bluuebunny/crossref_metadata_2025 huggingface dataset:
Macromolecular physiology of plastids: VI. changes in membrane structure associated with shifts in the absorption maxima of the chlorophyllous pigments
Authors: Henningsen K. W.
Published: April 2021

Dispersal of the prolamellar body tubules into primary lamellar layers has been studied in leaves of dark-grown bean seedlings (Phaseolus vulgaris L.). Following complete photoconversion of protochlorophyllide the dispersal process can proceed in either light or darkness. The dispersal is reversibly inhibited in leaves at 0°C. Above o °C the time course for the dispersal is temperature dependent. The spectral shift of the in vivo absorption maximum of newly formed chlorophyllide from 684 to 672 nm is also inhibited at o °C and the time course is temperature-dependent in a manner similar to that of the membrane dispersal. A causal connexion between the 2 phenomena can thus be suggested. Derangement of the tubular membranes produced by heat treatment or by freezing and thawing in darkness causes the appearance of photoactive protochlorophyllide absorbing at 635 nm. Photoconversion of protochlorophyllide 635 nm results in chlorophyllide absorbing at 672 nm and further dissociation of the membranes. Denaturation of the prolamellar body membranes by heating or by slow thawing results in the formation of photoinactive protochlorophyllide absorbing at 632 nm.
An inverse relation between cell density and the number of microvilli in cultures of BHK21 hamster fibroblasts
Authors: O’neill C. H. , Follett E. A. C.
Published: April 2021

The relationship between culture density and the number of microvilli on the cell surface has been studied using BHK21 cells in established monolayer culture. The number of microvilli can be estimated quantitatively by a scanning electron-microscope technique. It can be increased by applying specific antiserum, but whether antibody is applied or not a significant number of cells regularly bear microvilli. This number is characteristic of the culture density. In sparse cultures above a certain minimum density it is relatively high, and in confluent cultures it is much reduced. These results indicate that microvilli may be inhibited by cell contact.
Spheroplasts of Lactobacillus casei and the cellular distribution of bactoprenol
Authors: Barker D. C. , Thorne Kareen J. I.
Published: April 2021

Spheroplasts of L. casei were prepared by the simultaneous action of trypsin and lysozymc in the presence of EDTA and 20 % sucrose. Electron microscopy of thin sections of cells during the formation of spheroplasts showed that the spheroplasts emerged from the cell wall through a weak point at one end of the cell. Electron microscopy of negatively stained preparations showed that the structure of spheroplast membranes obtained by disruption in hypotonic medium varied according to the nature of the hypotonic medium used. Disruption in water gave small, delicate membrane fragments; disruption in 0·1 M tris-Cl gave large pieces with tubulo-vesicular attachments. The effects of Mg2+ and EDTA were also investigated. Bacto-prenol, the C55 isoprenoid alcohol concerned in cell wall biosynthesis, was located by growing L. casei in [14C] mevalonic acid. Over 90% of the cellular bactoprenol was found in the sphero-plast membrane when this was obtained with 0·1 M tris-Cl, but only 42% when the sphero-plasts were burst in water. It is suggested that the missing bactoprenol may be solubilized with the membranes of mesosomal orgin, and that bactoprenol is therefore a component of both plasma and mesosomal membranes.
Germination of stress-tolerant Eucalyptus pollen
Authors: Heslop-Harrison J. , Heslop-Harrison Y.
Published: April 2021

Earlier reports have indicated that the pollen of Eucalyptus is mechanically robust and unusually resistant to the osmotic stress imposed by immersion in water. We have investigated some of the features of the germination mechanism in the pollen of E. rhodantha with a view to clarifying the role of pollen-wall specializations in determining this resistance. Cultured in vitro, the pollen showed erratic germination, with a scatter of germination times up to 24 h. This was associated with variation between individual grains in the rate of hydration and dispersal of the pectins of the oncus, the thickened outer component of the intine present at each aperture. The oncus is itself differentiated, with a refractive outer layer lying within a sporopollenin operculum and itself overlying the protein-bearing layer of the intine. The outer layer, interpreted as a compacted pectin, undergoes only slow dissolution in aqueous media after the lifting of the operculum, and it is this that apparently protects the grain from the effects of short-term osmotic stress. The rate of dissolution varies between grains, possibly as a consequence of minor differences in developmental rate in the final stages of differentiation in the anther, and this contributes to the wider scatter of germination times. The dehydrated pollen gave one-third of the potential germination after 24 h exposure to 60°C, and a small proportion survived 24 h at 70°C. This degree of heat tolerance must primarily reflect properties of the protoplast of the vegetative cell, not examined in the present study; but the wall specializations may well provide a guard against extreme desiccation, and it is noteworthy that the function of the germination mechanism is not prejudiced by exposure to high temperatures.
Manipulating chromosome structure and metaphase status with ultraviolet light and repair synthesis inhibitors
Authors: Mullinger Ann M. , Johnson Robert T.
Published: April 2021

DNA repair occurs in metaphase-arrested cells in response to ultraviolet irradiation. In the presence of the repair synthesis inhibitors hydroxyurea and 1-β-D-arabinofuranosylcytosine the chromosomes of such cells, as seen in Carnoy-fixed preparations, are decondensed. The extent of decondensation is related to both the u.v. dose and the duration of incubation in the presence of inhibitors. For any particular cell type there is a reasonable correlation between the amount of decondensation and the number of single-strand DNA breaks generated by the repair process under the same inhibitory conditions, though the chromosome changes continue after the number of single-strand breaks has reached a plateau.

