bluuebunny/crossref_metadata_2025_split huggingface dataset:
Drosocrystallin, a major 52 kda glycoprotein of the Drosophila melanogaster corneal lens purification, biochemical characterization, and subcellular localization
Authors: Komori Naoka , Usukura Jiro , Matsumoto Hiroyuki 
Published: April 2021

ABSTRACT
We have identified a 52 kDa protein, which is a potent substrate for cholera toxin-dependent ADP-ribosylation, in the compound eye preparation of the fruit fly, Drosophila melanogaster. We find that the 52 kDa protein is a glycoprotein and a Ca2+ binder bearing a high content of leucine, serine and glycine. By microsequencing we determined its 13 N-terminal sequence, AYL*PIDLNQLAK, with the asterisk representing an ambiguous signal. In order to study further the 52 kDa protein we have raised a polyclonal antibody against a synthetic oligopeptide representing the N-terminal 13 residues of the 52 kDa protein. By immunogold labelling with the antibody, the epitopes were localized at the EM level to the laminated corneal lens. The number of the gold particles per /un2 in the electron-dense layer of the corneal lens was 2.5 times higher than that of the electron-lucent layer. The pattern of the 52 kDa protein distribution in the corneal lens suggests that the 52 kDa protein is the major protein component that participates in the pattern formation of the alternate refractive indices of the D. melanogaster corneal lens. An X-ray dispersion analysis in situ revealed that the laminated corneal lens contained a higher concentration of Ca2+, supporting the hypothesis that the 52 kDa protein binds Ca2+in vivo. To the best of our knowledge, this is the first report that identifies the protein entity of an arthropod corneal lens. We propose to designate this 52 kDa protein drosocrystallin.
Distribution of a nuclear envelope antigen during the syncytial mitoses of the early Drosophila embryo as revealed by laser scanning confocal microscopy
Authors: Riparbelli Maria Giovanna , Callaini Giuliano 
Published: April 2021

ABSTRACT
The changing distribution of a nuclear envelope antigen recognized by a monoclonal antibody raised against human fibroblast vimentin during the syncytial mitoses of the Drosophila embryo has been studied with a confocal laser scanning microscope. The antigen appears very early as irregular aggregates in the peripheral cytoplasm of the preblastoderm embryo. As the first nuclei reach the periplasm the antigen is localized on the nuclear envelope and the cytoplasmic staining decreases. In addition to the perinuclear labeling we observed intense midzone and polar staining during the mitotic cycle. A possible relationship between polar localization of the antigen and centrosome position is discussed.
Cytokeratins and retinal epithelial cell behaviour
Authors: Robey Helen L. , Hiscott Paul S. , Grierson Ian 
Published: April 2021

ABSTRACT
The expression of cytokeratins 18 and 19 by human retinal pigment epithelial cells (HRPE) has been suspected of being associated with HRPE proliferation. We have investigated the involvement of these cytokeratin subtypes in the proliferative and migratory behaviour of cultured HRPE. Cell proliferation markers (bromodeoxyuridine and proliferating cell nuclear antigen) and the cytokeratins were identified using immunohistochemical techniques. In vitro, cytokeratins 18 and 19, as detected by the monoclonal antibodies RGE 53 and K4.62, were expressed in a subset of HRPE and this subset was significantly less likely to be proliferating. Micro-chemotaxis chambers were used to study migrating cells and immunohistochemical staining for cytokeratins 18 and 19 revealed that actively migrating cells always expressed these two cytokeratins, whereas stationary cells did not label for these cytokeratin subtypes. It was apparent that cytokeratins 18 and 19 were not markers of proliferation, but were involved in the mobility of HRPE in vitro. Cytokeratins 18 and 19 may be useful indicators of simple epithelial cell migration in tissues.
A functional model of adult human prostate epithelium the role of androgens and stroma in architectural organisation and the maintenance of differentiated secretory function
Authors: Hayward Simon W. , Buono Raffaele Del , Deshpande Nagesh , Hall Pete A. 
Published: April 2021

