bluuebunny/crossref_metadata_2025_split huggingface dataset:
Movements of fluorescent probes in the mechanism of cell fusion induced by poly(ethylene glycol)
Authors: Ahkong Quet F. , Desmazes Jean-Pierre , Georgescauld Dinu , Lucy Jack A. 
Published: April 2021

ABSTRACT
It has been claimed that purified poly(ethylene glycol) (PEG) is able only to aggregate cells and not to fuse them. In our hands, purified PEG 6000 (recrystallized/dialysed) induces both aggregation and fusion of human erythrocytes, and the mechanism of fusion by the purified polymer has been investigated with fluorescent probes. No movement of a carbocyanine probe or of octadecyl rhodamine B chloride from labelled to unlabelled cells occurred in the absence of PEG or with cells treated with concanavalin A, protamine or spermine. With 40 % PEG, however, both probes immediately started to diffuse into the membranes of unlabelled cells. This indicates that continuity between the phospholipid bilayer membranes of adjacent erythrocytes (i.e. membrane fusion) is established within seconds in concentrated solutions of the polymer, and precedes the cell fusion event that is induced by purified PEG. These observations are consistent with the idea that micro-regions of shared phospholipid bilayer may be formed in the membranes of cells when they are forced together as a consequence of the dehydrating action of PEG. Intact erythrocytes were cytoplasmically labelled with 6-carboxyfluorescein to avoid the possibility that loading the cells with a cytoplasmic marker by hypotonic haemolysis might modify their response to PEG. Unlike the lipid probes, carboxyfluorescein did not diffuse from labelled to unlabelled cells in the presence of 40% PEG, and there was little diffusion on subsequent dilution of the polymer solution to 13%. However, after the PEG solution had been replaced by an isotonic buffer, a rapid transfer of the cytoplasmic fluorophore to unlabelled cells often occurred. This is considered to be more consistent with the osmotic rupture of a membranous barrier, such as a shared bilayer, between the labelled and unlabelled cells than with the return of cytoplasmic viscosity to normal when the PEG is removed. Possible reasons are discussed for the reported inability of purified PEG to fuse fibroblasts with hypotonically loaded human erythrocytes.
Evidence for heterogeneity in the 160/165×103mr glycoprotein components of desmosomes
Authors: Jones Jonathan C. R. , Vikstrom Karen L. , Goldman Robert D. 
Published: April 2021

ABSTRACT
We have prepared both monoclonal and polyclonal antibody preparations directed against the 160/165 ×103Mr glycoproteins (desmogleins) of bovine tongue epithelial desmosomes. The polyclonal antibody preparation recognizes desmosomes in a number of mouse tissues, e.g. mouse skin, heart, bladder and trachea, as determined by immunofluorescence microscopy. Furthermore, the polyclonal antibodies recognize poly-peptide(s), present in the high salt, Triton-insoluble residues (‘cytoskeleton preparations’) of mouse skin, heart, bladder and trachea, which comigrate with the 160/165×103Mr glycoproteins of bovine tongue epithelial desmosomes as determined by ‘Western’ immunoblotting. Conversely, the monoclonal 160/165 × 103Mr antibody preparation recognizes desmosomes of stratified squamous epithelial tissues but not desmosomes in other tissue types. Moreover, whereas the monoclonal antibodies recognize 160/165 × 103Mr polypeptides in mouse skin cell cytoskeletons they show no immunoreactivity with the cytoskeleton preparations of mouse bladder, trachea and heart following immunoblotting. These results suggest therefore that although there are conserved epitopes of the 160/165 × 103Mr glycoproteins there are also epitopes of these molecules which vary from tissue to tissue.
Double label immunofluorescence observations of cryostat sections of mouse skin using the monoclonal antibodies and antibodies directed against desmoplakin, a plaque component of desmosomes, reveal that the monoclonal antibodies do not recognize certain desmosomes in basal cells which are recognized by desmoplakin antibodies. Indeed, double label observations of cryostat sections of mouse skin using the monoclonal antibodies and human autoantibodies which react with hemidesmosomal components suggest that the monoclonal antibodies stain desmosomes located along the apical surfaces of basal cells but fail to recognize desmosomes along the lateral surfaces of these same cells. The latter desmosomes, however, are recognized by the polyclonal 160/165 × 103Mr antibody preparation. We discuss the possibility that the inherent polarity of basal epithelial cells is manifested in modifications of the 160/165×103Mr glycoproteins in desmosomes located along different surfaces of basal cells.
Glass beads load macromolecules into living cells
Authors: Mcneil Paul L. , Warder Elizabeth 
Published: April 2021