The dose response of chromosome decondensation varies between different cell types but is in general correlated with differences in levels of single-strand breaks accumulated under comparable inhibitory conditions. Decondensation can be detected after 0·5 Jm−2 in repair-competent human cells. In human cells defective in excision repair there is much less chromosome decondensation in response to the same u.v. dose and time of repair inhibition. However, a simian virus 40-transformed muntjac cell displays pronounced chromosome decondensation but has limited incision ability.

Both chromosome decondensation and single-strand break accumulation in the presence of inhibitors are reversed when DNA precursors are provided, but reversal after higher u.v. doses and longer periods of incubation leads to recondensed chromosomes that are fragmented. Elution of the DNA from such cells through polycarbonate filters under non-denaturing conditions reveals that double-strand DNA breaks are generated during the period of incubation with inhibitors.

Although the chromosomes of repair-inhibited metaphase cells are decondensed in fixed preparations, their morphology appears normal in intact cells. The cells also retain a capacity to induce prematurely condensed chromosomes (PCC) when fused with interphase cells: compared with control mitotic cells, the speed of induction is sometimes reduced but the final amount of PCC produced is similar.
Three-dimensional behaviour of mitochondria during cell division and germ tube formation in the dimorphic yeast Candida albicans
Authors: Tanaka Kenji , Kanbe Toshio , Kuroiwa Tsuneyoshi
Published: April 2021

This study was done to correlate mitochondrial behaviour with nuclear behaviour and cell division as well as with the germ tube formation in the dimorphic yeast Candida albicans. Threedimensional reconstruction of electron micrographs of serially sectioned cells of the three strains was used to determine the morphological and quantitative relationships between the structures. The results suggested that at the time of entry into the bud a few mitochondria fused into a single giant one, which fragmented during mitosis and resumed a single giant form before cytokinesis, and was then partitioned into two parts. This tendency was also shown during germ tube formation. Quantitative analysis has established that growth of organelles such as the nucleus and mitochondria closely followed total cell growth, the ratio of organelle volume to total cell volume being held relatively constant.
Antigenic heterogeneity of breast cell lines detected by monoclonal antibodies and its relationship with the cell cycle
Authors: Edwards Paul A. W. , Skilton Robert A. , Payne Andrew W. R. , Ormerod Michael G.
Published: April 2021

Established cell lines were stained by immunofluorescence with four monoclonal antibodies to study the phenomenon of antigenic heterogeneity and its possible relation to the cell cycle. Five cell lines thought to be of breast origin, MCF-7, ZR-75–1, T47D, MDA-MB-231 and HBL-100, were stained with three monoclonal antibodies (LICR-LON-M8, LICR-LON-M18, LICR-LON-M24) that each stain a distinct subset of normal breast epithelial cells, i.e. demonstrate antigenic heterogeneity in normal breast epithelial cells. Contrasting monoclonal antibodies LICR-LON-FIB75 and LICR-LON-FIB86 to homogeneously expressed antigens were also used.

For the antibodies demonstrating heterogeneity distinct positive and negative fractions were not seen by flow cytometry; the intensity of fluorescence varied continuously from background to a hundred times stronger than background. Fluorescent DNA staining showed no obvious relation between antigen expression and the cell cycle. The essentially constant proportion of cells of a given antigenic phenotype in the various phases suggests that these antibodies do not distinguish a phenotype associated with a distinct proliferating population of cells.
Cell-cycle-specific induction of quiescence achieved by limited inhibition of protein synthesis: Counteractive effect of addition of purified growth factors
Authors: Larsson Olle , Zetterberg Anders , Engstrom Wilhelm
Published: April 2021

We have previously shown that Swiss 3T3 cells located in the first part of G| (post-mitotic G1 cells younger than 4·0 h or Giprn cells) were arrested after 9–10 h in the cell cycle by a short (1 -8 h) exposure to serum-free medium or by a short (2–4 h) exposure to low doses of the protein synthesis inhibitor cycloheximide (CH). Kinetic data indicate that such Giprn cells rapidly return to G1 during this brief treatment and thereafter require a preparatory period of 8h before continuing to G1

Cells older than 4h, i.e. cells in mid or late G1 are already committed to DNA synthesis (presynthesis or G1 ps cells). These cells as well as S and G2 cells were consequently unaffected by the brief serum starvation or the brief treatment with cycloheximide.

In the present paper we show that the 10-h intermitotic delay that follows a 1–2 h exposure to serum-free medium can be completely counteracted by the presence of any one of the purified growth factors, epidermal growth factor (EGF), insulin or platelet-derived growth factor (PDGF). In contrast, the intermitotic delay following a longer exposure (8 h) to serum-free medium could no longer be counteracted by EGF or insulin. However, PDGF was still active in this respect.

