bluuebunny/crossref_metadata_2025 huggingface dataset:
Mitosis in Aspergillus Nidulans
Authors: Robinow C. F. , Caten C. E.
Published: April 2021

Mitosis in growing hyphae of haploid and diploid Aspergillus nidulans has been studied with the light microscope in Helly-fixed preparations. At the periphery of the nucleus a sharply defined granule is invariably found which has strong affinity for acid fuchsin. Mitosis begins with the duplication of this granule and the development inside the nucleus of a gradually lengthening fibre between the daughter granules. This fibre is also visible in dividing nuclei of living hyphae examined by phase-contrast microscopy, when it appears as a grey bar traversing the non-nucleolar region. Division of the chromatin has been studied in preparations stained with aceto orcein, directly or after hydrolysis. It proceeds through four phases, beginning with the condensation of distinct chromosomes from the chromatin network of the resting nuclei. Later the chromosomes become arranged in two parallel rows of chromatinic masses in which individual chromosomes can no longer be distinguished. Division is completed by migration of the chromatin to the ends of each row, giving the appearance of a transverse break in the double bar. The chromatin accumulating at the ends of the double bar condenses to form the two daughter nuclei. The number of chromatinic elements appears to be the same in both haploid and diploid nuclei, although the individual elements are larger in the latter.

Successive staining of the same dividing nucleus, first for the fibre (acid fuchsin) and then for the chromosomes (HCl-aceto orcein or HCl-Giemsa) has established that the fibre lies among the chromosomes, and that its elongation is closely related to the anaphase-telophase movements of the chromosomes. This suggests that the fibre is the equivalent of a mitotic spindle. That the fibre is connected with the chromosomes is further suggested by the observa-tion that, in stained preparations, the fibre is considerably thicker in dividing diploid than in haploid nuclei.

Electron microscopy of dividing nuclei has revealed that mitosis is carried out within the intact nuclear envelope. The mitotic spindle is composed of a bundle of fibrils which traverses the nucleus between two plaques of dense material associated with the envelope.

In order to explain the configurations seen during the division of the chromatin it is proposed that the chromosomes become attached to the central spindle at various points along its length. With the elongation of the spindle the sister chromatids separate and pass, asynchronously, to opposite ends of the spindle along two preferred lines, thereby producing the characteristic double bar figure. The absence of a metaphase plate is thus accounted for by the scattered points of attachment of the chromosomes to the spindle and by their asynchronous division.
A study of granule formation in the bat parafollicular cell
Authors: Nunez E. A. , Gould R. P. , Holt S. J.
Published: April 2021

The fine structure of the bat thyroid parafollicular cell has been examined at monthly intervals throughout the hibernating period. During November and December parafollicular cells appear either partly or totally degranulated and intact dense secretory granules are relatively sparse. The degranulated cells exhibit an inconspicuous Golgi complex and relatively few lysosome-type bodies. Few degranulated parafollicular cells are present in thyroid glands from bats collected in January. When found they are characterized by the presence of whorls of cytoplasmic agTanular membranes which enclose a central mass of cellular debris. January bat thyroids are characterized by the presence of three different types of parafollicular cell. One type contains no secretory granules. The cytoplasmic matrix of this type is rich in granular endoplasmic reticulum and free ribosomes and its small Golgi complex consists of several slightly dilated saccules. In close proximity to the Golgi complex are numerous small to medium-sized vesicles which often appear to merge with Golgi elements. Such vesicles are considered to represent the vehicle by which secretory product is transferred from the endoplasmic reticulum to the Golgi complex. The second type of parafollicular cell differs from the first in containing large numbers of intact dense secretory granules. It is also characterized by an extensive Golgi complex which appears to be forming new secretory granules, and by a less extensive granular endoplasmic reticulum. The third type of parafollicular cell shows a structure intermediate between the first two. The cytoplasm of all three types of January parafollicular cells contains many structures belonging to the lysosomal-vacuolar system, including autophagic vacuoles, vacuolated dense bodies and multivesicular bodies. By February and March only parafollicular cells of type 2 are observed. They contain few lysosome-like structures. It is concluded that during mid-hibernation (January), parafollicular cells undergo a series of intracellular changes during which new dense secretory granules are produced. Accompanying granule formation is an augmentation of lysosome-like structures which probably serve as a means of digesting debris from previous secretory cycles.
Fine-structural changes in rat liver microsomes treated with phospholipase c
Authors: Higgins Joan A.
Published: April 2021

