bluuebunny/crossref_metadata_2025 huggingface dataset:
Rotary movements and fluid membranes in termite flagellates
Authors: Tamm S. L. , Tamm Signhild
Published: April 2021

We previously described a remarkable type of cell motility that provided direct, visual evidence for the fluid nature of cell membranes. The movement involved continual, unidirectional rotation of one part of a protozoan, including the plasma membrane and cytoplasmic organelles, in relation to a neighbouring part. The cell membrane in the ‘shear zone’ appeared continuous with that of the rest of the cell. The rotary motor consisted, at least in part, of a non-contractile, microtubular axostyle which extended centrally through the cell. The protozoan was a devescovinid flagellate found in the hindgut of a Florida termite.

In this paper, we have confirmed earlier reports of this type of motility in other kinds of devescovinids from Australian termites. By demonstrating continuity of the plasma membrane in the shear zone of the Australian devescovinids as well, we have obtained additional examples that provide direct, visual evidence for fluid membranes. A comparative analysis of rotational motility in various devescovinids revealed 2 different kinds of rotary mechanisms. Hyperdevescovina probably have an internal motor, in which one part of the cell exerts forces against another part, as in the Florida termite devescovinid. Devescovina species, on the other hand, have an external motor, in which flagellar and/or papillar movements exert forces against the surrounding medium. The structure of the axostyle in different devescovinids was compared, and its role in rotational motility discussed with respect to the behavioural data.
Flagellar regeneration in Chlamydomonas reinhardtii: evidence that cycloheximide pulses induce a delay in morphogenesis
Authors: Farrell K. W.
Published: April 2021

The behaviour of a pool of flagellar precursors, assayed by the ability of cells to regenerate flagella in the absence of de novo protein synthesis, has been examined during organelle morphogenesis in the biflagellate alga Chlamydomonas. The results demonstrate that flagellar elongation can continue even when this pool is apparently empty and suggest that 2 sources of precursors are available to the regenerating flagella: those pre-existing in the cellular pool and those synthesized de novo. Further evidence for this was obtained by subjecting regenerating cells to pulses of cycloheximide. Cells exposed to this drug during the first 60 min post deflagellation formed only half-length (5-μm) flagella, whereas a pulse administered after this point allowed the formation of longer flagella and suggested that some de novo protein synthesis was required for the formation of full-length flagella, although it was not a prerequisite for the initiation of regeneration. In addition, it was found that, subsequent to the removal of the cycloheximide, flagellar regeneration did not recommence immediately, but was delayed for a period of approximately 45 min, irrespective of length of flagella formed prior to drug inhibition.

The nature of this cycloheximide-induced delay is unclear and certain alternatives, based on the exhaustion of structural/regulatory components are considered. Although it is not possible to distinguish between these alternatives, tubulin is not the limiting component, since a pool of this protein is present when flagellar elongation is prevented by cycloheximide.
Cell organelles at uncoated cryofractured surfaces as viewed with the scanning electron microscope
Authors: Woods P. S. , Ledbetter Myron C.
Published: April 2021

A method of direct visualization of cell organelles by scanning electron microscopy (SEM) is described. Plant and animal tissues fixed in glutaraldehyde and osmium tetroxide are treated with the ligand thiocarbohydrazide and a second osmium tetroxide solution, to increase their osmium content. Tissues are then dehydrated, infiltrated with an epoxy monomer, and together solidified with dry ice and fractured. The pieces are transferred to pure acetone, critical-point dried, attached to stubs with silver paint and viewed by SEM. The ligating procedure increases the osmium concentration at its original bonding site sufficiently to render the tissue electrically conductive, thus obviating the need for metallic coating. The organelles at the fractured surface are revealed in relation to their osmium incorporation rather than by surface irregularities as with coating methods. The image derived from the uncoated surface approaches in resolution that of transmission electron micrographs of thin sections. A portion of the image arising from a small distance below the surface, while at progressively lower resolution, provides some 3-dimensional information about cell fine structure.
The crystalline glycoprotein cell wall of the green alga Chlorogonium Elongatum: a structural analysis
Authors: Roberts K. , Hills G. J.
Published: April 2021

Members of the Chlamydomonaceae, mostly single-celled green algae, have been shown to contain a crystalline glycoprotein cell wall component. Most of the species examined fall into a class of algae whose walls have an identical crystalline unit cell. Chlorogonium elongatum has been chosen as a representative of this class in order to investigate in more detail its cell wall structure. The alga has a spindleshaped cell wall which retains its asymmetric shape on isolation. Sections from walls fixed in the presence of tannic acid clearly reveal a regular subunit monolayer, about 20 nm thick, within the wall. Sodium dodecylsulphate (SDS) polyacrylamide gel electrophoresis shows the presence of at least 2 major glycoprotein species in the wall. Negatively stained purified cell walls demonstrate the crystalline nature of the cell wall. Optical diffraction of bright-field images and direct electron diffraction both give clear diffraction patterns whose spacings extend out to 3 nm and fall on a reciprocal lattice whose vectors describe a 2-dimensional unit cell within the wall 2 · 15 nm × 7 · 0 nm and an included angle of 80°. Lattice defects within the cell wall are revealed by both negative staining and surface replication. Through-focal series were used to choose images with the optimal degree of underfocus for image processing. Linear integration and optical filtering of such images gave essentially the same result. A similar image was also obtained by computing the autocorrelation function of the amplitudes in the electron-diffraction pattern and the optical-diffraction pattern of the in-focus image. On the basis of these data a 2-dimensional model of the crystalline cell wall layer is presented.
Some effects of positively charged surface groups on cell aggregation
Authors: Maslow D. E. , Weiss L.
Published: April 2021