Most interestingly, the 12h gross intermitotic delay induced by a 4h exposure to CH could be efficiently counteracted by EGF, PDGF or insulin. However, this effect on CH-treated cells could be counteracted by the growth factor only in the presence of 10% serum. This indicates the existence of a cooperative effect between PDGF, EGF or insulin and an unidentified serum factor.

The effects on the cell cycle time of brief serum starvation and exposure to CH were compared with the effects on rate of protein synthesis and degradation. Although the effects of serum starvation on protein synthesis and degradation were found to be partially normalized by growth factors, we suggest that growth factors prevent cells from leaving the cell cycle by another mechanism and not merely by affecting the level of overall protein accumulation.
Glycogen metabolism and the nuclear envelope-annulate lamella system in the early chick embryo
Authors: Eyal-Giladi Hefzibah , Feinstein Naomi , Friedlander Michael , Raveh Dina
Published: April 2021

The intracellular sites of glycogen degradation in the mid to late uterine chick embryo were determined by cytochemical localization of glucose-6-phosphatase at the ultrastructural level. Enzyme activity was found between the two membranes of the nuclear envelope, in the annulate lamellae and in specialized glycogen-containing membrane scrolls. Annulate lamellae and glycogen scrolls were most frequent during the stages of intensive glycogen degradation. Annulate lamellae appear to be formed from the nuclear envelope. During the early post-laying stages annulate lamellae disappeared and were replaced by endoplasmic reticulum that appeared initially in scroll-like formations.
Behaviour of neutrophil leucocytes in uniform concentrations of chemotactic factors: Contraction waves, cell polarity and persistence
Authors: Shields James M. , Haston Wendy S.
Published: April 2021

The essential component of any hypothesis of random or directed cell movement is the mechanism of cell polarity. In this paper we describe the polar behaviour of human neutrophil leucocytes in uniform concentrations of chemotactic factors both in suspension and while moving across surfaces. Neutrophils exposed to uniform concentrations of chemotactic factors in suspension around the dissociation constant (Kd) for the receptor rapidly become distinctly bipolar; neutrophils exposed to supraoptimal uniform concentrations (100-foldKd) of chemotactic factors in suspension, although morphologically active, never reached the same degree of polarity as cells in optimal concentrations. These differences in polarity were shown to be the direct result of equatorial contraction waves stimulated on the cell surface by interaction with chemotactic factors. In optimal concentrations of chemotactic factors, contraction waves were initiated from one region of the cell, whereas in supraoptimal concentrations of chemotactic factors contraction waves emanated from all areas of the cell surface. Asymmetry in the distribution of surface receptors for Fc and C3b were observed in neutrophils polarized in uniform concentrations of chemotactic factor. In neutrophils, motile but not well polarized (in 10-6 M-jV-formylmethionyl-leucyl-phenylalanine (fMLP), receptors were uniformly distributed. In neutrophils polarized in concentrations of fMLP near the for the receptor (10-8 M) receptors for C3b and Fc were localized in the anterior region of the moving cell.

The link between contraction waves, cell polarity and receptor redistribution and their initiation by chemotactic peptides is discussed in the context of neutrophil locomotion and response to chemical signals.
Immunofluorescence microscopy of microtubules in intact cell lineages of the moss, Physcomitrella patens: I. normal and cipc-treated tip cells
Authors: Doonan John H. , Cove David J. , Lloyd Clive W.
Published: April 2021

Monoclonal antibodies to yeast tubulin have been used to visualize the distribution of microtubules in the intact filamentous protonemata of the moss Physcomitrella patens. Protonemata were prepared for immunofluorescence by fixation in formaldehyde and cells were made permeable with Driselase. Extensive cell files were preserved by ‘blotting’ the moss onto glutaraldehydederi vatized coverslips. Problems due to fluorescence from chloroplasts were obviated by extraction with dimethyl sulphoxide and the non-ionic detergent, Nonidet NP4O. These improvements allowed us to determine that microtubules were present throughout the cell cycle in the apical dome of caulonemal tip cells, that there was a pronounced association of microtubules with the nucleus, that ‘astral’ microtubules were associated with the mitotic spindle and during anaphase may be involved in reorientation of the spindle before an oblique cytokinesis in caulonemata and that the cytokinetic phragmoplast appeared identical to the structure described for higher plants.

Microtubules appeared to converge at the very tip of apical caulonemal cells and this was studied further by treating cells with CIPC - a drug that is known to produce multiple microtubuleorganizing centres — and which here produces multiple foci for microtubules at the tip.

