bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tubulin Post-Translational Modifications And The Construction Of Microtubular Organelles In Trypanosoma Brucei
Authors: Sasse Rosemary , Gull Keith 
Published: April 2021

ABSTRACT
We have used specific monoclonal antibodies to facilitate a study of acetylated and tyrosinated α-tubulin in the microtubule (MT) arrays in the Trypanosoma brucei cell. Acetylated α-tubulin is not solely located in the stable microtubular arrays but is present even in the ephemeral microtubules of the mitotic spindle. Moreover, there is a uniform distribution of this isoform in all arrays. Studies of flagella complexes show that acetylation is concomitant with assembly of MTs. There is no subsequent major modulation in the content of acetylated α-tubulin in MTs. Conversely, polymerizing flagellar MTs have a high tyrosinated α-tubulin content, which is subsequently reduced to a basal level at a discrete point in the cell cycle. The MTs of the intranu-clear mitotic spindle appear never to contain tyrosinated α-tubulin, suggesting that they are actually constructed of detyrosinated α-tubulin or that detyrosination is extremely rapid at this time in the cell cycle. T. brucei therefore, represents a cell type with extremely active mechanisms for the post-translational modification of α-tubulin. Our analyses of the timing of acquisition and modulation in relation to MT construction in T. brucei, suggest that acetylation and detyrosination of α-tubulin are two independently regulated post-translational modifications, that are not uniquely associated with particular subsets of MTs of defined lability, position or function. Post-assembly detyrosination of α-tubulin may provide a mechanism whereby the cell could discriminate between new and old MTs, during construction of the cytoskeleton through the cell cycle. However, we also suggest that continuation of detyrosination, allows the cell, at cell division, to partition into daughter cells two equivalent sets of cytoskeletal MTs.
The Role Of Hmg Coa Reductase And Dolichol Synthesis In The Control Of 3t6 Cell Proliferation: Effects Of Cell Crowding, Serum Depletion And Addition Of Epidermal Growth Factor
Authors: Larsson Olle 
Published: April 2021

ABSTRACT
The proliferation of 3T6 cells was substantially decreased when the monolayer cultures were allowed to reach confluency. This growth inhibition (so-called density-dependent inhibition) was of the same magnitude as that following serum depletion in non-confluent cultures. Each type of growth inhibition was correlated to a depression of the activity of 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG CoA) reductase, an enzyme that regulates the biosynthesis of cholesterol and isoprenoid derivatives (e.g. dolichol) by catalysing the reduction of HMG CoA (which is derived from acetyl-CoA) into mevalonate. However, the depression of enzyme activity was more substantial in cells exposed to cell crowding than that in serum-depleted cells (87 and 48 %, respectively). On the other hand, there was a 60–65 % inhibition of the incorporation of mevalonate into dolichol due to serum deprivation, while it remained at normal level in confluent cultures, which implies that the inhibitory effects on dolichol synthesis due to these two experimental conditions were approximately equipotent. Addition of epidermal growth factor (EGF) to the cell cultures, whose proliferation was inhibited due to serum depletion, restored DNA synthesis completely, and these effects were related to anormalization of the activity of HMG CoA reductase and of the incorporation of mevalonate into dolichol. In contrast, in confluent cells addition of EGF only caused a slight increase in DNA synthesis and activity of HMG CoA reductase, and there was no significant increase in the incorporation of mevalonate into dolichol either. Supplementation with mevalonate as well as dolichol to confluent cell cultures induced a significant increase in DNA synthesis, while only dolichol was capable of increasing DNA synthesis in serum-depleted cells.
Taken together, these results propose: (1) the existence of two regulatory sites in the biosynthesis of dolichol, the first one of which is constituted by HMG CoA reductase, and the second one by an unknown enzyme in the distal part of the pathway; (2) that the activity of HMG CoA reductase is stimulated by serum and growth factors, and is inhibited by cell crowding; (3) that the second step is independent of the culture density, but is stimulated by serum and growth factors; (4) that these two regulatory sites are involved in the control of cell proliferation in 3T6 cells.
Effects Of Phorbol Esters On Shape And Locomotion Of Human Blood Lymphocytes
Authors: Wilkinson P. C. , Lackie J. M. , Haston Wendy S. , Islam Laila N. 
Published: April 2021

