bluuebunny/crossref_metadata_2025_split huggingface dataset:
Postmitotic reassembly of the cell nucleus in whole cells: an electron-spectroscopic study
Authors: Stracke Sylvia , Martin Rainer 
Published: April 2021

ABSTRACT
By electron-spectroscopic imaging it is possible to visualize selectively the distribution of phosphorusrich structures such as nucleosomes, ribosomes or other ribonucleoprotein particles. Using this method we re-examined assembly of the nucleus in telophase of dividing onion root cells and human HeLa cells. Our observations disagree considerably with conclusions drawn from work with cell-free systems. We consistently observed reassembly of nuclear envelope cisternae from vesicles in the cytoplasm without direct contact with chromatin. The preassembled envelope cisternae then enclosed the telophase chromosome mass, contacting the chromatin in some tracts, but also trapping cytoplasmic material such as ribosomes between chromosomes and envelope. Until a late stage in telophase the re-forming nuclear envelope left large gaps between the nuclear and the cytoplasmic compartments. Exclusion of cytoplasmic material from the re-assembling nucleus was facilitated by prenucleolar material, which accumulated in the deep furrows of the chromosomes and interchromosomal spaces. This material expanded considerably while the envelope was still open, in this way displacing cytoplasm non-selectively from the future nucleus. The model we propose for reassembly of the nucleus in whole cells does not postulate contact with and complete enclosure of chromosomes by the re-forming envelope, and suggests a decisive role for expanding prenucleolar material in the process of nucleocytoplasmic compartmentalization.
Pollen development in orchids. 5. A generative cell domain involved in spatial control of the hemispherical cell plate
Authors: Brown R. C. , Lemmon B. E. 
Published: April 2021

ABSTRACT
The unequal first pollen mitosis in moth orchids (Phalaenopsis) is followed by an unusual form of cytokinesis that isolates a small lens-shaped generative cell from a large vegetative cell. No preprophase band of microtubules predicts the division plane and the new cell plate grows completely around the generative cell rather than fusing with the parental wall. Development of the phragmoplast cytoskeleton consisting of fusiform bundles of microtubules and F-actin occurs in three major stages: (1) the initial asymmetrical phragmoplast conforming to the shape of the interzonal region, which tapers from the broad mass of chromosomes at the generative pole to the rounded mass at the vegetative pole; (2) the symmetrical plate-like phragmoplast; and (3) the hemispherical phragmoplast, which curves around the generative nucleus. Microtubules of the generative half of the hemispherical phragmoplast are nuclearbased, while those on the vegetative side terminate in endoplasmic reticulum. The path of the phragmoplast appears to outline a cytoplasmic domain defined by a radial system of microtubules emanating from the generative nucleus.
The effects of dimethyl sulfoxide and retinoic acid on the cell growth and the phenotype of ovarian cancer cells
Authors: Grunt T. W. , Somay C. , Pavelka M. , Ellinger A. , Dittrich E. , Dittrich Ch. 
Published: April 2021

ABSTRACT
We have compared the in vitro effects of the differentiation inducers dimethyl sulfoxide (DMSO) and retinoic acid (RA) on a polyclonal human ovarian cancer cell line (HOC-7). Density gradient fractionation of untreated cells reveals that a proportion of rapidly growing, polygonal cells with medium density is capable of spontaneous reversion into a slowly growing low-density phenotype with flattened morphology similar to non-transformed human ovarian surface epithelial cells. Clonal expansion of these low-density cells proves that the observed characteristics are stable for prolonged culture periods. Exposure of HOC-7 cells to DMSO and RA or removal of the serum from the medium is effective in enhancing the proportion of these low-density cells. Application of DMSO causes the cells to become flattened and elongated, and to develop rodlike protrusions. In these cytoplasmic extensions thick filament bundles are dominant. Immunofluorescence studies demonstrate that both untreated low-density subclones and DMSO-treated polyclonal cells are much more reactive for cytokeratin than medium-density subclones or untreated parental cells. Furthermore, immunocytochemistry and fixedcell ELISA reveal 2-to 5-fold greater amounts of desmoplakins I and II and of fibronectin in low-density subclones and in DMSO-treated cells as compared to medium-density subclones and control cultures. RA exerts weaker effects on the phenotype of the cells. Both inducers reduce DNA synthesis and inhibit the anchorage-dependent and the anchorageindependent cell growth in a dose- and time-dependent manner. The restoration of the original morphology and growth rate after removal of the differentiation-inducing agents proves that the observed changes are reversible; this indicates that the cells do not become terminally differentiated. The described phenotypic and growth kinetic reactions should prove useful for the future recognition of induced maturation of ovarian cancer cells.
The complex cell cycle of the dinoflagellate protoctistCrypthecodinium cohniias studiedin vivoand by cytofluorimetry
Authors: Bhaud Yvonne , Salmon Jean-Marie , Soyer-Gobillard Marie-Odile 
Published: April 2021