Phospholipase C hydrolyses 50% of the phospholipids of rat liver microsomes. Our previous observations suggest that the hydrolysis is of the outer leaflet of the membrane bilayer. In the present investigation the products of hydrolysis and the effect of phospholipase C treatment on the morphology of total, rough and smooth microsomes were investigated. The only product of phospholipase C treatment of microsomes was diglyceride. After hydrolysis of half the phospholipid, the microsomes retained their vesicular structure; however, the average diameter of total and smooth microsomes was reduced by approximately half, and that of rough microsomes by a third. Profiles were observed that suggested that small vesicles may form by pinching off from large vesicles. About 10–15% of the phospholipase-treated microsomes had associated with them an amorphous droplet yielding a ‘signet-ring’-like structure. The rest of the vesicle membrane retained a bilayer structure. About 10–15% of the treated vesicles adhered in groups of two or three. In these groups the shared membrane was trilaminar. Freeze-fracture replicas of phospholipase C-treated microsomes exhibited a similar morphology to untreated microsomes having both concave and convex surfaces, the former exhibiting a greater density of intramembranous particles. These observations suggest that microsomal vesicles remain closed after treatment with phospholipase C and that the bilayer structure is retained. However, there are morphological changes that are possibly related to maintaining the stability of the membranes in which the outer leaflet consists essentially of diglyceride.
Mechanisms of displacement of sperm basic nuclear proteins in mammals. an in vitro simulation of post-fertilization results
Authors: Rodman Toby C. , Pruslin Fred H. , Allfrey Vincent G.
Published: April 2021

A standardized cytological preparation of mature mouse sperm has been devised to serve as an in vitro system for probing the intra-ooplasmic molecular events of transformation of the fertilizing sperm. Two parameters of the early phase of transformation in vivo are defined at the resolution of the light microscope : deletion of sperm-unique nuclear proteins, detectable by immunofluorescence, and retention of homogeneity of the residual DNA complex, with intact chromatin boundaries detectable by ethidium bromide staining. These studies show that both parameters are conserved when in vitro sperm preparations are treated with NaCl under reducing conditions. The deletion of 2 different classes of the unique basic proteins of mouse sperm nuclei is specified by the NaCl concentration : 0·7 M-NaCl displaces the non-protamine class but not the protamines, while 1 M-NaCl displaces both. On the other hand, the effects of treatment with trypsin at various concentrations and intervals are less consistent with the in vivo parameters, indicating fragmentation and displacement, not only of the sperm-unique basic proteins, but also of structural proteins believed to maintain the fundamental cohesive organization of the DNA matrix. These observations suggest that mechanisms other than proteolysis, e.g. localized changes in ionic concentrations, may participate in the post-fertilization displacement of the sperm-unique nuclear proteins in vivo. This study also supports the validity of the in vitro simulation as a model with which to probe the progression of transformation of the sperm nucleus to the zygote pronucleus.
Regional mapping of the gene coding for enolase-2 on human chromosome 12
Authors: Law Martha Liao , Kao Fa-Ten
Published: April 2021

Enolase-2 (ENO2), previously termed 14-3-2 protein, is an isozyme of enolase that is enriched in neuronal tissue. The gene coding for ENO2 was previously assigned to human chromosome 12. The present study presents data for a regional mapping of gene ENO2 using cell hybrids containing various deletions of human chromosome 12. These deletions were produced by treatment with chromosome-breaking agents. Cytogenetic analysis has allowed assignment of ENO2 to the short arm of chromosome 12, in the region of pter-pizos. This assignment is consistent with the segregation pattern of the 93 hybrid clones analysed. The segregation pattern has also established the linear order of 6 genes on chromosome 12: pter — TPI — GAPD — LDHB — ENO2— centromere — SHMT — PEPB — qter. Estimation of the relative distances between the 6 genes on chromosome 12 has been made by a statistical mapping analysis of the segregation data of the hybrid clones. A set of deletion hybrids containing various combinations of these 6 markers has been established for a rapid regional mapping of genes in one of these regions on chromosome 12.
Phylogenetic relationships within the invertebrata in relation to the structure of septate junctions and the development of ‘occluding’ junctional types
Authors: Green Colin R. , Bergquist Patricia R.
Published: April 2021

The structures of 13 variants of invertebrate septate junction are reviewed on the basis of freeze-fracture, lanthanum tracer and thin-section studies. In addition, a simple type of occluding junction in the phylum Porifera, a variation of tight junction in the phylum Tunicata and the vertebrate tight junction are covered. All the junctions considered form a belt around the apical circumference of cells lining a lumen or an exterior surface. The large number of these junctions now recognized permits discussion relating to invertebrate classification and suggested phylogenetic relationships, and to the development of intercellular junctions. The relationships revealed are discussed under three headings: Coelenterates and lower invertebrates, Proterostomia (the annelid, molluscan and arthropod lineage) and the Deuterostomia (the echinoderm and chordate lineage).

It is proposed that the pleated septate junction of the lower invertebrates resembles that of the hydrozoan rather than anthozoan Coelenterates. This lower invertebrate pleated septate junction occurs in several lower invertebrate phyla including the Annelida (of the proterostome lineage), but also occurs in the Sipunculoidea, a group supposedly on the deuterostome lineage. The proterostome line includes the molluscs and the arthropods, which have the molluscarthropod pleated septate junction. Several variations of the smooth septate junction are also seen in Arthropoda. Among the deuterostomes the Chaetognatha have both a paired septate junction and a pleated junction and are therefore considered to be not very far removed from the Sipunculoidea. The echinoderms and hemichordates also have double-septum septate junctions. In addition however, these two phyla have anastomosing septate junctions that are very similar, varying only in their final configuration. Of the two, the echinoderm anastomosing septate junction most closely resembles the tight junction seen in the tunicates, and the Hemichordata are therefore considered to be a lateral development from the main line of chordate evolution. The tunicates have a tight junction similar to that seen in vertebrates; it is however more ‘leaky’ and has distinctive freeze-fracture characteristics.