ABSTRACT
The effects of phorbol esters on shape change and locomotion of human blood lymphocytes were studied both immediately after separating the cells from blood and after overnight culture. Phorbol myristate acetate (PMA), phorbol dibu-tyrate (PDB) and related esters produced complex shape changes in lymphocytes at both times. These shapes were analysed quantitatively using objective measurements derived from the moments of cell shapes. Immediately after removal from blood, many lymphocytes (20–60% of the total) protruded and retracted veils or spikes at more than one point on the cell surface. The morphology of these cells was not typical of locomotor cells. Usually, formation of a veil was not followed by a contraction wave moving down the cell, though some cells did show contraction waves, and some moved into collagen gels or filters. After overnight culture, a high proportion (70–80 %) of cells had changed shape in PMA and PDB. Although the shapes were still atypical, they resembled classical locomotor morphology moreclosely; veils formed at one point on the cell surface tended to persist, and contraction waves and constriction rings were seen in many cells. These cells moved in large numbers into collagen gels or filters. Comparison of the paths traversed by PMA-treated lymphocytes in collagen gels suggested that cells cultured in PMA for 24 h pursued more persistent paths than those in short-term culture, but the difference was not marked. We suggest that phorbol esters induce immediate shape change without inducing the complete sequence of motor events necessary for efficient locomotion, whereas after prolonged culture in phorbol esters, locomotion is more efficient, possibly because phorbol esters, like other growth activators, stimulate events during the G| phase of growth that are necessary for full expression of locomotor capacity.
Contact-mediated triggering of lamella formation by Dictyostelium amoebae on solid surfaces
Authors: Owens Norman F. , Gingell David , Bailey Juliet 
Published: April 2021

ABSTRACT
Amoebae of the slime mould Dictyostelium discoid-eum form broad ultrathin cytoplasmic lamellae by a centripetal contractile process soon after they have spread on certain solid surfaces. We have investigated the surface requirements for initial triggering of this contact-mediated signalling system. The lamellar response is not normally evoked by glass, but is seen on glass covalently derivatized with paraffinic chains, as well as on glass covalently derivatized with amine groups and on glass bearing adsorbed polylysine. We have recorded the frequency of the lamellar response on these surfaces as a function of ionic strength and pH, and have measured the electrostatic potentials of the surfaces by the streaming potential method. Using these data we have concluded that the general trigger for the lamellar response is not a ‘simplE′ physical or chemical property of the substrata : it is not dependent on specific chemical groups, degree of hydrophobicity, electrostatic potential, or charge density, taken as isolated factors. It seems likely that triggering is dependent on the overall energetics of cell-substratum interaction.
Intermicrotubular actin filaments in the transalar cytoskeletal arrays of Drosophila
Authors: Mogensen M. M. , Tucker J. B. 
Published: April 2021

ABSTRACT
Rabbit muscle myosin subfragment SI decorates 6 nm diameter filaments in Drosophila wing epidermal cells in the arrowhead fashion characteristic of the binding of subfragment SI to actin filaments. The filaments in question are concentrated between microtubules that are mostly composed of 15 protofilaments and form cell surface-associated transcellular bundles. There are indications that the majority of the actin filaments have the same polarity and that, like the microtubules, they may elongate from sites at the apical surfaces of the cells.
The bundles of F actin and microtubules occur in dorsal and ventral epidermal cell layers of a wing blade. They are joined in dorso-ventral pairs by attachment desmosomes. These transalar cytoskeletal arrays may provide an example of a situation where actin filaments operate as stiffeners rather than active generators of force in conjunction with myosin. The arrays probably function as non-contractile pillars to maintain basal cell extensions and keep haemocoelic spaces open in the highly folded and expanding wing blades of late pupae.
Changes in the number and volume of fibrillar centres with the inactivation of nucleoli at erythropoiesis
Authors: Zatsepina Olga V. , Chelidze Pavel V. , Chentsov Yuri S. 
Published: April 2021

