bluuebunny/crossref_metadata_2025 huggingface dataset:
The three-dimensional arrangement of intergranal lamellae in chloroplasts
Authors: Paolillo D. J.
Published: April 2021

The intergranal lamellae, or frets, are helically arranged around each granum. All helices within a plastid are co-directional. The helices wind in the same direction from plastid to plastid, from cell to cell, and from species to species, and are right-handed. The continuity of the intergranal membranes in 3-dimensional space is facilitated by the fact that all the helices within a plastid wind in the same direction. Because there are multiple frets around each granum, the fretwork consists of a highly interlocked series of membranes. The incompleteness of the integration of the fretwork suggests that continuity of the membranes from granum to granum is obtained, to a large degree, by fusions of membranes rather than by continuous growth and ramification of a simpler membrane system.
The formation of filopodium-like processes by Trypanosoma (Trypanozoon) brucei
Authors: Wright K. A. , Lumsden W. H. R. , Hales H.
Published: April 2021

Bloodstream forms of Trypanosoma (Trypanozoon) brucei were observed in vitro to possess long filaments arising from both the posterior end and the tip of the flagellum, and also to become attached to various substrates, including themselves. The filaments were examined by negative staining and thin-section techniques and found to have a structure resembling filo podia. However, as their functional significance is still questionable, their designation as filopodia must be considered provisional. The possible functions of these structures, such as mediation of attachment, and release of antigenic material, are discussed.
Cytochemical localization of transferase activities: carnitine acetyltransferase
Authors: Higgins Joan A. , Barrnett R. J.
Published: April 2021

Two methods for the cytochemical detection of free CoA and their utilization in the fine-structural localization of carnitine acetyltransferase in rat heart are described. The first utilizes the reducing property of the SH group of CoA to reduce potassium ferricyanide to potassium ferrocyanide, which in the presence of uranyl ions forms an electron-dense precipitate of uranyl ferrocyanide. The second utilizes the mercaptide-forming property of the free SH group of CoA, which forms a precipitate with cadmiumions.

Using the uranyl-ferrocyanide method, reaction product due to endogenous enzymic activity was found on and between the cristae and between the inner and outer membranes of the mito- chondria in fresh heart muscle. In aldehyde-fixed tissue activity was recorded only between the inner and outer membranes. Endogenous activity was removed by preincubation of the tissue in a solution of ferricyanide. On addition of acetyl CoA and carnitine to the incubation medium, fresh tissue, which had been preincubated in ferricyanide, showed reaction product between and on the cristae and between the inner and outer membranes of the mitochondria, while fixed tissue showed reaction product in the latter position only. In both cases the activity between the outer and inner mitochondrial membranes was dependent on both acetyl CoA and carnitine, while the cristae reaction occurred in the absence of carnitine, but required acetyl CoA. All activity was inhibited by mercuric chloride. Acetyl carnitine reduced the activity in the fixed tissue and had severe effects on the structure of fresh mitochondria. These results suggest the presence of carnitine acetyltransferase, which survives aldehyde fixation, on the inner surface of the outer mitochondrial membrane and/or the outer surface of the inner mitochondrial mem- brane. A second enzyme which released CoA from acetyl CoA occurred in relation to the cristae of unfixed mitochondria.

The cadmium method was less satisfactory than the uranyl-ferrocyanide method but with fixed tissue gave confirmatory results.
Localization of intranucleolar dna in root meristematic cells ofAllium cepa
Authors: Chouinard L. A.
Published: April 2021

The problem of the localization of intranucleolar DNA in root meristematic cells of Allium cepa has been investigated under both light and electron microscopy following formaldehyde fixation coupled with deoxyribonuclease extraction. The observations reveal that most, if not all, of the intranucleolar DNA is located inside small lacunar spaces confined to the fibrillar regions of the nucleolar mass. These results would be consistent with the view that the DNA-containing lacunar spaces in question correspond, in fact, to cross-sections of a meandering channel containing the extended nucleolar organizing segment of the nucleolar chromosome.
The structure of sycamore callus cells during division in a partially synchronized suspension culture
Authors: Roberts K. , Northcote D. H.
Published: April 2021

Sycamore suspension callus cells have been partially synchronized to give a culture with a rmtotic index of 15 %. Living dividing cells of the culture have been examined with Nomarski differential interference optics and a comparable study made on fixed cells with the electron microscope.

An organized band of reticulate cytoplasm partially encircles the nucleus at mitosis. The cell divides by the formation of a phragmosome which grows across the large vacuole; this allows the organization of the cytoplasm which forms the cell plate to be examined separately from the more general cytoplasm of the cell. The cell plate grows from one side of the cell to the other and down its length a complete developmental sequence can be seen.

The Golgi bodies and the endoplasmic reticulum are probably involved in the formation of material for the construction of the cell plate and young cell wall. Microfibrils are formed within the plate in the more mature regions, while material contained within vesicles is incor porated at the young growing edge. At the edge of the plate microtubules are found and these correspond to the fibnllar appearance of the phragmoplast seen with the optical microscope.

