bluuebunny/crossref_metadata_2025 huggingface dataset:
Neuromuscular junctions in the body wall muscles of the earthworm, Lumbricus terrestris linn
Authors: Mill P. J. , Knapp M. F.
Published: April 2021

The fine structure of the neuromuscular junctions in the body wall muscles of the earthworm 13 described. The segmental nerves send branches into the muscle layers. Axons in the nerve branches contain numerous synaptic vesicles and contact is established between these axons and muscle fibres or muscle tails; the latter may extend for a considerable distance from the muscle fibre. The cleft between the axolemma and sarcolemma is 85–120 nm wide and contains basement membrane material. At intervals small aggregations of electron-dense material are attached to the axonal membrane and synaptic vesicles are associated with these. The sarcolemma bears rather larger masses of dense material and is also specialized extracellularly.
The rapid intermixing of cell surface antigens after formation of mouse human heterokaryons
Authors: Frye L. D. , Edidin M.
Published: April 2021

Cells from established tissue culture lines of mouse (CIID) and human (VA-2) origin were fused together with Sendai virus, producing heterokaryons bearing both mouse and human surface antigens which were then followed by the indirect fluorescent antibody method. Within 40 min following fusion, total mixing of both parental antigens occurred in over 90 % of the heterokaryons.

Mouse H-2 (histocompatibility) and human surface antigens were visualized by successive treatment of the heterokaryons with a mixture of mouse alloantiserum and rabbit anti-VA-2 antiserum, followed by a mixture of fluorescein-labelled goat anti-mouse IgG and tetramethyl- rhodamine-labelled goat anti-rabbit IgG(Fc).

The CIID × VA-2 fusions were carried out in suspension and maintained at 37°C in a shaking water bath; aliquots were removed at various intervals and stained with the above reagents. The heterokaryon population was observed to change from an initial one (5-min post-fusion) of non-mosaics (unmixed cell surfaces of red and green fluorescence) to one of over 90 % mosaics (total intermixing of the 2 fluorochromes) by 40 min after fusion. Mouse-human hybrid lines, derived from similar fusions, gave fluorescence patterns identical to those of the mosaic heterokaryons.

Four possible mechanisms would yield such results: (i) a very rapid metabolic turnover of the antigens; (ii) integration of units into the membrane from a cytoplasmic precursor pool; (iii) movement, or ‘diffusion ‘of antigen in the plane of the membrane; or (iv) movement of existing antigen from one membrane site into the cytoplasm and its emergence at a new position on the membrane.

In an effort to distinguish among these possibilities, the following inhibitor treatments were carried out: (1) both short- and long-term (6-h pre-treatment) inhibition of protein synthesis by puromycin, cycloheximide, and chloramphenicol; (2) short-term inhibition of ATP formation by dinitrophenol (DNP) and NaF; (3) short- and long-term inhibition of glutamine- dependent pathways with the glutamine analogue 6-diazo-5-oxonorleucine; and (4) general metabolic suppression by lowered temperature.

The only treatment found effective in preventing the mosaicism was lowered temperature, from which resulted a sigmoidal curve for per cent mosaics versus incubation temperature. These results would be consistent with mechanisms iii and/or iv but appear to rule out i and ii. From the speed with which the antigen markers can be seen to propagate across the cell membrane, and from the fact that the treatment of parent cells with a variety of metabolic inhibitors does not inhibit antigen spreading, it appears that the cell surface of heterokaryons is not a rigid structure, but is ‘fluid’ enough to allow free ‘diffusion’ of surface antigens resulting in their intermingling within minutes after the initiation of fusion.
Observations on the fine structure and development of the spindle at mitosis and meiosis in a marine centric diatom (Lithodesmium undulatum): IV. the second meiotic division and conclusion
Authors: Manton Irene , Kowallik K. , Stosch H. A. Von
Published: April 2021

The second meiotic division is shown to be of critical importance for interpretation of structures and events already seen at other divisions. The development of flagellar bases at interkinesis is demonstrated in relation to precursor material seen to accumulate near each pole at meiosis I. The name ‘paracentrosome’ is suggested for this material, which is used up in forming the flagellar bases and spindle precursor. The spindle at late prophase, metaphase and telophase II is shown to resemble those of other divisions except that it is consistently smaller; this fact is numerically demonstrated from serial sections of metaphase II cells. The greatly reduced size and unusual shape of the polar plates present in addition to flagellar bases at metaphase II suggest that these are in a sense equivalent structures with a mutually competitive relation to the paracentrosome.

Preliminary observations with the light microscope on the relatively large nuclei of oogonia during meiosis I have shown that the haploid chromosome number is not less than 19 nor more than 23 and that chromatid separation in relation to the kinetochores at anaphase I is normal; the bearing of these findings on interpretation of the spindle is discussed.

