bluuebunny/crossref_metadata_2025_split huggingface dataset:
Commentary structure and dynamics of transcriptionally active chromatin
Authors: Ausio Juan 
Published: April 2021

ABSTRACT
Chromatin is a highly dynamic macromolecular complex that undergoes continuous structural modification during the various stages of genetic activity. Such dynamic behavior is the result of a complex and perfectly balanced network of interactions involving histones, DNA and ions in an aqueous environment. In the past eighteen years since the discovery of the nucleosome particle, a great deal of information has been obtained on interactions responsible for maintaining the static three-dimensional structure of inactive chromatin (see Van Holde, 1988, for a recent review on chromatin). More recently, structural studies of chromatin have focused on the analysis of the molecular events involved in chromatin activation. A variety of models have been proposed, primarily to explain the structural transitions undergone by chromatin during transcription. In the following sections, I will briefly review what is known about the structure of transcriptionally active chromatin and propose a model for “activation” that would account for the chemical and ionic interactions that are possibly involved. I hope that this model will stimulate design of new experimental approaches with which to understand better the structure-function relationship in chromatin.
Nuclear distribution of PCNA during embryonic development in Xenopus laevis: a reinvestigation of early cell cycles
Authors: Leibovici Michel , Monod Guillaume , GÉraudie Jacqueline , Bravo Rodrigo , Méchali Marcel 
Published: April 2021

ABSTRACT
The immunocytological distribution of the proliferating cell nuclear antigen (PCNA), a protein involved in DNA replication, has been examined during the early development of Xenopus laevis. The protein is uniformly detected in nuclei during early stages up to the neurula stage. PCNA is detected by its distinctive cyclical pattern during early development, remaining detectable only during the period of S phase of each cell cycle. Immunological detection of PCNA is therefore a useful and specific non-isotopic marker of S-phase cells in the embryo. PCNA associates with typical karyomeric structures, suggesting that DNA replication starts before the nuclear compartment is entirely formed. At the midblastula transition, a new pattern of PCNA staining becomes apparent. First, a new type of PCNA staining is detected at the nuclear periphery. Second, mitotic clusters with different PCNA distributions suggest that the onset of desynchronization of the cell cycle at this stage is not random.
Extrajunctional distribution of N-cadherin in cultured human endothelial cells
Authors: Salomon Daniela , Ayalon Oran , Patel-King Ramila , Hynes Richard O. , Geiger Benjamin 
Published: April 2021

ABSTRACT
Human endothelial cells contain prominent Ca2+-dependent intercellular adherens-type junctions (AJ), which are associated, at their cytoplasmic surfaces, with actin, vinculin and plakoglobin. The transmembrane adhesion molecules present in these sites are members of the cadherin family, which are recognized by a pan-cadherin serum, directed against the conserved C terminus of these molecules. Immunoblotting analysis of cultured human endothelial cells using these antibodies revealed three immunoreactive bands with apparent molecular masses of 135,130 and 120 kDa. Cloning and sequencing of the 135 kDa cadherin from an endothelial cDNA expression library indicated that this molecule is a typical cadherin, essentially identical to N-cadherin. Transfection of cDNA encoding this molecule into CHO cells resulted in the induction of AJ formation and an apparent epithelialization of the cells. Immunofluor-escent labeling with antibodies to chicken N-cadherin indicated that the molecule is associated with intercellular junctions in the transfectants. In contrast, cultured human umbilical cord endothelial cells exhibited a largely diffuse N-cadherin labeling over the entire cell surface with only occasional enrichment in cell-cell junctions. Comparison of this pattern with the discrete junctional labeling obtained with the pan-cadherin antibody suggests that different cadherins, co-expressed in the same endothelial cells, may undergo differential surface distribution.
Modulation of anaphase spindle microtubule structure in stamen hair cells of Tradescantia by calcium and related agents
Authors: Zhang D. H. , Wadsworth P. , Hepler P. K. 
Published: April 2021

