bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cytoskeletal elements, cell shaping and movement in the angiosperm pollen tube
Authors: Heslop-Harrison J. , Heslop-Harrison Y. , Cresti M. , Tiezzi A. , Moscatelli A. 
Published: April 2021

ABSTRACT
The ellipsoidal generative cell of the pollen grain of Endymion nonscriptus usually elongates further following germination and entry into the tube, producing attenuated extensions the forward one of which may reach into the vicinity of the vegetative nucleus. This shape change is accompanied by the stretching of the microtubule cytoskeleton of the cell, identified in the present work by immunofluorescence using monoclonal antibodies to tubulin. Complementary observations of living generative cells of Iris pseudacorus showed that they undergo slow undulatory movements accompanied by variation in shape and length during passage through the tube. Such changes must presumably be accompanied by modifications of the microtubule cytoskeleton.
Colchicine at 1 mM eliminated microtubules from tubes and most generative cells of E. nonscriptus,but did not radically affect pollen-tube shape or extension growth, nor arrest the movements of the vegetative nucleus and generative cell into and through the tube. Generative cells in colchicine-treated pollen of Galanthus nivalis rounded up and failed to undergo the usual changes in shape during passage through the tube. Secondary consequences were changes in precedence in movement through the tube, and a greater dispersal along its length. On the assumption that no other cytoskeletal elements remain to be discovered, it seems likely that microfilaments rather than microtubules provide the motive force for movement in the tube, although the latter are involved in shaping the generative cell and adapting it to its passage.
Cytoplasmic calcium measurements in intact higher plant cells: Results from fluorescence ratio imaging of fura-2
Authors: Clarkson David T. , Brownlee Colin , Ayling Sarah M. 
Published: April 2021

ABSTRACT
Roots of seedling tomato (Lycopersicon esculenturn) and oilseed rape (Brassica napus) plants were grown on agarose slopes formed on microscope slides. Root hairs were used for microelectrode insertions, for observations of the rate of cytoplasmic streaming and for measurements of fluorescence of the calcium indicator fura-2
The resting potential across the plasma membrane of root hair cells was between −140 and − 160 mV in both species. Insertion of microelectrodes caused immediate cessation of cytoplasmic streaming; 1–4 min later streaming restarted and quickly recovered its normal velocity of 1–3 μm s−1. Once the electrode had been inserted, current pulses had no obvious effect on streaming. Thus, fura-2 injected by iontophoresis was quickly distributed throughout the length of the hair and the epidermal cell body. Injections were made into the cap of cytoplasm at the tip of the hair. Movement of fura-2 from the cytoplasm into the vacuole was observed; this was marked by a great increase in its fluorescence after excitation at 350 nm and occurred after about 15–30 min in tomato, but more rapidly (5–10 min) in oilseed rape hairs. No fluorescence was associated with the cell walls; this was demonstrated by plasmolysing the cells.
The distribution and concentration of Ca2+ was estimated from the ratio of fura-2 fluorescence excited at 350 and 385 nm and from digital image analysis. Resting levels of calcium in the cytoplasm near the tip were in the range 30·90 nM, while much higher levels, which eventually saturated the fura-2 response (&gt;5 μM), were seen in vacuoles once the indicator had crossed the tonoplast. This distribution was rapidly perturbed by the addition of the calcium channel blocker, verapamil, to the outer solution. There was a return to the original distribution as the verapamil was diluted. Separate experiments showed that verapamil inhibited cytoplasmic streaming over a time-course similar to these perturbations in cytoplasmic Ca2+ and caused a small depolarization (by approx. 30 mV) of the membrane potential. The application of 3×10−5 or 3 ×10−4 M-N-ethyl maleimide to the hairs strongly and irreversibly depolarized the plasma membrane electric potential, caused cytoplasmic streaming to stop or slow down and resulted in a very rapid rise in cytoplasmic Ca2+, which became uniformly distributed with saturating 350:385 nm ratios.
‘blebbing’ of the nuclear envelope of mouse zygotes, early embryos and hybrid cells
Authors: Szöllösi Maria S. , Szöllösi Daniel 
Published: April 2021

