ◽  
Voltage Electron ◽  
High Voltage Electron Microscope ◽  
Spindle Microtubules ◽  
Diploid Cells

Mitosis in yeast Saccharomyces cerevisiae was investigated in thick (0-25-I mum) serial sections with a high voltage electron microscope and in preparations of spheroplasts spread on a water surface. Spindle microtubules originate from a plaque-like structure called the spindle pole bosis the SPB duplicates and a set of long and short microtubules develops on each SPB. The spindle arises as the SPBs separate on the nuclear membrane adense and are not individually visible. Genetic studies, however, have indicated that there are 17 linkage groups. The number of microtubules was determined in diploid and haploid spindles on serial stereo micrographs. In diploid mitosis about 40 microtubules issue from a SPB. Most are non-continuous and often they are visibly associated with a chromatin fibre. The spindle in haploid cells is similar except that the number of microtubules is about half that in diploid cells and the SPB is smaller. The pole-to-pole microtubules vary in number from spindle to spindle, but in each case enough microtubules are present to account for each linkage group being associated with a single non-continuous microtubule. We conclude that mitosis in yeast is comparable in its general aspect to that observed in typical eukaryotes.

Centrosomes and mitotic spindle poles: a recent liaison?

2015 ◽  
Vol 43 (1) ◽  
pp. 13-18 ◽  
Author(s):  
Pavithra L. Chavali ◽  
Isabel Peset ◽  
Fanni Gergely
Keyword(s):  
Cell Division ◽  
Mitotic Spindle ◽  
Potential Benefit ◽  
Cell Types ◽  
Daughter Cells ◽  
Spindle Poles ◽  
Pericentriolar Material ◽  
Mitotic Spindle Assembly ◽  
Spindle Microtubules ◽  
Ordered Assembly

Centrosomes comprise two cylindrical centrioles embedded in the pericentriolar material (PCM). The PCM is an ordered assembly of large scaffolding molecules, providing an interaction platform for proteins involved in signalling, trafficking and most importantly microtubule nucleation and organization. In mitotic cells, centrosomes are located at the spindle poles, sites where spindle microtubules converge. However, certain cell types and organisms lack centrosomes, yet contain focused spindle poles, highlighting that despite their juxtaposition in cells, centrosomes and mitotic spindle poles are distinct physical entities. In the present paper, we discuss the origin of centrosomes and summarize their contribution to mitotic spindle assembly and cell division. We then describe the key molecular players that mediate centrosome attachment to mitotic spindle poles and explore why co-segregation of centrosomes and spindle poles into daughter cells is of potential benefit to organisms.

A New 1000kv Electron Microscope

1969 ◽  
Vol 27 ◽  
pp. 100-101
Author(s):  
J.L. Williams ◽  
K. Heathcote ◽  
E.J. Greer
Keyword(s):  
Electron Microscope ◽  
Direct Observation ◽  
High Voltage ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Specimen Thickness ◽  
Voltage Electron ◽  
High Voltage Electron Microscope ◽  
Dynamic Experiments ◽  
Conventional Instrument

High Voltage Electron Microscope already offers exciting experimental possibilities to Biologists and Materials Scientists because the increased specimen thickness allows direct observation of three dimensional structure and dynamic experiments on effectively bulk specimens. This microscope is designed to give maximum accessibility and space in the specimen region for the special stages which are required. At the same time it provides an ease of operation similar to a conventional instrument.

A New 1,000 kV Electron Microscope

1967 ◽  
Vol 25 ◽  
pp. 260-261
Author(s):  
M. Nishigaki ◽  
S. Katagiri ◽  
H. Kimura ◽  
B. Tadano
Keyword(s):  
Electron Microscope ◽  
High Voltage ◽  
Basic Research ◽  
Chromatic Aberration ◽  
High Accuracy ◽  
Biological Specimen ◽  
Voltage Electron ◽  
High Voltage Electron Microscope ◽  
High Voltage Electron

The high voltage electron microscope has many advantageous features in comparison with the ordinary electron microscope. They are a higher penetrating efficiency of the electron, low chromatic aberration, high accuracy of the selected area diffraction and so on. Thus, the high voltage electron microscope becomes an indispensable instrument for the metallurgical, polymer and biological specimen studies. The application of the instrument involves today not only basic research but routine survey in the various fields. Particularly for the latter purpose, the performance, maintenance and reliability of the microscope should be same as those of commercial ones. The authors completed a 500 kV electron microscope in 1964 and a 1,000 kV one in 1966 taking these points into consideration. The construction of our 1,000 kV electron microscope is described below.