A study was made of the effects of 2,3-dimethylmaleic anhydride (DMA), a reagent removing positive charges, on the aggregation and surface charge of embryonic chick neural retina cells. Neural retina cells, recovered from the dissociation procedure, were cultured on a gyratory shaker and the aggregate diameters formed in the presence of DMA or DMA-serum dialysate, following DMA-pretreatment, or in appropriate control cultures measured. The electrophoretic mobilities of similarly treated cells were also determined. In addition, cellulose acetate electrophoresis was carried out on samples of serum containing DMA, and the incorporation of 14C-amino acids into DMA-treated cells studied.

Aggregates formed in the presence of DMA, or following DMA-pretreatment, were significantly smaller than aggregates from control cultures. The electrophoretic mobility of DMA-treated cells was significantly increased in serum-containing medium, but not serum-free Hanks’ solution. At 24 h after removal of DMA-containing medium, the mobilities of pretreated cells were similar to those of controls.

The electrophoretic pattern of DMA-treated serum was changed only with concentrations of DMA many times that affecting cell aggregation or mobility. DMA-serum dialysate did not significantly reduce aggregate size. The incorporation of 14C-amino acids in DMA-treated cells and the structure of aggregates were unchanged from controls.

It is concluded that positively charged constituents of the cell periphery play a demonstrable, but not limiting, role in cell aggregation, while a minor role for positive charges on serum protein cannot be totally excluded.
The actions of restriction endonucleases on lampbrush chromosomes
Authors: Gould D. C. , Callan H. G. , Thomas C. A.
Published: April 2021

Lampbrush chromosomes from oocytes of Notophthalmus viridescens were dispersed in media containing restriction endonucleases isolated from Haemophilus and E. coli. These endonucleases cleave duplex DNAs at specific palindromic sequences of nucleotides, and several sensitive sites occur per micron of DNA. The overwhelming majority of the lateral loops of lampbrush chromosomes are extensively fragmented by these endonucleases, but an occasional pair of loops is refractory. A notable example of loops showing this refractory property are the giant loops on chromosome II in the presence of Hae. These loops, whose DNA-containing axes are several hundred micra long, are sensitive to other nucleases such as EcoB, endonuclease I and pancreatic DNase I; their refractory behaviour towards Hae therefore indicates that the sequence sensitive to this particular endonuclease is systematically absent. This anomalous property can be comprehended if it be assumed that the axial DNA of the giant loops consists of tandem repeats of a sequence which happens not to include the sensitive site.
Synthesis of the inner mitochondrial membrane and the intercalation of respiratory enzymes during the cell cycle of Chlorella
Authors: Forde B. G. , Gunning B. E. S. , John P. C. L.
Published: April 2021

The ratio of inner to outer mitochondrial membrane area remains close to 1·8 throughout the cell cycle in synchronized cells of Chlorella fusca var. vacuolata 211-8p. Using estimates of this ratio, together with our previous estimates of mitochondrial surface area, to calculate the absolute area of inner mitochondrial membrane, it is demonstrated that growth of the inner mitochondrial membrane during the cell cycle occupies an extended period and parallels the growth of the whole cell. In contrast, the synthesis of succinate dehydrogenase and cytochrome oxidase is restricted to the last third of the cell cycle. It is concluded that mitochondrial growth involves the intercalation of periodically synthesized respiratory enzymes into membranes made earlier in the cycle, with consequent 5-fold changes in the density of active enzyme molecules in the membrane. These observations are discussed in relation to the control of mitochondrial membrane synthesis, membrane assembly and respiration rate during the cell cycle.
45Calcium localization in islets of langerhans, a study by electron-microscopic autoradiography
Authors: Howell S. L. , Tyhurst Margaret
Published: April 2021

Attempts were made to localize the sites of uptake of 45calcium in B cells of islets of Langerhans by electron-microscopic autoradiography. Despite the potential sources of error inherent in the partial loss of radioactivity during fixation, and in the relatively high energy of emission of this isotope, distribution of silver grains differed significantly from random in all experiments. Grains were concentrated over mitochondria and to a lesser extent over storage granules. Incubation of islets in the presence of 10 mM glucose and isobutylmethyl-xanthine before fixation and autoradiography resulted in a small but not statistically significant reduction in silver grains associated with the mitochondria. These results further indicate a dominant role of mitochondria in the regulation of cytosolic calcium concentrations in pancreatic B cells.
Spindle birefringence of isolated mitotic apparatus: further evidence for two birefringent spindle components
Authors: Forer Arthur , Kalnins V. I. , Zimmerman A. M.
Published: April 2021

We studied sea-urchin zygote mitotic apparatus (MA) isolated in hexylene glycol, transferred immediately to a glycerol-dimethylsulphoxide medium, and stored for 2 weeks at room temperature. Treatment with 0·5 M KC1 caused loss of 45% of the birefringence, but microtubules remained intact (as seen electron microscopically in glutaraldehyde-fixed MA), and tubulin was not extracted (as determined by polyacrylamide gel electrophoresis). These results suggest that a non-tubulin component which is extracted by the KC1 contributes 45% of the MA birefringence.