In the living cell an active movement of organelles along the peripheral cytoplasm can be seen and with fixed cells viewed with the electron microscope microtubules are often found adjacent to the plasmalemma and lying close to mitochondria, crystal-containing bodies and plastids.

The appearance of crystal-containing bodies and plastids containing phytoferntin is described.
Temperature-induced orientation instability during meiosis: An experimental analysis
Authors: Henderson S. A. , Nicklas R. B. , Koch Carol A.
Published: April 2021

Low-temperature (10 °C) treatments alternated with periods at room temperature (25 °C) were found to induce grossly abnormal kinetochore orientation behaviour during metaphase I in spermatocytes of the grasshopper Melanoplus differentialis. These orientation abnormalities involved multiple reorientations before a stable bipolar co-orientation was finally achieved. Similar orientation abnormalities have been induced in the past by heat-treatments in the locust Schistocerca. The unstable orientation behaviour of bivalents in eight cold-treated living spermatocytes was analysed with the aid of time-lapse cinematographic records. Analyses made included the relationship between kinetochore position within the spindle and reorientation, the frequency of reorientation and the velocity of chromosome motion after reorientation. It had been shown previously, in untreated cells, that physical tension applied by micromanipulation could stabilize experimentally induced unipolar bivalents and inhibit their reonentation. Two experiments of this type with cold-treated spermatocytes were successful in demonstrating the importance of tension in the maintenance of stability, even in unstable, cold-treated material. In discussing the interpretation of these results it is proposed that bivalents have an inherent tendency to reorient periodically. In untreated cells the time between reorientations is long enough to enable interkinetochoric tensions, and resulting stable bipolar orientation, to be achieved, but in cold-treated cells this time is reduced, thereby hindering the establishment of tension and stability.
Cell surface components and growth regulation in cultivated arterial smooth muscle cells
Authors: Nilsson Jan , Ksiazek Tomasz , Thyberg Johan , Wasteson Ake
Published: April 2021

The surface of rat arterial smooth muscle cells was characterized with respect to some of its chemical and functional properties. The effects of selective enzymic degradations (hyaluronidase, chondroitinases, heparitinase or neuraminidase) on [35S]sulphate-prelabelled cells and on binding sites for cationized ferritin (CF) were examined to assess the presence and relative importance of individual species of macromolecules on the cell surface. The results indicate that about half of the strongly anionic sites on the cell surface (binding CF at pH 2·0) could be ascribed to sulphate groups of glycosaminoglycans and about half to carboxyl groups of sialic acid residues in glycoproteins and/or glycolipids. Weaker anionic sites (binding CF at pH 7·0) largely originated from carboxyl groups of glycosaminoglycans. Chondroitin sulphate and heparan sulphate were the main glycos aminoglycans. The surface of cells from young animals showed a higher glycosaminoglycan and a lower sialic acid content than that of cells from adult animals.

Continuous treatment of the cultures with neuraminidase stimulated serum-induced initiation of DNA synthesis, while treatment with hyaluronidase or heparitinase inhibited it. Addition of hyaluronic acid, heparin or heparan sulphate to the culture medium inhibited initiation of DNA synthesis as well as cell proliferation. The effect was more marked in cultures of cells from young animals than from adults, although the latter cells were found to grow at a higher rate and to higher densities. These results suggest a role for cell-surface and pericellular glycoconjugates in growth regulation. A possible mechanism of action is that these molecules, due to their anionic charge or by steric exclusion, interfere with the binding of platelet-derived growth factor, a highly cationic polypeptide, to its cell-surface receptor.
The interval of the cytoplasmic cycle observed in non-nucleate egg fragments is longer than that of the cleavage cycle in normal eggs of Xenopus laevis
Authors: Shinagawa Atsunori
Published: April 2021

Non-nucleate fragments of animal eggs are known to show a cyclic change in rigidity similar to the change observed in the cleavage of normal eggs. The interval of cyclic change was compared with that of the cleavage cycle in Xenopus eggs. Procedures employed by both Hara and co-workers and Sakai & Kubota to obtain non-nucleate fragments were carefully repeated. Non-nucleate fragments produced by either of those procedures had a longer duration of cycle than the normal cleavage cycle (about 30%). By injection of colchicine or vinblastine the cycle of the nucleate fragment was lengthened to coincide with that of the non-nucleate partner. It is suggested that the basic cycle length intrinsic to egg cytoplasm is modulated by assembly-disassembly of the mitotic apparatus, as demonstrated by Sluder in the chromosome cycle of sea urchin eggs.
Units of chromosome replication and packing
Authors: Mullinger Ann M. , Johnson Robert T.
Published: April 2021