Design of Sensitive Photographic Materials for the High-Voltage Electron Microscope

1975 ◽  
Vol 33 ◽  
pp. 156-157
Author(s):  
Murray Vernon King ◽  
Donald F. Parsons
Keyword(s):  
Electron Microscope ◽  
Radiation Damage ◽  
High Voltage ◽  
Radiation Sensitivity ◽  
Fast Electrons ◽  
Voltage Electron ◽  
High Voltage Electron Microscope ◽  
High Voltage Electron ◽  
Biological Studies ◽  
Low Sensitivity

Effective application of the high-voltage electron microscope to a wide variety of biological studies has been restricted by the radiation sensitivity of biological systems. The problem of radiation damage has been recognized as a serious factor influencing the amount of information attainable from biological specimens in electron microscopy at conventional voltages around 100 kV. The problem proves to be even more severe at higher voltages around 1 MV. In this range, the problem is the relatively low sensitivity of the existing recording media, which entails inordinately long exposures that give rise to severe radiation damage. This low sensitivity arises from the small linear energy transfer for fast electrons. Few developable grains are created in the emulsion per electron, while most of the energy of the electrons is wasted in the film base.

In-situ electrical-resistivity measurements in the high-voltage electron microscope

1978 ◽  
Vol 36 (1) ◽  
pp. 68-69
Author(s):  
W. E. King
Keyword(s):  
Electrical Resistivity ◽  
Electron Microscope ◽  
High Voltage ◽  
Resistivity Measurements ◽  
Voltage Electron ◽  
High Voltage Electron Microscope ◽  
High Voltage Electron ◽  
Wide Range ◽  
Helium Gas

A side-entry type, helium-temperature specimen stage that has the capability of in-situ electrical-resistivity measurements has been designed and developed for use in the AEI-EM7 1200-kV electron microscope at Argonne National Laboratory. The electrical-resistivity measurements complement the high-voltage electron microscope (HVEM) to yield a unique opportunity to investigate defect production in metals by electron irradiation over a wide range of defect concentrations.A flow cryostat that uses helium gas as a coolant is employed to attain and maintain any specified temperature between 10 and 300 K. The helium gas coolant eliminates the vibrations that arise from boiling liquid helium and the temperature instabilities due to alternating heat-transfer mechanisms in the two-phase temperature regime (4.215 K). Figure 1 shows a schematic view of the liquid/gaseous helium transfer system. A liquid-gas mixture can be used for fast cooldown. The cold tip of the transfer tube is inserted coincident with the tilt axis of the specimen stage, and the end of the coolant flow tube is positioned without contact within the heat exchanger of the copper specimen block (Fig. 2).

Solute redistribution during in-situ irradiation of alloys in the high voltage electron microscope

1978 ◽  
Vol 36 (1) ◽  
pp. 370-371
Author(s):  
P. R. Okamoto ◽  
N.Q. Lam ◽  
R. L. Lyles
Keyword(s):  
Electron Microscope ◽  
High Voltage ◽  
Driving Forces ◽  
Solute Diffusion ◽  
Solute Redistribution ◽  
Voltage Electron ◽  
High Voltage Electron Microscope ◽  
High Voltage Electron ◽  
Thin Foils ◽  
Solute Atoms

During irradiation of thin foils in a high voltage electron microscope (HVEM) defect gradients will be set up between the foil surfaces and interior. In alloys defect gradients provide additional driving forces for solute diffusion since any preferential binding and/or exchange between solute atoms and mobile defects will couple a net flux of solute atoms to the defect fluxes. Thus, during irradiation large nonequilibrium compositional gradients can be produced near the foil surfaces in initially homogeneous alloys. A system of coupled reaction-rate and diffusion equations describing the build up of mobile defects and solute redistribution in thin foils and in a semi-infinite medium under charged-particle irradiation has been formulated. Spatially uniform and nonuniform damage production rates have been used to model solute segregation under electron and ion irradiation conditions.An example calculation showing the time evolution of the solute concentration in a 2000 Å thick foil during electron irradiation is shown in Fig. 1.

Sign in / Sign up

Export Citation Format

Share Document