ABSTRACTThe complete cell cycle of the dinoflagellate Crypthecodinium cohniiBiecheler 1938 was observed in vivo in a synchronized heterogeneous population, after DAPI staining of DNA. In a given population, the relative nuclear DNA content in each class of cell was measured using a new numerical image-analysis method that takes into account the total fluorescence intensity (FI), area (A) and shape factor (SF). The visible degree of synchronization of the population was determined from the number of cells with a nuclear content of IC DNA at ‘synchronization’, time 0. One method of synchronization (method 1), based on the adhesiveness of the cysts, gave no better than 50% synchronization of the population; method 2, based on swimming cells released from cysts cultured on solid medium, gave 73% of cells with the same nuclear DNA content. A scatter plot of data for FI versus A in the first few hours after time 0 showed that the actual degree of synchronization of the population was lower.The length of the C. cohnii cell cycle determined in vivo by light microscopy was 10, 16 or 24 h for vegetative cells giving two, four or eight daughter cells, respectively. Histograms based on the FI measurements showed that in an initially synchronized population observed for 20 h, the times for the first cell cycle were: G1 phase, 6 h; S phase, 1 h 30 min; G2+M, lh 30 min, with the release of vegetative cells occurring 1 or 2h after the end of cytokinesis. The times for the second cell cycle were G1+S, 3h; G2+M, 2h.FI and A taken together, suggested that the S phase is clearly restricted, as in higher eukaryotes. A and SF, taken together, showed that the large nuclear areas were always in cysts with two or four daughter cells. FI and SF, taken together, showed that the second S phase always occurred after completion of the first nuclear division.Our data concerning the course of the cell cycle in C. cohnii are compared with those from earlier studies, and the control of the number of daughter cells is discussed; this does not depend on the ploidy of the mother cell.
The membrane skeleton of Pseudomicrothorax: II. biochemical and immunological characterization of the glycosylated cytoskeletal proteins
Authors: Huttenlauch Irm , Peck Robert K. 
Published: April 2021

ABSTRACT
The membrane skeleton, or epiplasm, is part of the structurally complex ciliate cortex. It is thought to have skeletal functions concerning the spatial organization of cortical elements such as the basal bodies.
Here we report the biochemical and immunological characterization of some components of the purified epiplasm of Pseudomicrothorax dubius. The epiplasm proteins consist of two quantitatively major groups of proteins, one of 76-80 ×103Mr, the other of ll-13×103Mr, which appear to be the principal structural elements of the epiplasm, and a series of minor components of 62-18×103Mr. Based upon lectin labeling and glycosidase treatment, some of the latter have been identified as glycoproteins. Using affinity-purified antibodies specific for individual glycoproteins or groups of glycoproteins, we were able to localize them in situ by immunoelectron microscopical methods. This in situ localization demonstrates that the glycosylated epitopes, unlike the glycoresidues of membrane proteins, are distributed throughout the entire epiplasmic layer rather than being restricted to regions adjacent to the cortical membranes. Thus, these proteins represent glycosylated, cytoskeletal elements. At least one of these glycoproteins (Mr 62×103) shows positive immunoreactivity with a monoclonal antibody (Pruss anti-IFA) recognizing most intermediate filament (IF) proteins, indicating that IF proteins might be present in protozoan cytoskeletons.
Distribution of membranes and the cytoskeleton during cell plate formation in pollen mother cells of Tradescantia
Authors: Schopfer Christel R. , Hepler Peter K. 
Published: April 2021