Further evidence for this conclusion came from indirect immunofluorescence experiments. Non-extracted (control) MA were fixed with formaldehyde and reacted with antibody against tubulin: there was intense staining of the spindle fibres and astral rays. Electron microscopically, however, microtubules were not present in formaldehyde-fixed MA. Since formaldehyde fixation caused breakdown of microtubules but the tubulin remained in the MA (as judged by reaction with antibodies) we suggest that after microtubule breakdown the tubulin remains in the MA because it is bound to a peri-microtubule spindle component (which we call ‘substance γ’). When KCl-extracted MA were fixed with formaldehyde and reacted with antibody against tubulin there was very little staining of spindle fibres and astral rays. Electron microscopically, formaldehyde caused microtubule breakdown, and since the tubulin is lost from formaldehyde- fixed, KCl-extracted MA (as judged by reaction with antibodies), we suggest that the tubulin- binding component, substance γ, is extracted by the 0·5 M KCl.

Pressure treatment caused the asters not to stain with antibody against tubulin, suggesting that the stability of substance γ is different in different regions of the mitotic apparatus.
The ordered columnar structure of mouse filiform papillae
Authors: Hume William J. , Potten Christopher S.
Published: April 2021

Examination of carefully oriented 1-μm and 5-μm sections of mouse dorsal tongue together with scanning electron microscope observations indicates a high degree of cellular organization in the papillae. It has been suggested that each filiform papilla consists of 2 dominant and 2 minor columns of cells. Labelling patterns of the basal cells have been investigated in relation to these columns.
Synthesis of a liver enzyme in hybrid cells
Authors: Riddle V. G. H. , Harris H.
Published: April 2021

Rat hepatoma cells were fused with cells of an established mouse lymphoma line, with normal diploid mouse macrophages, lymphocytes and fibroblasts and with normal diploid rat macrophages and lymphocytes. The liver-specific enzyme tyrosine aminotransferase was produced by almost all the hybrid cells, but usually at a lower level than in the parental hepatoma cells. Most of the hybrids also showed increased levels of this enzyme after exposure to dexamethasone. In the rat x mouse hybrids, the electrophoretic mobility of the enzyme indicated that only the rat hepatoma enzyme was produced. The findings are difficult to explain in terms of simple models involving a single diffusible repressor or activator of tyrosine aminotransferase synthesis.
Cyclic adenosine monophosphate and the development of polysphondylium
Authors: Jones M. E. , Robertson A.
Published: April 2021

Centre formation in Polysphondylium violaceum is delayed for 2 h on buffered agar containing 10−3 M c-AMP, and for up to 22 h on unbuffered agar with the same c-AMP concentration. With ambient c-AMP concentrations as low as 10−8 M, P. pallidum forms numerous, small, atypical aggregates which do not fruit. This effect is independent of whether the agar is buffered.

P. violaceum amoebae are weakly attracted to the tip of a microelectrode containing 10−3 or 10−4 M c-AMP, but the electrode cannot compete when natural centres form nearby. P. pallidum amoebae are not attracted.

Aggregates of P. violaceum and of P. pallidum are strongly attracted to a microelectrode releasing c-AMP. The observation of Shaffer that Polysphondylium grex switch over from secreting an acrasin that attracts homologous amoebae to one that attracts the larger Dictyo- stelium species suggests that the second acrasin might be c-AMP. The above results strengthen this conjecture.

As c-AMP inhibits centre formation, the secretion of c-AMP by older aggregates may explain an inhibition of centre formation in the ‘ overlay’ experiments of Shaffer.
A theoretical study of the effects of cyclic amp phosphodiesterases during aggregation in Dictyostelium
Authors: Nanjundiah Vidyanand , Malchow Dieter
Published: April 2021

During aggregation the larger Dictyostelium species use cAMP as a chemoattractant and possibly also as a transmitter. In passage from cell to cell, cAMP levels are modulated by diffusion and by enzyme hydrolysis. It appears that the important cAMP-hydrolysing enzyme is a phosphodiesterase bound to the cell membrane, the main roles of which are (1) very fast hydrolysis of cAMP and (2) steepening of spatial cAMP gradients. An extracellular phosphodiesterase has no function, so far as can be conjectured from present data.
Differentiation and positioning of nuclei in eggs of the cecidomyid Heteropeza pygmaea
Authors: Meats Marian , Tucker J. B.
Published: April 2021

During the first three cleavage divisions of the egg nuclei a precise sequence of spindle orientation and elongation parallel to the longitudinal axis of the egg is apparently involved in positioning one nucleus among the polar granules at the posterior pole of the egg. The size of this nucleus, and the position at which the egg cleaves when pole cell formation occurs, appear to constitute part of the mechanism which ensures that only one nucleus is included in the first pole cell.