ABSTRACT
In the mouse zygote and in two-cell stage embryos the inner leaflet of the nuclear envelope of pronuclei and that of blastomere and polar body II nuclei evaginate, forming multiple blebs within the perinuclear space, which contains a granular material. Blebbing exists only in oocytes activated by sperm in vivo or in vitro, or parthenogenetically by treatment with ethanol or puromycin. The germinal vesicle and an interphase nucleus formed after treatment of the oocyte at metaphase I by puromycin do not form blebs. Formation of blebs is specifically located in the cell cycle. The burst of the blebbing activity occurs during the first half of the cell cycle in one-cell embryos and in the earliest interphase period in the second cell cycle. Blebbing ceases from the beginning of the third cell cycle. The occurrence in the cytoplasm of ‘doublelayered’ vesicles containing granular material resembling bleb contents and the disappearance of blebs from the nuclear envelope by the end of the cell cycle provide evidence that blebs represent a step in the transport of some material from the nucleus to the cytoplasm. Ethanolic phosphotungstic acid does not stain blebs, suggesting the absence of basic protein in their contents.
Blebbing can be induced in somatic (thymocyte) and embryonic (blastomere of 8-cell stage embryo) nuclei following cell hybridization with activated oocytes. Their response to the oocyte cytoplasm by initiating blebbing depends on: (1) the position of the host cell in its cell cycle at the moment of hybridization, and (2) the time spent by the foreign nuclei in the host cytoplasm following cell fusion. If donor nuclei are introduced close to the time of activation, they start to produce blebs at the time corresponding to the initiation of blebbing by the female pronucleus in the first cell cycle. If foreign nuclei are introduced a few hours after activation they must be incubated in the host cytoplasm for some time before initiation of bleb formation, provided that the host pronucleus has initiated blebbing by that time.
The existence of blebbing in nuclei formed only after oocyte activation, and the timing and the general occurrence of this event during the earliest cleavage stages of almost every mammalian embryo, suggest that this special nucleocytoplasmic transport plays a specific role at the beginning of development.
The organization of the cytoskeleton during meiosis in eggplant (Solarium melongena (L)): microtubules and F–actin are both necessary for coordinated meiotic division
Authors: Traas J. A. , Burgain S. , Vaulx R. Dumas De 
Published: April 2021

ABSTRACT
Because two division planes form at right angles, male meiosis in higher plants provides striking examples of both division control and spatial programming.
To investigate these processes we have stained microtubules and actin filaments during male meiosis in the eggplant. Our results indicate the following.
(1) That microtubules and their nucleation sites are involved in the establishment of polarity; this is supported by our observation that the drug CIPC affects spindle polarity.
(2) That actin microfilaments are involved in spindle formation and integrity, but not in the establishment of polarity: cytochalasin B and D affect the organization of the spindle microtubules, but not their polarized distribution.
(3) That microtubules radiating from the daughter nuclei at the cell poles during interkinesis probably establish the future division plane by concentrating actin in that plane (cf. the proposed role of asters in positioning the contractile ring in animal cells).
(4) That this concentration of F-actin in the division plane may be involved in preparing the cytoplasm for cytokinesis and in memorizing the division plane (much as the preprophase band observed in polarized tissues does).
(5) That phragmoplast formation is a two-step process. No phragmoplast forms after metaphase I, but a four-way phragmoplast forms after metaphase U, indicating that mitosis and cytokinesis are not obligatorily coupled.
These studies demonstrate that actin and microtubules are jointly involved in the spatial coordination of the division process.
Lectin binding of F9 embryonal carcinoma cells: evidence for population heterogeneity and developmentally regulated high–Mr cell surface proteins
Authors: Tienari Jukka , Virtanen Ismo , Lehtonen Eero 
Published: April 2021

