bluuebunny/crossref_metadata_2025 huggingface dataset:
Fine structure and function of the cytopharyngeal basket in the ciliate Nassula
Authors: Tucker J. B.
Published: April 2021

The fine structure of the cytopharyngeal basket is described in detail. A circular palisade of cytopharyngeal rods is encircled at certain levels by a sheath and two annuli; crest-shaped structures project outwards from the basket. The rods, crests and sheath are largely composed of cytoplasmic microtubules of approximate diameter 240 Å. One of the annuli is mostly composed of densely staining inter-tubular material; the other consists mainly of fine fibres ranging in diameter from 40 to 90 Å.

Ingestion of algal filaments by Nassula and the sequence of movements of the rods which occurs during feeding are described. Evidence is presented which indicates that the crests and fibrous annulus both contract during feeding and act antagonistically to displace the apparently rigid rods. The initial displacement of the rods is spatially related to the orientation of the algal filament which is to be ingested. It is suggested that certain cilia respond to contact with the filament by transmitting some form of excitation to some of the crests along small bundles of microtubules which interconnect them. The suction force drawing the filament into the organism apparently acts only in the lumen of the basket. The basket seems to function as a fairly rigid structure which, acting in conjunction with the suction force, guides the filament into the organism and manipulates it to some extent. The basket also apparently grips the partly ingested filament to prevent it from slipping out of the organism during pauses in the action of the suction force.
Perfusion fixation with glutaraldehyde and post-fixation with osmium tetroxide for electron microscopy
Authors: Van Harreveld A. , Khattab F. I.
Published: April 2021

The conductivity of cerebral cortex drops during perfusion with glutaraldehyde in 5 min to about 60% of the original value, to remain unchanged during the subsequent 10–15 min of perfusion. Circulatory arrest causes a similar drop in the tissue conductivity. Perfusion of asphyxiated tissue with glutaraldehyde does not produce additional major changes in the conductivity. Perfusion of the cortex with an osmium tetroxide solution causes an initial drop in conductivity. However, after about 3 min this trend is reversed and the conductivity increases again to close to the pre-perfusion value. Perfusion of asphyxiated cortex with OsO4 causes a marked increase of the conductivity. So does perfusion with an OsO4 solution of tissue previously treated with glutaraldehyde.

One interpretation of these impedance changes is that glutaraldehyde perfusion causes, like asphyxiation, a transport of extracellular material into the intracellular compartment and that during OsO4 perfusion an extracellular space is again created. This possibility is supported by electron micrographs made of this material. Cerebral cortex perfused with glutaraldehyde and post-fixed with OsO4 shows electron-transparent dendritic elements and to a lesser extent pre-synaptic terminals, which seem to be swollen. When the cortex is flooded with a salt solution during glutaraldehyde perfusion the dendrites exhibit ballooning in the surface layer of the cortex, suggesting that the fluid on the cortex participates in the swelling. The tissue elements in the glutaraldehyde-perfused and OsO4 post-fixed cortex are separated by narrow extracellular spaces. The latter may have been produced by the OsO4 perfusion as is suggested by a comparison of micrographs prepared by freeze substitution (which tends to preserve the water distribution) of glutaraldehyde-perfused but not post-fixed cortex with micrographs of cortex treated with OsO4 after the glutaraldehyde perfusion. In accordance with the conductivity changes, the former micrographs showed very little extracellular space, and in many places tight junctions, whereas the latter showed clefts between the tissue elements.
The atp-dependent concentration of calcium by a golgi apparatus-rich fraction isolated from rat liver
Authors: Hodson Stuart
Published: April 2021

Fractions rich in Golgi apparatus were isolated from female rat liver and incubated with different concentrations of MgATP and calcium. Calcium was sequestered by the fraction in the presence but not in the absence of ATP. The kinetics of the uptake process showed an exponential type accumulation to a maximum value which could be explained by a ‘pump-leak’ mechanism where Vmax was estimated at 6·7 nmol calcium/mg protein/min. Apparent Km for ATP was 1 mM and for calcium was 85μ. The uptake is not inhibited in 5 mM azide, nor is it enhanced in 5 mM oxalate, suggesting that the sequestration is not caused by contamination from mitochondria or microsomes. Parallel experiments on a fraction rich in plasma membrane show at least a 10-fold reduction of activity over the Golgi apparatus-rich fraction, which is interpreted as evidence that plasma membrane contamination in the Golgi apparatus-rich fraction is not primarily responsible for the calcium sequestration activity. Morphometric analysis showed that about 85% of the recognizable membrane profiles in the fraction was associated with elements of the Golgi apparatus. Further evidence is given which suggests that at least 90% of the fraction was uniform in its ability to sequester calcium. In preliminary experiments it was not found possible to separate galactosyltransferase activity (a specific enzyme associated with the Golgi apparatus) from the calcium-sequestration mechanism. The stoichiometry of the hydrolysis of ATP associated with the uptake of calcium is the same as that shown by other isolated membrane systems.
The role of spindle pole bodies and modified microtubule ends in the initiation of microtubule assembly in Saccharomyces cerevisiae
Authors: Byers Breck , Shriver Kathleen , Goetsch Loretta
Published: April 2021

The spindle poles of the budding yeast, Saccharomyces cerevisiae, have been removed from mitotic and meiotic cells by osmotic lysis of spheroplasts. The spindle pole bodies (SPBs) -diskoidal structures also termed ‘spindle plaques’ -have been analysed for their ability to potentiate the polymerization of microtubules in vitro.