ABSTRACT
We have previously shown that changes in the intracellular free calcium concentration ([Ca2+]i) modulate the rate of anaphase chromosome motion in stamen hair cells of Tradescantia. Elevations between 0.8–1.0 μM accelerate motion, while increases above 2.0 μM or decreases below resting level inhibit motion. The related signaling agent GTPγS also accelerates motion, however, by a mechanism that does not appear to involve changes in [Ca2+)i. To explore further the mechanism by which Ca2+ and GTPγS regulate chromosome motion we have analyzed the direct effect of these agents on the structure of the spindle microtubules (MTs). First, we injected carboxyfluorescein-derivatized brain tubulin and allowed it to incorporate into spindle MTs. Then, during appropriate times of anaphase, we injected Ca2+ or related agents and monitored their effect on spindle MT fluorescence using a confocal laser scanning microscope. A high level of Ca2+ (10 μM), known to inhibit motion, causes extensive degradation of spindle MT structure. An intermediate level (2 μM), which slows but does not stop movement, produces a distinct decay of fluorescence. A level of Ca2+ (0.8–1 μM) known to accelerate motion, however, generates only a small change in which the kinetochore fibers appear less distinct, and the overall spindle fluorescence is more diffuse. The Ca2+ buffer EGTA, which transiently blocks motion, has no detectable effect on spindle structure. GTPγS, which enhances motion also has no discernible effect on spindle structure. The results with Ca2+ support the idea that the ion facilitates anaphase motion through depolymerization of kinetochore MTs. However, if the depolymerization is extensive then inhibition of motion ensues. GTPγS, on the other hand, accelerates motion without inducing a detectable effect on spindle MT structure.
Expression of tau protein in non-neuronal cells: microtubule binding and stabilization
Authors: Lee Gloria , Rook Susan L. 
Published: April 2021

ABSTRACT
The microtubule-associated protein tau is a developmentally regulated family of neuronal phosphoproteins that promotes the assembly and stabilization of microtubules. The carboxy-terminal half of the protein contains three copies of an imperfectly repeated sequence; this region has been found to bind microtubules in vitro. In addition, a fourth copy of the repeat has been found in adult-specific forms of tau protein. To examine the structure and function of tau protein in vivo, we have transiently expressed fetal and adult forms of tau protein and tau protein fragments in tissue culture cells. Biochemical analysis reveals full-length products with heterogeneity in post-translational modification synthesized in the cells. Immunofluorescent staining of transfected cells shows that, under our conditions, sequences on both sides of the repeat region are required for in vivo microtubule co-localization. These additional regions may be required either for enhancing micro tubule contacts or for proper protein folding in the cell. In our expression system, the bundling of cellular microtubules occurs only in transfections using four-repeat tau constructs; any four-repeat construct capable of binding is also able to induce bundling. Our data suggest that the presence of bundles is correlated with enhanced microtubule stability; factors that increase stability such as higher levels of tau protein expression or the presence of the fourth repeat, increase the fraction of transfected cells showing bundles. Finally, the presence of tau protein in the cell allows all interphase microtubules to become acetylated, a post-translational modification usually reserved for a subset of stable cellular microtubules.
Investigation of the role of microtubules in protein secretion from lactating mouse mammary epithelial cells
Authors: Rennison Michelle E. , Handel Susan E. , Wilde Colin J. , Burgoyne Robert D. 
Published: April 2021

ABSTRACT
Disruption of microtubules has been shown to reduce protein secretion from lactating mammary epithelial cells. To investigate the involvement of microtubules in the secretory pathway in these cells we have examined the effect of nocodazole on protein secretion from mammary epithelial cells derived from the lactating mouse. Mouse mammary cells have extensive microtubule networks and 85% of their tubulin was in a polymeric form. Treatment with 1 μg/ml nocodazole converted most of the tubulin into a soluble form. In a continuous labelling protocol it was found that nocodazole did not interfere with protein synthesis but over a 5 h period secretion was markedly inhibited. To determine whether the inhibition was at the level of early or late stages of the secretory pathway mammary cells were pulse-labelled for 1 h to label protein throughout the secretory pathway before nocodazole treatment. When secretion was subsequently assayed it was found to be slower and only partially inhibited. These findings suggest that the major effect of nocodazole is on an early stage of the secretory pathway and that microtubules normally facilitate vesicle transport to the plasma membrane. An involvement of microtubules in vesicle transport to the plasma membrane is consistent with an observed accumulation of casein vesicles in nocodazole-treated cells. Exocytosis stimulated by the calcium ionophore ionomycin was unaffected by nocodazole treatment. We conclude from these results that the major effect of nocodazole is at an early stage of the secretory pathway, one possible target being casein vesicle biogenesis in the trans-Golgi network.
Regulation of nuclear envelope precursor functions during cell division
Authors: Vigers G. P. A. , Lohka M. J. 
Published: April 2021