ABSTRACT
The cellular pattern and distribution of membranes have been analyzed during cytokinesis in pollen mother cells of Tradescantia and compared with those of actin microfilaments (MFs) and microtubules (MTs). Membranes have been stained with DiOC6(3) and MFs with rhodamine-labeled phal-loidin (RP); analysis has been carried out on the confocal laser scanning microscope. MTs have been visualized as birefringent elements in the polarized light microscope. The results show that when the interzone first appears in mid anaphase it contains an even distribution of membranes. However, by late anaphase these elements have been cleared away, leaving the interzone largely devoid of DiOC6(3)-positive material. MTs are found throughout this zone, while MFs appear in two non-overlapping sets on both sides of the cell equator. Thereafter membrane elements reappear in the interzone, but only along the equatorial line of the forming cell plate. Presumably these equatorial elements are composed of endoplasmic reticulum and Golgi vesicles, since the larger organelles, including amyloplasts and mitochondria, are excluded from the phragmoplast. MFs, like MTs, arrange preferentially normal to the cell plate, forming a dense array on both sides, but being absent from the zone occupied by the membranes. By contrast, the parallel set of MTs, while excluding larger organelles from the phragmoplast, intermingle with the membrane elements in the cell equator. As cytokinesis proceeds membranes continue to concentrate on the cell plate as indicated by its marked increase in staining with DiOC6(3). From a consideration of spatial and temporal organization of the phragmoplast components it is reasonable to suggest that both cytoskeletal components participate in the aggregation of vesicles that give rise to the cell plate. Membranes, on the other hand, through the provision of surface binding sites and/or through the regulation of the cytoplasmic calcium ion concentration, might be involved in the assembly and stabilization of the cytoskeleton.
Distribution of PCNA in Drosophila embryo during nuclear division cycles
Authors: Yamaguchi Masamitsu , Date Takayasu , Matsukage Akio 
Published: April 2021

ABSTRACT
An immunocytochemical method using a specific antibody was employed to detect the proliferating cell nuclear antigen (PCNA) in Drosophila embryos during the first 13 nuclear division cycles. Strong nuclear staining with the anti-PCNA antibody was observed at interphase throughout 13 cycles. Metaphase chromosomes were not stained throughout these cycles. The chromosomal (nuclear) staining reappeared at anaphase until cycle 10 and at telophase in cycle 11. During cycles 12 and 13, nuclear staining was detected exclusively at interphase. Relatively uniform staining of syncytial cytoplasm was observed throughout mitotic phases until cycle 9. In the following cycles, strong staining in both the central yolk mass and the cortical layer of cytoplasm was detected at metaphase and telophase. During interphase of cycles later than the 9th, staining in the central yolk mass got much fainter and that in the cortical cytoplasm completely disappeared. These results suggest that the PCNA dissociates from chromosomes at metaphase; then in later mitotic phases, it is transported from the syncytial cytoplasm into nuclei to participate in formation of the active DNA-replication enzyme complexes.
Calcium sequestration in the golgi apparatus of cultured mammalian cells revealed by laser scanning confocal microscopy and ion microscopy
Authors: Chandra Subhash , Kable Eleanor P. W. , Morrison George H. , Webb Watt W. 
Published: April 2021