Free SPBs were completely deprived of any detectable native microtubules by incubation in the absence of added tubulin and were then challenged with chick neurotubulin, which had been rendered partially defective in self-initiation of repolymerization. Electron microscopy revealed that these SPBs served as foci for the initiation of microtubule polymerization in vitro. Because the attached microtubules elongated linearly with time but did not increase in numbers after the first stage of the reaction, it is apparent that there are a limited number of sites for initiation. The initiating potential of the SPBs was found to be inhibited by enzymic hydrolysis of protein but not of DNA. The microtubule end proximal to the site of initiation on the SPB is distinguished by a ‘closed’ appearance because of a terminal component which is continuous with the microtubule wall, whereas the distal end has the ‘open’ appearance characteristic of freely repolymerized neurotubules. SPBs which were partially purified on sucrose gradients retained their ability to initiate the assembly of microtubules with the same structural differentiation of their ends. The occurrence of closed proximal ends on native yeast microtubules suggests that closed ends may play a role in the initiation of microtubule polymerization in vivo, as well as in vitro.
Formation and growth of gap junctions in mouse myocardium during ontogenesis: A freeze-cleave study
Authors: Gros D. , Mocquard J. P. , Challice C. E. , Schrevel J.
Published: April 2021

The freeze-cleave technique demonstrates the presence of gap junctions at early stages of mouse cardiac muscle ontogenesis. The formation and growth of these junctions were studied at 4 stages of development: 10, 14, 18 days post-coitum (dpc) and at the adult stage. The diverse aspects of the gap junctions are interpreted as different steps in their formation. The first indication of this formation seems to be the presence of linear arrays of 9-nm particles on PF faces. At one end of these arrays a small aggregate of particles appears which acts as nucleation site and grows by incorporating individual gap particles and/or linear arrays. Nexuses with arms and/or central particle-free zones would represent intermediate steps in the formation of junctions. The largest nexuses could be formed by fusion of smaller ones and/or by accretion of gap particles. Analysis of the size distribution of gap junctions shows their growth during their development. At 10 dpc the surface area (.S) of nexuses ranges from 01 to 3 × 10−2μm2, at 14 dpc from 01 to 15 × 10−2μm2, at 18 dpc from 0·1 to 26 ×10−2μm2, and at the adult stage from 01 to 54 ×1o−2μm2. The percentage of large nexuses (S>0·5× 10−2μm2) steadily increases from 10 dpc to the adult stage.

Fixation by glutaraldehyde before glycerol infiltration does not induce any modification in the size distribution of adult heart gap junctions.
Common features of coated vesicles from dissimilar tissues: Composition and structure
Authors: Woods John W. , Woodward Michael P. , Roth Thomas F.
Published: April 2021

Coated vesicles were purified and characterized from porcine brain and chicken oocyte. Electrophoresis on sodium dodecylsulphate (SDS) gels showed that coated vesicles from either source have three major proteins in common with apparent molecular weights of 180000, 120000, and 55000 Daltons. Negatively stained specimens from both sources appear to consist of a highly ordered array of short interconnected rods or ridges of material on the exterior surface of a membrane vesicle. Coated vesicles purified from porcine brain and chicken oocyte have mean external diameters of 75·0 and 85·o nm, respectively. In coated vesicles of the appropriate orientation, portions of the coat material appear to be organized into hexagons and pentagons. Based on the observed variability of size and apparent structure, it is postulated that the basic structural subunits of coated vesicles are the short rods or ridges of material observed on the exterior membrane vesicle surface. It is suggested that multiples of these sub-units can be assembled into many arrangements to yield coated vesicles of different sizes and coat structure. It is also proposed that the structural subunit is a complex of 3 proteins of molecular weight of 180000, 120000 and 55000 Daltons.
Regulation of ribosomal rna synthesis in Tetrahymena pyriformis
Authors: Keiding J. , Andersen H. A.
Published: April 2021

Ribosomal RNA is synthesized at constant rate during most of the cell cycle in heat-shock synchronized populations of Tetrahymena pyriformis. Early in each macronuclear S-period the rate of synthesis increases abruptly, concomitant with replication of the genes coding for ribosomal RNA. The increase is prevented by inhibitors of DNA replication, added prior to the S-period. Similarly, in cultures synchronized by starvation/refeeding, inhibition of DNA replication, at the time when the rDNA is replicated, will prevent the normal increase in rate of RNA synthesis which follows refeeding.

We conclude that inhibition of rDNA replication interferes with the synthesis of rRNA, and we suggest that with respect to rRNA synthesis a gene dosis effect is operating in fastgrowing Tetrahymena cells.
Follicle cell bridges in the mosquito ovary: syncytia formation and bridge morphology
Authors: Fiil Annelise
Published: April 2021

In the mosquito, Culex pipiens quinqefasciatus, the follicle cells enveloping the oocyte and the nurse cells are connected by intercellular bridges. The bridges are formed by incomplete cytokinesis, and they persist for more than 30 h after their formation. Reconstructions from serial sections showed that one syncytial group contained at least 32 cells; several cells continued outside the series. The cells in a syncytium divide asynchronously; this results in an irregular, branched organization. The bridges may be either embedded in the cytoplasm of the cells, or they may form an extracellular connexion.
Ouabain binding during plasma membrane biogenesis in duck salt gland
Authors: Hossler Fred E. , Sarras Michael P. , Barrnett Russell J.
Published: April 2021