These observations emphasize the involvement of microtubules in tip growth, alignment of the cell plate and nuclear migration - processes that are fundamental to the morphogenesis of filamentous organisms.
X chromosome inactivation during induced differentiation of a female mouse embryonal carcinoma cell line
Authors: Paterno Gary D. , McBurney Michael W.
Published: April 2021

P10 is a line of embryonal carcinoma cells with a euploid female karyotype. By making use of variant alleles of X-linked genes and of cytogenetic procedures, we have previously shown that the P10 cells have two genetically active X chromosomes. We show here that P10 cells rapidly differentiate into a cell type resembling extraembryonic endoderm when cultured in the presence of retinoic acid. This differentiation is accompanied by X chromosome inactivation as determined by the appearance of a late-replicating X chromosome. Analysis of the X-linked variant alleles indicated that the P10-derived endoderm did not preferentially inactivate paternally derived X chromosomes. This result is in contrast to the situation in normal extraembryonic endoderm, and suggests that the X inactivation process in differentiating P10 cultures resembles that which occurs in normal embryonic rather than extraembryonic tissues.
Thylakoid formation from coiled lamellar bodies during carposporogenesis in Faucheocolax attenuata setch. (rhodophyta, rhodymeniales)
Authors: Delivopoulos Stylianos G. , Kugrens Paul
Published: April 2021

Chloroplast development during carposporogenesis in the parasitic red alga Faucheocolax attenuata Setch. was studied by electron microscopy. Proplastids are usually found in the peripheral cytoplasm of young carpospores and are characterized by the presence of portions of a peripheral thylakoid and coiled lamellar bodies that range in size up to 0·5μm. One type of coiled lamellar body occurs in the peripheral region of the proplastid and is continuous with the peripheral thylakoid, while the other type is found in the central portion of the stroma. These coiled lamellae separate and expand, adding membranes to both thylakoid systems, thereby functioning as thylakoid-forming bodies. As each coiled lamella unravels, it forms an undulated double-membraned structure having the same width as a thylakoid. After substantial expansion, the developing thylakoids begin to straighten and assume a parallel orientation to each other, thus becoming mature thylakoids. Small coiled lamellae often persist in mature carpospore chloroplasts, and are utilized in additional thylakoid formation during carpospore germination.
Microtubules are at the tips of root hairs and form helical patterns corresponding to inner wall fibrils
Authors: Lloyd Clive W. , Wells Brian
Published: April 2021

Root hairs have sometimes provided contradictory evidence for microtubule/microfibril parallelism. This tissue was re-examined using optimized conditions for the fixation, before immunofluorescence, of root hairs. In phosphate buffer, microtubules did not enter the apical tip of radish root hairs and were clearly fragmented. However, in an osmotically adjusted microtubule-stabilizing buffer, microtubules were observed within the apical dome and appeared unfragmented. Microtubules are not, therefore, absent from the region where new cell wall is presumed to be generated during tip growth. A spiralling of microtubules was seen at the apices of onion root hairs.

Using shadow-cast preparations of macerated radish root hairs, it was confirmed that steeply helical microtubules matched the texture of the inner wall. In onion, the 45° microtubular helices are accompanied by similarly wound inner wall fibrils.

Results do not support the view that microtubules are not involved in the oriented deposition of fibrils in root hairs. Instead, they are interpreted in terms of a flexible helical cytoskeleton, which is capable of changing its pitch but is sensitive to fixation conditions.
Use of aqueous two-phase partition to detect cell surface changes during growth of Dictyostelium discoideum
Authors: Sharpe Paul T. , Watts D. J.
Published: April 2021

Changes in the cell surface properties of amoebae of Dictyostelium discoideum during growth in different culture conditions have been studied by aqueous two-phase partitioning on a thin-layer countercurrent distribution apparatus. Changes in cell surface properties were not dependent on the source of nutrients but only on cell density. There was a progressive increase in cell surface hydrophobicity with cell density in both axenic cultures and cultures grown with a bacterial substrate. It is proposed that it is these cell-density-related surface changes that account for the ability of amoebae grown in different conditions to sort out during subsequent development in a manner related to cell fate.
Flow cytometer study of anterior-like cells in Dictyostelium discoideum
Authors: Voet Ludwig , Krefft Marianne , BrÜderlein Martina , Williams Keith L.
Published: April 2021

The Dictyostelium discoideum asexual fruiting body consists of spores, stalk and basal disk cells. Recently, a fourth cell class has been proposed. It has been suggested that these cells originate from anterior-like cells that remain undifferentiated. Anterior-like cells are randomly distributed among prespore cells in the posterior part of the slug. Here monoclonal antibodies that recognize the surface of prespore cells (MUD1), and spores (MUD3) are used in a quantitative flow cytometer assay to demonstrate that this fourth cell class does not exist in the mature fruiting body. However, the tip cells are slow to differentiate, and hence immature fruiting bodies contain a small population of undifferentiated tip cells. We confirm that anterior-like cells represent a large percentage of the nonprespore cell population in the slug. In this report we were unable to distinguish these anterior-like cells from prestalk cells on the basis of size or monoclonal antibody staining.
Tetrazolium salt and electron-microscopic studies of cellular degeneration and necrosis in the interdigital areas of the developing chick limb
Authors: Hammar S. P. , Mottet N. K.
Published: April 2021

Cellular degeneration and necrosis were studied in the interdigital areas of developing hind limb buds of normal chick embryos by means of enzyme-specific tetrazolium salts and electron microscopy.