Blastoderm formation occurs without a well-defined migration of nuclei to the egg surface. Nuclei are so large in relation to the size of the egg that uniform spacing and distribution of nuclei ensures that a large proportion are situated near the egg surface. Those nuclei which are near the egg surface divide synchronously to form a layer of blastoderm nuclei, while membranous cleavage furrows invaginate from the egg surface between them. Nuclei in the central region of the egg chamber condense to form yolk nuclei before blastoderm nuclei have been separated from the rest of the egg by the completion of the cleavage membranes.

Polar granules provide the only evidence of fine-structural differences in different regions of the egg chamber cytoplasm. They are found near the posterior pole of the egg from an early stage of oogenesis. They undergo a specific sequence of structural changes and increase in size as the egg grows. No microtubular or microfibrillar arrays have been found in the egg chamber which might form a cytoskeletal basis for spindle orientation or for the spatial differences which develop during differentiation of the uncleaved egg cytoplasm.
Conformational constraints in nuclear dna
Authors: Cook P. R. , Brazell I. A.
Published: April 2021

We have investigated DNA superstructure in a wide range of nuclei of higher cells by gently lysing cells to release structures that resemble nuclei but are depleted of nuclear proteins. The sedimentation properties of these structures, which we call nucleoids, have been examined in sucrose gradients containing the intercalating agent, ethidium. The sedimentation rate of nucleoids derived from the growing cells of mammals, birds, amphibians and insects varies in the manner characteristic of circular and superhelical molecules of DNA. These characteristic changes in sedimentation rate are abolished by irradiating the nucleoids with low doses of γ-rays, a procedure known to introduce single-strand scissions into DNA. We have also investi gated by similar means DNA superstructure in nucleoids derived from a variety of different chick cells. Nucleoids derived from adult hen erythrocytes differ from the other nucleoids studied in that their sedimentation rate does not vary in the manner characteristic of super-coiled DNA.
Characterization of nuclear structures containing superhelical DNA
Authors: Cook P. R. , Brazell I. A. , JOst E.
Published: April 2021

Structures resembling nuclei but depleted of protein may be released by gently lysing cells in solutions containing non-ionic detergents and high concentrations of salt. These nucleoids sediment in gradients containing intercalating agents in a manner characteristic of DNA that is intact, supercoiled and circular. The concentration of salt present during isolation of human nucleoids affects their protein content. When made in 1·95 M NaCl they lack histones and most of the proteins characteristic of chromatin; in 1·0 M NaCl they contain variable amounts of histones. The effects of various treatments on nucleoid integrity were investigated.
Purification of gibberellic acid-induced lysosomes from wheat aleurone cells
Authors: Gibson R. A. , Paleg L. G.
Published: April 2021

Using isopycnic density gradient centrifugation, lysosomes were concentrated in a single region of a sucrose-Ficoll gradient (ρ = 1·10 g cm−3), well separated from most other cell organelles. Gibberellic acid-induced lysosomes were found to be rich in α-amylase and protease but not ribonuclease. The lysosomal band also contained a majority of the NADH2-cytochrome c reductase, a marker enzyme for endoplasmic reticulum, found in the gradient. Examination of electron micrographs revealed that a purified band of lyosomes contained at least 3 vesicle types, ranging in size from 0·1 to 0·5 μm. The significance of these findings to proposed mechanisms of action of gibberellic acid is discussed.
Barium accumulation in rat pancreatic B cells
Authors: Howell S.L. , Tyhurst Margaret
Published: April 2021

Barium has been used as an electron-opaque substitute for calcium in a study of the distribution of divalent cations between organelles in homogenates or intact rat islets of Langerhans. These were incubated in the presence of barium acetate. Accumulation of electron-opaque deposits was stimulated during incubation of islets in the presence of high glucose concentrations and was diminished in conditions in which intracellular cyclic AMP levels were raised. Mitochondria were found to be the principal sites of accumulation of electron-opaque deposits. Addition of dinitrophenol to homogenates or intact islets abolished mitochondrial barium accumulation. X-ray microanalysis of the deposits in frozen sections showed them to consist predominantly of barium and phosphate.

These experiments serve to emphasize further the critical role of mitochondria in the regulation of divalent cation accumulation in B cells, and to confirm that a direct effect on intra-cellular distribution of divalent cations may represent one important mechanism of action of cyclic AMP in regulating insulin secretion.
Molecular regulation of Snai2 in development and disease
Authors: Zhou Wenhui , Gross Kayla M. , Kuperwasser Charlotte
Published: April 2021

The transcription factor Snai2, encoded by the SNAI2 gene, is an evolutionarily conserved C2H2 zinc finger protein that orchestrates biological processes critical to tissue development and tumorigenesis. Initially characterized as a prototypical epithelial-to-mesenchymal transition (EMT) transcription factor, Snai2 has been shown more recently to participate in a wider variety of biological processes, including tumor metastasis, stem and/or progenitor cell biology, cellular differentiation, vascular remodeling and DNA damage repair. The main role of Snai2 in controlling such processes involves facilitating the epigenetic regulation of transcriptional programs, and, as such, its dysregulation manifests in developmental defects, disruption of tissue homeostasis, and other disease conditions. Here, we discuss our current understanding of the molecular mechanisms regulating Snai2 expression, abundance and activity. In addition, we outline how these mechanisms contribute to disease phenotypes or how they may impact rational therapeutic targeting of Snai2 dysregulation in human disease.
On the asymmetric partitioning of nucleocytoplasmic transport – recent insights and open questions
Authors: Kalita Joanna , Kapinos Larisa E. , Lim Roderick Y. H.
Published: April 2021