In the phylum Porifera a form of simple parallel membrane junction appears to serve an occluding function. This junction has regular intercellular spacing in the absence of any septa and it is suggested that the spacing in septate junctions is probably not dictated by the septa. This interpretation is reasonable particularly when the diversity of septal types in conjunction with stable intercellular spacing is considered. Finally, a theory is put forward suggesting that in evolution a change from the septate to the tight junction could simply involve a modification of a ‘membrane spacing factor’, which allows the membranes of adjacent cells to come together at intervals, in the normal tight junction pattern.
The fusion of trout spermatozoa with chinese hamster fibroblasts
Authors: Yip J. H. K. , Bols N. C.
Published: April 2021

Polyethylene glycol (PEG) was used to fuse trout sperm with Chinese hamster fibroblasts (CHW-1102). As judged by light microscopy, PEG significantly increased the number of associations between the two cell types. Electron microscopy revealed that the sperm nuclei were in the cytoplasm of CHW-1102 in cultures that had been treated with PEG and in phagosomes of CHW-1102 in cultures not treated with PEG.

In the cytoplasm many sperm nuclei had an extensive network of fibres whereas before fusion the sperm nuclei contained chromatin blocks which were packed together tightly. In phagosomes they consisted of dispersed blocks of chromatin. The chromatin of the few sperm that were found outside CHW-1100 cells but in the same culture that was treated with PEG also lacked fibres and consisted of loosely packed blocks. Most nuclei were unaltered by PEG treatment alone although in a few the packing of the chromatin blocks was loosened. Thus the decondensation of sperm nuclei within CHW-1102 cells appears to be brought about by the mammalian cytoplasm rather than by digestion in phagosomes, the culture conditions, or the treatment used to induce fusion.
Intake of phosphatase-active plasma membrane into the cytoplasm of oocytes from sexually immature hamsters
Authors: Weakley Brenda S. , Webb Pauline , James John L.
Published: April 2021

When thiamine pyrophosphate is used as substrate, heavily stained complexly branching cytoplasmic structures are seen in developing oocytes of the hamster between 8 and 12 days after birth. This coincides with the onset of the dictyate stage of first meiotic prophase. The structures appear to arise from the oocyte plasma membrane in two ways: (1) by proliferation and infolding of the membrane, and (2) by membrane being pushed into the cytoplasm by invaginating follicle cell processes. This appears to be followed by proliferation of the 1n-vaginated plasma membrane within the oocyte cytoplasm.

Incubations with ATP or β-glycerophosphate, with and without the inhibitor L-p-bromo-tetramisole, indicate that the staining is not due to the action of either ATPase or alkaline phosphatases.

The origin and possible developmental significance of the reactive structures are discussed.
Heterogeneity of the kinetics of proliferation within human skin fibroblastic cell populations
Authors: Azzarone B. , Macieira-Coelho A.
Published: April 2021

Human adult fibroblasts from the papillary layer of the skin showed increased survival in vitro when compared to reticular fibroblasts from the same biopsy. Fibroblasts of the papillary layer were also analysed in vitro to study the kinetics of proliferation between subcultivation and resting phase. It was found that the growth fraction and maximum cell densities vary as a function of the initial inoculum. The results are compatible with the presence of two cell populations with different growth potentials. The heterogeneity of these populations has to be taken into account when studying cellular aging.
Osteoblasts release osteoclasts from calcitonin-induced quiescence
Authors: Chambers T. J.
Published: April 2021

There is an increasing body of indirect evidence that suggests that the localization and activity of osteoclasts, the major agents of bone resorption, are controlled by osteoblasts. In this paper I provide direct evidence that osteoblasts are indeed able to alter the behaviour of osteoclasts. I used calcitonin (CT) to suppress the cytoplasmic activity of osteoclasts isolated on glass or plastic substrates, and found that, while isolated osteoclasts remain quiescent in CT for as long as the hormone is present, they regain activity when osteoblasts are added to the culture. ‘Escape’ from CT-induced quiescence could not be transmitted by supernatants from cultures of osteoblasts, was not due to inactivation of CT by osteoblasts, and did not occur if osteoblasts were separated from osteoclasts by a Millipore filter. When osteoblasts and osteoclasts were cultured together at low cell density in CT, I found that while those osteoclasts in the culture that remained isolated also remained inactive, osteoclasts that were in contact with osteoblasts showed a return to activity. This return to activity was most marked in the portion of osteoclast periphery in direct contact with an adjacent osteoblast. Although contact with osteoblasts did not abrogate the sensitivity of osteoclasts to added calcitonin, osteoclasts were found to escape inhibition by calcitonin at a rate proportional to the number of osteoblasts with which they were in contact.
Intermediate (10 nm) filament proteins and the Ca2+-activated proteinase specific for vimentin and desmin in the cells from fish to man: an example of evolutionary conservation
Authors: Nelson W. James , Traub Peter
Published: April 2021