ABSTRACT
The number and volume of fibrillar centres, the structural components of interphase cell nucleoli on the surface of which rRNA is synthesized, have been studied in differentiating erythroblasts of mouse embryo liver. Complete series of ultrathin sections of erythroblast nuclei have been used at the main stages of differentiation: proerythroblast, basophilic erythroblast, polychromatophilic erythroblast and normoblast. It has been shown that in the active nucleoli of proerythroblasts the number of fibrillar centres is correlated with cell ploidy and exceeds by several-fold the number of nucleolusorganizing regions of chromosomes. The total volumes of fibrillar centres in 2C (0 ·369 μm3) and 4C (0 ·749 μm3) proerythroblasts are proportional to number of nucleolus- organizing regions. With the maturation of erythroblasts the total number of fibrillar centres declines and in normoblasts it is 3-to 10-fold less than that of the nucleolus- organizing regions. The total volume of fibrillar centres in normoblasts (0-102 μm3) is threefold smaller than that in proerythroblasts (0 ·369 μm3), even though the mean volumes of individual fibrillar centres are significantly higher (0·0042 μm3 in proerythroblasts and 0 ·039 in normoblasts). The optical density of fibrillar centres in normoblasts can be higher compared with that of proerythroblasts. It has been suggested that the inactivation of nucleoli at erythropoiesis is accompanied by the fusion of individual fibrillar centres and, possibly, by the compaction of their material.
Ultraviolet microbeam irradiation of chromosomal spindle fibres shears microtubules and permits study of the new free ends in vivo
Authors: Wilson Paula J. , Forer Arthur 
Published: April 2021

ABSTRACT
Irradiation of birefringent chromosomal spindle fibres in cranefly spermatocytes in metaphase I or anaphase I produces an area of reduced birefringence (ARB) on the fibre. This ARB moves poleward and is lost at the pole. Ultrastructural and immunofluorescence analysis of ARBs obtained by irradiation with monochromatic ultraviolet light of wavelength 260 nm shows that the microtubules in the irradiated area are depolymerized, though the rest of the spindle appears unaffected. The area of microtubule depolymerization moves poleward with the ARB, and once the ARB reaches the pole the irradiated half-spindle appears normal. The motion of the ARB, therefore, appears to be due to the behaviour of the sheared microtubules in the chromosomal spindle fibre. The relative stability of the sheared microtubules shows that chromosomal fibre microtubules are not dynamically unstable, as are microtubules under certain conditions in vitro. However, ARB motion may be due to a moderated version of dynamic instability. Possible models for ARB motion are discussed.
Functionally polarized layers formed by epidermal cells on a permeable transparent collagen film
Authors: Yoshizato Katsutoshi , Nishikawa Akio , Taira Toshio 
Published: April 2021

ABSTRACT
Rat epidermal cells were cultured on a transparent collagen film, which was permeable to low Mr substances. Then the cells were bathed in both media (apical and basal), which were separated by the collagen membrane. The cells formed a multilayered epidermal sheet with well-developed structures of desmosomes. This sheet on a permeable support was found to be an effective permeability barrier for glucose and amino acids. The epidermal layer showed functional polarity for the uptake and excretion of nutrients, metabolites and newly synthesized proteins: glucose and amino acids were taken up exclusively from the basal medium and lactate was secreted selectively into the same medium, whereas ammonia was secreted into the apical medium. The apical media became more acidic than the basal ones, presumably due to the preferential distribution of H+ transport systems on the apical side of the epidermal layer. The epidermal cells that expressed functional polarities in vitro as described above were able to proliferate and differentiate, and remained healthy for as long as at least 40 days even using a conventional culture medium with foetal calf serum, but without any special growth factors and feeder cells.
Dissociation and re-assembly of the endoplasmic reticulum in live cells
Authors: Koch G. L. E. , Booth C. , Wooding F. B. P. 
Published: April 2021