ABSTRACT
We describe and characterize an exceptionally rapid and simple new technique for loading large numbers of cultured cells with large macromolecules. The culture medium of the cell monolayer is replaced by a small volume of the macromolecule to be loaded. Glass beads (75-500 μm diameter) are then sprinkled onto the cells, the cells are washed free of beads and exogenous macromolecules, and ‘bead-loading’ is completed. The conditions for bead-loading can readily be modified to accommodate cell type and loading objectives: for example, the amount of loading per cell increases if bead size is increased or if beads are agitated after sprinkling onto the monolayer, but at the expense of increased cell loss. As many as 97% of a population of bovine aortic endothelial (BAE) cells were loaded with a 10000Mr dextran; and 79% with a 150 000Mr dextran using beadloading. Various cell lines have been loaded using glass beads. Moreover, bead-loading has the advantage of producing loaded cells that remain adherent and well-spread, thus minimizing recovery time and permitting immediate microscopic examination.
The gap junction
Authors: Warner Anne 
Published: April 2021

ABSTRACT
Gap junctions are intercellular structures that link cells and allow them to exchange ions and small metabolites without recourse to the extracellular space (for reviews and early references, see Goodenough, 1979; Loewen-stein, 1981). They are found in every organ including the nervous system, where they form electrical syn-apses; the only exception being fully differentiated skeletal muscle. Gap-junctional communication is present between all cells in early embryos, regardless of eventual developmental fate, at times when cellular interactions are known to be taking place (Potter et al. 1966). The widespread occurrence of gap junctions has led to the proposal that they play an important part in coordinating cellular signals, including those involved in growth control and embryogenesis.
Cytolocalization of prosomes as a function of differentiation
Authors: De Sa M.F. Grossi , De Sa C. Martins , Harper F. , Coux O. , Akhayat O. , Pal J. K. , Florentin Y. , Scherrer K. 
Published: April 2021

ABSTRACT
Prosomes, ubiquitous ribonucleoprotein (RNP) particles of defined biochemical and morphological structure, first isolated as a subcomplex of the repressed globin mRNP in avian and mouse erythroblasts, were also found in the cytoplasm of other vertebrates associated with other mRNAs. Here we show that prosomes are also present in the cell nucleus and, furthermore, that the cytolocalization of specific prosomal peptides is a function of differentiation. Four monoclonal antibodies, raised against the duck prosomal proteins, p27K, p28K, p29K and p31K (K = 103Mr) react to variable degree with prosomes of chicken, mouse, and human cells. Immunocytochemical and biochemical analyses show that all four antigens are present in both the cytoplasm and the nucleus of avian erythroblasts and avian erythroblastosis virus (AEV)-transformed erythroleukaemic cells. Interestingly, the prosomes disappear in the course of the terminal differentiation of erythroblasts to mature erythrocytes. Although all the four prosomal antigens tested are present in both the nuclear and cytoplasmic compartments, slight differences in the immunofluorescent patterns indicate that each antigen may have a particular cytological distribution that varies in the course of differentiation.
Culture patterns and sorting of rat Sertoli cell secretory proteins
Authors: Ueda Hiroshi , Tres Laura L. , Kierszenbaum Abraham L. 
Published: April 2021