Comparisons with other organisms are carried out in a preliminary way and the investigation ends with a résumé of the more important externally visible events in male gametogenesis adjusted to a common time scale in the course of one day.
Specificity of pseudopodium induction by the action of cations on Amoeba proteus
Authors: Brewer J. E. , Bell L. G. E.
Published: April 2021

Some substituted cholines, long-chain aliphatic substituted amines and simple inorganic salts have been tested at different concentrations for their ability to induce pseudopodia from Amoeba proteus. The ability of substituted amines to induce pseudopodia is inversely related to their ability to bind to acid polysaccharides. The reaction of compounds with the surface polysaccharide is probably not in itself the only requirement for pseudopodium induction. A mechanism is proposed in which the properties of the cell membrane are altered by the reaction of compounds with bulky cationic groups with the membrane lipid. The formation of new pseudopodia is not the direct result of a local reduction in the surface charge density or in surface potential.
Studies on the mechanism underlying the inhibition by puromycin of cell aggregation in vitro
Authors: Dunn M. J. , Owen E. , Kemp R. B.
Published: April 2021

Cells dissociated with 0·25 % crude trypsin from the muscle tissue of 9-day-old chick embryos were employed to investigate the effect of puromycin on cellular metabolism. Parallel studies were also made, using the gyratory shaker, to confirm the effectiveness of puromycin in inhibiting cell aggregation and protein synthesis.

Puromycin when introduced at a concentration of 10μg/ml into a suspension of cells in Eagle’s MEM did not completely inhibit cell aggregation. Small aggregates were formed in the first 4 h of the experiment. Protein synthesis of the rotated cells, as measured by the incorporation of L-[α -14C]leucine into proteins, was arrested by 91 · 7% within 15 min of introducing puromycin into a cell suspension. The antibiotic retained its inhibitory effect on protein synthesis for the 24-h period of rotation.

Puromycin inhibited the cellular oxygen uptake and carbon dioxide evolution of the rotated cells by 40 % within 4 h of its introduction. However, treated cells were still respiring, though at a much reduced rate, at the end of the 24-h experimental period. The release of radioactive carbon dioxide by puromycin-treated cells was also inhibited by 40% at the 4-h stage but after 8 h no further14CO2 was evolved. The presence of the antibiotic markedly inhibited the uptake of glucose by trypsin-dissociated cells.

The level of glycogen and lactate in cells suspended in Eagle’s MEM was reduced very considerably over a 24-h period. The presence of puromycin accelerated glycogen utilization over the first 6 h of rotation but at 24 h there was a difference of only 0 · 6 % between the glycogen content of treated cells and controls. At 24 h 11·3 % less lactate remained in the puromycin-treated cells than in the controls.

The ATP/ADP ratio of trypsin-dissociated cells decreased from an initial value of 2 · 59 to 1 · 45 after rotation for 24 h. In the presence of puromycin the ATP/ADP ratio was 0 · 62 at 4 h and had further declined to 0 · 48 by 24 h.

The effects of puromycin on the aggregation, protein synthesis and cellular metabolism of trypsin-dissociated cells are discussed in relation to cellular adhesive mechanisms.
Axonal microtubules of crayfish and spiny lobster nerve cords are decorated with a heat-stable protein of high molecular weight
Authors: Warren Robert H.
Published: April 2021

Axons of crayfish and spiny lobster ventral nerve cords contain large numbers of microtubules that are decorated with fine filaments. These microtubules can be stabilized in permeabilized axons using buffers that contain either polyethylene glycol or glycerol/dimethyl sulphoxide. In the former, the stabilized microtubules retain their filaments and their normal spacing; in the latter, the filaments are stripped off and the bare microtubules collapse onto one another. This observation has been used as the basis for a method of identifying some of the proteins that make up the filaments. Axons are first permeabilized and stabilized in either buffer and then treated with a microtubuledepolymerizing buffer. The axons treated first with polyethylene glycol release tubulin and significant quantities of microtubule-associated proteins (MAPs), while the axons pre-treated with glycerol release tubulin and only traces of associated proteins. One of the proteins released in largest quantity along with tubulin from polyethylene glycol-treated axons is a high molecular weight, heatstable MAP that co-electrophoreses with MAP-2 from mammalian brain. This same protein copurifies with tubulin that is obtained from crayfish nerve cords by two cycles of polymerization and depolymerization. It is concluded that this protein is a component of the filaments that decorate the axonal microtubules of the crayfish and spiny lobster.
Possible translocation of a gene for thymidylate kinase from the chloropast to the nuclear genome during evolution
Authors: Groot Egon J. De , Schweiger Hans-Georg
Published: April 2021

On the basis of the fact that in Acetabularia three out of four enzymes that are involved in deoxyribonucleotide metabolism are translated on 70 S ribosomes, while a fourth one is synthesized on 80 S ribosomes, the suggestion was made that translation on 80 S ribosomes might reflect an evolutionarily more recent situation while translation on 70 S ribosomes might be of more ancient origin. This hypothesis prompted experiments aimed at defining the site of translation of thymidylate kinase in a species closely related to Acetabularia but long ago separated from it: Batophora oerstedii. It was demonstrated that in this species, in contrast to a number of Acetabul·ariae, thymidylate kinase is translated on 70 S ribosomes. A possible, appealing interpretation of this finding is that during evolution the coding site for thymidylate kinase has been translocated from the chloropast to the nuclear genome.
X-ray analysis of Lymnaea stagnalis muscle fibres does not suggest that the sarcolemma is permeable to lanthanum ions
Authors: Swales L. S. , Gardner D. R.
Published: April 2021