ABSTRACT
The endoplasmic reticulum (ER) of a typical interphase 3T3 fibroblast consists of a compact perinuclear arrangement of cistemae and lamellae which can be observed by immunofluorescence with antiendoplasmin. During mitosis the reticulum dissociates into small fragments from which it appears to re-assemble in the daughter cells. When interphase 3T3 cells are exposed to calcium ionophores, but not other ionophores, there is a similar dissociation of the ER into small uniform fragments, which are dispersed throughout the cytoplasm. Electron microscopy shows that the fragments consist of small vesicular structures and that essentially all of the rough ER except the nuclear envelope is dissociated. The dissociation of the ER by calcium ionophore is a relatively specific process since other organelles and supramolecular assemblies remain unaffected. When cells with dis sociated ER are returned to normal medium, there is a rapid reassembly of the fragments into the continuous reticulum. In a proportion of the cells it is possible to observe linear arrays of the fragments, which probably represent intermediates in the re-assembly process. These observations demonstrate that the ER in interphase 3T3 cells can be dissociated into, and re-assembled from, small fragments. Re-assembly of the ER from the fragments is dependent on the presence of millimolar levels of calcium in the external medium. In the presence of calcium, re-assembly is inhibited by the calcium channel blocker, verapamil. Thus calcium ions appear to play an important role in ER structure and assembly.
Microtubule and microfilament distribution during the secretory activity of an insect gland
Authors: Dallai R. , Marchini D. , Callaini G. 
Published: April 2021

ABSTRACT
The distribution of microfilaments and microtubules was studied in the secretory cells of the accessory glands of the medfly (Ceratitis capitata) female reproductive system. Actin, demonstrated by rhodamine-phalloidin and heavy meromyosin decoration, is present around the extracellular storage cavity at the level of microvilli and the subapical cytoplasm of the cell. The size of such a cavity varies in relation to the secretory cycle of the cells and also the distribution of microfilaments. In the early stage of activity microtubules detected using an antibody against a-tubulin are distributed at the periphery of the cell and around the extracellular cavity. During secretion they occupy the whole of the cytoplasm and disappear at the end of the secretory cycle. The mechanism of the discharge of the secretion from the extracellular cavity is discussed.
Localized osmotic swelling and cell fusion in erythrocytes: Possible implications for exocytosis
Authors: Ahkong Quet F. , Lucy Jack A. 
Published: April 2021

ABSTRACT
Factors that govern the experimentally induced fusion of erythrocytes with one another may generally be relevant to whether or not osmotic forces drive membrane fusion in exocytosis because, under appropriate conditions, osmotic swelling can drive the fusion of erythrocytes. It is now reported that these cells fuse when they are subjected to osmotic swelling caused by exposure to small permeant molecules. The behaviour of erythrocytes in fusion induced by treatment with a concentrated solution of high molecular weight poly(ethylene glycol) (PEG) is also of specific interest in relation to exocytosis because the haemoglobin of erythrocytes that are dehydrated by concentrated solutions of the polymer may be regarded as a model for the tightly packed, dehydrated contents of the granules in secretory cells. We have observed that, under certain conditions of rehydration, the swelling of aqueous microdroplets between the dehydrated haemoglobin and the plasma membrane is closely associated with the fusion of partially rehydrated but still shrunken, PEG-treated erythrocytes. It is therefore apparent that osmotic forces, acting locally at the sites of aqueous microdroplets, can drive the fusion of membranes that encapsulate a dehydrated, concentrated protein, even though gross osmotic swelling at the level of the light microscope is absent. This finding is consistent with the possibility that osmotic swelling may play a role in exocytotic membrane fusion if it is restricted to a small zone immediately under the granule membrane.
Gradient and temporal signal perception in chemotaxis
Authors: Vicker Michael G. 
Published: April 2021

ABSTRACT
In a recent Commentary, Lauffenburger et al. (1988) argued that chemotaxis depends on the perception of a chemoattractant (CA) concentration (CCA) gradient across the length of a polymorphonuclear (PMN) neutrophil leucocyte. Haston & Wilkinson (1987) agreed with this view, pointing out that PMN polarize after contact with a CA. In the light of the important role expected of concentration gradients in taxis and in morphogenesis, it seems appropriate to continue this dialogue; first, by replying to the remarks of Lauffenburger et al. (1988) about our failure to detect taxis in spatial gradients and, second, by introducing into it some rudimentary facts about cell behaviour and signals, which have apparently attracted little attention in discussions of gradient perception.
Ki-67 detects a nuclear matrix-associated proliferation-related antigen : I. Intracellular localization during interphase
Authors: Verheijen R. , Kuijpers H. J. H. , Schlingemann R. O. , Boehmer A. L. M. , Driel R. Van , Brakenhoff G. J. , Ramaekers F. C. S. 
Published: April 2021