ABSTRACT
A functional model of adult human prostate epithelium is described. This model shows that stromal cells, but not an androgenic stimuli, are required for architectural organisation of prostate epithelium. Within an organised structure, androgenic stimulation is required for the establishment of secretory epithelial cell morphology and associated function. In the absence of stromal cells but in the presence of androgens architectural organisation and secretory function are lost. Epithelial parenchymal units (organoids) from human prostate tissue were isolated, cultured within a three-dimensional collagen matrix, and xenografted subcutaneously into athymie mouse hosts. The grafted gels were rapidly invaded by host fibroblasts. Epithelial organisation initially disappeared but was re-established concurrently with the stromal cell invasion. In intact male hosts, cuboidal and columnar cells that expressed human prostate-specific secretory markers were found. In castrated male and in female hosts epithelial structures were lined with flattened epithelium with no secretory function. This phenomenon could be reversibly replicated by treating intact male hosts with the anti- androgen Flutamide. Gels containing organoids grafted within 0.45 /mi Millipore chambers were not invaded by stromal cells and rapidly lost all epithelial organisation and secretory function. When organoids cocultured with human foreskin fibroblasts were grafted within chambers, structural organisation of the epithelium was supported. These results indicate that both heterologous human fibroblasts and mouse stromal cells are capable of permissively supporting adult human prostate epithelial function.
Mechanisms of epibolic tissue spreading analyzed in a model morphogenetic system roles for cell migration and tissue contractility
Authors: Armstrong Margaret T. , Armstrong Peter B. 
Published: April 2021

ABSTRACT
The processes responsible for epithelial spreading during wound healing and embryonic morphogenesis were investigated in an organ culture model in which an epithelial tissue (chick embryo pigmented retinal epithelium) spread over the surface of an aggregate of mesenchyme cells (chick embryo cardiac mesenchyme). The heart mesenchyme aggregate is differentiated into a core of stellate cells associated with a fibronectin-poor matrix surrounded by a cortical zone, 2-5 cells in thickness, of flattened cells embedded in a fibronectin-rich extracellular matrix. Envelopment of the mesenchyme aggregate is accompanied by a movement of the cells and the fibronectin-rich extracellular matrix of the cortex over the core tissue in advance of the spreading pigmented retina tissue. Three distinct processes were identified as contributing to epithelial spreading in this system: (1) active migration of the pigmented retinal epithelium; (2) active contraction of the cortical cells of the mesenchyme aggregate to tow the attached epithelial tissue over the mesenchyme aggregate; and (3) ingression of surface-located cells of the mesenchyme aggregate to decrease the exposed surface area by decreasing the number of cells at the surface.
Poly ADP-ribosylation: a histone shuttle mechanism in DNA excision repair
Authors: Us Felix R. Altha 
Published: April 2021

ABSTRACT
In DNA excision repair of mammalian cells, the processing of ADP-ribose by the poly ADP-ribosylation system of chromatin is stimulated several thousand-fold. Most of this turnover is associated with the automodification reaction of the nuclear enzyme poly(ADP-ribose) polymerase and the degradation of polymerase-bound polymers by the enzyme poly(ADP-ribose) glycohydrolase. The automodification cycle catalyzes a temporary dissociation from and reassociation of histones with DNA. It is proposed that this mechanism, termed “histone shuttle”, may guide specific proteins to sites of repair. In addition, histone shuttling driven by the poly ADP-ribosylation system seems to be involved in nucleosomal unfolding of chromatin in DNA excision repair.
Synchronization of hair cell regeneration in the chick cochlea following noise damage
Authors: Stone Jennifer S. , Cotanche Douglas A. 
Published: April 2021

ABSTRACT
Pure-tone overstimulation for prolonged time leads to hair cell death in frequency-specific regions of the cochlear epithelium. Unlike mammals, birds replace missing hair cells by stimulating mitosis in an uncharacterized precursor cell. Regenerated hair cells, initially identifiable by their immature stereocilia and small surface areas, differentiate into mature cells in a manner which parallels embryonic development. In the current study, we examined whether hair cell regeneration is initiated during noise exposure or after the end of acoustic trauma. We exposed 7- to 15-day-old chicks to a 1500 Hz pure tone at 120 dB SPL (re 20 μPa) for 4, 12, and 24 hours and examined the recovering cochlear epithelium with scanning electron microscopy to determine when regenerated hair cells were first identifiable. The earliest evidence of new hair cells appeared roughly 96 hours after the onset of 4-, 12-, and 24-hour exposures. Our previous studies initially identified new hair cells 96 hours after the start of a 48-hour exposure. Therefore, hair cell regeneration follows a similar time course relative to the onset of noise exposure, regardless of the ultimate duration of exposure. Since we estimate that hair cells take at least 48 hours after their genesis to form immature stereocilia, the signal which induces hair cell precursors to re-enter the cell cycle and to divide probably has its initial effects very early during the exposure period. (A previous report of these data was given at the 1991 American Society for Cell Biology conference.)
Antennapedia homeobox as a signal for the cellular internalization and nuclear addressing of a small exogenous peptide
Authors: Perez F. , Joliot A. , Bloch-Gallego E. , Zahraoui A. , Triller A. , Prochiantz A. 
Published: April 2021