ABSTRACT
Previously, we have shown that nuclear envelope assembly in cell-free extracts of Xenopus eggs requires two distinct vesicle-containing fractions, called Nuclear Envelope Precursor Fractions A and B (NEP-A and NEP-B). These fractions are characterized further in this paper and the manner in which they are regulated during metaphase is examined. Antisera against the NEP-B fraction recognized several proteins common to NEP-B and Xenopus oocyte or liver nuclei, but not to NEP-A or cytosol. A known glycoprotein component of the nuclear pore complex, p62, also co-fractionated with NEP-B, whereas the Xenopus egg lamin LUI did not. Together, these results provide further evidence that the NEP-B fraction contains precursors of the nuclear envelope.
The regulation of NEP-A and -B function during metaphase, when the nuclear envelope is disassembled, was examined by treating each fraction with metaphase cytosol or purified protein kinase preparations isolated from metaphase-arrested eggs. Treatment of NEP-B with metaphase cytosol, under conditions where proteins are irreversibly phosphorylated, inhibited the subsequent assembly of the nuclear envelope by preventing the binding of NEP-B to chromatin. In contrast, similar treatment of NEP-A did not affect its ability to form nuclear envelopes. The changes in NEP-B during metaphase did not appear to be regulated directly by either p34cdc2/cyclin B, S6 kinase II or MAP kinase.
Regulation of fibroblast-mediated collagen gel contraction by platelet-derived growth factor, interleukin-1 α and transforming growth factor-β1
Authors: Tingström Anders , Heldin Carl-Henrik , Rubin Kristofer 
Published: April 2021

ABSTRACT
We have examined the effects of three macrophage-derived cytokines, platelet-derived growth factor (PDGF), transforming growth factor-β1 (TGF-β1) and interleukin-la (IL-1 a) on the contraction of collagen type I gels populated by human foreskin fibroblasts. Contraction was quantified as loss in gel weight. Both PDGF-A A and PDGF-BB were found to induce a rapid collagen-gel contraction. TGF-β1 also stimulated gel contraction but with a delayed onset and at a slower rate than the PDGF-stimulated contraction. Rabbit polyclonal IgGs recognizing PDGF-A A and PDGF-BB, respectively, specifically inhibited the effects of the corresponding PDGF isoforms. However, the stimulatory effect of TGF-JÎ1 was not affected by any of the anti-PDGF antibodies. The ability of PDGF to stimulate contraction became less pronounced in collagen gel cultures grown in the absence of growth factors over periods of several days. Under the same conditions, the stimulatory effect of TGF-β1 was not reduced. The reduced response to PDGF may be due to reduced tension on fibroblasts growing in collagen gels, since fibroblasts on free-floating gels showed a marked reduction in PDGF-BB-induced PDGF β-receptor aggregates when compared to fibroblasts on attached collagen gels. IL-1α inhibited initial collagen gel contraction, and at later stages induced a visible degradation of the collagen gels, presumably due to the generation of collagenase activity. The combination of IL-la and PDGF-BB stimulated initial collagen gel contraction, although less effectively than PDGF-BB alone. At later stages, collagen gel degradation was stimulated by this combination of cytokines. In contrast, the combination of IL-la and TGF-β1 did not stimulate collagen gel contraction, or any visible collagen gel degradation. Our data suggest that fibroblast-mediated collagen gel contraction can be modulated by cytokines via different mechanisms. Our data are of importance in the understanding of the modulatory roles of cytokines in connective tissue cell activities in inflammatory processes, such as wound healing.
Evidence for an inverse relationship between the differentiation of adipocytic and osteogenic cells in rat marrow stromal cell cultures
Authors: Beresford J. N. , Bennett J. H. , Devlin C. , Leboy P. S. , Owen M. E. 
Published: April 2021