Cultured cells from representative species of the five main Classes of Chordates (Mammalia, Aves, Reptilia, Amphibia and Pisces) were examined for the presence of intermediate (1o nm) filament proteins. Using a combination of morphological and immunological methods and detailed protein-chemical characterization it is demonstrated that intermediate filament proteins, as shown also for actin and tubulin, are highly conserved during evolution. This conclusion is based on the following observations: (1) intermediate (10 nm) filaments were found in all the cell lines examined from taxonomically distant species; (2) the intermediate filaments were stable in buffers of low ionic strength in the presence of 4 mM-Mg2+ and Triton X-100; (3) under these ionic conditions, detergent-resistant residual cell structures were formed in which the intermediate filament morphology was highly preserved; (4) the intermediate filaments could be completely solubilized in the absence of Mg2+; (5) all intermediate filament proteins were quantitatively precipitated by (NH4)2SO4 at 25% saturation; (6) the intermediate filament proteins, with the exception of some of the proteins from Xenopus and viper cells, all bound to single-stranded DNA-cellulose, and could be eluted at various NaCl concentrations depending on the individual proteins; (7) the intermediate filament proteins had similar molecular weights (50 000 to 58 000), isoelectric points (pl 5·6–5·3) and isoelectric variants, probably due to different degrees of phosphorylation; (8) the major intermediate filament proteins were degraded by the vimentin-specific, Ca2+-activated proteinase and had similar peptide patterns; (9) all the intermediate filament proteins had an antigenic determinant in common and cross-reacted with the monoclonal antibody α-IFA.

In addition, the presence of intermediate filament proteins in taxonomically distant species was indirectly supported by the finding that a Ca2+-activated proteinase activity could be detected in the cells from fish to man. This proteinase had identical characteristics in all the cells examined. It had a molecular weight of 100000, a pl of 5·2, Ca2+ dependence, neutral pH optimum and degraded Ehrlich ascites tumour cell-vimentin with the production of a specific and characteristic peptide pattern. The proteinase was also classified as a thiol-type enzyme. The proteinases from the taxonomically distant species had, therefore, identical properties to the vimentin-specific, Ca2+-activated proteinase purified from Ehrlich ascites tumour cells.
Locomotory invasion of human cervical epithelium and avian fibroblasts by hela cells in vitro
Authors: Stephenson E. M.
Published: April 2021

The locomotory invasive ability of HeLa cells was tested against: (a) embryonic chick heart fibroblasts (CHF); and (b) normal epithelial cells from human cervix (HCE) in explant confrontations. Data for analyses were obtained from replicate cultures fixed 24 h after junction and from 24-h time-lapse films.

The mean invasion index for HeLa versus CHF did not indicate significant obstruction but analyses of hourly radial advance and orientation frequencies showed that obstruction eventually developed as postjunctional incubation time increased. Early contacts between HeLa and CHF demonstrated non-reciprocity of type I contact inhibition of locomotion by the tumour cells, which continued moving in their original direction to underlap contact-inhibited fibroblasts and eventually to occupy spaces vacated by them. When CHF population density increased and free space diminished, HeLa cells displayed directional and probably substratedependent contact inhibition.

The high invasion index of HeLa versus HCE was largely due to occupation of previous HCE territory by tumour cells and only occasionally to actual infiltration of the epithelial sheet. After contact with HeLa, ruffling substrate-adherent marginal epithelial cells displayed contractile, type 1 contact inhibition of locomotion. After orientation changes, they gradually retreated. Against HCE, HeLa cells exhibited non-reciprocity of type I contact inhibition and continued radially forward, following the retreating epithelial margin. They did not move onto exposed upper surfaces of epithelial cells and did not underlap marginal cells firmly adherent to the substratum. Invasion of the epithelial sheet was seen only when initial access beneath a cell with a non-adherent margin was available. The contact relationships of isolated invading HeLa cells with their epithelial neighbours suggested successive non-reciprocal contact inhibition reactions.
Structure of Drosophila polytene chromosomes : Evidence for a toroidal organization of the bands
Authors: Mortin Lawrence I. , Sedat John W.
Published: April 2021

Avoiding acid fixation or squashing, the structure of Drosophila salivary gland polytene chromosomes has been examined in detail in nuclei, with special emphasis on the organization of the DNA in the chromosome bands. Cut serial sections, optical serial sections, scanning electron microscopy (SEM) on whole mounts, high-voltage electron microscopy (HVEM) on whole mounts, and pancreatic DNase I digestion monitored by fluorescent microscopy have been used to complement one another in this analysis. With all five of these techniques, stereo pairs were used to aid in the three-dimensional reconstruction of chromosomal structures.

Evidence is presented that most, if not all, of the polytene chromosome bands are torusshaped. The DNA of these bands is largely confined to the rim, with the interior essentially DNA-free. The chromatin in each polytene band is also seen to have an extremely regular and highly ordered substructure. This substructural organization is largely radially symmetric in the bands and generally parallel to the chromosome axis. In addition, each band appears to be a distinct architectural entity with regard to its exact structural features and dimensions. A model is presented that follows these organizational boundary conditions.
Growth and observations of chinese hamster seminiferous epithelium in vitro
Authors: Ellingson D. J. , Yao K. T. S.
Published: April 2021