ABSTRACT
Ki-67 is a commercially available mouse monoclonal antibody, which reacts with a nuclear antigen in proliferating cells. The antibody can be used to determine the growth fraction of human tumours in situ and has been shown to be of prognostic importance.
In this study it is shown that in interphase cells Ki-67 reacts with an antigen, mainly present in the nucleoli. Confocal scanning laser microscopy and immunoelectron microscopy on human MR65 monolayer cells revealed that this nucleolar antigen is predominantly localized in the nucleolar cortex and in the dense fibrillar components.
The Ki-67 antigen appeared to be preserved in nuclear matrix preparations obtained after in situ fractionation of MR65 cells. Despite many efforts, we could not identify the antigen in immunoblotting or immunoprecipitation assays.
Testing of cell cultures of different species by means of indirect immunofluorescence revealed that the antibody reacted with human cells and with the Rhesus monkey kidney-derived cell line LLC-MK2.
Oligodendrocytes repel axons and cause axonal growth cone collapse
Authors: Fawcett J. W. , Rokos J. , Bakst I. 
Published: April 2021

ABSTRACT
We have examined the interactions between axons regenerating from dorsal root ganglia (DRGs) derived from newborn rats and oligodendrocytes cultured by three different techniques. Cultures examined after 2 days have a profuse outgrowth of axons from the DRGs, forming a dense mat on the culture surface. However, the axons avoid growing on oligodendrocytes; axons are seen all around these cells, but do not grow over them. We have also performed time-lapse video studies of the interactions between axonal growth cones and oligodendrocytes. Axons grow normally until their growth cone comes into direct contact with an oligodendrocyte, following which the growth cone remains motile for 30-60 min, but without making any progress over the cell. The growth cone then suddenly collapses, and the axon retracts, leaving a thin strand in contact with the cell. After this a new growth cone is usually elaborated and the process repeated. Oligodendrocytes are therefore inhibitory to axonal growth, and this may partially explain the failure of axons to regenerate in the mammalian central nervous system.
Glycophorin expression in murine erythroleukaemia cells
Authors: Ulmer Jeffrey B. , Dolci Elizabeth D. , Palade George E. 
Published: April 2021

ABSTRACT
We have identified mature and putative precursor forms of glycophorins expressed in a virus-transformed murine erythroleukaemia (MEL) cell line and compared them with their normal erythroblast counterparts. The following differences were found: (1) the two major MEL cell glycophorins (apparent Mr values 29–30 and 43(×103)) have greater mobility on polyacrylamide gels than their normal gp-3 and gp-2 counterparts, due at least in part to differences in their oligosaccharide sidechains; (2) MEL cell gp-3 consists of two discrete proteins; and (3) there are more potential glycophorin precursors in MEL cells than in normal mouse erythroblasts. Four proteins, with apparent Mr values of 21, 23, 26 and 27(×103), have tentatively been identified as glycophorin precursors, based on the following findings: (1) they are immunologically related to the glycophorins; and (2) their synthesis was induced by dimethyl sulphoxide coincidentally with that of gp-3 and gp-2. They do not appear to be glycoproteins, as evidenced by their lack of incorporation of [3H]galactose, [3H]glucosamine or [3H]mannose. In contrast, gp-3 and gp-2 incorporated [3H]galactose and [3H]glucosamine but not [3H]mannose. Partial characterization of the glycan moieties of MEL cell glycophorins indicates that they consist mostly of tri- and tetrasaccharides, with no indication of any N-linked chains. Hence, the glycans of MEL cell glycophorins are mostly (if not all) O-linked. Furthermore, treatment with N-glycanase did not change their electrophoretic mobility on polyacrylamide gels. MEL cell glycophorins were also shown to be modified by phosphoryl and fatty acyl groups.