The cross-striated muscle from the heart ventricle and the smooth penis retractor muscle of the freshwater snail Lymnaea stagnalis have been investigated by X-ray microanalysis to establish whether lanthanum can cross the plasma membrane, as has been reported by other investigators. Tissues were incubated in 1 mM ionic lanthanum before fixation in phosphate- or cacodylate-buffered fixative. X-ray mapping for emissions in the lanthanum energy range indicates a concentration of emissions that coincided only with the network of sub-surface transverse tubules formed by the invagination of the plasma membrane and with the plasma membrane/extracellular space interface. X-ray energy spectra were collected from various cell compartments; peak-to-background ratios were obtained and analysed statistically. Cacodylate buffer is less effective than phosphate buffer in precipitating lanthanum, but no evidence to suggest the redistribution of lanthanum in cacodylate-buffered preparations was found. Lanthanum is precipitated only in the sub-surface transverse tubules and at the plasma membrane/extracellular space interface in both heart ventricle muscle and penis retractor muscle, fixed in either phosphate or cacodylate buffer. There was no evidence of lanthanum precipitation in the background cytoplasm or on any cytoplasmic organelle. These results confirm our hypothesis that lanthanum does not cross the plasma membranes in these molluscan tissues.
Surface charge of insect haemocytes, examined using cell electrophoresis and cationized ferritin-binding
Authors: Takle Garry , Lackie Ann M.
Published: April 2021

Differences in the negative surface charge of haemocytes from Periplaneta americana and Schistocerca gregaria have been revealed using cell electrophoresis and cationized ferritin-binding. Although haemocyte populations from both insect species exhibit ranges of negative surface charge, both techniques show that Schistocerca haemocytes are significantly more negative than Periplaneta haemocytes. The results may help to explain why Schistocerca haemocytes adhere poorly to negative substrata, both in vitro and in vivo, and suggest that an electrostatic mechanism may be involved, at least in part, in adhesion of insect haemocytes to substrata.
Further ultracytochemical analysis of rat adenohypophyseal cells: Detection of two distinct enzyme activities within a single cell
Authors: Khokhlov S. Ye. , Nevorotin A.F.
Published: April 2021

In an effort to ascertain whether a lysosomal enzyme, aryl sulphatase (ArSase), might share the same cell and, or, the same intracellular structure(s) with a histochemical Golgi apparatus marker, thiamine pyrophosphatase (TPPase), in rat pituitary mammotrophs and somatotrophs a technique for a selective demonstration of both enzyme activities within a single specimen has been developed. The technique is based on the selective dissolution of TPPase-related precipitates with 50% sulphuric acid in an ultrathin section of a pituitary specimen processed consecutively for ArSase and TPPase. The analysis of pairs of micrographs displaying the same structures before and after treatment with sulphuric acid has shown that TPPase-related precipitates are located mainly in the trans-Golgi lamellae while ArSase-related precipitates (resistant to sulphuric acid) could be found in both GERL and its derivatives (lysosomes, immature secretion granules) and in the TPPase-reactive trans-Golgi pole in both cell types studied. When seen within the same dictyosome each enzyme activity resided generally within an individual Golgi lamella. Direct continuity between ArSase- and TPPase-related precipitates was also occasionally encountered. The results indicate that ArSase and TPPase activities can share the trans-cisternae of the Golgi apparatus in mammotrophs and somatotrophs of rat pituitary.
A new vacuum-operated stress-providing instrument that applies static or variable duration cyclic tension or compression to cells in vitro
Authors: Banes Albert J. , Gilbert Jerome , Taylor Duane , Monbureau Olivier
Published: April 2021

An instrument providing cyclic stress to cells cultured in vitro has been developed. The unit uses a vacuum to deform a plastic Petri dish yielding 0·13% compression to cells on the inner surface, measured by strain gauge recordings. A regimen of 25 s stress and 5 min relaxation induced no significant change in synthesis of a 45×103Mr protein that comigrates with actin, whereas a 52× 103Mr protein that comigrated with tubulin decreased from 12·7 ± 0·451% of the total protein synthesized in control, static cells to 8·53 ± 0·182% in stressed cells. The unit may have a broad application in monitoring biochemical changes in response to stress in cells such as muscle, lung, tendon, ligament and bone that are normally subjected to tension or compression.
Mutual desmosome formation between all binary combinations of human, bovine, canine, avian and amphibian cells: Desmosome formation is not tissue- or species-specific
Authors: Mattey D. L. , Garrod D. R.
Published: April 2021