Seminiferous tubules from 1- to 3·5-month-old Chinese hamsters were cultivated under dialysis membranes in Rose chambers. The growth and development of the germinal cells was followed daily with phase-contrast microscopy and time-lapse cinemicrography. Spermatogonia lived for 2 or 3 weeks and underwent frequent mitoses. Spermatocytes in metaphase at culture initiation completed their meiotic division. These cells remained healthy for 3–4 days. Such phenomena as germinal cell/Sertoli cell association, nuclear rotation, multinucleated cell formation and spermatid formation were studied and photographed.
Pollen-wall proteins: localization and enzymic activity
Authors: Knox R. B. , Heslop-Harrison J.
Published: April 2021

Cytochemical methods have been used to examine the distribution of acid phosphatase, ribonuclease, esterase, amylase and protease activity and protein in the walls of pollen grains and spores. Enzyme activity was detected in the walls of 50 angiosperm pollens, in pine, and in the spore wall of a species of Equisetum. Activity was absent from the 2 species of ferns examined. The survey covered all major structural pollen types. In all cases enzyme activity was associated principally with the intine, the inner cellulosic part of the wall. In all aperturate grains activity was concentrated in or around the apertural intine; that is, over or near the potential sites of emergence of the pollen tube. Protein concentrations in the intine at these sites could be demonstrated in several pollen types by staining methods and by absorption at 285 nm.

Developmental study showed that the enzymes are incorporated in the intine during the early period of wall growth following the release of the spores from the meiotic tetrads. During this period, stratified ribosomal endoplasmic reticulum lies adjacent to the inner spore wall over the areas of incorporation. In Cosmos bipinnatus, a composite, the material is incorporated as ribbons or leaflets, which interleave with cellulose lamellae. In other species the wall protein may take the form of granules, tubules or vesicles, embedded in the intine cellulose. At maturity the intine is separated from the spore cytoplasm by an intact plasmalemma, so the wall enzymes are to be regarded as being extracellular.

In some species enzyme activity was detected in materials on the surface of the pollen exine derived probably from the surrounding tapetal tissue; however, this was usually insignificant compared with that shown by the intine.

The intine enzymes are very readily leachable, and their function is probably connected with the early nutrition of the pollen tube and the penetration of the stigma. It is likely that the wall proteins make up a very large proportion of mobile protein of the pollen grain, and they may therefore be important in pollen allergenicity.
The development and structure of pyrenoids in Bulbochaete hiloensis
Authors: Retallack B. , Butler R. D.
Published: April 2021

Chloroplasts in zoospore mother cells, zoospores, and sporelings of Bulbochaete are shown to possess spherical granular areas which are interpreted as incipient pyrenoids, which arise de novo within the stroma. Very young pyrenoids are often seen adjacent to pale, fibril-containing areas of the chloroplast. Later, the incipient pyrenoids are often associated with a single thylakoid and become invested with starch grains and cytoplasmic channels as they develop into forms morphologically indistinguishable from the mature pyrenoids of the vegetative cell. In young oogonia, degenerating pyrenoids are seen to possess much starch. Infrequently, the matrix of a degenerating pyrenoid is invaded by the stroma, its dimensions decreasing to those of an incipient pyrenoid. The appearance of young pyrenoid-like structures within lateral outgrowths of the chloroplasts of young oogonia and sporelings may represent a further method of pyrenoid formation.
Relationships between intramembranous particles and surface coat carbohydrates in cells of a compact tissue
Authors: Murphy C. R. , Swift J. G.
Published: April 2021

The structural relationships between intramembranous particles (IMPs) and surface carbohydrates have been studied in cells of a compact tissue — uterine epithelial cells – using an in vivo technique. This involves introducing small amounts of glycerol into the uterine lumen of anaesthetized rats. The treatment results in extensive aggregation of IMPs in the apical plasma membrane of the epithelial cells, but no change in the distribution of several surface carbohydrates could be detected.

Our findings indicate that, in this case, the carbohydrates are not the surface expression of the IMPs, which are generally believed to represent integral membrane proteins. We suggest that the structural relationships between IMPs and surface carbohydrates in the plasma membrane of this compact tissue cell are more similar to those in membranes of free-floating nucleated cells than to those in the much-studied erythrocyte membrane.
Thymidylate kinase from Acetabularia I. Properties of the enzyme
Authors: Groot Egon J. De , Schweiger Hans-Georg
Published: April 2021

The occurrence of a dTMP kinase as well as a dTDP kinase in Acetabularia mediterranea has been demonstrated. A test system was developed by which it was possible to estimate the enzyme activity in an individual Acetabularia cell or even in a cell fragment.