The conditions necessary for optimal ouabain binding in the avian salt gland were examined. Binding was enhanced by ATP and Mg2+ and was decreased by K+, but was unaffected by added Na+. Both maximal binding and complete inhibition of Na,K-ATPase activity were obtained at 1 × 10−6 M ouabain. Half maximal binding and half maximal inhibition of Na,K-ATPase activity were obtained at 1·7×10−7 M ouabain. Ouabain binding increased in parallel with increasing specific activity of the Na,K-ATPase during salt-induced salt gland specialization. The ratio of Na,K-ATPase activity to ouabain-binding sites remained constant during the salt stress as well as after removal of the salt diet. Autoradiography indicated binding to partially and fully differentiated secretory cells of the salt gland. The ouabain binding assay appeared to be a more useful indicator of membrane amplification than Na,K-ATPase activity since it is rapid, essentially irreversible, less sensitive to tissue fixatives, and quantitatively measured the number of enzyme molecules.
Osmoregulation in the algaVacuolaria virescens. structure of the contractile vacuole and the nature of its association with the golgi apparatus
Authors: Heywood Peter
Published: April 2021

The contractile vacuole of the chloromonadophycean alga Vacuolaria virescens is a permanent structure that possesses a specialized membrane: subunits of this membrane have a diameter of 21—24 nm and in places are arranged in a regular hexagonal pattern. The lateral walls of these subunits form regularly spaced bristles or pegs which extend inwards from the trilaminar membrane for a distance of 13–15 nm. The contractile vacuole is situated immediately above an extensive Golgi apparatus that covers most of the anterior surface of the nucleus. Vesicles of Golgi origin give rise to subsidiary vacuoles which in turn empty into the contractile vacuole. Golgi vesicles, subsidiary vacuoles and the contractile vacuole contain similar electron-dense material. It is suggested that this material might be a highly hydrophilic substance which will attract water from the cytoplasm into the Golgi vesicles, subsidiary vacuoles and contractile vacuole from whence it is discharged from the cell. This method of osmoregulation is compared to that occurring in other algae and protozoa.
New staining method for the detection of sister-chromatid exchanges in brdu-labelled chromosomes
Authors: Alves P. , Jonasson J.
Published: April 2021

A direct Giemsa technique is described for the selective staining of BrdU-substituted chromatin. It can advantageously be used for sister-chromatid exchange analysis and for the detailed cytological study of the mouse kinetochores. Presumptive inversions within the centromeric region of chromosomes 4 and 18 in the C57Black/6J mouse are described.
Developmental stages in the formation of inverted gap junctions during turnover in the adult horseshoe crab, Limulus
Authors: Lane Nancy J.
Published: April 2021

Stages leading to the formation of inverted gap junctions between certain basal replacement or interstitial cells in the mid-gut of adult Limultts can be followed by freeze-fracturing. Free, 13-nm EF intramembranous particles first appear to be organized into short linear arrays or small clusters of particles, which then become transformed into anastomosing particulate networks covering a considerable surface area. These subsequently become concentrated into smaller, more nearly circular, macular plaques of EF particles or PF pits. These EF particles, both when free or assembled into macular arrays, possess a central channel or pore. Numerous formed gap junctions are present in Limulus mid-gut, which suggests that cell-to-cell communication is an important feature of the mature tissue. The results show that arthropod tissues can be used to study the development of gap junctions not only in differentiating systems but also in adult tissues during normal cell turnover.
The effects of surfactants on cell aggregation
Authors: Owen E. , Clifford J. , Marson A.
Published: April 2021

Trypsin-dissociated HeLa and human amnion cells were employed to investigate the effects of an anionic, a cationic, and a non-ionic surfactant on cell reaggregation. All the surfactants inhibited the reaggregation process, the effect increasing with surfactant concentration, and becoming significant at concentrations of about 10−6 M for the anionic surfactant and about 10−5 M for the others. The effects of temperature and ionic strength on HeLa cell aggregation, and of surfactants on the electrophoretic mobility of HeLa cells and the release of sialic acid from their surfaces by neuraminidase, were also examined. The results indicate that although electrostatic effects occur and are likely to be of importance in cell reaggregation, the effect of surfactants cannot be accounted for on this basis alone, and it is suggested that interactions involving the cell-surface glycoproteins, possibly indirectly via effects on the fluidity of the cell membrane, are also significant in this respect.
Invasive locomotory behaviour between malignant human melanoma cells and normal fibroblasts filmed in vitro
Authors: Stephenson E. M. , Stephenson N. G.
Published: April 2021

Explants of human malignant melanoma (MM96), normal adult human skin fibroblasts (HSF) and embryonic chick ventricle were confronted in pairs. The 2 outwandering populations in each confrontation eventually met in a situation where each could potentially invade the other. The human explants were artificially prepared from dissociated cells. The primary objective of the study was to compare the relative invasive capacities of the malignant and non-malignant human cell populations against a standard population of chick heart fibroblasts (CHF). Relative invasiveness was also compared for (a) malignant human melanoma cells against human and avian fibroblasts; (b) the 2 different fibroblast populations against MM96; and (c) the 2 fibroblast populations against each other.