Fusion between mitotic and S-phase cells induces the formation of prematurely condensed chromosomes (PCC) in the interphase partner. Viewed in the light microscope, S-phase PCC derived from the Indian muntjac appear to be fragmented and heterogeneous. In scanning electron micrographs prepared by an osmium impregnation technique, which avoids the need to sputter-coat the specimen, the S-phase fragments derived from an individual cell are resolved into about 1000 fibre aggregates, together with more dispersed fibres. Aggregates are roughly spherical and vary in diameter between about 0 · 25 and 1 · 6 μm. The spatial distribution of the aggregates shows some order: chains of single aggregates and, less commonly, duplicated chains occur. Regions of the PCC where the fibres are more dispersed are considered to be likely candidates for sites of replication at the time of fusion. The relationship between the condensed aggregate structure of the S-phase PCC and replication clusters is discussed, and also the assembly of aggregates to form metaphase chromosomes.
Glycoproteins of the AKR leukaemia cell surface and their relevance to leukaemiaspecific surface antigens
Authors: McClure Myra O. , Cook Geoffrey M. W.
Published: April 2021

An attempt has been made to prepare antibodies against leukaemia-specific surface antigens by immunizing (C57 Bl/6 × C3H/He)F1 mice with formaldehyde-stabilized AKR leukaemic cells. The presence of antibodies was examined by indirect immunofluorescence microscopy (IFM) and the indirect antiglobulin resetting reaction (IARR). Galactose oxidase treatment destroyed the ability of leukaemic cells to react with antibodies prepared in the hybrid mice, an effect that was reversed by treating the enzyme-modified cells with borohydride. Analysis by immunoprecipitation and polyacrylamide gel electrophoresis of leukaemic cells, labelled by the galactose oxidase/[3H]-NaBFU technique, indicated that a group of glycoproteins of apparent molecular weight >70 000 was involved. Antibodies could be raised in AKR mice to the same group of glycoproteins by immunization with irradiated leukaemic cells or irradiated neuraminidase-treated leukaemic cells. The level of antibody raised in AKR mice had no effect on the growth of leukaemic lymphoblasts introduced subcutaneously into the host.

Antibodies prepared in hybrid mice against leukaemic cells also were absorbed by lymphoid cells of pre-leukaemic 6-month-old AKR mice, indicating that contrary to previous claims in the literature antigens detected by such antisera are not related to malignancy. Hybrid mouse serum cross-reacted with antigens from purified RNA virus isolated from Abelson lymphoma, as demonstrated by the immunoelectrophoretic blotting technique. The pattern of reactivity was not appreciably altered following the absorption of antibodies directed against leukaemic cells. It is concluded that the glycoproteins detected by us may not be viral antigens but normal high molecular weight lymphoid glycoproteins with altered glycosylation patterns that are induced when the viral genomes are expressed.
Thymidylate kinase from Acetabularia ii. regulation during the life cycle
Authors: Groot Egon J. De , Schweiger Hans-Georg
Published: April 2021

The activity of dTMP kinase (EC 2.7.4.9) was estimated during the development of Acetabularia. Enzyme activity was increased at the beginning of the generative phase. Regulation of the dTMP kinase activity was observed even in the absence of the nucleus. More than 30 days after enucleation the enzyme activity increased two- to threefold. The increase in activity was inhibited by puromycin and cycloheximide but not by rifampicin and chloramphenicol.

These results indicate that the enzyme is coded by the nuclear genome and translated on 80 S ribosomes. From mixing experiments with low-activity and high-activity cell extracts it is concluded that the regulation is due to de novo synthesis of the enzyme.
Residual cell division measurements are unreliable as indicators of the timing of events in the Saccharomyces cerevisiae cell cycle
Authors: Richmond K. M. V. , Williamson D. H.
Published: April 2021

We report here an analysis of the execution point of the temperature-sensitive Saccharomyces cerevisiae cell cycle mutant, cdc27-47. When a logarithmically growing culture was shifted from standard growth conditions (strain 27.8B growing in YEPD at 25 °C) to the restrictive temperature cell division ceased abruptly and reproducibly within one population doubling time, the extent of cell division indicating an execution point early in the cell cycle. Approximately 50% of stationary-phase cells were able to divide when refed with fresh medium at 37 °C, showing that the execution point could be passed before ‘start’. This makes the sharp cut-off in cell division difficult to explain. This difficulty was compounded by observations of the cell cycle stage at which individual cells acquired the capacity to divide at 37 °C. Half the cells that were budded at the time of a temperature shift-up formed three division-blocked cells, and in 11 of these 13 cases, two were descended from the original mother cell and one from the original bud. Thus, mother and daughter cells pass the execution point independently; daughters usually during G1, and mothers usually in the budded phase of the previous cycle. The sharp cut-off in cell division is therefore spurious, and a mechanism is proposed to account for it, which has implications for the interpretation of the execution points of other cdc mutants.