ABSTRACT
Co-localization of the elements calcium, potassium, sodium and magnesium with sequestering organelles has been achieved by application of two microscopy techniques on the same cell. Organelles were first localized by laser scanning confocal microscopy (LSCFM) using fluorescent organelle stains. The same cells were then analyzed for elemental distribution with ion microscopy. This approach has identified a perinuclear region of prominent total calcium concentration with the Golgi apparatus. Live cells were fluorescently stained with C6-NBD-cera-mide for labeling the Golgi apparatus prior to cryogenic preparation and freeze-drying, and imaged with LSCFM for Golgi localization; identical cells were then analyzed with ion microscopy to image subcellular distributions of total calcium, potassium, sodium and magnesium. In three cell lines, LLC-PKi porcine kidney epithelial cells, Swiss 3T3 mouse fibroblast cells and L5 rat myoblast cells, the Golgi regions contained significantly higher total calcium concentrations than any other region of the cell (as measured at the spatial resolution of ion microscopy of about 0.5 /an). Intracellular potassium, sodium and magnesium were homogeneously distributed throughout the cell and did not show this pattern. Measurements of depletion of calcium by exposure to calcium-free medium showed that the Golgi apparatus was substantially more resistant to calcium depletion than all other regions of these cells, but sequestered Ca2+ could be released from the Golgi by exposing the cells to calcium ionophore A23187. The Golgi apparatus appears to sequester about 5% of the total cell calcium in LLC-PKi cells, about 2.5% in 3T3 cells and L5 cells.
The effect of permeant and impermeant osmoticants on exocytosis in guinea pig sperm
Authors: Green D. P. L. 
Published: April 2021

ABSTRACT
Guinea pig sperm were suspended in calcium-containing medium supplemented with various concentrations of the tetrasaccharide, stachyose. At concentrations up to and including 0.6 M, stachyose was without effect on the A23187-induced acrosome reaction. At 1.0 M stachyose, &gt;97% of sperm retained their acrosome after exposure to A23187, as judged by light microscopy. Electron microscopy demonstrated, however, that exocytotic membrane fusion had occurred, although with substantial retention of the acrosomal matrix. Sperm incubated in 1.0 M stachyose solutions also underwent exocytotic membrane fusion in the absence of A23187 and external calcium.
Sperm suspended in 0.175 M ammonium chloride solution progressively lost motility over 30 min, but without acrosomal swelling. By contrast, sperm in 0.19 M ammonium acetate underwent substantial swelling of the acrosome within 2-5 min. 70-80% of these sperm were able to exclude the vital dye propidium iodide with their acrosomes swollen. These sperm underwent acrosomal shrinkage if resuspended in normal medium within 5-10 min, and the majority (60-70%) recovered some motility. These sperm could undergo an A23187-induced acrosome reaction. Electron microscopy indicated that swelling in ammonium acetate solution solubilizes much of the acrosomal matrix and causes internal fusion between adjacent regions of the outer acrosomal membrane. There was no exocytotic membrane fusion in ammonium acetate solution, however.
The evidence suggests that there is no stachyose osmolality for guinea pig sperm which will suppress the membrane fusion associated with exocytosis, and that sufficiently high osmolalities cause exocytotic membrane fusion in the absence of calcium. The evidence also suggests that release of the normal osmotic compression of the acrosomal matrix does not cause local blebbing of the acrosomal membrane, nor does it render the membrane fusigenic. The data are consistent with the model for exocytosis in which no granule swelling occurs prior to membrane fusion.
Assembly of actin-containing cortex occurs at distal regions of growing neurites in pc12 cells
Authors: Sanders Mitchell C. , Wang Yu-Li 
Published: April 2021

ABSTRACT
Although actin filaments are known to be localized in the cortex of axons and in the growth cones of nerve cells, it is unclear how actin-containing structures are assembled during nerve growth. We have studied the formation of actin structures in growing neurites by microinjecting fluorescent phalloidin or actin into PC 12 neuron-like cells to label endogenous actin filaments. Upon stimulation of neurite growth in cells microinjected with fluorescent phalloidin, little or no fluorescence was detected in nascent growth cones and adjacent neurites despite the presence of actin filaments in these regions, suggesting that actin filaments were primarily formed by de novo assembly rather than the transport and reorganization of pre-existing, phalloidin-labeled actin filaments. Time-lapse observations of the distribution of phalloidin-labeled actin filaments during neurite elongation confirmed that fluorescence associated with pre-existing neurite cortex spread out more slowly than the elongation of neurites. Furthermore, when a dark spot was photobleached with a laser microbeam along neurites of cells microinjected with either fluorescent phalloidin or actin, the spot showed no appreciable translocation during active neurite elongation. Taken together, these results suggest that de novo assembly of actin filaments plays a crucial role in the formation of growth cones and adjacent cortex in the distal region of neurites, but does not appear to require the anterograde or retrograde transport of cortical filaments, or the passive stretching of the proximal segment of the neurite cortex.
Laminin and basement membrane-associated microfilaments in wild-type and mutant Drosophila ovarian follicles
Authors: Gutzeit Herwig O. , Eberhardt Wolfgang , Gratwohl Eugen 
Published: April 2021