Time-lapse films were prepared for each confrontation. Each cell population was also filmed in a free or unconfronted state. The films were analysed in terms of (a) cell speeds in relation to numbers of contacts; (6) the net radial outward velocity; (c) directional frequencies of movements; and (d) cell behaviour resulting from heterologous collisions. Replicate cultures were fixed and stained 24–40 h after junction. Measurements from these indicated relative distances travelled by each population towards the opposing population and towards free space. Nuclear overlap and population density measurements were also recorded.

MM96 cells invaded the standard chick fibroblast population but eventually encountered moderate obstruction. Invasion was mainly by oriented movement between successive pairs of chick cells. Contact paralysis of ruffling of MM96 was not seen. Human fibroblasts did not invade the CHF population. They were contact inhibited by chick cells and typically reversed their direction of movement. MM96 cells invaded the human fibroblast population without obstruction. They were not usually contact inhibited by HSF and several different pathways for locomotory invasion, including overlapping, were involved. Cells of both fibroblast populations were contact inhibited by leading-edge collisions with MM96 cells. Fibroblast invasion of MM96 was by underlapping of orthogonally oriented, bipolar melanocytes or dendritic processes. Human fibroblasts were less obstructed than chick fibroblasts during invasion of MM96. Initiation of invasion of HSF by chick cells was apparently orientation dependent. Invasion by CHF was assisted by the alignment, morphology and contact-mediated withdrawal of the human cells. Heterologous contact inhibition was not apparently defective for either cell type. In all confrontations involving both malignant and non-malignant cells, the extent and pattern of invasion could be related to apparent deficiencies in heterologous contact inhibition of locomotion.
SV40-transformed hamster cells resistant to 100–250 μg/ml of ethidium bromide
Authors: Wolf Gudrun , Tejmar Liana , Borell Sabine , Klietmann Wolfgang
Published: April 2021

SV4o-transformed hamster cells were selected for resistance to ethidium bromide (EB). Several cell lines were established, which grew in the presence of up to 250 μg/ml EB. The EB resistance is genetically stable. The cloned resistant cells show no difference in morphology, with the exception of the mitochondrial ultrastructure, which exhibits condensed cristae formation. The tumorigenicity of these cells in Syrian gold hamsters is considerably reduced. Incorporation of radioactive labelled thymidine into mitochondrial DNA is not influenced by the presence of the drug. Gel electrophoresis with mitochondrial proteins from wild-type and resistant cells reveals significantly different patterns. The mechanism of EB resistance is discussed.
Directionality of microtubule assembly: an in vivo study with the ciliate Tetrahymena
Authors: Ng Stephen F.
Published: April 2021

A temperature-sensitive mutant homozygous for the recessive gene molb in Tetrahymena thermophila offers opportunity for studying the direction of microtubule assembly in vivo. At 39 °C the mutant fails to divide properly; the 2 daughter animals remain attached and bend over each other. As revealed by protargol staining, the bending results in acute turning and breaking of some of the longitudinal microtubular bands close and parallel to the surface. Hence, 2 broken microtubular ends are available for study of the problem of directionality of microtubule assembly, by assessing which of the 2 ends regenerates. In most cases the posterior portion of the longitudinal microtubular band regenerates. The present study hence supports the conclusion based on in vitro observation in other systems that microtubule assembly is predominantly unidirectional.
The cell surface of Trypanosoma cruzi: cytochemistry and freeze-fracture
Authors: De Souza W. , Martfnez-Palomo A. , González-Robles A.
Published: April 2021

The ultrastructure of epimastigotes of Trypanosoma cruzi, obtained from, acellular cultures, and bloodstream trypomastigotes, isolated from infected mice, were studied by thin-sectioning and freeze-fracturing techniques. Epimastigotes showed a thin (5 nm) surface coat when stained with ruthenium red, while the surface coat of trypomastigotes was more prominent (15 nm thick). Both P and E faces of the plasma membrane of T. cruzi had roughly the same number of intramembranous particles (IMP) as seen by freeze-fracture. The plasma membrane of bloodstream trypomastigotes had less IMP than epimastigotes. Several differentiations of the plasma membrane were observed. In epimastigotes a cytostome appears as a particle-poor region delimited by a pallisade-like row of adjacent IMP. Bloodstream trypomastigotes did not have a cytostome. Instead, abundant pinocytic vesicles were observed. At the base of the flagellum of epimastigotes a ciliary necklace was found. At this region, the surface coat was differentiated as long, hair-like projections after staining with ruthenium red. The flagellar membrane had less IMP than the body membrane. Clusters of IMP were present on both faces of the flagellar membrane at the flagellar-body adhesion zone of epimastigotes. Linear arrays of IMP were also seen. In bloodstream trypomastigotes clusters of particles were observed both on the flagellar and cell body membranes. Our observations demonstrate the presence of considerable structural variations of the T. cruzi plasma membrane at the two stages of the life cycle studied.
Labelling of human resting lymphocytes by continuous infusion of [3H]thymidine. I. characterization of cytoplasmic label
Authors: Schick P. , Trepel F. , Maisel K. H. , Past W. , Reisert I. , Begemann H. , Pilgrim Ch.
Published: April 2021