In addition, the expression of the cdc27-47 execution point was modified by both genetic and environmental factors, being affected by carbon source, by the petite condition, and by genetic background. This illustrates the difficulties of interpreting execution point data and the dangers of extrapolation of cell cycle parameters between strains and growth conditions.
Nuclear structure: determination of the fate of the nuclear envelope in Drosophila during mitosis using monoclonal antibodies
Authors: Fuchs J.P. , Giloh H. , Kuo C.H. , Saumweber H. , Sedat J.
Published: April 2021

Libraries of monoclonal antibody against nuclear proteins of Drosophila melanogaster have been established recently to investigate nuclear structure and function. Some of the antibodies have been characterized as being directed against the nuclear envelope. Further studies detailed in this paper describe the fate of the nuclear envelope during mitosis. Indirect immunofluorescence staining of whole developing Drosophila embryos has been used as a system in which nuclear events can be studied both synchronously and in a longitudinal gradient of mitotic structures. The results show a pattern of breakdown and reconstruction of the nuclear envelope in which the antigen is always present in particulate structures. In addition, the processes of antigen rearrangement are shown to be spatially determined throughout mitosis.
Nuclear crystalloids in sieve elements of boraginaceae: A protein digestion study
Authors: Thorsch Jennifer , Esau Katherine
Published: April 2021

Nuclear crystalloids have been found in sieve elements of several Boraginaceae. Nuclei of differentiating sieve elements of Echium and other genera except Amsinckia contain one or more crystalloids composed of thin rods densely packed in parallel arrangement. After the nuclei disintegrate in the maturing sieve element the crystalloids are released into the cell lumen where they persist intact. In Amsinckia the crystalloid consists of two components: a dense component, similar to the crystalloid in the other genera and a loosely arranged paracrystalline component. The proteinaceous nature of the nuclear crystalloids and their possible similarity to P-protein was investigated by enzyme digestion techniques. Three proteolytic enzymes were employed in this study: protease, pepsin and trypsin. Successful digestion of the dense crystalloid in both Echium and Amsinckia was obtained with each enzyme tested. P-protein plugging the sieve plate pores was also digested. The loose component’m Amsinckia and the aggregated and dispersed P-protein were not affected by the enzyme digestion procedures. These results seemed to indicate that the density or compactness of the proteinaceous inclusions may play a role in the differential response.
Comparison of quick-frozen and chemically fixed sea-urchin eggs: Structural evidence that cortical granule exocytosis is preceded by a local increase in membrane mobility
Authors: Chandler Douglas E.
Published: April 2021

Eggs of the purple sea-urchin, Strongylocentrotuspurpuratus, were fertilized and fixed with 2% glutaraldehyde at various stages during cortical granule exocytosis. Fixation resulted in membrane blebs being formed precisely at the point of incipient granule fusion. These blebs pinched off to form the membranous vesicles frequently seen in exocytic pockets and in the perivitelline space. In contrast, eggs that were fixed with osmium tetroxide or were quick-frozen without chemical fixation, showed no signs of bleb or vesicle formation. Rather, fusion of each granule appeared to begin at a single minute pore, 30 –50 nm in diameter, which then enlarged. We suggest that formation of blebs during glutaraldehyde fixation is an artifact that is caused by a highly localized and transient increase in membrane mobility. Normally, this increased mobility facilitates fusion of granule and plasma membranes, but in the presence of glutaraldehyde it leads to large-scale distortions of these fusing membranes.
Cloned human teratoma cells differentiate into neuron-like cells and other cell types in retinoic acid
Authors: Thompson S. , Stern P. L. , Webb M. , Walsh F. S. , Engstrom W. , Evans E. P. , Shi W. K. , Hopkins B. , Graham C. F.
Published: April 2021

Single cell clones were isolated from the human teratoma line, Tera-2. The cells of three of these clones were studied. The progressively growing cells were shown to be tumorigenic, and they were characterized by the lack of expression of β 2β2-microglobulinand HLA-A,B,C determinants on the cell surface. The majority of the cells expressed Thy-1 antigen and a 90 × 103 molecular weight protein recognized by the monoclonal antibody F10.44.2; between a third and half of the cells expressed the sugar specificities detected by the anti-SSEA-1 monoclonal antibody. In response to 5 × 10−5 M-retinoic acid applied to cells in monolayer culture, the cells differentiated into a population of flat static cells arrested in the GI phase of the cell cycle. A substantial proportion of these differentiated cells expressed β 2-microglobulin and 43 × 103 molecular weight HLA-A,B,C polypeptides, Thy-1, SSEA-1 sugar determinants, and the 90 × 103 protein recognized by F10.44.2. The apparent molecular weight of fibronectin secreted by the cells decreased by about 5 × 103Mr to 235 × 103Mr after differentiation. The progressively growing cells lacked reactivity with reagents that mark cells in the nervous system. Following aggregation and retinoic acid treatment, neuron-like cells were formed. These cells reacted with reagents that also react with human neurons in culture: they reacted with tetanus toxin, the anti-neurofilament antibodies BF10 and RT97, the anti-ganglioside, GQlc antibody F12 A2B5, and anti-Thy-1.