Using succinic acid as a substrate and the tetrazolium salt, nitro blue tetrazolium, succinate dehydrogenase was specifically demonstrated via a colour reaction in which degenerating and necrotic cells—those with no enzyme activity—did not stain, while those with enzyme activity stained deeply blue-black.The interdigital cells exhibited near absent levels of succinate dehy-drogenase as early as stage 26–27, one to two days prior to when morphological evidence of degeneration and necrosis was present. It was postulated that the mechanism of cell death resulted from decreased activity and/or loss of strategic cellular enzymes such as succinate dehydrogenase, with a subsequent fall in the cellular adenosine triphosphate (ATP) level and a resultant compromise in vital cellular processes, eventually leading to cell death. The evidence indicated that ‘biochemical degeneration’ occurred prior to morphological changes in cells.

The ultrastructural events of degenerating and necrotic cells were also studied. Most ‘dying’ cells observed had already been phagocytosed and were observedin various stages of degenera-tion. The degenerating cells exhibited both nuclear and cytoplasmic changes. There was evidence of active intracellular digestion within the phagocytes. Numerous lysosomes were observed within these cells and some appeared fused with the digestive vacuole’s membranes.The origin of the phagocytes was not determined.

No definite information was obtained concerning the utility of cellular degeneration and necrosis in the interdigital areas of the hind limb buds. It appeared to be at least partially causal in separation of the digits.
The effect of high hydrostatic pressure on the mechanical properties of the surface of the sea-urchin egg
Authors: Wolpert L. , Marsland D. , Hirshfield M.
Published: April 2021

There is evidence that high hydrostatic pressure can solate and weaken gel-like structures within cells. It was thus of interest to determine its effect on the mechanical properties of the cell surface since it has been suggested that there is a gel-like cortex beneath the plasma mem brane which makes a significant contribution to its mechanical properties. The mechanical property measured was the resistance to deformation when the eggs were compressed mechanic ally. The eggs of Arbacia were compressed by placing a small plate made either from a fragment of coverslip or Melinex plastic sheet on a small group of eggs (20–100) so that the eggs sup ported the plate. The deformation of the eggs was determined by measuring their increased diameter. The mechanical properties of the cell membrane of unfertilized eggs appear to be unaffected by high hydrostatic pressure. When compressed fertilized eggs, from which the fertilization membrane had been, removed by urea treatment, were subjected to pressure of about 7 × 104 kN m−2 (10000 lb/in.2), there was a significant increase in cell diameter. This increase was such that it appeared as if the resistance to deformation of the membrane were about quartered. Surprisingly after a few minutes the diameter decreased again indicating a contraction at the membrane even while under high hydrostatic pressure. Release of the hydrostatic pressure resulted in a further reduction in diameter and thus contraction at the surface, the final diameter sometimes being less than that before the hydrostatic pressure was applied.
A biological system producing a self-assembling cholesteric protein liquid crystal
Authors: Neville A. C. , Luke B. M.
Published: April 2021

The protein in the oothecal glands of praying mantids (Sphodromantis tenuidentata, Mio mantis monacha) exists in the form of lamellar liquid crystalline spherulites, which coalesce asthey flow out of a punctured gland tubule. Electron micrographs of sections of these spherulites after fixation show parabolic patterns of an electron-light component, set in a continuous matrix of protein. Such patterns arisein helicoidal systems (e.g. arthropod cuticle) and microdensito metric scans of the matrix show a rhythmical electron-density variation consistent with helicoidal structure. Double spiral patterns identical to those seen in liquid crystal spherulites are illustrated. These properties resemble those of cholesteric liquid crystals. The constructional units appear to be molecular rather than fibrillar as described by previous authors. The helicoidal architecture arises by self-assembly in the gland lumen. Lamellar surface structures self-assembled spontaneously on glass coverslips when the protein was left to stand for several days. When heated to 55 °C, the birefringent liquid crystalline protein abruptly changes to an isotropic gel, with associated loss of parabolic patterning in electron micrographs and of the rhythmical electron-density variation on microdensitometric scans. This behaviour is compared to the formation of gelatin from collagen, in terms of the randomization of an originally ordered secondary structure.
On the function and fate of phage progeny rna in infected bacteria
Authors: Fenwick M. L.
Published: April 2021

Spheroplasts of Escherichia coli infected with RNA phage R17 were treated with actinomycin D to inhibit the synthesis of host RNA and protein. Ribosomes occurred predominantly as monomers, dimers and trimers which were active in viral protein synthesis. Partly finished strands of viral RNA in lysates sedimented at a rate characteristic of groups of 6–12 ribosomes and their buoyant density suggested that they were in fact attached to ribosomes. Newly completed strands were associated with single (70-s) ribosomes, forming a 75-S complex, and with ribosome dimers and trimers. Some were probably loosely bound to 30-s ribosomal subunits.