After continuous 3H-TdR infusion in vivo or incubation with 3H-TdR in vitro human blood lymphocytes were examined by light-microscopic and electron-microscopic autoradiography. Using relatively long autoradiographic exposure times (50–300 days) not only nuclear but also cytoplasmic labelling was visualized, the cytoplasmic label being present in up to 96 % of the cells. The cytoplasmic label was predominantly associated with the mitochondria and was removed from the cells nearly completely by treatment with DNase but not with RNase or cold perchloric acid. It is concluded that this cytoplasmic label mainly represents 3H-TdR incorporated into mitochondrial DNA which is continuously renewed in an average turnover time of 14 days or less. This value is compatible with a turnover time of 11 days for mitochondrial DNA in mammalian cells reported in the literature.
An experimental study of the interaction between the soil amoeba Naegleria gruberi and a glass substrate during amoeboid locomotion
Authors: Preston T. M. , King C. A.
Published: April 2021

The amoeboid locomotion of the soil protozoon Naegleria gruberi has been studied using reflexion-interference microscopy. Two types of contact are made with a planar glass substrate. One, formed at a considerable distance from the substrate in deionized water (≏100 nm) has been termed ‘associated contact’ and usually involves a considerable surface area (of the order of 100 μm1), i.e. about a third of the cell profile. From this broad platform filopodia are produced which form close contacts (‘focal contacts’). In locomotion the area of associated contact is very mobile, in contrast to the focal contacts which, once established, are stable. Focal contact sites are left behind on the glass surface (‘footprints’) when the amoeba moves away. The cell-substrate gap in the associated contact is greatly affected by the ionic strength of the medium and particularly the valency of the cation component. This suggests that long-range forces of attraction play an important role in keeping the amoeba close to a substrate and thus allow the production of filopodia from the ventral surface to form focal contacts.
Long-term maintenance of reaggregated hypothalamic cultures developed from embryonic rat hypothalamus: prostaglandin release during synaptogenesis in vitro
Authors: Gyévai Angela , Chapple Paul J. , Douglas William H. J.
Published: April 2021

Hypothalamic aggregate cultures were developed from hypothalami taken from rat embryos at 17—19 days gestation. The aggregate cultures exhibited a prominent morphological differentiation during 3-4 weeks in culture. The fine structure of the synapses formed in the aggregates resembled synapses in the adult animal. During synaptogenesis the aggregates spontaneously released prostaglandin E2 (PGE2). The amount of PGES released in the media was reversed upon the morphological differentiation of the hypothalamic cultures. Media containing a higher PGE2 concentration increased the extracellular prolactin accumulation in monolayer cultures developed from adult rat hypophysis.
Acetylcholinesterase: A useful marker for the isolation of sarcolemma from the bivalve (Modiolus demissus demissus) myocardium
Authors: Watts John A. , Pierce Sidney K.
Published: April 2021

The presence of cholinesterase activity in M. demissus hearts was demonstrated by light- and electron-microscopic histochemistry and by enzymic assay. The enzyme proved to be acetylcholinesterase (AChE) since acetylthiocholine was the preferred substrate, and eserine or BW284c51inhibited the enzyme activity, while isoOMPA was without effect. The AChE was localized and uniformly distributed along the cell surface membranes of the cardiac muscle cells. A fraction 8-fold enriched in AChE was isolated from pooled ventricles by a combination of differential and sucrose density gradient centrifugation. This sarcolemmal fraction contained little mitochondrial contamination as determined by electron microscopy and by succinate cytochrome c reductase activity. In addition, this fraction stained uniformly for AChE, indicating that it was free of other membrane types (for example sarcoplasmic reticulum which did not stain for ACHE). Therefore, this fraction contained purified cell surface membrane free of contamination by other membranous organelles.
Nuclear divisions with reduced numbers of microtubules in Tetrahymena
Authors: Jaeckel-Williams Ruth
Published: April 2021

Ultrastructural observations on micronuclei of Tetrahymena thermophila, WH-6 dividing in the presence of colchicine suggest that the microtubules present at the time of drug addition persist, but no new microtubules are formed. It is suggested that the great amount of elongation seen in these drug-treated micronuclei may not be due to the persisting microtubules alone. On the basis of ultrastructural and light-microscopic observations, it is proposed that at least part of the motive force responsible for the elongation which occurs in colchicine may be due to an active sliding of the micronuclear membrane on some element of the cell cortex. In addition, the macronucleus of this species of Tetrahymena was found to be capable of dividing in colchicine in the absence of internal nuclear microtubules.
The pattern of appearance of enzymic activity during the development of the golgi apparatus in amoebae
Authors: Flickinger Charles J.
Published: April 2021