The progressively growing cells of these Tera-2 clones are therefore capable of forming at least two types of cell: the flat cells in monolayer cultures and the neuron-like cells. None of the cell populations reacted with the monoclonal antibody against SSEA-3 and these cloned cells are therefore distinct from previous isolates from Tera-2.
Accumulation of smooth cisternal in the tapetal cells of Ulex europaeus l. (papilionoideae)
Authors: Misset M. T. , Gourret J. P.
Published: April 2021

In addition to the events occurring in the tapetai cells of most angiosperm species (cell wall lysis, endoplasmic reticulum development and sporopollenin deposition on the outer face of the cells) the tapetum of Ulex europaeus L. exhibits two characteristics. No orbicules, generally associated with the locular face of the secretory tapetum of angiosperms, are found, but some intracellular macro-orbicules are formed and then released in the anther loculus.The great diversity of reticular structures is emphasized by the presence of accumulations of smooth and flattened ciβternae in the mature cells. Their similarity to those observed in the sieve elements of differentiating phloem suggests a possible functional interpretation in the light of the ‘holocrine’ behaviour of the tapetai cells.
Nuclear pores and interphase chromatin: High-resolution image analysis and freeze etching
Authors: Nicolini C. , Vernazza G. , Chiabrera A. , Maraldi I. N. , Capitani S.
Published: April 2021

Computer-enhanced analysis of electron micrographs of thin-sectioned rat liver nuclei, combined with three-dimensional reconstruction of the same Feulgen-stained nuclei, points to a unique clustering of chromatin DNA fibres near the nuclear border.

Computer-enhanced image analysis has been applied to electron micrographs of the envelopes of the same rat liver nuclei prepared by freeze etching and a few essential geometrical parameters characterizing the pores and their distribution have been determined. During interphase, clusters of nuclear pores, closely paralleling the clustering of membrane-attached chromatin fibres, have been identified on the envelope, the number of these being similar to the number of homologus pairs of metaphase chromosomes. Furthermore, rapid changes induced in chromatin distribution appear to be associated with rapid changes in pore number, but not in the number of pore clusters.
A pathway of plasma membrane biogenesis bypassing the golgi apparatus during cell division in the green algaCylindrocapsa geminella
Authors: Sluiman Hans J.
Published: April 2021

Cell division in Cylindrocapsageminella, in particular the mode of septum membrane biogenesis, has been studied with the transmission electron microscope. Septum formation takes place in a narrow layer of cytoplasm separating post-mitotic nuclei. First, each daughter nucleus develops a wide cytoplasmic pocket (invagination) containing numerous strands of rough endoplasmic reticulum (ER). Next, a proliferation of rough ER is observed in the equatorial zone of cytoplasm, which invariably contains a small number of widely scattered microtubules. The equatorially aligned cisternae of rough ER produce smooth-membraned vesicles, interpreted as smooth ER, which subsequently coalesce to form the membranous transverse septum. Thus, primary septum formation does not follow any of the two previously known basic cytokinetic patterns in green plants (i.e. plasma membrane furrowing and cell-plate formation), but instead represents a novel type of membrane flow, which effectively bypasses the Golgi apparatus. This pathway of membrane flow has remained largely ignored in current concepts of endomembrane structure and function in eukaryotes. However, it appears to be more widespread than has previously been recognized, especially in autospore-producing green algae and in red algae during the formation of tetraspores. It may represent an evolutionary intermediate type of cell division between the supposedly primitive method of plasma membrane furrowing and the more advanced cell-plate system.
Extracellular matrix interactions with the apical surface of vascular endothelial cells
Authors: Kramer Randall h.
Published: April 2021

Cultured aortic endothelial cells, like their in vivo counterparts, form highly organized, confluent monolayers of polarized epithelioid cells that secrete, exclusively at their basal surface, an extracellular matrix to which they then attach. The influence of isolated subendothelial matrix preparations on cell polarity and monolayer organization was studied by presenting fragments of the matrix to confluent bovine aortic endothelial cell cultures. The matrix particles were immediately bound to the apical aspect of the cell monolayer and induced rapid reorganization of the monolayer into cells with a fibroblastoid morphology. To determine if fibronectin, the major glycoprotein of the subendothelial matrix, could be involved in the observed apical cell surface – matrix interactions, latex beads or small discs of Nucleopore filters were coated with the glycoprotein and presented to confluent monolayers. In a fashion similar to that observed with matrix fragments, materials coated with fibronectin caused focal reorganization of the cell layer. After contact with the coated beads, the underlying endothelial cells flowed upward and spread over the entire bead, forming a canopy of confluent cells that draped the particle. Contact of confluent monolayers with the coated filters induced similar behaviour, except that monolayer reorganization into the fibroblastoid phenotype was followed by emigration of the majority of underlying cells through the pores to the upper filter surface, where they formed a new organized cell monolayer with the typical endothelial cell mor-phology. Thus contact of the apical surface of endothelial cells with structures to which they adhere initiates a rapid disruption of the organized cell monolayer, followed immediately by a concerted effort of the local population to re-establish both cell polarity and monolayer contiguity. The expression of this behaviour may be important during tissue remodelling that occurs in neovascularization and during interactions with thromboemboli.
In vivo transcription of ribosomal rna in relation to the mitotic cycle in Physarum polycephalum*
Authors: Hunter G. J. , Turnock G.
Published: April 2021