Our previous work has suggested that the molecular components of desmosomes are highly conserved between different tissues and different vertebrate species. In order to determine whether the adhesion recognition mechanism of desmosomes is also conserved we have examined the specificity of desmosome formation between different epithelial cell types by co-culturing binary combinations of cells from different species and from epidermal and non-epidermal origin. The following cell types were used: human (HeLa, cervical carcinoma), bovine (Madin Darby bovine kidney, MDBK), canine (Madin Darby canine kidney, MDCK), avian (chick embryonic corneal epithelium) and amphibian (Rana pipiens, adult corneal epithelium). Different cells in co-culture were identified on the basis of at least one of the following criteria: (1) morphology by phase-contrast microscopy; (2) presence or absence of staining of cytokeratin with monoclonal antibody LE6l; (3) morphology at the electron microscope level. Mutual desmosome formation between different cell types was assessed using fluorescent antibody staining with anti-desmoplàkin antibodies and confirmed using electron microscopy. We have found that mutual desmosome formation occurred between all binary combinations of human, bovine, canine, avian and amphibian cells. Thus there is complete non-selectivity of desmosome formation between five different epithelial cell types from three vertebrate classes. Our results suggest that desmosome formation is not tissue- or speciesspecific and that the mechanism for intercellular binding involved in desmosomal adhesion is highly conserved.
The analysis of malignancy by cell fusion: III. hybrids between diploid fibroblasts and other tumour cells
Authors: Wiener F. , Klein G. , Harris H.
Published: April 2021

Diploid fibroblasts were fused with cells of two highly malignant near euploid tumours, a sarcoma induced by polyoma virus and a spontaneous carcinoma; the resulting hybrid cells were tested for their ability to grow progressively in vivo. In the case of the sarcoma/fibroblast hybrids, 15 clonal populations, each derived from a separate primary fusion, were examined. Most of these populations already showed substantial losses of chromosomes by the time enough cells had been generated to permit chromosomal analysis; but a few clones were isolated with chromosomal constitutions approximating to the sum of the 2 parental chromosome sets. Those clones that had undergone substantial chromosomal losses were highly tumorigenic, but some of the clones that contained the complete, or almost complete, chromosome sets of both parent cells, showed a greatly reduced take incidence. Initially these clones produced very few tumours, but the take incidence rose as continued cultivation of the cells in vitro resulted in progressive loss of chromosomes. In the case of the carcinoma/fibroblast hybrids, clonal populations with very high chromosome numbers were selected for special study. These were also found to have a very low take incidence, comparable to that of the hybrids formed by the fusion of the tumour cells with L cell derivatives. None of the tumours produced by the injection of either the sarcoma/fibroblast or carcinoma/fibroblast hybrids were composed of cells containing chromosome complements corresponding to the sum of the 2 parental chromosome sets; all the tumours were formed by selective overgrowth of cells from which some chromosomes had been eliminated. These results indicate that the malignancy of the tumour cells can be suppressed by diploid fibroblasts as well as by the L cell derivatives, but that the rapid chromosome losses characteristic of hybrids between these tumour cells and diploid fibroblasts generate malignant segregants with much greater frequency. In the range of material examined in the present experiments malignancy thus behaves as if it were a recessive character.
Perinuclear dense bodies: Characterization as DNA-containing structures using enzyme-linked gold granules
Authors: Berger Sigrid , Schweiger Hans-Georg
Published: April 2021

Cytoplasmic structures of the Acetabularia cell that appear close to the nucleus during the vegetative phase of the life cycle have been characterized cytochemically. The results obtained by using RNase and DNase linked to gold granules indicate that RNA and also DNA occur in these cytoplasmic structures.
Cell proliferation in Allium cepa L. meristems under 8-hydroxyquinoline, a chelating agent that affects DNA and RNA polymerases
Authors: Ferrero M. L. , Torre C. De La
Published: April 2021

8-Hydroxyquinoline (HQ) chelates Mg2+ and Mn2+ and, secondarily, affects the activities of DNA and RNA polymerases. The in vivo effect of HQ has been estimated in Allium cepa L. meristems growing under new growth kinetics in the presence of this agent. HQ (at both 5 × 10−5 M and 10−4M) depressed incorporation of [3H]uridine much more effectively than that of [3H]-thymidine. Cycle kinetics in meristems behaved as if they were independent of the rates of synthesis or accumulation of RNA since, under HQ, cycle time was only moderately modified and the new cycle kinetics achieved could be explained by the new rates of [3H] thymidine incorporation. Lengthened S periods were partially compensated for by shortened G2 phases, suggesting that, in these cells, both the growth cycle and its coupling with the DNA-division cycle were not disturbed by a decreased amount of RNA.

Finally, the nucleolar cycle during mitosis, but not the interphase nucleolus, was modified under the new rates of RNA synthesis.
Clonal analysis of expression of epithelial antigens in cultures of normal human breast
Authors: Edwards Paul A. W. , Brooks Isobel M. , Bunnage Hilary J. , Foster A. Virginia , Ellison Morag L. , O’Hare Michael J.
Published: April 2021

Cells from normal human breast epithelium were cloned in monolayer culture and the clones were stained with monoclonal antibodies. Tissue was from reduction mammoplasty operations. Cloning efficiences were 5-30%.

Two types of clone were identified: 10 to 30% were of relatively spread cells whose boundaries were often difficult to see by phase-contrast microscopy but where they were visible they appeared as dark lines. The edges of the clones usually appeared to be under tension. These clones were stained by two monoclonal antibodies, LICR-LON-M8 and M24, that stain luminal epithelial cells in the intact tissue, but not myoepithelial or stromal cells. Within a clone the cells showed a full range of antigenic phenotypes. This was confirmed for clones grown from single cells that had been isolated manually. The second type of clone was more compact with little evidence of tension at the edges, and cell boundaries were clearly visible and bright under phase contrast. These clones were not stained by antibodies M8 or M24. Both types of clone stained with a third monoclonal antibody that is specific for luminal epithelial cells in the intact tissue, LICR-LON-M18, but the distribution of staining was different in the different types of clone.