Macromolecular cargoes are asymmetrically partitioned in the nucleus or cytoplasm by nucleocytoplasmic transport (NCT). At the center of this activity lies the nuclear pore complex (NPC), through which soluble factors circulate to orchestrate NCT. These include cargo-carrying importin and exportin receptors from the β-karyopherin (Kapβ) family and the small GTPase Ran, which switches between guanosine triphosphate (GTP)- and guanosine diphosphate (GDP)-bound forms to regulate cargo delivery and compartmentalization. Ongoing efforts have shed considerable light on how these soluble factors traverse the NPC permeability barrier to sustain NCT. However, this does not explain how importins and exportins are partitioned in the cytoplasm and nucleus, respectively, nor how a steep RanGTP–RanGDP gradient is maintained across the nuclear envelope. In this Review, we peel away the multiple layers of control that regulate NCT and juxtapose unresolved features against known aspects of NPC function. Finally, we discuss how NPCs might function synergistically with Kapβs, cargoes and Ran to establish the asymmetry of NCT.
RACK1 plays a critical role in mast cell secretion and Ca2+ mobilization by modulating F-actin dynamics
Authors: Filho Edismauro G. Freitas , da Silva Elaine Z. M. , Ong Hwei Ling , Swaim William D. , Ambudkar Indu S. , Oliver Constance , Jamur Maria Célia
Published: April 2021

Although RACK1 is known to act as a signaling hub in immune cells, its presence and role in mast cells (MCs) is undetermined. MC activation via antigen stimulation results in mediator release and is preceded by cytoskeleton reorganization and Ca2+ mobilization. In this study, we found that RACK1 was distributed throughout the MC cytoplasm both in vivo and in vitro. After RACK1 knockdown (KD), MCs were rounded, and the cortical F-actin was fragmented. Following antigen stimulation, in RACK1 KD MCs, there was a reduction in cortical F-actin, an increase in monomeric G-actin and a failure to organize F-actin. RACK1 KD also increased and accelerated degranulation. CD63+ secretory granules were localized in F-actin-free cortical regions in non-stimulated RACK1 KD MCs. Additionally, RACK1 KD increased antigen-stimulated Ca2+ mobilization, but attenuated antigen-stimulated depletion of ER Ca2+ stores and thapsigargin-induced Ca2+ entry. Following MC activation there was also an increase in interaction of RACK1 with Orai1 Ca2+-channels, β-actin and the actin-binding proteins vinculin and MyoVa. These results show that RACK1 is a critical regulator of actin dynamics, affecting mediator secretion and Ca2+ signaling in MCs.

This article has an associated First Person interview with the first author of the paper.
Control of multiciliogenesis by miR-34/449 in the male reproductive tract through enforcing cell cycle exit
Authors: Wu Yu-Jie , Liu Yue , Hu Yan-Qin , Wang Li , Bai Fu-Rong , Xu Chen , Wu Jing-Wen
Published: April 2021

Multiciliated cells (MCCs) are terminally differentiated postmitotic cells that possess hundreds of motile cilia on their apical surface. Defects in cilia formation are associated with ciliopathies that affect many organs. In this study, we tested the role and mechanism of the miR-34/449 family in the regulation of multiciliogenesis in EDs using an miR-34b/c−/−; miR-449−/− double knockout (dKO) mouse model. MiR-34b/c and miR-449 depletion led to a reduced number of MCCs and abnormal cilia structure in the EDs starting from postnatal day (P)14. However, abnormal MCC differentiation in the dKO EDs could be observed as early as P7. RNA-seq analyses revealed that the aberrant development of MCCs in the EDs of dKO mice was associated with the upregulation of genes involved in cell cycle control. Using a cyclin-dependent kinase inhibitor to force cell cycle exit promoted MCC differentiation, and partially rescued the defective multiciliogenesis in the EDs of dKO mice. Taken together, our results suggest that miR-34b/c and miR-449 play an essential role in multiciliogenesis in EDs by regulating cell cycle exit.
An ancient glaucophyte c6-like cytochrome related to higher plant cytochrome c6A is imported into muroplasts
Authors: Kleiner Friedrich Hans , Vesteg Matej , Steiner Jürgen Michael
Published: April 2021

Cytochrome c6 is a redox carrier in the thylakoid lumen of cyanobacteria and some eukaryotic algae. Although the isofunctional plastocyanin is present in land plants and the green alga Chlamydomonas reinhardtii, these organisms also possess a cytochrome c6-like protein designated as cytochrome c6A. Two other cytochrome c6-like groups, c6B and c6C, have been identified in cyanobacteria. In this study, we have identified a novel c6-like cytochrome called PetJ2, which is encoded in the nuclear genome of Cyanophora paradoxa, a member of the glaucophytes – the basal branch of the Archaeplastida. We propose that glaucophyte PetJ2 protein is related to cyanobacterial c6B and c6C cytochromes, and that cryptic green algal and land plant cytochromes c6A evolved from an ancestral archaeplastidial PetJ2 protein. In vitro import experiments with isolated muroplasts revealed that PetJ2 is imported into plastids. Although it harbors a twin-arginine motif in its thylakoid-targeting peptide, which is generally indicative of thylakoid import via the Tat import pathway, our import experiments with isolated muroplasts and the heterologous pea thylakoid import system revealed that PetJ2 uses the Sec pathway instead of the Tat import pathway.
Physiological requirements for ciliary reactivation of bracken fern spermatozoids
Authors: Wolniak Stephen M. , Cande W. Zacheus
Published: April 2021