We have investigated the transcription of ribosomal RNA in plasmodia of Physarum polycephalum by a combination of pulse-labelling with [3H]uridine and RNA; DNA hybridization. The DNA used for the hybridization was a Hindlll restriction fragment (cloned in bacteriophage λ) of Physarum ribosomal DNA that carries a substantial fraction of the rRNA genes, enabling the ribosomal transcripts in the newly synthesized RNA to be measured. We found that ribosomal RNA constituted about 40 % of the pulse-labelled RNA at all times during the synchronous mitotic cycle.
Changes in the shape of mitochondria following osmotic stress to the unicellular green alga Chlamydomonas reinhardii
Authors: Morris G. J. , Coulson G. E. , Leeson E. A.
Published: April 2021

The effects of various stresses on mitochondrial activity and structure within the unicellular green alga Chlamydomonas reinhardii were investigated using the fluorescent probe rhodamine 123. Within control cells, treatment with rhodamine 123 stained an intense fluorescent network, which was considered to be mitochondrial from the similarity in structure to models of mitochondria reconstructed from serial-section electron microscopy, and because this pattern of staining was abolished following the addition of metabolic inhibitors.

Following osmotic shrinkage and rehydration, fragmentation of the mitochondrial network was observed within potentially viable cells. This was reversible within 1 h of resuspension in isotonic medium. Exposure of cells to hypertonic solutions of rapidly permeating compounds did not induce similar structural alterations. These changes in the mitochondria were confirmed by thin-section electron microscopy. In the presence of higher osmolalities of non-permeating compounds, which induce a greater loss of viability, rhodamine 123 stained cells uniformly. Following the osmotic stresses induced by slow rates of freezing and subsequent thawing no fragmentation in mitochondrial staining was observed. These findings demonstrate that shrinkage and rehydration may induce alterations to the structure and function of organelles and may be factors in determining cellular viability following osmotic stress.
Defective adhesion to extracellular matrix leads to altered social behaviour in cultured fibroblasts
Authors: Aplin John D. , Foden Lynn J.
Published: April 2021

We describe the properties of variant mouse fibroblasts selected for poor adhesion to growth substratum containing subcellular matrix accumulated by adherent cells at confluence. The variant cells adhere to virgin plastic and grow normally to confluence in the presence of serum. After subculture and reseeding onto the same surface the cells initially adhere, but after a further 2 days of growth they retract into aggregates and detach. If the aggregates are dispersed and cells reseeded onto the same surface they remain rounded. However, if the same cells are added back to virgin plastic they adhere and grow normally. The retraction can be abolished by treating the subcellular matrix-coated plastic with papain. This behaviour therefore reflects the ability of the cells to modify the composition of the underlying substratum during growth. The variant cells also exhibit retraction 2 days after seeding on a surface previously containing wild-type cells at confluence, while wild-type cells do not retract on subcellular matrix deposited by variants. This shows that the variant behaviour arises not from a deficiency in the subcellular matrix, but from an alteration in the adhesive capacity of the cells. The results are interpreted in terms of three putative adhesion mechanisms: cell–cell adhesion in confluent monolayers and aggregates; ‘early’ type cell–substratum adhesion, which occurs during culture on virgin plastic; and ‘late’ type cell–substratum adhesion occurring on surfaces containing accumulated subcellular matrix. The variant phenotype is characterized by a deficiency in the last of these. It is also associated with an increased ability to grow in suspension culture at high dilution.
Measurements of intracellular ph and its relevance to cell differentiation in Dictyostelium discoideum
Authors: Inouye Kei
Published: April 2021

A method was developed in this study to measure the intracellular pH (pHi) of Dictyostelium discoideum cells with a pH-sensitive fluorescence dye, carboxyfluorescein dibutyrate, and the pHi values of cells on the stalk and spore pathways were compared. The pHi of prestalk cells was lower than that of prespore cells by approximately 0·3 pH unit. In monolayer cultures of sporogenous mutants, which can differentiate into stalk cells and spores without cell contact, the pHi of the amoebae depended on the medium: media in which the majority of cells eventually become stalk cells reduced the pHi while conditions favouring spore formation increased the pHi. Addition of weak acids lowered the pHi.

These results are in good accordance with the model presented by Gross and coworkers, which proposes that the choice between alternative pathways of cell differentiation is regulated via pHi and that low pHi favours stalk differentiation whereas high pHi favours spore formation.
Binding and uptake of the toxic lectin modeccin by baby hamster kidney (BHK) cells: Isolation of mutants defective in the internalization of modeccin
Authors: Gleeson Paul A. , hughes R. Colin
Published: April 2021

The effects of the toxic lectins, ricin and modeccin, on baby hamster kidney (BHK) cells have been compared. Modeccin is about 20–50 times more toxic to BHK cells than ricin. Binding studies showed that there are 105 to 2 × 105 modeccin binding sites/cell compared with approximately 107 binding sites for ricin. Inhibition studies with galactosides indicate that both N- and O-glycans with terminal galactosyl residues are effective inhibitors of modeccin binding. Surface-bound modeccin, at O°C, was found to be very rapidly endocytosed (75–80% in 10 min) when cells were warmed to 37°C.