The simplest interpretation of the two types of clone is that luminal epithelial cells give rise to the spread type of clone while the myoepithelial cells give rise to the more abundant and vigorous compact clones. Alternatively, the compact clones may be from luminal epithelial cells that have lost differentiated characteristics.
Significance of a common epitope of plant and animal endomembranes
Authors: Bolwell G. Paul
Published: April 2021

A rat monoclonal antibody that was raised against a common epitope on bean (Phaseolus vulgaris) endomembranes has been shown to cross-react with microsomal polypeptides from a number of plant and animal species. Immunoblotting has shown that the epitope is present on a large subset of polypeptides on microsomes of five animal species. The antigenic site appears to be accessible on intact bean membranes since it is readily digested by trypsin. The epitope is probably not derived post-translationally since the same Mr range is immunoprecipitated from polypeptides newly synthesized in vivo and in vitro. The polypeptides in bean appear to be regulated independently, one of Mr 58000, in particular, was highly induced by treatment of suspension cultures with fungal elicitor.

Preincubation of membranes enriched with endoplasmic reticulum and Golgi apparatus with the antibody blocks transfer of radioactivity from one compartment to the other in vitro. The common antigenic site could possibly be concerned in recognition or some fusion event during membrane trafficking within the cell.
Transplantation studies to investigate mesoderm-ectoderm adhesive cell interactions during gastrulation
Authors: Johnson Kurt E.
Published: April 2021

Experiments involving transplantation of the roof of the blastocoel in Rana pipiens embryos reveal that the inner surface of the roof of the blastocoel must be coated with a fibrous extracellular matrix (F-ECM) to serve as a substratum for mesodermal cell migration. When the roof of the blastocoel is inverted the original outer surface, now projecting toward the blastocoel, does not become coated with F-ECM and does not support mesodermal cell migration. When the roof of the blastocoel is removed from a normal embryo and transplanted into an interspecific arrested hybrid embryo known to be deficient in F-ECM synthesis, the grafted ectodermal fragment does not become coated with F-ECM and does not support normal mesodermal cell migration. When a hybrid graft is placed in a normal embryo, the grafted ectodermal fragment becomes coated with F-ECM and supports mesodermal cell migration. In normal control embryos migrating mesodermal cells are polarized due to formation of lamellipodia on their leading but not their trailing edges. These cells are arranged in overlapping layers. The leading cells form lamellipodia on the roof of the blastocoel and trailing cells form lamellipodia on one another.
Kinetics and differentiation of marrow stromal cells in diffusion chambers in vivo
Authors: Bab I. , Ashton B. A. , Gazit D. , Marx G. , Williamson M. C. , Owen M. E.
Published: April 2021

Rabbit marrow cells inoculated into diffusion chambers (107 cells/chamber) were implanted intraperitoneally into athymic mouse hosts and cultured in vivo for 20 days. A connective tissue consisting of bone, cartilage and fibrous tissues is formed by the stromal fibroblastic cells of marrow within the chambers. Cell kinetics and tissue differentiation have been studied using histo-morphometric and biochemical analyses. Haemopoietic cell numbers decrease to less than 0·05% of the initial inoculum during the 20-day period. At 3 days an average of 15 stromal fibroblastic cells only are identifiable within the chambers. After 3 days there is a high rate of stromal cell proliferation with a doubling time of 14·5 h during the period from 3 to 8 days and an increase in the total stromal cell population by more than six orders of magnitude from 3 to 20 days. Thirteen to fourteen population doublings occur before expression of the first observable differentiation parameter, alkaline phosphatase activity. The data demonstrate that the mixture of connective tissues formed within the chamber is generated by a small number of cells with high capacity for proliferation and differentiation. This is consistent with the current hypothesis that stromal stem cells are present in bone marrow.
Evidence for the existence in the nuclei of friend cells of a new type of ribonucleoprotein network
Authors: Yancheva Nina , Markov Dimiter , Djondjurov Lalio
Published: April 2021

We have isolated fragments of a novel nuclear structure exhibiting the morphological and biochemical characteristics of a ribonucleoprotein network. Under transmission electron microscopy it is visualized as irregularly interconnected branches assembled by tightly packed particles with sizes between 100Å and 300Å. RNA extracted from this structure shows a complex pattern: as well as ribosomal 28S and 18S RNA, small amounts of heterogeneous nuclear RNA and small nuclear RNA are also present. The protein composition of the network indicates a strong domination of ribosomal polypeptides. This fact, considered together with the sedimentation characteristics of the prevailing type of particles isolated directly from the network, supports the conclusion that ribosomal particles are the representative particulate elements. Further electrophoretic analysis of the protein has pointed out that it also contains a significant number of acidic polypeptides. Control experiments have suggested that the site of origin of the network fragments studied is not nucleolar: the fragments are released during extraction, most probably from the nuclear periphery. It is not yet clear whether this structure is localized only at the nuclear periphery or represents an extensive structure, occupying the entire volume of the nucleus. It is speculated that the network is involved in the extranucleolar transport and maturation of ribosomes.
Widespread occurrence of anti-troponin T crossreactive components in non-muscle cells
Authors: Lim Soo-Siang , Hering Gordon E. , Borisy Gary G.
Published: April 2021