The enzyme catalyses the phosphorylation of dTMP in the presence of ATP. In the test system described, dTTP is formed as well as dTDP. This indicates that there is also a dTDP kinase present in the enzyme preparation. Characteristics of the enzyme such as pH optima at pH 7·0 and 8·75, a temperature optimum at 45 °C, a Km value of 3·3 × 10−6M and a high specificity for ATP were established. In homogenates that were preserved at −70°C the enzyme activity was retained even after many weeks. Freezing at −70°C and then thawing resulted in an increase in enzyme activity. The enzyme was inhibited by low concentrations of dTTP. After centrifugation of homogenates the greater part of the enzyme activity was found in the sediment. From the observation that purified chloroplast preparations contained most of the dTMP kinase activity, and that chlorophyll and enzyme activity cosedimented in a linear sucrose gradient, it was concluded that the enzyme is located in the chloroplasts.
Infection of macronuclear anlagen of Paramecium caudatum with the macronucleus-specific symbiont Holospora obtusa
Authors: Fujishima M. , Gortz H.D.
Published: April 2021

The gram-negative bacterium Holospora obtusa is an endonuclear symbiont of Paramecium caudatum, which is incorporated into the host cells via the food vacuoles and infects their macronucleus exclusively, but never the micronucleus. Since these two kinds of nuclei originate from a fertilization nucleus, it is assumed that the macronucleus acquires a property necessary for it to be recognized by the bacterium at a certain time during the nuclear differentiation process. We found that this property is acquired by four of the eight postzygotic nuclei as soon as the four nuclei differentiate morphologically into the macronuclear anlagen.
The positions of centromeres on the somatic metaphase plate of grasses
Authors: Heslop-Harrison J. S. , Bennett M. D.
Published: April 2021

Centromere positions on the metaphase plate of 48 root-tip cells of four grass species were analysed using metaphases reconstructed from electron micrographs of serial thin sections.

Centromere alignment was almost perfect on a plane in 15 untreated metaphases of cereals where 2n = 14; only 2% of the total variance in centromere position was out of the plane of the metaphase plate. In 23 similar cells pretreated with ice-water, the mean out-of-plate variance was 9%, compared to an expectation of 18% if centromeres were positioned randomly in space. In cold-treated cells of Zea mays (2n = 20), the out-of-plate variance (14%) was significantly less than a random expectation of 20%.

The distances of centromeres from the mean centromere position (MCP) were also analysed. They showed that the centromeres tended to be normally distributed about a circle around the MCP in all the species. Thus centromeres tend towards a ring arrangement in metaphase cells.

Analysis of separation distances of all pairs of centromeres in each reconstructed cell showed that the distribution of distances between centromeres is not random: there are significantly fewer centromeres that are close together and more that are at medium separation distances than is expected on a random basis. This is different from previously published assumptions about centromere disposition in squashed metaphases.
Changes in the synthesis, distribution and sulphation of glycosaminoglycans of cultured human skin fibroblasts upon ascorbate feeding
Authors: Edward Michael , Oliver Roy F.
Published: April 2021

The effect of ascorbic acid on the synthesis, distribution and sulphation of glycosaminoglycans by human skin fibroblasts has been examined. Medium was supplemented with ascorbate over several days, and cultures incubated with [* 3H]glucosamine and Na235SO4 for 48 h, followed by analysis of the glycosaminoglycans in the medium, in collagenase and trypsin extracts, and in cell fractions. Ascorbate feeding resulted in a reduction in hyaluronate synthesis, which was the main 3H-labelled component and was distributed mainly in the medium fractions. Sulphated glycosaminoglycans showed a reduction in incorporation of 3H label, but increased sulphation following ascorbate feeding. In control cultures 53% of 3H-labelled sulphated glycosaminoglycans and 63% of 35S-labelled glycosaminoglycans were present in the medium fraction, while in ascorbate-fed cultures, 41% of 3H label and 38% 35S label were incorporated into medium-sulphated glycosaminoglycans. Ascorbate also caused an increase in cell density and in collagen production and deposition.
Toroidal bands in polytene chromosomes of Drosophila
Authors: Sorsa Veikko
Published: April 2021

Results obtained from the thin-section electron microscopy of salivary gland chromosomes of Drosophila melanogaster mainly support the concept of cable-like organization of polytene chromosomes, with disk-like bands composed of parallel bundles of homologous chromomeres. Outward orientation of loop fibres may generally cause a toroidal bending in the chromomere bundles. Both longitudinal and transverse sections of polytene chromosomes indicate that the bands may contain toroidal subunits. Torus-shaped bands were only found in thin sections of the most distal and most proximal regions, as well as in certain heavy bands at the late-replicating regions of polytenized interphase chromosomes. This suggests that an incomplete duplication of chromomeres may be a reason for torus formation, by preventing the separation of sister chromatids at the earliest phases of the polytenization process. The appearance of more numerous, but smaller, subunits in thin-sectioned faint bands is interpreted as a consequence of more complete segregation of sister chromatids in those bands during polytenization.
Dimethylsulphoxide induction of the murine macrophage-like line p388d1: Change of phagocytic ability and cell surface properties
Authors: Capo Christian , Mishal Zohair , Balloy Vincent , Benoliel Anne-Marie , Bongrand Pierre
Published: April 2021