ABSTRACT
Undifferentiated F9 embryonal carcinoma (EC) cells bound fluorochrome-coupled Helix pomatia agglutinins (HPA) and peanut agglutinins (PNA) homogeneously, but were distinctly heterogeneous in their binding of Dolicbos biflorus agglutinin (DBA) conjugates. Upon chemically induced differentiation the proportion of cells binding the DBA conjugates increased, but a distinct heterogeneity in the intensity of binding remained among the parietal endoderm (PE)-like F9 derivatives. These cells were heterogeneous in their binding of HPA conjugates as well, and many of them failed to bind PNA conjugates, apparently due to sialylation of the PNA-binding sites.
Electrophoretic analysis of lectin-binding glycol-proteins in the detergent-soluble fraction of the cells revealed the appearance of a doublet of polypeptides of Mr 300 000-400 000 upon ddferentiation induced by retinoic acid (RA). In addition, an Mr 220 000 polypeptide appeared upon ddferentiation induced by RA and dibutyryl cyclic AMP (dbcAMP). These polypeptides were obtained from both metabolically labelled and surface-labelled cells. A major secreted glycoprotein, which comigrated with laminin, bound to DBA. This suggests that laminin secreted by the ddferentiated F9 derivatives contains O-glycosidic saccharides.
The results show that even though differentiation of F9 cells leads to changes in their binding of fluorochrome-coupled lectins, these lectin conjugates reveal distinct population heterogeneity among unddferentiated and differentiated F9 cells and are hence likely to be of limited value in the characterization of individual cells. At the whole cell population level, on the other hand, affinity binding to lectins reveals the appearance of high-Mr. cell surface proteins in differentiating F9 cells.
Characterization of proteins immunologically related to brain microtubule-associated protein MAP-1B in non-neural cells
Authors: Díaz-nido Javier , Avila Jesús 
Published: April 2021

ABSTRACT
Brain microtubule-associated protein MAP-1 is composed of at least two polypeptides, MAP-1A and MAP-1B, which are among the main components of the neural cytoskeleton. Specific monoclonal and polyclonal antibodies against MAP-1B stain nuclei, mitotic spindles, centrosomes and the cytoplasmic microtubule network of different non-neural cells studied by immunofluorescence microscopy. It appears that these cells contain two proteins of 325K and 22OK (K = 10?Mτ), which are immunologically related to brain MAP-IB. The 325K protein, which is localized to the cytoplasmic microtubule network, the centrosome and the mitotic spindle, seems to be structurally related to neural MAP-1B, as judged from their similar peptide maps and phosphorylation patterns. The 220K protein, which is localized to the nuclear matrix in interphase cells and to the mitotic spindle in dividing cells, has a proteolytic profile different from that of neural MAP-IB and is phosphorylated to a much lesser extent than the 325K protein. Both proteins bind tubulin in viuo, which suggests that they may participate in microtubule assembly in vivo, the 325K protein could perform such a role during the entire cell cycle, while the 22OK protein could be implicated in the formation of the mitotic spindle.
Cingulin: Characterization and localization
Authors: Citi Sandra , Sabanay Helena , Kendrick-Jones John , Geiger Benjamin 
Published: April 2021

ABSTRACT
Cingulin, a protein component associated with the tight junctions of chicken intestinal epithelium, has been purified to homogeneity by a new procedure and characterized. Purified cingulin is a heat-stable elongated dimer, composed of two polypeptides of Mr 108000 (cingulin-108), with a Stokes’ radius of ∼15nm, and a molecular length of 130 nm ± 32 nm. Monoclonal antibodies were used to determine the tissue distribution and subcellular localization of cingulin in a variety of avian tissues and cultured cells. Indirect immunofluorescence analysis of semi-thin frozen sections demonstrated that cingulin is localized in the junctional complex of various polarized epithelia and in the endothelium, whereas it is essentially absent from mesenchymal and myogenic cells. Ln permeabilized and fixed cultured chick embryo kidney cells, the antibodies stained solely the regions of contacts between the epithelial cells. Double immunofluorescent labeling of these cells with anti-cingulin and anti-vincu-lin antibodies showed that cingulin is localized close to the vinculin-rich cytoskeletal belt associated with adherens junctions, but is absent from focal contacts and stress fibers. In cultured kidney cells, actin was detected mainly in stress fibers and in the peripheral junctional regions, where it showed a distribution similar to that of cingulin, suggesting that actin filaments may be part of the submembrane cytoskeleton at the level of the tight junction. Indirect immunoelectron microscopic labeling of ultrathin frozen sections of chicken intestine showed that cingulin is localized along the endofacial surfaces of the tight junction (fzonula occludens), and is apparently excluded from the more basal zonula adhaerens, and from the desmosomes.
Intracellular localization of the glycosyl-phosphatidylinositol-specific phospholipase C of Trypanosoma brucei
Authors: Bülow Roland , Griffiths Gareth , Webster Paul , Stierhof York-Dieter , Opperdoes Fred R. , Overath Peter 
Published: April 2021