Two modeccin-resistant BHK cell lines have been isolated and characterized. These mutants bind normal levels of modeccin and retain sensitivity to the toxic action of ricin. Both cell lines were shown to be defective in their ability to internalize modeccin. Also, these mutants are morphologically different from parental cells as they align and elongate very prominently at low cell densities. These mutants will be very useful in understanding the mechanism of uptake and transport of modeccin.
Microtubules in root hairs
Authors: Traas Jan A. , Braat Paul , Emons Anne Mie C. , Meekes Herman , Derksen Jan
Published: April 2021

The microtubules of root hairs of Raphanus sativus, Lepidium sativum, Equisetum hyemale, Limnoblum sloloniferum, Ceratopteris thalictroides, Allium sativum and Urtica dioica were inves-tigated using immunofluorescence and electron microscopy. Arrays of cortical microtubules were observed in all hairs. The microtubules in the hairs show net axial orientations, but in Allium and Urtica helical microtubule patterns are also present. Numerical parameters of microtubules in Raphanus, Equisetum and Limnobium were determined from dry-cleave preparations. The results are discussed with respect to cell wall deposition and cell morphogenesis.
Regulation of cell growth by vitreous humour
Authors: Lutty Gerard A. , Mello Robert J. , Chandler Carol , Fait Carolyn , Bennett Alonzo , Patz Arnall
Published: April 2021

Extracts of normal vitreous have been found to inhibit angiogenesis in two animal models: tumour-induced neovascularization in the rabbit corneal micropocket and retinal extract-induced angiogenesis in the chick chorioallantoic membrane assay. Using in vitro assays, we have found recently that an extract of bovine vitreous, free of hyaluronic acid, inhibits proliferation of cells in the aortic wall, i.e. endothelium and smooth muscle cells, as well as capillary and corneal en-dothelium. The inhibition is dose-dependent, as determined by either cell count or [3H]thymidine incorporation, and not due to cytotoxicity, as demonstrated with a double-label thymidine assay. The inhibitor is trypsin-sensitive and heat-stable (95°C for 10 min).

Conversely, proliferation of pericytes, lens epithelium and fibroblasts (dermal and corneal) was stimulated by the vitreous extract. This mitogenic activity was heat-labile. Growth of pigment epithelium and several tumour cell lines was unaffected.

The data demonstrate that normal vitreous contains a heat-stable growth inhibitor specific for endothelium and smooth muscle cells, and a non-specific heat-labile mitogen. The paradoxical effect of this antiangiogenic factor on arterial and capillary contractile cells, smooth muscle and pericytes, suggests a basic difference in the regulation of the two vasculatures. The results suggest that a substance in normal vitreous may be important in controlling neovascularization that results from diabetic and other retinopathies, and could be useful for inhibiting tumour-induced angiogenesis.
Nucleolar activation and vacuolation in embryo radicle cells during early germination
Authors: Deltour R. , barsy T. De
Published: April 2021

The activation of the nucleolus of primary root cells of Sinapis alba embryos during the first 72 h of germination was monitored by autoradiographic, ultrastructural and microstereological methods. Autoradiographs showed that within 48 h, the nucleolus progressively resumed the capacity to synthesize pre-rRNA molecules at a high rate. In quiescent embryos the nucleolus was small, compact and composed of mixed granular and fibrillar components. Within the first 6 h of germination a strong nucleolar vacuolation occurred, accompanied by a decrease in the volume of the nucleolus and a concomitant high loss of its ribonucleoproteins (RNPs). From 6 to 24 h, nucleolar vacuolation decreased to reach a stable level. During this last period the volume of the nucleolus increased by the accumulation of the fibrillar component resulting from a slow pre-rRNA processing. At 24 h the nucleolus presented a predominantly fibrillar texture. After 24 h, nucleolus growth continued but was due to the accumulation of the granular component, indicating that pre-rRNA processing occurred at a higher rate than during the first day of germination. F rom 48 h the nucleolus was composed of well-delineated granular and fibrillar areas. Dense nucleolus-associated chromatin as well as fibrillar centres were always observed during the whole period of observation. In addition, previous studies on the nucleolus of radicle cells of Zea mays embryo during early germination were completed by studying changes in the nucleolar volume and in the density of pre-ribosomal subunits of the granular component. On the basis of the data obtained with both species we suggest that a possible function for the nucleolar vacuoles is the increase in the nucleolus–nucleoplasm exchange interface in response to a rapid increase in the output of nucleolar RNPs. The nucleolar growth pattern during early germination is also discussed.
Gap-junctional communication in a communication-defective and in a communication-competent teratocarcinoma cell line
Authors: Lo Cecilia W. , Fang Daniel , Hooper Martin l.
Published: April 2021

We examined the gap-junctional communication properties of a communication-defective cell line R5/3 and its communication-competent revenant H2T12. For these studies, we carried out microelectrode impalements to monitor ionic coupling and dye coupling. Our dye-injection experiments revealed that the H2T12 cells are much more efficient in dye coupling than the R5/3 cells. This latter observation is in agreement with the previous finding that the H2T12 cells are much better metabolically coupled than the R5/3 cells. With ionic coupling measurements, however, both cell lines exhibited similar levels of cell–cell coupling. The R5/3 cells demonstrated an ionic coupling coefficient of 0·19 ±0·011 (S.E.M.) and H2T12 a coupling coefficient of 0·25 ± 0·009 (S.E.M.). These results in conjunction with observations from other studies indicate that the different experimental approaches for monitoring gap-junctional communication may have different levels of sensitivity for detecting as opposed to measuring the level of cell—cell coupling.
Isolation of pure pellicles containing intact basal bodies of Tetrahymena pyriformis
Authors: Tiedtke Arno
Published: April 2021

A new procedure for mass isolation of pure pellicles containing intact basal bodies of Tetrahymena pyriformis is reported.