Newly made RNA began to accumulate in mature virions after a lag of about 5 min and rose to 16 % of the total labelled viral RNA by the end of the growth cycle, either with continuous labelling or after a 5-min pulse followed by a non-radioactive chase. It is suggested that most, if not all, new viral RNA forms a rather stable association with ribosomes and that a small proportion of this RNA may subsequently pass into progeny virus.
Calcium ions initiate the selective depolymerization of the pellicular microtubules in bloodstream forms of Trypanosoma brucei
Authors: Dolan Michael T. , Reid Colin G. , Voorheis H. Paul
Published: April 2021

Calcium ions (100μM) were found to initiate the selective and complete depolymerization of the pellicular microtubules of Trypanosoma brucei. The Ca2+-dependent release of tubulin was found to occur without the detectable mediation of calmodulin. The released, depolymerized, pellicular tubulin from T. brucei cross-reacted with a monoclonal antibody raised against yeast tubulin. The pellicular tubulin was found to be composed of two alpha isotypes (apparently equal amounts) and one beta isotype. No other proteins were released from the plasma membrane-microtubule complexes during treatment with Ca2+. The released pellicular tubulin was capable of reassembly into microtubules with normal ultrastructure. The observations reported here suggestthat a special process may be required to accommodate the cleavage furrow during cytokinesis. Thia process would either be the Ca2+-dependent depolymerization of at least two of the cross-linked pellicular microtubules or the detachment of the cross-bridges between two pairs of pellicular microtubules on opposite sides of the cell.
Pollen-stigma interactions in Brassica: IV. Structural reorganization in the pollen grains during hydration
Authors: Elleman Carole J. , Dickinson H.G.
Published: April 2021

With the aid of osmium tetroxide vapour, dry pollen and pollen at various stages of hydration has been fixed anhydrously for examination with the transmission electron microscope (TEM). In addition to establishing features characteristic of grains at different states of hydration, this technique has enabled the detection of a superficial layer investing both the exine and the pollen coating. This layer, some 10 nm in depth, binds both lanthanum and Alcian Blue and is shown to be the first component of the pollen grain to make contact with the stigmatic pellicle. The use of vapour fixation has also rendered it possible to chart the passage of water into the pollen grains with great accuracy, for each level of hydration displays a strikingly different cytoplasmic organization. For example, dry pollen is characterized by the presence of unusual structures at the protoplast surface and large numbers of spherical fibrillar bodies, whilst the protoplast of hydrating pollen is conspicuously stratified and contains a peripheral layer of membranous cisternae, subjacent to which is a fibrillar matrix derived from the spherical bodies found in the dry grains. Vapour-fixed, fully hydrated pollen resembles conventionally fixed grains.

The pollen coating appears electron-translucent after anhydrous fixation and contains discrete, slightly rounded bodies some 50 nm in diameter. The uptake of water by grains on the stigma is accompanied by conspicuous structural changes in this layer for, after a short period in contact with the papillar surface, the spherical bodies rapidly disappear and the coat becomes electron-opaque. Close examination of this ‘converted’ coating reveals the presence of membranous vesicles and other structural components.
The synaptonemal complexes of Caenorhabditis elegans: Pachytene karyotype analysis of hermaphrodites from the recessive him-5 and him-7 mutants
Authors: Goldstein Paul
Published: April 2021

The him-5 and him-7 mutants (high incidence of males) of Caenorhabditis elegans both showed increased rates of X chromosome non-disjunction (16% and 3%, respectively) but him-7 also had a high frequency of autosomal non-disjunction (34%). Synaptonemal complex (SC) karyotype analysis revealed a haploid chromosome number of six in each strain. Alterations in him-7 nuclear morphology were observed but there were no aberrations in SC structure that could account for the increased frequency of autosomal non-disjunction. However, the frequency of X-chromosome nondisjunction occurred at predicted rates on the basis of the number of disjunction regulator regions (DRRs) present on the SCs. The observation that the levels of X-chromosome non-disjunction were not influenced by the increase in the frequency of autosomal non-disjunction supports the notion that the X chromosome is subject to separate controls during meiosis. The him-7 mutant is nested within the rad-4 map region on linkage group V, however, SC analysis did not reveal the physical position on the chromosome because of synaptic adjustment.
Comparison of the relative importance of tyrosine-specific vinculin phosphorylation and the loss of surface-associated fibronectin in the morphology of cells transformed by rous sarcoma virus
Authors: Kellie S. , Patel B. , Mitchell A. , Critchley D. R. , Wigglesworth N. M. , Wyke J. A.
Published: April 2021

We have investigated the relative importance of tyrosine-specific phosphorylation of vinculin and the loss of surface-associated fibronectin in the maintenance of the rounded morphology characteristic of chick embryo fibroblasts (CEF) transformed by Rous sarcoma virus (RSV). To address this question we have examined the interaction of CEF and RSV-CEF in vitro with exogenously added fibronectin in both 3-day culture experiments and short-term, 3-h spreading experiments. We report that the addition of human plasma fibronectin to cultures of RSV-CEF results in the restoration of a near-normal morphology, as has been described previously, with the added fibronectin incorporated into an extracellular matrix. However, the phosphotyrosine content of vinculin in these cells was unchanged from that of control, untreated RSV-CEF despite the change in morphology. In short-term spreading experiments RSV-CEF were unable to adopt a fully spread morphology on fibronectin substrates, with defects in the formation of adhesion plaques and microfilament bundles compared with untransformed CEF. pp60v-src was present in the newly formed adhesion plaques of RSV-CEF spreading on fibronectin substrates. The relevance of these results to the maintenance of the transformed phenotype is discussed.
Organization of stigma surface components inBrassica:A cytochemical study
Authors: Gaude T. , Dumas C.
Published: April 2021