Using a monoclonal antibody generated against striated muscle troponin T, we previously noted the presence of crossreactive components in smooth muscle and non-muscle cells. Since the presence of troponin T in tissues other than striated muscle is controversial, we sought to establish the nature of the crossreaction and to determine the extent of its occurrence. For this study, indirect immunofluorescence microscopy and immunoblot analyses were performed. Crossreactive material was found in diverse cells from the animal, plant and fungal kingdoms. On the basis of morphological distributions, both microtubule-associated and non-microtubule-associated components were revealed. Microtubule-associated components of animal cell lines included a 35×103Mr protein, similar in electrophoretic mobility to skeletal troponin T (37×103Mr). Reactive components of comparable mobility were observed in immunoblots of brain and cerebellar homogenates. Filamentous staining was observed in a variety of mammalian cells in culture and in cells of vertebrate tissues. Chick cerebellar tissue showed reactions in the neurites of the molecular layer and granule cell bodies. In the plant kingdom, examination of the onion root-tip cells indicated an association of crossreactive components with interphase cortical microtubules, preprophase bands, the mitotic spindle and phragmoplast microtubules. In the fungal kingdom, both interphase and mitotic spindle microtubules in a cellular slime mould were reactive. Nonmicrotubule-associated components were observed in the centrosphere regions of mitotic seaurchin eggs, in mitotic and interphase plasmodia of Physarum polycephalum, and in trichocysts and basal bodies of Paramecium tetraurelia. In all systems examined, the troponin T crossreactive components were located in regions or on structures of possible Ca2+ or calmodulin activity, suggesting a possible functional similarity to troponin T.
Splitting and internalization of the desmosomes of cultured kidney epithelial cells by reduction in calcium concentration
Authors: Mattey D. L. , Garrod D. R.
Published: April 2021

Desmosome assembly may be induced in simple epithelial (MDBK and MDCK) cells maintained in low calcium medium (LCM: [Ca2+] <0·05mM) by raising [Ca2+] to that of standard culture medium (SM: [Ca2+] = l·8mM). Here it is shown that if cells in SM are simply returned to LCM, their desmosomes split in the intercellular region within 15 min and the desmosomal halves are internalized within 30 min. This is the first time that desmosome splitting has been shown to occur in response to a reduction in [Ca2+] rather than Ca2+ chelation. Fluorescent antibody staining shows that the desmosomal glycoproteins as well as the plaque constituents are internalized, although a pool of the glycoproteins known as desmocollins remains at the cell surface, apparently unassociated with other desmosomal components. Desmosomal halves that have been recently internalized in response to LCM treatment do not return to the cell surface to participate in new desmosome formation. MDCK cells are able to form new desmosomes rapidly (15—30 min) while old desmosomes continue to be internalized.

The desmosomes of MDBK cells remain sensitive to splitting and internalization in response to reduction in [Ca2+] for up to 14 days of culture in SM. In contrast, the desmosomes of MDCK cells become resistant to reduction in [Ca2+], as well as Ca2+ chelation by EGTA, after 4–5 days in SM. When treated with LCM or EGTA, MDCK cells with ‘stabilized’ desmosomes partially separate but remain attached to each other at some points. Regions of attachment stain brightly with anti-demosomal antibodies and are characterized by ‘giant’ desmosomes, up to 4/tm long, roughly 20 times larger than those formed in cells in SM. These giant desmosomes may form by lateral fusion of small desmosomes.
Ascorbate induced changes in glycosaminoglycan synthesis and distribution of normal and sv40-transformed fibroblasts
Authors: Edward Michael
Published: April 2021

The effect of ascorbate on the glycosaminoglycans synthesized by normal and simian virus 40(SV40)-transformed human skin fibroblasts was examined. Cells were incubated in the presence or absence of ascorbate, and radiolabelled with [3H]glucosamine and [35S]sulphate for 48 h, 3 days after reaching confluence. Glycosaminoglycans were analysed in the medium, a collagenase extract, and in the trypsin/cell-associated fraction. Hyaluronic acid was the main 3H-labelled glycosaminoglycan in all but the collagenase extracts, and showed a large decrease in normal fibroblast cultures, but a significant increase in SV40-transformed fibroblast cultures following feeding with ascorbate. Incorporation of [3H]glucosamine into sulphated glycosaminoglycans was reduced in normal fibroblast cultures but increased slightly in SV40-transformed cultures following ascorbate supplementation. [35S]sulphate incorporation remained essentially unaltered in both cell cultures. Ascorbate stimulated the deposition of glycosaminoglycans into the insoluble matrix of normal fibroblasts while reducing the deposition in SV40-transformed fibroblast cultures. The observed changes may in part be related to ascorbate-induced deposition of collagen in normal fibroblast cultures and the inability of the transformed fibroblast cells to deposit an extensive extracellular matrix, in addition to possible changes in the specific activity of the UDP-iV-acetyl- [3H]hexosamine pool.
Calcium-induced desmosome formation in cultured kidney epithelial cells
Authors: Mattey D. L. , Garrod D. R.
Published: April 2021

Previous work has shown that cultured kératinocytes do not form desmosomes at low [Ca2+] (<0·l mM) but may be induced to do so by raising [Ca2+] to physiological levels (l·8–2mM). Here, fluorescent antibody staining with specific anti-desmosomal antibodies and electron microscopy have been used to determine whether Ca2+-induced desmosome formation also occurs in simple epithelial cells.