ABSTRACT
Glycosyl-phosphatidylinositol-specific phospholipase C (GPI-PLC) is a membrane-bound enzyme of bloodstream forms of Trypanosoma brucei, which cleaves the GPI-membrane anchor of the variant surface glycoprotein forming diacylglycerol and 1,2-cyclic phosphate on the inositol ring. The cellular localization of the enzyme was studied by fractionation of sub-cellular organelles and immunofluorescence microscopy and was found to be primarily cytoplasmic. This was confirmed by _mmune-electron microscopy using cryo-sections, which showed that the labelling was predominantly on the cytoplasmic side of intracellular membranes but was absent from the plasma membrane includeing the region lining the flagellar pocket. The significance of these results for the possible function of the phospholipase is discussed.
Conformation and movement of the vegetative nucleus of the angiosperm pollen tube: association with the actin cytoskeleton
Authors: Heslop-Harrison J. , Heslop-Harrison Y. 
Published: April 2021

ABSTRACT
Actin is present in the cytoplasm of the vegetative cell of angiosperm pollens in numerous fusiform, spiculate or toroidal bodies, and also as a sheath enveloping the vegetative nucleus. During activation following hydration, the compact cytoplasmic bodies are translated into skeins of extended fibrils, and circulatory movements begin in the cytoplasm. Throughout this period the vegetative nucleus, with fibrillar actin now associated with the surface, undergoes a continuous change of shape. In the extending tube following germination the actin cytoskeleton consists of numerous mainly longitudinally oriented fibrils. After entry into the tube the vegetative nucleus remains associated with the fibrils, usually extending greatly in length and developing attenuated, often pointed extensions. The observed conformations, which change continuously, suggest that varying local tensions are applied to the vegetative nucleus during passage through the tube. Cytochalasin D breaks up the actin fibril system and brings about a rapid contraction of the nucleus, at the same time eliminating the elastic extensions of the nuclear envelope. Nuclei isolated physically from unfixed tubes also contract in length as the fibrillar components of the cytoskeleton are detached. These findings indicate that the movement of the vegetative nucleus depends on local associations of the nuclear envelope with the actin cytoskeleton of the vegetative cell.
Effects of leukotrienes on phenotypic properties and growth of arterial smooth muscle cells in primary culture
Authors: Palmberg Lena , Claesson Hans-Erik , Thyberg Johan 
Published: April 2021

ABSTRACT
During the first few days in primary culture arterial smooth muscle cells (SMCs) go through a transition from a contractile to a synthetic phenotype. Morphologically, this process includes loss of myofilaments and formation of an extensive rough endoplasmic reticulum and a large Golgi complex. Functionally, it leads to the cells losing their contractility, beginning to secrete extracellular matrix components, and dividing in response to growth factor stimulation. Similar changes in the structure and function of the SMCs occur in the initial stages of atherogenesis. The object of the present investigation was to study the effects of leukotrienes on the differentiated properties and growth of rat aortic SMCs in primary culture. Enzymically isolated cells were seeded directly on a plastic surface in serum-containing medium or on a substratum of plasma fibronectin in serum-free medium. The change in cell morphology was followed by transmission electron microscopy, and the activation of cell growth by thymidine autoradiography and cell counting. The results demonstrate that 10pM-LTB4, -LTC4, -LTD4 and -LTE4 all speeded up the shift of the SMCs into a synthetic phenotype, whereas 5S,12S-DHETE (an isomer of LTB4) lacked effect. Further, LTB4, LTC4 and LTD4 stimulated the SMCs to enter the cell cycle earlier than in the controls, enhanced the proliferative response to serum mitogens, and under serum-free conditions induced DNA synthesis by themselves. Indomethacin did not interfere with the effect of LTB4 on the structural transformation of the cells but blocked its effect on DNA replication, suggesting that only the latter involved endogenous production of a cyclo-oxygenase product. When seeded on a substratum of laminin, a component of the basement membrane that normally surrounds smooth muscle, the cells were initially retained in a contractile phenotype and no clear effect of the leukotrienes was noted. Taken together, the findings indicate that leukotrienes are able to stimulate the phenotypic modulation and growth of arterial SMCs in primary culture.
Serotonin regulates cytosolic Ca2+ activity and membrane potential in a neuronal and in a glial cell line via 5-HT3 and 5-HT2 receptors by different mechanisms
Authors: Reiser Georg , Donié Frédéric , Binmöller Franz-Josef 
Published: April 2021