The nature of secretion products forming the pellicle of the dry-type stigma surface has been investigated in Brassica by an electron-microscopic cytochemical study. In order to determine whether the pellicle is organized like a membrane, we used a series of cytochemical methods to visualize cell membranes. Three groups of techniques presenting different degrees of specificity in terms of cell surface markers have been used: non-specific labelling of cell-surface components, characterization of glycoconjugates and localization of enzymic activities. We demonstrated that the pellicle of Brassica stigmas, although not possessing a trilamellar structure, presents numerous characteristics encountered in biological membranes. In particular, it possesses cytochemically demonstrable enzyme activities, including ATPase and adenylate cyclase, whose role remains to be elucidated in relation to the pollen-stigma interactions.
Structure of dictyosomes in Scenedesmus acuminatus observed by the freezesubstitution method
Authors: Ueda Katsumi , Noguchi Tetsuko
Published: April 2021

The ultrastructure of dictyosomes in Scenedesmus acuminatus was studied by the freezesubstitution method. The cisternal membranes of dictyosomes clearly exhibited three-layered structures. Electron-dense particles were visible on or in the proximal cisternal membranes. The upper and lower membranes of the distal cisternae were in close contact with each other at the cisternal centre. The electron density of transfer vesicles, secretory vesicles and cisternal margins was very high. These findings differ considerably from those obtained by conventional chemical fixation, and suggest that freeze-substitution is superior to chemical fixation for preserving structures, without distortion and preventing dissolution of constituent substances.
Contraction of collagen lattice by peritubular cells from rat testis
Authors: Ehrlich H. P. , Borland K. M. , Muffly K. E. , Hall P. F.
Published: April 2021

Peritubular cells from 15- and 25-day-old rat testis trapped in collagen lattices caused those lattices to contract. Contraction proceeded more rapidly and to a greater extent using cells from younger rats. When 36000 cells from 15- and 25-day-old rats were trapped in 800 mm2 lattices, the areas were reduced to 28 mm2 and 170 mm2, respectively, within 24h. The cells from older rats were less effective at contracting the lattice than cells from younger rats. Cytochalasin B (5μg ml−1) inhibited lattice contraction and caused disruption of actin filaments as seen by fluorescent staining with Rh-phalloidin. Cholera toxin (10μgml−1), and 1 mM-dibutyryl cAMP inhibited lattice contraction, as did 10 μM-trifluoperazine, commonly an inhibitor of calmodulin. The total intracellular concentration of cAMP was greater in peritubular cells from 25-day-old rats than in those from 15-day-old rats: 427 ± 34 and 120 ± 16pmolmg−1 cell protein, respectively. When peritubular cells in monolayer were permeabilized with glycerol, the addition of ATP caused the cells to contract. Cell contraction was greater in cells from 15-day-old rats than 25-day-old rats. When cells were grown on silicone rubber, they caused that surface to wrinkle. Peritubular cells from 15-day-old rats caused the onset of wrinkling at 4 h. At the same time, no wrinkling was observed with cells from 25-day-old rats. Studies of lattice contraction and cell contraction were also made using cells from 20-day-old rats. In each case, contraction was intermediate between that of cells from 15- and 25-day-old rats. The possibility exists that lattice contraction, cell contraction and wrinkling of silicone film result from a mechanism of actin filament sliding, generated by myosin ATPase activity, and is inhibited by cAMP. The reduced rate of contraction in cells from 25-day-old rats may be related to their higher intracellular levels of cAMP. Evidence exists to show that cAMP blocks myosin ATPase activity by inhibiting the phosphorylation of its regulatory peptide, myosin light chain.
Increase in the concentration of major boar sperm surface proteins during maturation in the epididymis
Authors: Saxena N. K. , Saxena N. , Hunt W. , Peterson R. N. , Henry L. , Russell L. D.
Published: April 2021

High-resolution two-dimensional polyacrylamide gel electrophoresis analyses have indicated that several major boar sperm plasma membrane polypeptides (PMPs) increased in concentration during maturation in the epididymis. To investigate this further,monoclonal antibodies (MAbs) to two of these PMPs/glycoproteins referenced as 4·85 and 5·0 and polyclonal antisera (PCA) raised against PMP 5·0 were used to quantify these changes. ELISA assays for both PMPs, using solubilized plasma membrane (PM) and intact PM as antigens, indicated that both PMPs were present in greater concentration in cauda PM than in caput PM. ELISA assay using PCA against 5·0 and whole sperm from the cauda and caput epididymis also showed an increase in the concentration of this protein during transit through the epididymis. Indirect FITC fluorescence microscopy showed that MAbs reacted specifically with the luminal aspect of secretory cells of the corpus epididymis and, with greater intensity, with secretory cells of the cauda segment. When MAb tags were used the fluorescence technique indicated that PMP 5·0 was restricted to a narrow area of PM overlying the principal segment of the head of caput sperm but fluorescence extended further into the principal segment and into the flagellum of cauda PM. Since MAb binding sites appear to be partially masked, PCA were also used to localize antigenic sites. PCA to PMP 5·0 showed antigen localization in both head and flagellar PM of cauda and caput sperm. The fluorescence of cauda sperm PM, however, was significantly more intense than that of caput sperm PM. MAb to PMP 4·85 also bound to antigen more extensively on cauda sperm. The sum of these analyses suggests that major sperm PMPs are secreted by the epididymis and absorbed by sperm. The magnitude of surface protein changes occurring during epididymal transit in boar sperm appears to be greater than heretofore recognized.
Characterization of cystathionine synthase as a selectable, liver-specific trait in rat hepatomas
Authors: Goss S. J.
Published: April 2021