The murine macrophage-like cell line P388D1 ingests immunoglobulin-coated sheep red cells (IgG-SRC) poorly, but after 3 days incubation in the presence of 1–5% dimethyl sulphoxide (DMSO), it becomes highly phagocytic. We used this model to correlate triggering of phagocytosis with some surface properties of P388D1 cells, possibly involved in recognition or engulfment of particles. The accessibility of Fc receptors on the cell membranes did not seem to be affected by DMSO treatment since the binding of IgG-SRC to cells was the same before and after treatment with DMSO. A technique allowing quantitative determination of the stability of cell-particle binding showed that binding strength was not the only essential factor in triggering ingestion. Hydrophobicity and surface charge were postulated to play a role in phagocytic recognition. No change in hydrophobicity, as assayed by contact-angle measurement, and in net surface charge evaluated by cell electrophoresis, was observed for normal and DMSO-treated P388D1 cells. On the other hand, the fluorescence polarization of a membrane lipophilic probe (1,6-diphenyl-1, 3,5-hexatriene) was significantly (P < 0’01) decreased when P388D1 cells were treated with DMSO. Since the effect of DMSO on P388D1 function was delayed, our results were consistent with the view that enhancement of phagocytosis might be a consequence of some modification of membrane dynamics, due to changes in lipid composition.
Effects of respiratory-burst stimuli on the flow properties of pig blood neutrophils
Authors: Higson F. K. , Kikuchi Y. , Jones O. T. G. , Hughes G. M.
Published: April 2021

Stimulants of the neutrophil respiratory burst, both soluble and crystalline, greatly inhibit the flow of cells through a Nuclepore filter with 5 μm diameter pores. The stimuli appear to reduce cell deformability by a mechanism independent of the activation of respiration, since inhibitors of the respiratory burst (N-phenyl maleimide, quercetin) are without effect on the change. It is suggested that stimuli reduce membrane deformability rather than cause aggregation, since slowing of flow is independent of cell density and is much less when the filter pore size is increased to 8μm. The technique, recently used to study alterations in erythrocyte deformability, may be appropriate for investigating leucocyte flow in some clinical disorders.
Plasma membrane-associated filament systems in cultured cells visualized by dry-cleaving
Authors: Mesland Dick A. M. , Spiele Hermina
Published: April 2021

Substrate-attached critical-point-dried cells cleave along the level of the substrate-adherent membrane if removed by means of adhesive tape. The remaining membrane fragments on grids can be visualized three-dimensionally by means of stereo transmission electron microscopy. Attachment of cells may be achieved by active spreading of the cell, or artificially by poly-L-lysine adherence of prefixed cells. In 11 different cell types a filamentous network appears to remain associated with the cytoplasmic face of the membrane. In one hepatoma cell type virtually no filamentous network could be detected. Two general network morphologies are described: the hepatocytic network and the lymphoid network. Since no correspondence could be found between cytoplasmic structure and the structure of the membrane-associated network, and since cells generally cleave along the level of this network, excluding cell organelles, we conclude that it comprises a distinct structural system, analogous to the membrane skeleton of the red cell membrane.
Prometaphase forces towards opposite spindle poles are not independent: An on/off control system is identified by ultraviolet microbeam irradiations
Authors: Sillers Peggy J. , Wise Dwayne , Forer Arthur
Published: April 2021

Individual spindle fibres in prometaphase spermatocytes of the cricket, Neocurtilla hexadactyla, were irradiated with an ultraviolet microbeam. The stretched heteromorphic bivalent (X2Y) contracted to about 75% of its pre-irradiation length after irradiation of either of its two oppositely directed spindle fibres (i.e., the X2 or Y spindle fibre). The X2Y bivalent also contracted after irradiation of the connection between the kinetochores of the univalent X1 chromosome and the Y chromosome but it did not contract after irradiation of autosomal spindle fibres or of the spindle fibre of the X1 univalent sex chromosome. The spindles sometimes shrank after irradiation, but contraction of the X2Y bivalent was independent of spindle shrinkage.

The data strongly suggest that the oppositely directed forces on a bivalent are not independent. One reason is that the X2Y contractions were asymmetrical: during contraction one kinetochore remained fixed in position while the other kinetochore (facing the opposite pole) moved towards the equator. In 75% of the cases the stationary kinetochore was associated with the irradiated spindle fibre. Thus, we suggest that the irradiation of a spindle fibre produces a state analogous to rigor in the irradiated spindle fibre and, because of effects on the control system, produces relaxation of tension in the oppositely directed non-irradiated spindle fibre, so that the kinetochore associated with the non-irradiated spindle fibre moves towards the equator. These experiments have identified a control system that coordinates force production to opposite poles.
Anchoring filaments of the amphibian epidermal-dermal junction traverse the basal lamina entirely from the plasma membrane of hemidesmosomes to the dermis
Authors: Ellison Janice , Garrod D. R.
Published: April 2021

An electron microscopical study of the epidermal-dermal junction in the axolotl and adult Rana pipiens has been carried out. This shows that filaments of about 12 nm in diameter, known as anchoring filaments, pass from the hemidesmosomes at the base of the epidermal cells across the basal lamina to the dermis. There they may unite to form broader fibres, known as anchoring fibrils, or may simply form bundles.

In the axolotl, particularly, the anchoring fibrils or bundles of anchoring filaments, enmesh with the collagen fibres of the dermis.

Removal of epidermal cells with EDTA results in separation along a plane in the lamina rara of the basal lamina, i.e. between the plasma membrane of the cells and the lamina densa. The anchoring filaments remain inserted into the lamina densa. Hemidesmosomal plaques are no longer visible in regions of the plasma membrane that have been separated from the basal lamina by EDTA, and no evidence was found that plaques are engulfed by the cells.