Physiological parameters affecting reactivated ciliary beat in spermatozoids of bracken fem (Pteridium aquilinum) were studied using a Triton/glycerol permeabilized cell model system. Reactivation frequencies of polylysine-tethered cells equalled in vivo rates at neutral pH. Frequency was dependent on ATP and Mg!+ concentration, and reactivation was inhibited by millimolar or greater free calcium. Reactivation was reversibly inhibited by micromolar concentrations of sodium ortho-vanadate, while intact cells were not affected by millimolar levels of the inhibitor. This is the first characterization of in vitro ciliary beat in a non-algal plant cell and demonstrates that the nucleotide and ionic requirements for reactivation of bracken cilia are similar to those of other systems.
Karyotype analysis of the plant-parasitic nematode Heterodera glycines by electron microscopy: II the tetraploid and an aneuploid hybrid
Authors: Goldstein Paul , Triantaphyllou A. C.
Published: April 2021

In the plant-parasite nematode Heterodera glycines, 2 forms, in addition to the diploid (9 bivalents), have been isolated and analysed: a tetrapioid (18 bivalents) and an aneuploid (14 bivalents, hybrid between the diploid and the tetrapioid). Observations on the formation of their karyotypes indicates normal and non-homologous pairing. Eighteen normal synaptonemal complexes (SC) are present in pachytene nuclei of the tetrapioid. Two of the SCs are enclosed in a large heterochromatin mass that is displaced to one side of the nucleus. Such a mass has not been observed in the diploid or the aneuploid. Another 2 normal SCs of the tetrapioid have each a ‘modified SC region’ (MSC) within which the SC appears disorganized. The aneuploid has 14 SCs that are unattached at either end. Only 25% of the karyotype length is normal in the appearance of the SCs. The rest can be traced by the presence of disorganized SC material and condensed chromatin. Four MSCs are present in the hybrid nuclei. The possible role of the MSCs in the sex determination system is discussed.
Quantitative demonstration of cell surface involvement in a plant-animal symbiosis: Lectin inhibition of reassociation
Authors: Meints R. H. , Pardy R. L.
Published: April 2021

The freshwater hydra, Hydra viridis is normally associated with Chlorella like, algal symbionts which inhabit the host’s digestive cells. Under experimental conditions bleached hydra will reassociate with algae harvested from green hydra, but not from our cultures of wild type Chlorella or strain NC64A which when originally isolated from Paramecium bursaria was symbiotically competent. Because of its demonstrated selectivity, the reassociation process is hypothesized to involve a recognition interface whose active participants are the algae cell wall and the digestive cell membrane. The data presented here confirm the hypothesis and suggest some potential molecular characteristics of the interacting partners.

Concanavalin A (Con A), a plant lectin, used widely for similar studies in other systems totally inhibits reassociation; Wheat Germ Agglutinin (WGA), ricin and Lens culinaris lectin do so to a lesser degree. These results are consistent with the hypothesis that glycoproteins on the cells’ peripheries are involved in cell-cell recognition in this system.
Protein labelling with 3H-NSP (N-succinimidyl-[2,3-3H]propionate)
Authors: Müller G. H.
Published: April 2021

A tritium compound with low molecular weight and diameter, N-succinimidyl-[2,3-3H]-propionate (3 H-NSP), was used to label histones and nonhistone proteins from calf thymus, and nuclear and total salivary gland proteins from larvae of the midge Chironomus thummi. Labelled proteins were visualized after electrophoresis in 10% polyacrylamide slab gels by fluorography. The method is much more sensitive than staining procedures, allowing to discern approx, 0.1 μg protein.
The relationship between the excess-delay phenomenon and temperature-sensitive periods in Tetrahymena thermophila
Authors: Frankel Joseph , Mohler Jym , Frankel Anne Koopmans
Published: April 2021

Although temperatures of 37·5 and 39 °C allow continuous and rapid exponential growth of wild type Tetrahymena thermophila, sudden shifts up to these temperatures can bring about long excess-delays of cell division with accompanying resorption of developing oral primordia. A characteristic parameter of this delay-phenomenon is the physiological transition point, before which delays are maximal and after which they are negligible. When measured at a restrictive temperature that does not induce excess delays (36 °C), the end of the temperature-sensitive period of the cell division arrest of mutant cdaAI precedes the physiological transition point, that of cdaHI roughly coincides with it, while the entire temperature-sensitive period of cdaC2 comes after the physiological transition point. When cdaAI cells are exposed to 37·5 °C or above, the manifestations of temperature sensitivity are drastically affected: the estimate of the end of the temperature-sensitive period (the execution point) becomes spuriously late, and the characteristic division arrest following heat shocks is not manifested. The differential effects of the higher restrictive temperatures on cdaHI are more subtle, whereas those on cdaC2 are negligible. We conclude that the excess-delay phenomenon involves a set-back of gene-mediated processes occurring at specific stages of the cell cycle.
Characterization and localization of adenylyl cyclase in membrane vesicles and intact boar and human spermatozoa
Authors: Peterson R. N. , Russell L. , Hook L. , Bundman D. , Freund M.
Published: April 2021