Cell growth using homocysteine as a source of cysteine-sulphur requires two enzymes, cystathionine synthase (CS) and γ-cystathionase (CT). The second of these enzymes, CT, is apparently present in most cell lines regardless of their tissues of origin, since most cells can grow m vitro in the absence of cystine if they are provided with cystathionine, the intermediate in the pathway. Likewise, homocysteine will support the growth of many human cells. However, of a wide range of rodent cells, only well-differentiated rat hepatoma cells were found to grow using homocysteine in place of cystine. It is shown that cell growth in homocysteine-medium correlates well with the presence in the cells of detectable levels of CS. Furthermore, in cells able to grow in homocysteine-medium, it is possible to demonstrate the homocysteine-dependent transsulphuration of serine to cysteine. Growth in homocysteine-medium is not dependent on the release of preformed cysteine from disulphide complexes with serum proteins. In cell hybrids, and in ‘dedifferentiated’ variants of rat hepatomas, CS, but not CT, is subject to extinction coordinately with well-characterized liver-specific traits. For rodent cells, homocysteine-medium thus acts as a selective medium requiring the expression of a single liver-specific trait, CS. In addition it is shown that, in certain hepatoma variants, CS is regulated co-ordinately with a urea-cycle enzyme (carbamoyl phosphate synthetase I) by glucocorticoids and cyclic-AMP.

Cell death through cysteine starvation is briefly considered. The immediate cause of death is apparently an insufficient supply of reduced glutathione. Selenium and vitamin E assist cell growth when the supply of cysteine is limiting.
The primary culture of epithelia from the secretory coil and collecting duct of normal human and cystic fibrotic eccrine sweat glands
Authors: Lee C. M. , Carpenter F. , Coaker T. , Kealey T.
Published: April 2021

Isolated human eccrine sweat glands have been microdissected into their secretory and reabsorptive components. Complete separation of these epithelia was confirmed by differential uptake of Neutral Red stain by an intermediate section of gland containing the junction between the secretory coil and the collecting duct. Primary cultures were obtained from explants of both tissues in medium RPMI-1640 or Williams E supplemented with foetal calf serum, insulin, transferrin, epidermal growth factor and hydrocortisone. The cells in the initial coil cultures had an elongated morphology while those of ductal origin were polyhedral. After 10 days both cultures were composed of polyhedral cells of varying diameter. All these morphological types were of epithelial lineage, as demonstrated by the binding of a monoclonal antibody to cytokeratin, the intermediate filament specific for epithelial cells.

Outgrowth from both secretory and reabsorptive epithelia were multilayered, with plentiful desmosomal connections and an underlying basal lamina. Ultrastructural features typical of the epithelial cell types present in intact eccrine sweat glands were absent in a high proportion of the proliferating cells but domes, indicative of transepithelial active ion transport, were present in dense cultures from the reabsorptive duct.

Outgrowth was also obtained from the secretory and reabsorptive epithelia of sweat glands from two cystic fibrotic patients. Since the most characteristic malfunction of cystic fibrosis is the impaired ion transport in the eccrine sweat gland, the availability of cultured epithelia should provide a useful model for study of the disease.
Basal body loss during fungal zoospore encystment: evidence against centriole autonomy
Authors: Heath I. Brent , Kaminskyj Susan G. W. , Bauchop Tom
Published: April 2021

The controversial question of the possible autonomy of centrioles, as shown by the persistence of all or part of them in the generative cell line throughout the life cycle of organisms, remains unresolved. All previous reports on shedding or withdrawal of cilia and flagella showed that their basal bodies (= centrioles) were retained in the cells where they may, or may not, subsequently disassemble. We show that in the fungusNeocallimastix sp. the basal bodies are discarded with the flagella when zoospores encyst. This shedding of basal bodies argues against centriolar persistence in any form and thus against their autonomy and endosymbiotic origin.
Pattern of dna increase in macronuclear anlagen of Tetrahymena
Authors: Roth Joachim , Cleffmann Günter
Published: April 2021

By combining cytophotometry with autoradiography, five stages of macronuclear anlagen can be discriminated by their DNA content until the end of the first cell cycle after conjugation in Tetrahymena. DNA increases from 2C to about 32C. Each S-phase is followed by a non-synthetic period. Comparing the mean nuclear DNA content after and before each S-phase revealed that 16C anlagen contain significantly less DNA than twice the amount of 8C anlagen. This is unlike the situation in other S-phases during which the amount of DNA is precisely doubled. In the second cell cycle after conjugation some of the cells increase their macronuclear G2 DNA content beyond the 64C stage, while the majority show a mean G2 content of about 64C.