It is proposed that the hemidesmosome-anchoring filament system provides a structural link between the collagenous filament system of the dermis and the intracellular cytokeratin filament system of the epidermis, which, in turn, is linked between cells by desmosomes.
Pollen-pistil interactions in interspecific crosses of Populus (sections aigeiros and leuce): Pollen adhesion, hydration and callose responses
Authors: Gaget M. , Saï C. , Dumas D, C. , Knox R. B.
Published: April 2021

Intersectional crosses between species of sections Aigeiros (Populus deltoides, P. × euramericana or P. nigra) and Leuce (P. alba or P. tremuloides) are known to be reciprocally incompatible. The site of pollen tube arrest is on the stigma surface in pollinations between pistils of section Leuce and pollen of section Aigeiros; tubes failed to penetrate the stigma surface. In reciprocal matings, pollen of section Leuce germinated and tubes penetrated the stigma and style, where arrest occurred. Rejection may be accompanied by swelling of the tube tips, and callose plug formation. In the cross between P. deltoides and P. alba a callose response was detected in the cell walls of the transmitting tissue, adjacent to the rejected pollen tubes.
Centrioles in the mammary epithelium of the rat
Authors: Dylewski Daniel P. , Keenan Thomas W.
Published: April 2021

Through serial thin-section analysis of rat mammary epithelial cells, the number of centrioles per cell and their intracellular location were determined. In all developmental stages (e.g. virginal, pregnant, lactation, involution), each epithelial cell contained a single centriole that was located in the apical region. Centrioles were 200-220 (ݲ = 210) nm in transverse section, and exhibited the typical 9 × 3 ‘pinwheel’ configuration of microtubules. In longitudinal section, centrioles were 330—380 (ݲ = 360) nm in length. Each centriole was surrounded by a homogeneous pericentriolar matrix. During mitosis in pregnant animals, centrioles were paired at the nuclear poles and oriented at right angles (90°) to each other. At the completion of mitosis a single diploβome (pair of centrioles) was associated with each interphase nucleus. Because all postmitotic cells contained only a single centriole, it was assumed that one of the two diplosomal centrioles had disintegrated. There appeared to be a correlation between centriole location and cell polarity. When centrioles were located near the apical plasma membrane, epithelial cells exhibited polarity. However, when centrioles were associated with the nuclear poles during mitosis, epithelial cells were typically apolar. These observations suggest that centrioles may function as determinants in cell polarity.
Acquisition of a limited lifespan by differentiating cells derived from pc13 embryonal carcinoma cells
Authors: Rayner Michael J. , Pulsford James A. J.
Published: April 2021

Retinoic acid (RA) has previously been shown to induce the differentiation of mouse embryonal carcinoma (EC) cells to endoderm-like cells that have a slower rate of proliferation and are non-tumorigenic. These cells also acquire the ability to respond to a range of exogenous growth factors. Wc have analysed the change in growth phenotype for PC13 EC cells using video recordings and autoradiography. We have shown that the endoderm-like cells have a longer cell cycle time than their undifferentiated counterparts (five cell divisions after exposure to RA the differentiated cells had a median cell cycle time of 1800 min compared to 800 min for control cells). The endoderm-like cells also have a progressively decreasing probability of dividing again and this indicates that the differentiation process is accompanied by the acquisition of a limited life-span. The characteristics of mortal cells are well documented, and the endoderm-like cells demonstrate the properties of such cells.

In addition, we have confirmed the observation that epidermal growth factor (EGF) can stimulate the proliferation of the endoderm-like cells and have shown, using autoradiography, that 92% of these cells express EGF receptors. Using video recordings, we have demonstrated that the effect of EGF is to shorten the cell cycle of the differentiating cells. We have also shown that EGF can enhance the survival of the endoderm-like cells and thereby prolong their life-span. It is known that EGF and other growth factors can prolong the life-span of mortal cells derived from normal tissues, but we have demonstrated that EGF can have this effect on the differentiated derivatives of a tumour cell
The associated reactivation of two x-linked genes the spontaneous and azacytidine-induced re-expression of ornithine transcarbamoylase and glucose-6-phosphate dehydrogenase in a rat hepatoma
Authors: Goss S. J.
Published: April 2021

Two X-linked genes, specifying ornithine transcarbamoylase (OTC) and glucose-6-phosphate dehydrogenase (Glc-6-PD), are reversibly suppressed in certain derivatives of the rat H4-II-E hepatoma. Either gene can become reactivated spontaneously, and it is shown that both can be reactivated by azacytidine treatment. This gene reactivation has been investigated by enzyme assay and by the use of selective growth media (‘ornithine-medium’ to select for OTC, and medium containing diamide to select for Glc-6-PD). There is a clear tendency for both genes to be reactivated together, though either can become active alone. Since OTC is an enzyme of the urea-cycle, and Glc-6-f*D is an enzyme of the hexose monophosphate shunt, and since these two pathways are normally under quite separate control, it would seem that the coupled regulation of the two genes in these hepatomas is abnormal. It is suggested that the suppression of the two genes resembles X-inacti vation: in both cases, azacytidine treatment induces gene reactivation with a high frequency and results in different clones of cells expressing widely varying amounts of enzyme activity. The association between the re-expression of OTC and Glc-6-PD might indicate that some phenomenon like the position-effect is occurring.