ABSTRACT
The differentiation of adipocytic and osteogenic cells has been investigated in cultures of adult rat marrow stromal cells. Adipocytic differentiation was assessed using morphological criteria, changes in expression of procollagen mRNAs, consistent with a switch from the synthesis of predominantly fibrillar (types I and HI) to basement membrane (type IV) collagen, and the induction of expression of aP2, a specific marker for differentiation of adipocytes. Osteogenic differentiation was assessed on the basis of changes in the abundance of the mRNAs for the bone/liver/kidney isozyme of alkaline phosphatase and the induction of mRNAs for bone sialoprotein and osteocalcin. In the presence of foetal calf serum and dexamethasone (10−8 M) there was always differentiation of both adipocytic and osteogenic cells. When the steroid was present throughout primary and secondary culture the differentiation of osteogenic cells predominated. Conversely, when dexamethasone was present in secondary culture only, the differentiation of adipocytes predominated. When marrow stromal cells were cultured in the presence of dexamethasone in primary culture and dexamethasone and 1,25-dihydroxyvitamin D3 (1,25(OH)2D3; 10−8 M) in secondary culture, the differentiation of adipocytes was inhibited whereas the differentiation of osteogenic cells was enhanced, as assessed by an increase in expression of osteocalcin mRNA. The results, therefore, demonstrate an inverse relationship between the differentiation of adipocytic and osteogenic cells in this culture system and are consistent with the possibility that the regulation of adipogenesis and osteogenesis can occur at the level of a common precursor in vivo.
Microinjection of antibodies against talin inhibits the spreading and migration of fibroblasts
Authors: Nuckolls Glen H. , Romer Lewis H. , Burridge Keith 
Published: April 2021

ABSTRACT
Talin is believed to be one of the key proteins involved in linking actin filaments to extracellular matrix receptors in focal adhesions. Our strategy for studying the function of talin has been to inactivate talin in living fibroblasts in tissue culture through the microinjection of affinity-purified, polyclonal anti-talin antibodies. The effect of the injected anti-talin antibodies on cell spreading was found to depend on how recently the cells had been plated. Cells that were in the process of spreading on a fibronectin substratum, and which had newly developed focal adhesions, were induced to round up and to disassemble many of the adhesions. However, if fibroblasts were allowed to spread completely before they were microinjected with the anti-talin antibody, focal adhesions remained intact and the flat morphology of the cells was unaffected. The percentage of cells that were able to maintain a spread morphology despite the injection of anti-talin antibodies increased during the first few hours after plating on fibronectin substrata. Fibroblasts that were allowed to spread completely before microinjection with the anti-talin antibody retained both intact focal adhesions and a flat, wellspread morphology, but failed to migrate effectively. Our experiments do not directly address the role of talin in mature focal adhesions, but they indicate that talin is essential for the spreading and migration of fibroblasts on fibronectin as well as for the development and initial maintenance of focal adhesions on this substratum.
An evaluation of cell separation techniques in a model mixed cell population
Authors: Murphy S. J. , Wait D. J. , Jones G. E. 
Published: April 2021