ABSTRACT
A cocultivation chamber and two types of permeable substrates have been used to study: (1) the culture patterns of rat Sertoli and peritubular cells, and Sertoli cells cocultured with spermatogenic cells or peritubular cells; and (2) the polarized secretion of Sertoli cell-specific proteins transferrin, S70 and S45-S35 heterodimeric protein. Substrates included a nylon mesh (with openings of 100 μm) coated with extracellular matrix (ECM) material and an uncoated microporous filter (with pores of O·45 μm). Sertoli cells cultured on ECM-coated nylon mesh organized a continuous sheet of multilayered epithelial cells essentially devoid of spermatogenic cells while peritubular cells formed a layer of squamous cells. Sertoli cells cultured on uncoated microporous substrate formed a continuous sheet of cuboidal epithelial cells with numerous basal cytoplasmic processes projecting into the substrate and abundant apically located spermatogenic cells, while peritubular cells organized one or two layers of loose squamous cells, [35S]methionine-labelled secretory proteins resolved by two-dimensional polyacrylamide gel electrophoresis and autoradiography displayed cell-specific patterns that were slightly influenced by the type of substrate. Sertoli cells cocultured with peritubular cells on uncoated microporous substrate under conditions that enabled separation of apical and basal surfaces, secreted proteins in a polarized fashion. While transferrin was released bidirectionally, S45 – S35 heterodimeric protein was released apically. S70 was detected in both apical and basal compartments. We conclude from these studies that: (1) the number of spermatogenic cells decreases when Sertoli-spermatogenic cell cocultures are prepared on ECM-coated nylon substrate; and (2) Sertoli cells in coculture with spermatogenic or peritubular cells on uncoated microporous substrate, organize continuous sheets displaying polarized protein secretion.
Evidence for septate junctions in the syzygy of the protozoon Gregarina polymorpha (Protozoa, Apicomplexa)
Authors: Dallai Romano , Talluri Maria Vegni 
Published: April 2021

ABSTRACT
A septate junction is described in reproductive pairs of the protozoon Gregarina polymorpha, using conventional thin sections, lanthanum tracer and freeze-fracture techniques. The septate junction is established between the plasma membranes at the tips of the joined epicytic folds. It is characterized by an intercellular space of 14–17 nm traversed by septa with a repeat of 15–25 nm. Lanthanum-treated material exhibits transparent curves forming a meshwork. Freeze fracture replicas show membrane modifications in the shape of short rows of intramembranous particles on the E fracture face of the plasma membrane. The significance of the finding of such a septate junction between protozoan cells is discussed.
Supercoils in plant DNA: Nucleoid sedimentation studies
Authors: Stoilov L. M. , Zlatanova J. S. , Vassileva A. P. , Ivanchenko M. G. , Krachmarov Ch. P. , Genchev D. D. 
Published: April 2021

ABSTRACT
Plant nuclei have been studied with respect to the three-dimensional structure of DNA. Nucleoids derived from nuclei by non-ionic detergent and high salt treatment were analysed by sedimentation in a series of sucrose gradients containing increasing amounts of the intercalating agent ethidium bromide. In addition the nucleoid sedimentation behaviour was investigated following gamma irradiation. The results show that plant DNA is supercoiled, as is the DNA from the other eukaryotes studied, and contains approximately the same concentration of superhelical turns but probably relatively fewer DNA superhelical loops. The plant nuclear populations in all cases studied give rise to two distinct nucleoid bands. These have been characterized by electron microscopy and by their DNA and protein content. The possible origin of the two bands is discussed.
Shedding of cytoplasmic actins by developing muscle cells
Authors: Denning G. M. , Kim I. S. , Fulton A. B. 
Published: April 2021

ABSTRACT
Young myotubes develop spots or macules of actin and alpha actinin under and adjacent to slender actin strands. Macules are found only before sarcomere formation. They contain cytoplasmic actin, small amounts of alpha actin, and alpha actinin but are devoid of tubulin, myosin and vimentin. In the electron microscope, they are seen to contain small filaments but no other organelles. Macules are found on the culture dish and in the culture medium, as well as on the lower and lateral surfaces of the cells. Both emetine and cycloheximide, at doses that inhibit protein synthesis, accelerate formation and shedding of macules. This presents the first observation of a cellular system in which the changeover from a generalized cytoskeleton to a tissue-specific cytoskeleton involves shedding of the cytoplasmic isoforms of cytoskeletal proteins.
Microtubules associate with actin-containing filaments at discrete sites along the ventral surface of Allogromia reticulopods
Authors: Bowser Samuel S. , Travis Jeffrey L. , Rieder Conly L. 
Published: April 2021