The success of the procedure depends on the elimination of the sticky mucocyst contents before fractionation of the cells, which is induced by Alcian Blue 8GS. Under appropriate ionic conditions > 95% of the cells are able to form a capsule by simultaneous extrusion of all mature mucocysts. About 50% of these cells are able to escape from their capsules, which are now devoid of mature mucocysts. These cells are separated from the empty capsules and encapsulated cells by passage through layers of gauze of 10 μm pore size. The fractionation of mucocyst-free cells in homogenization buffer yields pure pellicles, which are retained when the homogenate is sieved through steel sieves of 5 μm pore size.

Electron-microscopic controls show that the isolated pellicles are not contaminated with subcellular particles. Cells homogenized in the presence of low concentrations of Triton X-100 yield pellicles consisting of the known cell-surface-associated cytoskeletal elements, together with basal bodies. The cilia are detached just above the kinetosomal plate. The basal bodies of isolated pellicles are obviously undamaged, since all the known structures of native basal bodies are preserved. Even the granular matrix, a labile structure in the lumen of the basal body that probably contains RNA, is preserved.
Clonal separation of mature melanocytes from premelanocytes in a diploid human cell strain: Spontaneous and induced pigmentation of premelanocytes
Authors: Bennett Dorothy C. , Bridges Katherine , Mckay Ian A.
Published: April 2021

Strains of pigmented melanocytes can be derived reproducibly from normal human skin. Published procedures have been modified here to yield a strain, ‘Nohm-l’, comprising many unpigmented cells as well as cells with various degrees of pigmentation observable by light microscopy. The unpigmented cells contain early melanosomes (pigment organelles) and the specific enzyme tyrosinase. They are on average smaller and less dendritic than the pigmented cells. Nohm-l cells show normal chromosomal banding patterns and normal proliferative behaviour, including senescence. They form no tumours in immunodeficient (nude) mice.

Nohm-l cells have been cloned and yield two distinct types of colony, depending on the progenitor cell. Well-pigmented melanocytes engender pure colonies of pigmented cells, but cells with little or no pigment can produce both unpigmented and pigmented progeny. Thus there is a separate cell type, or premelanocyte, which can differentiate spontaneously and stably in culture; this cell type includes both unpigmented and faintly pigmented cells. Usefully, most premelanocytes are viable after frozen storage, unlike well-pigmented melanocytes. Some components of the culture medium affect the proportion of pigmented cells in Nohm-l cultures, and hence probably the ratio of mature melanocytes to premelanocytes. Rapid pigmentation can be induced artificially and simply, by using a medium with increased extracellular pH and tyrosine concentration.
Successive asexual life cycles of starved amoebae in the cellular slime mould, Dictyostelium mucoroides var.Stoloniferum
Authors: Bonner J. T. , Joyner B. D. , Moore A. A. , Suthers H. B. , Swanson J. A.
Published: April 2021

Dense masses of spores of Dictyostelium mucoroides var. stoloniferum have the ability to germinate and aggregate rapidly in the absence of food. This is made possible by the presence of a dominant, self-produced spore germination activator. The germination—aggregation cycle can be repeated in as many as six successive generations. In each generation the spore size is reduced so that ultimately they are only a fraction of the size of those produced by the parental, bacteria-fed amoebae.
Regulation of cell shape in Euglena gracilis. IV. localization of actin, myosin and calmodulin
Authors: Lonergan Thomas A.
Published: April 2021

The immunofluorescence patterns for actin, myosin, calmodulin and tubulin were observed in Euglena gracilis Klebs strain Z during the biological clock-controlled shape changes observed with division-synchronized cells, and during two shock responses that induce cell rounding. The fluorescence patterns for actin, myosin, calmodulin and tubulin show a high degree of coincidence and are visualized as lines running parallel to, and having the same spacing as, the pellicle strips beneath the plasma membrane. The fluorescence patterns remain intact during the daily shape changes, implying that the shape changes do not result from cycles of polymerization and depolymerization of the microtubules and microfilaments. Resuspension of cells in Ca2+-free medium induces cell rounding of many of the cells. The actin and calmodulin patterns are partially disrupted by the Ca2+-free resuspension, while the myosin pattern is almost totally disrupted. Microtubules are unaffected by this treatment. Prior exposure of cells to the calmodulin antagonist trifluoperazine or to the microfilament-stabilizing peptide phalloidin stabilize the actin, myosin and calmodulin patterns against disruption by the Ca -free resuspension and other shock responses. The possibility of an actomyosin contractile system controlled by calmodulin is discussed.