ABSTRACT
Muscle precursor cells may act not only as a means of inserting normal genes into diseased muscle fibres, in order to correct or alleviate a genetically inherited myopathy, but recent demonstrations have shown they may prove an invaluable tool for the expression of, and systemic dissemination of, non-muscle gene products. If muscle precursor cells are proved to act as such widespread vectors in terms of gene therapy, then it is imperative that methods are properly elucidated to produce large populations of pure viable myogenic cells for such purposes. In the past, many methods of cell separation have been investigated but carry with them the problems of either a lack of myogenic purity of the population or poor percentage recovery of the original cell population.
In the present work we have investigated two methods for segregating myogenic from non-myogenic cells and have critically reviewed the efficiency of separation of the two techniques used. To obtain a quantitative measure of separation efficiency, segregation was carried out on a 1:1 mixture of murine C2 myogenic and murine 3T3 fibroblastic cells. To distinguish between C2 and 3T3 cells, the latter were prelabelled with the fluorescent strain carboxyfluoroscein diacetate succinimyl ester (CFSE). Once incorporated into the cell, CFSE remains there, thus preventing transfer of the label to C2 cells. Both methods of separation used depend on the affininty of myogenic cells for the monoclonal antibody Mab H28, which specifically binds to the mouse neuronal cell adhesion molecule N-CAM, but differ in that one method, “panning”, completes segregation by adherence of N-CAM positive cells to a dish precoated with secondary IgG antibody whereas in the other separation proceeds by the use of commercially available IgG-coated magnetic beads. Results indicate magnetic bead separation to be more efficient than panning if the beads are precoated with 0.1% gelatin.
Specific patterns of cortical and endoplasmic microtubules associated with cell growth and tissue differentiation in roots of maize (Zea mays L.)
Authors: BalukŠa F. , Parker J. S. , Barlow P. W. 
Published: April 2021

ABSTRACT
Indirect immunofluorescence using a-tubulin antibodies applied to sections of maize roots prepared using low melting point wax was found to give excellent visualisation of both cortical and endoplasmic microtubule (MT) arrays. This allows, for the first time, these arrays to be investigated in cells of the different tissues at various stages of their growth and development. Many cells in the zone between the meristem and the cell elongation region develop a highly ordered transversal bundling of cortical MTs, which we suggest is related to vacuolation of the cytoplasm and which may also be indispensable for the ensuing rapid cell elongation. On the other hand, there are subtle differences between the cells of the individual tissues regarding the arrangement of their cortical MTs in this zone. The possible physiological significance of these tissue-specific MT arrays is discussed. Endoplasmic MTs were seen to encircle and to connect the nucleus with the cortical MT arrays in both dividing and elongating cells. Even the G1 phase nuclei of the slowly dividing cells of the quiescent centre were encircled by endoplasmic MTs. The continuity of the two MT systems may provide the cell with an important signalling system whereby mechanical and physiological information is relayed from the exterior of the cell to the nucleus.
Organization and characterization of fibrillar collagens in fish scales in situ and in vitro
Authors: Zylberberg L. , Bonaventure J. , Cohen-Solal L. , Hartmann D. J. , Bereiter-Hahn J. 
Published: April 2021

ABSTRACT
The characterization of the fibrillar collagens and the cellular control of their spatial deposition were studied in fish scales using immunofluorescence, electron microscopy, electrophoretic and HPLC analyses, immunoprecipitation and hybridization with cDNA probes. This study was carried out on undisturbed and regenerating scales in situ and in organ and cell cultures from regenerating scales. The hyposquamal scleroblasts forming a pseudoepithelium show an apico-basal polarization and synthesize thick collagen fibrils (100 nm) organized in a plywood pattern as long as the integrity of the cell-cell and cell-collagenous matrix contacts are preserved. In culture, scleroblasts become fibroblast-like and produce an unordered meshwork of thin collagen fibrils (30 nm). Comparison of the synthesized collagens in culture with those extracted from the scales indicates that culture conditions modify fibrillogenesis but do not change the expression of fibrillar collagen genes. Type I collagen, the prédominent component, is associated with the minor type V collagen. Type HI collagen was not present. In type I collagen, a third chain, α3 chain, was identified. The ratio between the 3 chains suggests the coexistence of two heterotrimers (αl(I))2 α2(1) and αl(I) α2(1) α3(I). Analysis by HPLC and electrophoresis of the cyanogen bromide-derived peptides obtained from the purified α3 chain support the hypothesis that αl(I) and α3(I) chains are encoded by two different genes. The presence of the two types of heterotrimers in vivo as well as in vitro could correspond to an innate property of the goldfish scleroblasts. Despite the fact that teleost cyanogen bromide-derived peptides differ from those of higher vertebrates, homologies with the mammalian collagen genes (human, for example) are sufficient to allow the detection of mRNA transcripts for al(I), α2(I) and α2(V) from confluent scleroblast cultures with human probes.