ABSTRACT
We have investigated the distribution of actin and microtubules in pseudopodial networks (reticulopods) of the protozoan Allogromia sp., strain NF, in order to help elucidate the respective roles these components play in network organization and motility. Double-label fluorescence studies with tubulin antibodies and tetramethyl-rhoda-mine (TMR)-phalloidin reveal that microtubules and filamentous actin co-localize in regions where trunk pseudopods contact the substratum and splay to form the pseudopodial network; distal to these regions the network contains numerous microtubules but little or no F-actin. Similar results were obtained using various commercial actin antibodies. Correlative anti-actin immunofluorescence and high-voltage electron microscopy of serial 0-25 jtm sections reveal that actin is contained within discrete electronopaque, fan-shaped structures distributed along the cytoplasmic aspect of the ventral reticulopo-dial membrane. Electron microscopy of serial 100 nm sections from conventionally fixed specimens confirms that these actin-rich plaques are composed of a felt of roughly parallel, 5nm diameter filaments. A subset of parallel and often bundled microtubules is enmeshed within, or contacts the periphery of, these filament plaques. Upon leaving a plaque, bundled microtubules frequently splay into smaller bundles. These observations are consistent with the hypothesis that interactions between microtubules and actin-containing microfilaments, particularly at substratum adhesion points, are involved in various aspects of reticulopodial motility, particularly network morphogenesis and cell body locomotion.
Effects of elevated intracellular magnesium on cytoskeletal integrity
Authors: Prescott A. R. , Comerford J. G. , Magrath R. , Lamb N. J. C. , Warn R. M. 
Published: April 2021

ABSTRACT
Increasing the intracellular magnesium concentration of PtK2 cells by 1 mM or more resulted in the disassembly of the interphase microtubule array over a period of 5 min after microinjection. This effect was found to be both transient and fully reversible, with the microtubule arrays reforming after further incubation. These effects were studied using immunofluorescence microscopy of fixed cells, and also in living cells using rhodamine-tubulin or rhodamine-conjugated anti-tubulin antibodies and image intensification and enhancement techniques.
Simultaneously and accompanying the disassembly of the microtubule arrays the F-actin stress fibres also disappeared, usually leaving the peripheral and perinuclear F-actin microfilaments intact. In contrast, increasing intracellular magnesium appeared to have no effect on the vimentin-containing intermediate filaments of PtK2 cells.
These effects on the cytoskeleton were specific to magnesium and could not be mimicked by either microinjection of injection buffer of equivalent ionic strength or sham injection. Raising the intracellular free calcium to the same extent resulted in the disassembly of the microtubule network, but appeared to have no effect on the Factin stress fibres.
Estramustine binds map-2 to inhibit microtubule assembly in vitro
Authors: Stearns Mark E. , Tew Kenneth D. 
Published: April 2021

ABSTRACT
We have investigated the ability of estramustine to bind to rat brain microtubule-associated proteins (MAPs) and purified MAP-2 in vitro. [3H]estramustine’s relative affinity for tubulin and MAPs was assessed by gel filtration chromatography, immunoprecipitation and binding assays. Scatchard analysis demonstrated a specific affinity of the drug for MAP-2. Calculations from kinetic parameters and non-linear regression analysis gave a Kd of 15 μM, and a Bmax of 3·4×10−7 M ml−1. Extrapolation of this value suggested that each MAP-2 molecule binds approximately 20 molecules of estramustine.
Microtubule assembly studies and SDS-poly-acrylamide gel electrophoresis revealed that at 20–60 μM levels, estramustine inhibited the association of MAPs with taxol microtubules. Turbidity (Asso) studies further demonstrated that 20–60μM estramustine inhibited MAP-2-driven tubulin assembly and produced microtubule disassembly. Electron-microscopic studies confirmed the centrifugation and turbidity results. The data demonstrated that estramustine can bind MAPs and MAP-2 specifically, thereby inhibiting microtubule assembly.
The use of cell division cycle mutants to investigate the control of microtubule distribution in the fission yeast Schizosaccharomyces pombe
Authors: Hagan Iain M. , Hyams Jeremy S. 
Published: April 2021

ABSTRACT
We have characterized the changes in microtubule organization that occur through the cell division cycle of the fission yeast Scbizosaccbaro-tnyces pombe by indirect immunofluorescence microscopy. During interphase, groups of cytoplasmic microtubules, independent of the spindle pole body (SPB), form an array extending between the cell tips. These microtubules are involved in positioning the nucleus at the cell equator and in the establishment of cell polarity. At mitosis, the interphase array disappears and is replaced by an intranuclear spindle extending between the now duplicated SPBs. Elongation of the spindle sees the appearance of astral microtubules emanating from the cytoplasmic face of the SPBs. These persist until the end of anaphase whereupon the spindle microtubules depolymerize and two microtubule organizing centres (MTOCs) at the cell equator re-establish the interphase array. We have used the unique properties of various cell division cycle mutants to investigate further the function of these different microtubule arrays and their temporal and positional control.
Regulation of cell shape in Euglena gracilis v. time-dependent responses to ca2+ agonists and antagonists
Authors: Lonergan Thomas A. , Williamson Lura C. 
Published: April 2021