Both Madin-Darby canine and bovine kidney cells (MDCK and MDBK) exhibit Ca2+-induced desmosome formation, but there are significant differences between them. MDCK cells resemble kératinocytes in showing showing rapid desmosome formation characterized by the simultaneous appearance of four desmosomal antigens at the cell periphery within 15–20 min of raising the [Ca2+]. In contrast MDBK cells take between 7 and 8 h to form desmosomes after Ca2+ switching, and this is characterized by slow appearance of two desmosomal antigens, the 175–164 (×103)Mr glycoprotein and desmoplakin, at the cell periphery.

Differences in the pattern of staining for desmosomal antigens between the two cell types in low and high [Ca2+] are described and discussed in relation to desmosome formation and internalization. Triton X-100 extractability of desmosomal antigen staining is also considered. While most is non-extractable, staining for the glycoproteins known as desmocollins is completely extractable from MDCK cells in low [Ca2+], but that which reaches the cell periphery after Ca2+ switching becomes non-extractable. Although neither cell type forms desmosomes in low [Ca2+], both possess zonulae adhaerentes, suggesting a difference in Ca2+ requirement for formation of these two junctions.
Endoplasmic reticulum contains a common, abundant calcium-binding glycoprotein, endoplasmin
Authors: Koch G. , Smith M. , Macer D. , Webster P. , Mortara R.
Published: April 2021

The most abundant protein in microsomal membrane preparations from mammalian cells has been identified as a 100 × 103Mr concanavalin A-binding glycoprotein. The glycosyl moiety of the glycoprotein is completely sensitive to endoglycosidase H, suggesting a predominantly endoplasmic reticulum localization in the cell. Using a monospecific antibody it was shown by binding and immunofluorescence studies that the glycoprotein is intracellular. Immunoelectron microscopy showed that the glycoprotein was at least 100 times more concentrated in the endoplasmic reticulum than in any other cellular organelle. It was found to be substantially overexpressed in cells and tissues rich in endoplasmic reticulum. Since it is the major common protein component associated with the endoplasmic reticulum we refer to it as endoplasmin. Calcium-binding studies show that endoplasmin is a major calcium-binding protein in cells, suggesting that at least one of its roles might be in the calcium-storage function of the endoplasmic reticulum. The amino-terminal sequence of endoplasmin is identical to that of a 100 × 103Mr stress-related protein.
The lack of attachment of transformed embryonic lung epithelial cells to collagen I is corrected by fibronectin and FXIII
Authors: Paye M. , Lapière CH. M.
Published: April 2021

PER cells, a transformed pulmonary epithelial cell line that adhered to a large extent to a fibronectin substratum, were found to be attachment-deficient to collagen I. Although fibronectin can bind to collagen I monomers and polymers, the addition of exogenous fibronectin in the attachment medium induced the adhesion of these cells to collagen I polymers but not to monomers. By adding the transglutaminase of blood coagulation, FXIII, in the presence of fibronectin, the attachment of PER cells to collagen I monomers could be recovered while the minimal concentration of fibronectin needed to promote their adhesion to polymers was lowered. These studies indicate that FXIII enhances the fibronectin-mediated attachment of PER cells to collagen I.
Differential expression of murine macrophage surface glycoprotein antigens in intracellular membranes
Authors: Smith Michael J. , Koch Gordon L. E.
Published: April 2021

A monoclonal antibody recognizing a novel murine macrophage glycoprotein antigen (MAG) was prepared by the hybridoma technique. Using live and permeabilized macrophages to estimate surface and total MAG, respectively, it was found that at least 50% of the total antigen was intracellular. In contrast, another macrophage surface glycoprotein antigen, Mac-1, was undetectable in the intracellular pool. Immunofluorescence studies confirmed the existence of a substantial intracellular pool of MAG antigen. Similar results were obtained with a panel of cultured tumour cell lines. In one such cell line, it was shown that surface MAG existed in a distinct punctate pattern indicative of microdomains, whereas surface Mac-1 antigen gave a uniform distribution. The possible role of the surface microdomains in the differential expression of the two surface glycoproteins in intracellular membranes is discussed.
Disassembly of the mammalian metaphase chromosome into its subunits: Studies with ultraviolet light and repair synthesis inhibitors
Authors: Mullinger Ann M. , Johnson Robert T.
Published: April 2021

Metaphase chromosomes of a simian virus-transformed Indian muntjac cell line have been examined by scanning electron microscopy of material in which the fully packed metaphase structure is progressively relaxed. Such chromosomes are seen in standard, spread preparations of ultraviolet light-irradiated, metaphase-arrested cells, which have been incubated in the presence of inhibitors of DNA synthesis; they are processed for electron microscopy by trypsinization, further fixation and osmium impregnation.