The enzymic properties of adenylyl cyclase in purified membrane vesicles from human and boar spermatozoa are described. Plasma membrane vesicles, which appear to be right-side-out, show a marked increase in activity in the presence of the detergent Triton X-100, manganous ion and alkaline pH. Electron-microscope cytochemical assays indicated the presence of adenylyl cyclase in boar and human sperm plasma membranes and also within the axoneme of intact human spermatozoa. The significance and precautions in the evaluation of the cytochemical data are discussed.
Genetic dissection of the final exocytosis steps in Paramecium tetraurelia cells: cytochemical determination of Ca2+-ATPase activity over preformed exocytosis sites
Authors: Plattner H. , Reichel K. , Matt H. , Beisson J. , Lefort-Tran M. , Pouphile M.
Published: April 2021

In different Paramecium tetraurelia strains the occurrence of a Ca’+-ATPase (or p-nitro-phenylphosphatase) activity at the preformed attachment and exocytosis sites of the secretory vesicles (trichocysts) was analysed by electron-microscopic cytochemistry and X-ray micro analysis. In conjunction with freeze-fracture studies it was found that only those strains, which contain rosette particles, display this Ca,+-ATPase activity (7S, K 401, nd 9 (18 °C)), while other strains (nd 6, nd 9 (27 °C), tarn 38) are devoid of both these characteristics. The presence (absence) of rosette particles and of Ca,+-ATPase activity at the preformed exocytosis sites is correlated with the capability (incapability) of performing exocytosis in these strains. We discuss several possible interpretations of this structure-function correlation.
Cell migration through three-dimensional gels of native collagen fibres: collagenolytic activity is not required for the migration of two permanent cell lines
Authors: Schor Seth L. , Allen Terence D. , Harrison Christine J.
Published: April 2021

Three-dimensional gels of native type I collagen fibres have been used as a substratum for the growth and migration of Chinese hamster ovary cells (fibroblastoid cell line) and RPMI–3460 melanoma cells (tumorigenic cell line from Syrian hamster). Quantitative data concerning the migration of these cells from the gel surface into the 3-dimensional collagen gel matrix have been obtained. The migration of both cell types into the collagen gel matrix is not accompanied by the degradation of collagen fibres. The possible implications of these observations for tumour cell invasion in vivo are discussed.
Human diploid fibroblast-like cells (CF-3) as a model system for the study of Golgi biogenesis
Authors: Dell’orco R. T. , Terracio L. , Douglas W. H. J.
Published: April 2021

Human diploid fibroblast-like cells were maintained in an arrested, essentially non-mitotic state for extended periods of time in culture by lowering the serum concentration in the medium from 10 to 0·5 %. These arrested cells re-entered the proliferative state when subcultivated in medium containing 10 % serum. The morphological distribution and enzymic activities associated with the Golgi complex were examined during growth, arrest, and recovery. Cells grown in medium containing 10 % serum possessed a well-developed Golgi complex consisting of parallel arrays of membranes and associated vesicles. Galactosyl transferase activity was highest after 3 days growth (17·5 ±5·0 nmol galactose transferred/45 min/mg protein) and decfined to 9·8 ± 30 at day 7. When the serum concentration was reduced to 0·5 %, Golgi complexes were rarely observed by electron microscopy and galactosyl transferase activity was further reduced to 4·6 ± 1·2 and 3-2 ± 1·4 after 3- and 7-day arrests, respectively. Arrested cells subcultured into medium containing 10 % serum recovered from the arrested state and ultrastructurally resembled cells continuously cultured at the higher serum level. Numerous Golgi complexes reappeared and galactosyl transferase activity increased to 13·0 ± 3·34 days after subcultivation. These results indicate that the Golgi complex can be experimentally manipulated in human diploid fibroblast-like cells in a manner which may be useful for the study of the biogenesis of this organelle.
Three-dimensional reconstruction of organelles in Euglena gracilis Z.: II. Qualitative and quantitative changes of chloroplasts and mitochondrial reticulum in synchronous cultures during bleaching
Authors: Pellegrini Max
Published: April 2021

By ultrathin serial sectioning, morphological and volumetric changes of the plastidome and chondriome have been observed in Euglena gracilis Z during bleaching in darkness with addition of sodium acetate to the culture medium. In order not to introduce any modification to the synchronization pattern during bleaching, green cells were previously grown photoauto-trophically on Cramer&Myers medium under continuous illumination and synchronized by temperature cycles. They were then cultivated (1) photoheterotrophically with sodium acetate added to the culture medium and (2) heterotrophically by keeping the culture in darkness. This regime allowed observation of the influence (1) of synchronization by temperature cycles and (2) of sodium acetate and darkness on the plastidome and chondriome of photoautotrophic cells synchronized by light-dark cycles as described previously. In photoautotrophic cells, the plastidome, consisting of about ten diskoidal chloroplasts, occupies 15 % of the cell volume. The chondriome, in the form of one single giant mitochondrion branched throughout the cell, represents 6 % of the cell volume.