ABSTRACT
The daily changes in cellular shape observed in growth-synchronized cultures of Euglena gracilis Klebs strain Z, were altered by exposure to Ca2+ channel agonists and antagonists. The response of the cells to these pharmacological agents depended, in part, on the time in the growth cycle that the cells were exposed. The Ca2+ channel blockers verapamil and nifedipine and the intracellular Ca2+ antagonist TMB-8 all caused cell rounding when elongated cells from the middle of the light cycle were treated. These results were the same as with other methods used to deprive cells of extracellular Ca2+, such as exposure to EGTA or resuspension in Ca2+-free medium. The cell response in mid light cycle to the channel blockers was reversible by simultaneous exposure to CaCl2, and the nifedipine response was also reversed by simultaneous exposure to the structurally related Ca2+ agonist BAY-K 8644. Exposure of cells in the first hour of the light cycleto verapamil, nifedipine or TMB-8 caused an unexpected result. Instead of preventing the round cells from elongating in the first portion of the light cycle, as do LaCl3, EGTA or resuspension in Ca2+-free medium, a greater than expected percentage of elongated cells was found in the treated population. This represents the first instance in which the biological clock control over the rate and extent of cell elongation was accelerated. The calcium agonist CGP-28392 did not have an effect on cell elongation in the early portion of the light cycle but caused cell rounding in the middle of the light cycle. The calcium agonist BAY-K 8644 did not cause any shape changes alone, but was capable of reversing the effects of nifedipine in the middle of the light cycle.
Elongation of g1 phase by transient exposure of rat 3y1 fibroblasts to caffeine during the previous and present generations
Authors: Okuda Atsuyuki , Kimura Genki 
Published: April 2021

ABSTRACT
When density-arrested rat 3Y1 fibroblasts were stimulated to enter S phase by seeding sparsely in fresh medium, caffeine inhibited this stimuΛlation. When compared at the doses that gave the same levels of the inhibition of entry into S phase, caffeine inhibited protein synthesis to a far lesser extent than cycloheximide. This indicates that caffeine affects some event(s) specific to entry into S phase rather than general protein synthesis. When cells synchronized at early S phase were exposed to caffeine, progression of S and G2 phases was prolonged by only 1 h (from 6 h to 7 h). However, after removal of caffeine at mitosis, the G1 phase was prolonged for 5 h (from 11 h to 16h). These results are consistent with our model that the initiation of S phase is regulated throughout the period between the adjacent S phases. When cells were incubated with normal medium conΛtaining serum during the S and G2 periods, a subsequent 6-h pulse exposure to caffeine caused prolongation of G1 phase for 7h (from 11 h to 18 h). On the other hand, when cells were incuΛbated in the absence of serum during these periods, the prolongation was only 2h (from 16 h to 18 h). Similarly, entry into S phase was proΛlonged only 2h, when a 6-h pulse exposure to caffeine was given immediately after release from density arrest or serum-deprivation arrest. These results indicate the involvement of the relaxation process, which is not affected by cafΛfeine, when serum-deprived cells or density-arΛrested cells restore the process prerequisite for entry into S phase.
Characterization of a spore protein inducing factor from Dictyostelium discoideum
Authors: Gibson F. P. , Hames B. David 
Published: April 2021

ABSTRACT
Spore coat protein synthesis during development by submerged pseudoplasmodia of Dictyostelium discoideum requires a low molecular weight factor secreted by cells incubated at high density in buffer. The further characterization of this spore protein inducing factor (SPLF) is reported. Its behaviour during anion-exchange chromatography and the loss of activity upon esterification suggests the presence of a carboxylic acid group essential for biological activity. Gel permeation chromatography resolves a major SPIF activity with Mr ∼ 160-200 and a minor activity with Mr∼ 340-420. Anion-exchange HPLC further resolves the major SPIF activity into four components, one major and three minor. Methionine, analogues of methionine, and precursors of methionine biosynthesis are all effective in maintaining spore coat protein synthesis. Conditioned medium contains methionine at a concentration sufficient to account for its SPIF activity and this activity is abolished by cyanogen bromide treatment. These results indicate that SPIF is either methionine or a close analogue of methionine.