Decondensation is initially associated with a gradual elongation and loosening of the chromosome axis and, as loosening proceeds, the appearance of unexpected higher order structures - clusters of 20 –40 nm diameter fibres. The arrangement of the clusters shows much variation between spreads. In the most fully extended chromosomes clusters are arranged in two longitudinal series with pairing between sister chromatids; the diameter of the majority of clusters in such chromosomes is in the range 0 ·4 –0 ·6 μm. In the final stages of decondensation, clusters separate and individual chromosomes are no longer recognizable. Similar fibre clusters are found in interphase nuclei prepared by the same method.

We suggest that the clusters of chromatin fibres may assemble as intermediates in the construction of an axial structure, which is further compacted in the fully condensed metaphase chromosome.
Weak bases partially activate Xenopus eggs and permit changes in membrane conductance whilst inhibiting cortical granule exocytosis
Authors: Charbonneau Michel , Webb Dennis J.
Published: April 2021

At extracellular pH values close to their pKa values the weak bases, ammonia and procaine, elicited a series of events in non-activated Xenopus eggs, some of which resembled those normally occurring at fertilization. These included: (1) a transient increase in membrane conductance; (2) modification of the microvilli; (3) thickening of the cortical cytoplasm and displacement of the cortical granules; (4) pigment accumulation; (5) contractions and shape changes. However, these eggs did not undergo the cortical reaction nor emit the second polar body.

Cortical granule exocytosis of inseminated or artificially stimulated eggs was inhibited if the eggs had been previously treated for 15 min with the weak base and subsequently rinsed. Multiple sperm-entry sites were exhibited by the inseminated eggs, suggesting polyspermy. However, such eggs did not cleave and although sperm had fused with the egg membrane, they were not incorporated. Nevertheless, a transient increase in membrane conductance was evoked, which was longer in duration and had a slightly different form from that normally accompanying fertilization. In these eggs cortical granules were intact but displaced away from the plasma membrane. Prolonged contact with the weak base rendered eggs totally unresponsive to sperm or artificial stimuli but eggs recovered when rinsed sufficiently.

These effects of weak bases on unfertilized Xenopus eggs or during fertilization were completely absent at pH 7·4. Although changes in intracellular pH or Ca2+ may be involved in these phenomena, direct action by the weak base itself cannot be ruled out.
Temperature-induced modifications in size and pattern of microtubular organelles in a ciliate, Dileptus: I. supernumerary microtubules in axonemes of sensory cilia
Authors: Golinska Krystyna
Published: April 2021

Supernumerary microtubules were found in the so-called sensory cilia, in addition to a sensory axoneme. The supernumerary microtubules were not structurally connected to a basal body, but were probably anchored to clusters of dense material inside the ciliary shaft. The frequency of appearance of the supernumerary micro-tubules was found to be temperature-dependent: the higher the temperature during formation of sensory cilia, the greater was the number of supernumerary microtubules in cross-sections, and the more cross-sections contained them.

The possibility is discussed that the formation of the supernumerary microtubules is not due to formation of new nucleating sites inside the cilium. Instead, the microtubules may be remnants of a previously existing axoneme, separated from the basal body during the formation of a sensory cilium. Some of the microtubules of the released axoneme may persist as the super-numerary microtubules, if capped with dense material or some other structure within the ciliary shaft.
Onset of expression of the variant surface glycoproteins of Trypanosoma brucei in the tsetse fly studied using immunoelectron microscopy
Authors: Tetley L. , Turner C. M. R. , Barry J. D. , Crowe J. S. , Vickerman K.
Published: April 2021

The acquisition of the variant surface glycoprotein (variable antigen) coat by metacyclic stage Trypanosoma brucei in the salivary glands of the tsetse fly, Glossina mor si tans, has been studied in situ by transmission and scanning electron microscopy using monoclonal anti-bodies raised against metacyclic variable antigen types and complexed with horseradish peroxidase or colloidal gold. The coat is acquired after binary fission has ceased but while the parasite is still attached to the gland epithelium, i.e. before the mature metacyclic is released into the gland lumen. The variable antigen type heterogeneity previously observed in discharged mature metacyclics is here demonstrated in the nascent (attached) metacyclic population. The variant surface glycoprotein genes are thus not expressed in a fixed sequence since different metacyclic variable antigen types are present ab initio. The distribution of immunogold-marked nascent metacyclics of a particular variable antigen type, as shown by quadrat analysis of a scanning electron micrograph montage of the infected salivary gland epithelium, conforms to a Poisson series. This provides evidence that individual variant surface glycoprotein genes are stochastically activated and suggests that selective activation occurs after trypanosome division has ceased.
Order out of chaos
Authors: Downes C. S.
Published: April 2021

Those who puzzle over the origins of life are perplexed by the improbability of several independent functions arising together, in the absence of either a directing intelligence or a complete, selectable organism. As if to illustrate the inherent limitations of random assembly in producing a coherent whole, the Swedish Academy of Sciences have published, with minimal editing, almost all the papers presented at what must have been a stimulating but perhaps unsatisfactory conference.