The appearance of enzymic activity during the development of the Golgi apparatus was studied by cytochemical staining of renucleated amoebae. In cells enucleated for 4 days, there was a great decline in size and number of Golgi bodies, or dictyosomes. Subsequent renucleation by nuclear transplantation resulted in a regeneration of Golgi bodies. Samples of amoebae were fixed and incubated for cytochemical staining at intervals of 1, 6, or 24 h after renucleation. Enzymes selected for study were guanosine diphosphatase (GDPase), esterase, and thiamine pyrophosphatase (TPPase). All three were found in the Golgi apparatus of normal amoebae but they differed in their overall intracellular distribution. GDPase was normally present at the convex pole of the Golgi apparatus, in rough endoplasmic reticulum, and in the nuclear envelope. In amoebae renucleated for 1 h, light reaction product for GDPase was present throughout the small stacks of cisternae that represented the forming Golgi apparatus. By 6 h following the operation GDPase reaction product was concentrated at the convex pole of the Golgi apparatus. Esterase, which was distributed throughout the stacks of normal Golgi cisternae, displayed a similar distribution in the forming Golgi bodies as soon as they were visible. TPPase was normally present in the Golgi apparatus but was not found in the endoplasmic reticulum. In contrast to the other enzymes, TPPase reaction product was absent from the forming Golgi apparatus 1 and 6 h after renucleation, and did not appear in the Golgi apparatus until 24 h after operation. Thus, enzymes held in common between the rough endoplasmic reticulum and the Golgi apparatus were present in the forming Golgi apparatus as soon as it was detectable, but an enzyme cytochemically localized to the Golgi apparatus only appeared later in development of the organelle. It is suggested that Golgi membranes might be derived from the endoplasmic reticulum and thus immediately contain endoplasmic reticulum enzymes, while Golgi-specific enzymes are added later in development.
Action of a slowly hydrolysable cyclic amp analogue on developing cells of Dictyostelium Discoideum
Authors: Grossier C. , Gerisch G. , Malchow D. , Eckstein F.
Published: April 2021

Adenosine 3’,5’-cyclic phosphorothioate (cAMP-S) is a cyclic AMP (cAMP) analogue which is only slowly hydrolysed by phosphodiesterases of Dictyostelium discoideum. The affinity of cAMP-S to cAMP receptors at the cell surface is only one order of magnitude lower than that of cAMP. cAMP-S can replace cAMP as a stimulant with respect to all receptor-mediated responses tested, including chemotaxis and the induction of cAMP pulses.

cAMP-S does not affect growth of D. discoideum but it blocks cell aggregation at a uniform concentration of 5 × 10−7 M in agar plate cultures of strain NC-4 as well as its axenically growing derivative, Ax-2. Another wild-type strain of D. discoideum, v-12, is able to aggregate on agar plates supplemented with 1 mM cAMP-S. The development of Polysphondylium pallidum and P. violaceum is also highly cAMP-S resistant.

In Ax-2 both differentiation from the growth phase to the aggregation-competent stage and chemotaxis are cAMP-S sensitive, whereas in v-12 only chemotaxis is inhibited, v-12 can still form streams of cohering cells and fruiting bodies when chemotaxis is inhibited by cAMP-S.

Whereas cAMP induces differentiation into stalk cells at concentrations of 10−3 or 10 −4 M, cAMP-S has the same effect in strain v-12 at the much lower concentration of 10−8 M.
Analysis of the significance of a periodic, cell size-controlled doubling in rates of macromolecular synthesis for the control of balanced exponential growth of fission yeast cells
Authors: Barnes A. , Nurse Paul , Fraser R. S. S.
Published: April 2021

Mutant strains of the fission yeast Schizosaccharomyces pombe are available which divide at smaller mean sizes than wild type. Earlier work by the present authors has shown that all these strains double their rates of polyadenylated messenger RNA synthesis as a step once in each cell cycle. The smaller the cell, the later in the cycle is the doubling in rate of synthesis. Strains of all sizes, however, double their synthetic rate when at the same threshold size. We show here that the differences in cell cycle stage of doubling in rate of polyadenylated messenger RNA synthesis are enough to explain the reduced mean steady state polyadenylated messenger RNA contents of the smaller strains. The cell size-related control over doubling in rate of synthesis is also shown to maintain the mean polyadenylated messenger RNA content as a constant proportion of cell mass, irrespective of cell size. This control thus allows cells to maintain balanced exponential growth, even when absolute growth rate per cell is altered by mutation. It is also shown that the concentration of polyadenylated messenger RNA itself could act as a monitor of the threshold size triggering the doubling in rate of synthesis in each cell cycle.
Three-dimensional reconstruction of the chromatin bodies in the nuclei of mature erythrocytes from the newt Triturus Cristatus: the number of nuclear envelope-attachment sites
Authors: Murray A. B. , Davies H. G.
Published: April 2021

The arrangement of the chromatin bodies in the interphase nuclei of 6 erythrocytes has been investigated by means of 3-dimensi0nal reconstruction from electron micrographs of serial sections. When the borders of chromatin bodies are marked on the surface of each model, discrete areas of chromatin in contact with the nuclear envelope are revealed. The number of these areas is approximately equal to the number of chromosomes in the diploid set. The data suggest that each chromatin body corresponds to a condensed interphase chromosome and that each chromosome is attached to one discrete site on the nuclear envelope. The data are insufficient to show whether or not the condensed chromosomes are arranged in any orderly pattern in these nuclei.
A cinematographical study of the development of vitally stained Dictyostelium discoideum
Authors: Durston A. J. , York Frida
Published: April 2021

We report observations from time-lapse films of the development of Dictyostelium discoideum (Dd) stained with the vital dye neutral red. We have used this dye to enable us to track individual cells, as they move through multicellular tissues in later Dd stages. Our observations lead us to new conclusions about the control of cell movement and cell contact during Dd development, including the tentative conclusion that the aggregation control competences regulate cell behaviour throughout the whole of Dd. development. We are led to specific hypotheses for the mechanisms of later morphogenetic movements and pattern formation.
Quantitative immunoelectron-microscopic analyses of the distribution of cytochrome p-450 molecules on rat liver microsomes
Authors: Matsuura Shiro , Fujii-Kuriyama Yoshiaki , Tashiro Yutaka
Published: April 2021

Specific antibodies to phenobarbital-induced cytochrome P-450 were prepared by affinity chromatography and coupled to ferritin with glutaraldehyde. The ferritin antibody conjugates with molecular ratio of approximately one were isolated by gel filtration and were used for immunochemical and immunoelectron-microscopic analyses of the distribution of cytochrome P-450 on microsomes from untreated, phenobarbital- and methylcholanthrene-treated rats.