ABSTRACT
The mechanisms of action of two different serotonin receptors, found in a neuronal cell line (neuroblastoma x glioma hybrid cells) and in a non-excitable glioma cell line, were explored. In both cell lines, serotonin induced a dose-dependent, transient rise of cytosolic Ca2+ activity (measured by fura-2 or indo-1 fluorescence). Ca2+ channel blockers (Ni2+ and La3+, not nifedipine) suppressed the Ca2+ response to serotonin in the hybrid cells but not in the glioma cells. After application of Ca2+ ionophores (ionomycin and A23187) in order to short-circuit internal Ca2+ stores, serotonin was still able to induce a Ca2+ response in the hybrid cells but not in the glioma cells. Serotonin dose-dependently stimulated the rate of 45Ca2+ uptake several-fold in the hybrid cells, but hardly at all in the glioma cells. Thus, in the neuronal cell line cytosolic Ca2+ activity is raised through enhancement of Ca2+ entry into the cells from the extracellular environment via 5-HT3 receptors (blocked by ICS 205-930, MDL 72222 and GR 38032 F). The depolarization response caused by serotonin in the hybrid cells is due to activation of cation conductance), obviously allowing entry of extracellular Ca2+. In contrast to the neuronal cell line, in the glial cell line the rise of Ca2+ activity is mediated by ketanserin-susceptible 5-HT2 receptors (not affected by treatment with pertussis toxin) mainly liberating Ca2+ from internal stores. In the glioma cells the release of Ca2+ from internal stores leads to opening of Ca2+-dependent K+ channels, responsible for the hyperpolarizing response. Thus, the neuronal and the glial cell lines might provide suitable systems in which to study the diverse cellular functions triggered by the rise of cytosolic Ca2+ activity, which is caused by different serotonin receptors.
The shape of mitochondria and the number of mitochondrial nucleoids during the cell cycle of Euglena gracilis
Authors: Hayashi Yasuko , Ueda Katsumi 
Published: April 2021

ABSTRACT
The shape of mitochondria and the number of mitochondrial nucleoids in Euglena cells were examined throughout the cell cycle by fluorescence microscopy. Both photoheterotrophic and heterotrophic cells contained a network of mitochondria that did not divide into fragments at any stage of the cell cycle. Mitochondrial nucleoids could be clearly detected in the mitochondria by staining with ethidium bromide and with DAPI. Half of the mitochondrial nucleoids entered each daughter cell during cytokinesis. Nucleoids in the newly produced daughter cells increased in number as the cells increased in size. The number of nucleoids reached double the initial value in cells at the stage just prior to mitosis. The total length of the mitochondrial net was proportional to the cell volume.
Inhibitors of topoisomerases I and II arrest DNA replication, but do not prevent nucleosome assembly in vivo
Authors: Annunziato Anthony T. 
Published: April 2021

ABSTRACT
Specific inhibitors of eukaryotic DNA topoisomerases I and II (camptothecin and VM-26, respectively) were used to examine the involvement of topoisomerases in DNA replication and chromatin assembly in vivo. When used singly, either camptothecin or VM-26 inhibited DNA synthesis in HeLa cells by more than 80 %; when used simultaneously, the inhibitors effectively stopped replication, demonstrating that at least one class of topoisomerase must be active for fork propagation in vivo. To study nucleosome assembly during topoisomerase inhibition, three experimental strategies were employed: (1) pulse-chase experiments; (2) analyses of chromatin synthesized during residual replication in the presence of either camptothecin or VM-26; and (3) the assembly of previously replicated, unassembled DNA, generated in the presence of protein synthesis inhibitors. Using sensitivity to micrococcal nuclease and the maturation of non-nucleosomal replication intermediates as criteria, neither camptothecin nor VM-26, alone or in concert, inhibited nucleosome assembly under any experimental protocol tested. These data provide evidence that, although topoisomerase activity is essential for DNA replication, neither continuous fork propagation nor topoisomerase activity is required for chromatin assembly on new DNA.