The synchronization by temperature cycles in continuous illumination does not change the morphology and volume of these organelles. However, pyrenoids disappear. In photohetero-trophic culture with sodium acetate added, the plastidome fine structure does not vary but its volume decreases by 19–25 %. At that time, the plastidome thus occupies 12–13 % of the cell volume. Sodium acetate provokes, on the contrary, hypertrophy of the delicate threads of the mitochondrial reticulum which appears as a network with narrow meshes around other organelles. The chondriome thus comes to occupy 10–11 % of the cell volume. In heterotrophic cells, the combined effects of sodium acetate and darkness emphasize the regression of the plastidome volume and hypertrophy of the chondriome. The plastidome occupies 6–7 % of the cell volume while the chondriome appears as a fenestrated parietal shell occupying 15—16% of the cell volume. Maximal hypertrophy is obtained in 24 h. Total dedifferentiation of chloroplasts requires 6—9 successive generations in heterotrophic conditions.

These results are discussed in relation to numerous light-microscopic and ultrastructural observations. It has been demonstrated, as in photoautotrophic Euglena cells synchronized by light-dark cycles, that the plastidome of heterotrophic cells consists of about ten organelles, whereas the chondriome contains one single giant mitochondrion. Contrary to the opinion that the variations of the plastidome and chondriome are reciprocally related, it is proved here that dedifferentiation of chloroplasts and hypertrophy of the chondriome occur at different rates, and may be independent of one another.
A cell cycle study of the effects of Con A on synchronized mouse embryo fibroblasts: arrest at two specific stages of the cycle and dissociation between uptake of thymidine and DNA synthesis
Authors: Mallucci L. , Dunn M. , Wells V. , Delia D.
Published: April 2021

We have examined the effects of 50 µg m1−1 of Con A added to synchronized mouse embryo fibroblasts at different times during the cell cycle. We found that Con A caused arrest of growth not solely by preventing G1-G0 cells from entering the S-phase but also by exerting a G1 block. We also found that Con A, which prevented commencement of S-phase, did not arrest cells already in S from reaching the G2 stage but inhibited the S-phase associated process of thymidine uptake. The inhibition was greater when the Con A receptors were extensively clustered.
DNA is replicated at the nuclear cage
Authors: Mccready S. J. , Godwin J. , Mason W. , Brazell I. A. , Cook P.R.
Published: April 2021

Structures resembling nuclei are released when HeLa cells are lysed in a detergent and 2 M salt. These nucleoids, which lack any organized membrane structure, contain all the nuclear DNA packaged within a cage of RNA and protein. Their DNA is supercoiled so that the linear DNA must remain unbroken and looped during lysis. Following digestion with the restriction endonuclease, EcoRI, cages and associated DNA were filtered free of unattached DNA. Pulse-labelled (i.e. newly synthesized) DNA remains preferentially associated with the cages. This association has been confirmed by autoradiography. When nucleoids are prepared for electron microscopy by the Kleinschmidt procedure the DNA spills out to form a skirt around the flattened cage. Labelling, which is restricted to the region of the cage after short pulses, extends out into the skirt as the labelling time increases. A model, based on the premise that replication takes place at the nuclear cage, is presented in the Appendix. The results of the biochemical experiments and electron microscopy both indicate that the average size of the unit of replication is ∼ 20 μm. This is about one-quarter the size of the average structural unit -the loop. Therefore sequences in the loop must become attached to the nuclear cage prior to the initiation of DNA synthesis.
Genetic dissection of the final exocytosis steps in Paramecium tetraurelia cells: trigger analyses
Authors: Matt H. , Plattner H. , Reichel K. , Lefort-Tran M. , Beisson J.
Published: April 2021

A variety of trigger procedures were applied to analyse the exocytotic capability of different Paramecium tetraurelia strains. 7,S K 401, kin 241, nd g (18 °C) arecapable of exocytosis (permissive strains), in contrast to nd 6, nd 7, nd g (27 °C), tarn 38 and ftb A, although all procedures used enhance [Ca3+] in the cytoplasm of all strains tested and although strains nd 6, nd 7 and nd g (27 °C) contain a full set of morphologically normal trichocysts attached to the cell membrane. The results show that only those strains are permissive which were shown previously to contain a rosette of membrane-integrated particles and a Ca2+-ATPase activity in the cell membrane over the trichocyst attachment (exocytosis) sites. The results from trigger experiments with permissive and non-permissive strains would be compatible with a dual function of rosette particles as Ca2+ pumps and Ca2+ channels. Nevertheless, the latter aspect remains uncertain since we show that experiments along these lines published by others (introducing a Ca,+ ionophore from the outside) involve a solvent-induced artifact (pseudoexocytosis: matrix stretching in the absence of membrane fusion). In all strains, except for tarn 38 and ftb A (which have abnormal trichocysts incapable of being attached to the cell membrane), the isolated trichocyst matrix can be transferred from the contracted to the expanded state in vitro with certain trigger procedures. Our data clearly show that an increase of [Ca3+]t in the cytoplasm is not sufficient for exocytosis to occur and that non-permissivity is somehow due to an inability to perform membrane fusion. It remains open whether the lack of rosettes and Ca2+-ATPase activity at trichocyst attachment sites are the primary cause of non-permissivity’.