ABSTRACT
The localization of the extracellular matrix glycoprotein laminin was studied using polyclonal antilaminin antibodies. The laminin patterns of the basement membranes of the muscular epithelial sheath that envelops the ovariole were conspicuously different from those of the basement membrane of the follicular epithelium. In the latter structure laminin was stained in a pattern of parallel stripes oriented perpendicular to the long axis of the follicle; microfilament bundles at the adjacent basal side of the follicle cells have the same orientation. At late vitellogenic stages the orientation of the microfilaments remained the same while the laminin stripes were no longer visible. The orientation of laminin and F-actin was abnormal in follicles of the egg-shape mutant kugel, which produces shorter and thicker eggs than wild-type flies. This phenotype might result from the disturbance of the normal circular microfilament and/or laminin pattsern.
A 33 kda protein with sequence homology to the ‘laminin binding protein’ is associated with the cytoskeleton in hydra and in mammalian cells
Authors: Keppel E. , Schaller H. C. 
Published: April 2021

ABSTRACT
In hydra and in mammalian cells the monoclonal antibody V recognises an epitope which colocalises with cytoskeletal structures. Using this antibody for expression screening, a cDNA clone (955 bp) was isolated from hydra, which covers an open reading frame for a protein of 294 amino acids with a calculated molecular mass of 32.8 kDa. Northern blot analysis of hydra RNA resulted in a single mRNA species of 1.2 kb, and primer extension experiments proved this to be the full length message. 218 residues at the amino terminus of the hydra protein show extensive homology (73.5%) to a human protein designated ‘laminin binding protein’. The carboxyl-terminal 76 amino acids possess no significant similarity (20%). The monoclonal antibody V, which recognises an epitope in this carboxyl-terminal part, reacts in Western blots, both in hydra and in mammalian cells, with a protein of 33 kDa and not with the 45 kDa ‘laminin binding protein’. The 33 kDa protein is not extracellular or transmembrane, but has a strictly intracellular location as indicated by its amino acid sequence and by immunocytochemical and cell fractionation studies.
In non-dividing mammalian cells the 33 kDa protein colocalises with filamentous structures; in dividing cells it dissociates from it and concentrates centrally. Presence of the SPLR-sequence, which is the consensus phosphorylation motif for the p34cdc2 kinase, links this 33 kDa protein to events occurring during the cell cycle.
Abberant chemotaxis and differentiation in Dictyostelium mutant fgdc with a defective regulation of receptor-stimulated phosphoinositidase c
Authors: Bominaar Anthony A. , Kesbeke Fanja , Snaar-Jagalska B. Ewa , Peters Dorien J.M. , Schaap Pauline , Haastert Peter J. M. Van 
Published: April 2021

ABSTRACT
Dictyostelium cells use extracellular cyclic AMP both as a chemoattractant and as a morphogen inducing cell-type-specific gene expression. Cyclic AMP binds to surface receptors, activates one or more G-proteins, and stimulates adenylate cyclase, guanylate cyclase and phosphoinositidase C. Mutant fgdC showed aberrant chemotaxis, and was devoid of cyclic AMP-induced gene expression and differentiation. Both the receptor- and G-protein-mediated stimulation of adenylate cyclase and guanylate cyclase were unaltered in mutant fgdC as compared to wild-type cells. In wild-type cells phosphoinositi dase C was activated about twofold by the cyclic AMP receptor. In mutant fgdC cells, however, the enzyme was inhibited by about 60%. These results suggest that phosphoinositidase C is regulated by a receptor-operated activation/inhibition switch that is defective in mutant fgdC. We conclude that activation of phosphoinositidase C is essential for Dictyostelium development.