Binding assay showed that at the saturation level of the antibodies, microsomes from untreated, phenobarbital- and methylcholanthrene-treated rats bind 0·25, 0·41 and 0·14 mol of the antibody per mol of cytochrome P-450, respectively. From these data, the maximum number of the ferritin particles which can bind with microsomes was calculated. This number was in good agreement with the average number of ferritin particles bound per microsome which was determined by electron-microscopic observations of the microsomes incubated with the antibody conjugates at saturation level. Electron-microscopic observations also indicated that smooth microsomes can bind more conjugates than rough microsomes and this finding was consistent with the biochemical data that, on the protein basis, smooth microsomes contain more cytochrome P-450 than rough microsomes, even after correction for ribosomal proteins. The number of ferritin particles bound per smooth microsome was proportional to the diameter and non-random distribution of the ferritin particles on the microsomal vesicles, which was deduced simply by inspection in the previous paper from this laboratory, was confirmed by statistical analyses of electron micrographs of the labelled microsomes.
Capping of surface immunoglobulin on ‘hairy cells’ is independent of energy production
Authors: Splinter TH. A. W. , Collard J. G. , De Wildt Anke , Temmink J. H. M. , Décary Francine
Published: April 2021

Capping, independent of metabolic energy, of surface immunoglobulin (S-Ig) on ‘Hairy cells’ from patients with Hairy cell leukaemia (HCL) is described. As controls leukaemic cells from a patient with a prolymphocytic leukaemia (PLL) and blood lymphocytes from healthy individuals were used. The specificity of the energy-independent capping of the HCL-cells as compared to the controls was tested by incubation of the cells at 4 °C in the presence of 01 M· sodium azide with different FITC-labelled ligands.

In order to find an explanation for this phenomenon, the influence of cytochalasin B, colchicine and the combination of both drugs on capping of S-Ig and concanavalin (Con A)-receptors at 37 °C was investigated. Furthermore the effect of Con A on S-Ig capping and vice versa was studied. The results show that only S-Ig on HCL cells could form caps at 4 °C in the presence of sodium-azide. Cytochalasin B alone induced a strong inhibition of Con A capping on all 3 cell types, whereas S-Ig capping was unaffected. Colchicine alone had practically no effect. Anti-Ig inhibited subsequent patch and cap formation with Con A on both HCL cells and PLL cells, whereas Con A caps and patches were redistributed by anti-Ig on PLL cells, but not on HCL cells. Conversely, Con A could link S-Ig to other receptors, leading to inhibition of S-Ig capping at 4 °C on HCL cells and to co-capping of S-Ig at 37 °C on both cell types. In addition Con A induced redistribution of S-Ig caps. The combination of cocapping of S-Ig by Con A, followed by redistribution of the caps by FITC-anti-Ig simulated inhibition of S-Ig capping by Con A on PLL cells.

The major conclusions are: in some cases inhibition of capping may actually be caused by redistribution of caps; the energy-independent capping cannot be explained by free diffusion of S-Ig in the membrane through lack of any connexion with receptor-mobility regulating systems. It is proposed that the energy requirement of capping is needed to inactivate a specific mechanism, which restrains receptor mobility and which is non-operative in HCL cells.
Analysis of the time of utilization of proteins for the initiation and completion of division in synchronized Tetrahymena Pyriformis
Authors: Wheatley D. N. , Rasmussen L. , Zeuthen E.
Published: April 2021

We exposed division-synchronized Tetrahymena pyriformis to pulse treatments with the structural amino acid analogue p-fluorophenylalanine. We related the timing of the treatments to the delays in onset or completion of division, or both. The results enable us to distinguish between early and late phases in cell division in which the cells bring into action proteins which contribute to initiate and complete division, respectively. Normally, these events are separated by 25–30 min.
An experimental analysis of bivalent interlocking in spermatocytes of the newt Triturus Vulgaris
Authors: Callan H. G. , Pearce S. M.
Published: April 2021

Interlocked bivalents at 1st meiotic metaphase are relatively uncommon in spermatocytes of the newt Trituras vulgaris, but their frequency of occurrence can be significantly increased by subjecting newts to a 24-h heat shock. Newt spermatocytes are sensitive to a heat shock at any stage between the end of premeiotic S and mid to late pachytene. The heat shock does not cause evident desynapsis, nor does it significantly affect chiasma frequency; therefore the interlocked condition induced in spermatocytes which were subjected to a heat shock when they were in zygotene or pachytene is unlikely to be a consequence of synaptic trapping. By way of explanation it is suggested that a heat shock may cause telomeres to detach from the nuclear membrane, or from the synaptonemal complex where the latter is attached to the membrane, thus allowing non-homologous chromonemata to become intertwined before chiasmata have formed. If this explanation is valid, it is then further suggested that the recombination process which results in chiasma formation probably takes place in chromosomal regions lying outside the synaptonemal complex, rather than inside, between its 2 lateral elements.
Antibody staining of 10-nm (100-Å) filaments in cultured smooth, cardiac and skeletal muscle cells
Authors: campbell Gordon r. , chamley-campbell Julie , gröschel-stewart Ute , Small J. V. , Anderson Paul
Published: April 2021

Antibodies were prepared against the SDS-denatured 10-nm filament protein ‘skeletin’ extracted from chicken gizzard. The specificity of the antibody to the 10-nm filament protein was shown by immunodiffusion before and after purification of the protein on SDS gels by the enzyme-linked immunoabsorbent assay (ELISA) and by its specific absorption with purified skeletin. In immunofluorescence (where preimmune sera and antigen-absorbed antisera gave negative results), cultured cardiac, skeletal and smooth muscle cells and endothelial cells stained intensely. No staining was observed in fibroblasts present in these cultures, nor was there staining in glial cells or nerve cell bodies and fibres from sympathetic ganglion and Auerbach’s plexus cultures. Smooth muscle cells (regardless of their source and phenotypic state) and endothelial cells stained intensely in the perinuclear region and in a fine filamentous network that existed throughout the cytoplasm. In both chick and rat skeletal and cardiac muscle (cultures and frozen sections) filamentous network staining was observed, while in rat muscle the antibody was additionally localized in a regular pattern in the region of the Z-disk, and in the case of cardiac muscle associated with the intercalated disk. The addition of 10−6M colchicine to the culture medium of smooth and striated muscle and endothelial cells resulted in an aggregation of the filaments in the nuclear region.

Cultured smooth and striated muscle and endothelial cells and freshly isolated smooth muscle cells extracted of actomyosin and tubulin by high and low ionic strength solutions gave a staining pattern similar to non-extracted cells and in the electron microscope, exhibited filaments of predominantly 10 nm diameter.
Fine structure of meiotic prophase chromosomes and modified synaptonemal complexes in diploid and triploid Rhoeo Spathacea
Authors: Lin Yue J.
Published: April 2021

The synaptonemal complex (SC) in the diploid Rhoeo consists of 2 amorphous lateral elements, each about 46.0 nm thick, and one amorphous central element about 30.0 nm thick. The central region is about 115.0 nm wide. SC in the triploid have essentially the same dimensions as those of the diploid; both lateral (46.0 nm) and central (30.0 nm) elements are amorphous, and the central region is about 117.5 nm wide. The coil, observed in both diploid and triploid, is a modified short segment of SC with several twists at the end of a synapsed bivalent that is attached to the nuclear membrane. Serial sections in a diploid cell reveal that a coil extends inwards about 3.5 μm from the nuclear membrane and makes a complete turn at a distance of every 0.5 μm. There is a correlation between the modified ends of SC and terminal chiasmata in Rhoeo. The coils might have a positive role in the process of crossing over, or alternatively might be involved in ring formation by holding chromosome ends together while chiasmata are not involved. SC are present in chromocentres of both diploid and triploid. Chromocentres in diploid and triploid are indistinguishable, and appear to be formed from the aggregation of pericentromeric heterochromatin as a result of translocations which occurred close to the centromeres. 3-dimensional hypothetical pachytene configuration of the diploid is presented.
Ammonia plus another factor are necessary for differentiation in submerged clumps of Dictyostelium
Authors: Sternfeld John , David Charles N.
Published: April 2021

Differentiation of Dictyostelium amoebae can occur in submerged clumps of cells; under an oxygen atmosphere mature stalk cells and spores form, as has been shown in previous work. This report shows that at least 2 factors are released by the cells under these conditions, and that both, together, are required for differentiation of stalk cells and spores. One of the factors is ammonia (NH3 + NH4+). The other factor(s) is heat stable and dialysable but has not yet been further characterized. The factors can be collected in conditioned medium and, when added to cells, stimulate differentiation. Conditioned medium loses its biological activity upon the removal of the NH3 + NH4+. When NH3 + NH4+ is added back, activity is restored. Because NH3 + NH4+, alone, has no activity, a second factor(s) in the conditioned medium must be required for differentiation. It is also shown that calcium inhibits differentiation in submerged clumps and that in calcium-free medium the timing of differentiation is essentially the same as under aerial conditions.
A lectin which binds specifically to β-D-galactoside groups is present at the earliest stages of chick embryo development
Authors: Cook G. M. W. , Zalik S. E. , Milos N. , Scott V.
Published: April 2021

Extracts obtained from chick embryos at the pre-gastrula and gastrula stages are able to agglutinate trypsinized rabbit erythrocytes fixed with glutaraldehyde. Agglutination is inhibited by saccharides sharing a β -D-galactopyranoside configuration. Agglutinin activity is also inhibited by desialysed fetuin which bears terminal galactose residues but not by native fetuin, desialysed agalacto-fetuin, bovine submaxillary mucin and desialysed bovine submaxillary mucin. Lectin activity is present in extracts obtained from the embryonic area of the blastoderms as well as in extracts from extra-embryonic endoderm and ectoderm. In extracts subjected to gel filtration on ECD Sepharose, lectin activity eluted between ovalbumin (mol. wt 45000) and chymotrypsinogen (mol. wt 25000). Under some experimental conditions, thiodigalactoside, the most potent inhibitor of lectin activity, inhibited the aggregation of cells of the extra-embryonic endoderm of the primitive chick embryo.