ABSTRACT
In a previous study we demonstrated that a homeobox peptide corresponding to the 60 amino acid long DNA-binding region of the Drosophila antennapedia homeoprotein was capable of crossing the plasma membrane of cells in culture. This finding has led us to investigate whether chimeric molecules encompassing the homeobox would behave in a similar manner. We demonstrate here that a peptide of 93 amino acids composed of the homeobox and of the C terminus of Rab3, a small GTP-binding protein, crosses the membrane of myoblasts, myotubes and neurons and is conveyed to their nuclei.
This transport is highly efficient, is observed in all the cells present in the culture and occurs at 37°C and 12°C without quantitative peptide degradation. Beyond its theoretical implications for our current views on cellular interactions, this finding could have technical repercussions on the development of drugs with intracellular targets.
Stimulus-dependent relocation of the microtubule organizing center in human polymorphonuclear leukocytes
Authors: Chiplonkar Jayant M. , VandrÉ Dale D. , Robinson John M. 
Published: April 2021

ABSTRACT
Polymorphonuclear leukocytes (PMNs) exhibit extensive directional migration (chemotaxis) and phagocytic activities. We have developed an in vitro model to evaluate the organization of the microtubule organizing center (MTOC) in PMNs as the latter interact with various substrata, including immobilized antigen-antibody complexes. PMNs were layered on poly-L-lysine substrata containing ferritin (PL+F) or ferritin-antiferritin complex (PL+F+AF) and the location of MTOCs was determined by indirect immunofluorescence of tubulin using conventional epifluorescence microscopy and confocal laser scanning microscopy. The MTOCs in the majority of the PMNs attached to PL+F occupied an apical location (81.29% ± 3.34%), while in the majority of PMNs layered onto PL+F+AF, a basal location (79.37% ± 5.26%) was observed. Following disruption of microtubules (MTs) by nocodazole before layering the cells on the substrata, the proportions of PMNs with apical MTOCs were 65.2% ± 6.27% for PL+F and 47.2% ± 4.1% for PL+F+AF substrata, while the proportions of PMNs with basal MTOCs were 26.11% ± 8.89% for PL+F and 39.6% ± 4.4 for PL+F+AF substrata. The results indicate that MTOCs in human PMNs in vitro (i) occupied a ‘pre-defined’ apical location; (ii) translocated to a ‘newly defined’ basal location upon stimulation with immobilized antigenantibody complex; (iii) and depended on intact MTs for placement of MTOCs in both situations.
Mammalian skeletal muscle C-protein: purification from bovine muscle, binding to titin and the characterization of a full-length human cDNA
Authors: FÜrst Dieter O. , Vinkemeier Uwe , Weber Klaus 
Published: April 2021

ABSTRACT
We report a fast method for the isolation of homogeneous C-protein from bovine skeletal muscle. In electron micrographs C-protein appears as short rods with a relatively uniform length of about 50 nm. Protein sequencing shows a single N-terminal sequence. Radiolabelled C-protein strongly decorates titin II and myosin rods but not myosin heads. Binding to titin n is retained in preparations lacking titin-associated proteins. Antibodies to bovine C-protein were used to screen a λgtll cDNA library constructed from fetal human skeletal muscle. Clone HC38 is 3833 bp long and encodes a protein of 1138 amino acid residues. The start of the predicted sequence fits the N-terminal sequence of the bovine protein. All partial sequences obtained from the bovine protein (348 residues) and the sequence deduced from a partial chicken cDNA (Einheber and Fischman,1990can be aligned along the human sequence. The sequences of human and chicken C-proteins share 50% identity and 70% similarity. Along the repeat patterns of the human protein the fibronectin (Fn)-like domains are better conserved than the immunoglobulin (Ig)-like domains. Regions of strong divergence between chicken fast C-protein and human slow C-protein may represent differences in C-protein isoforms.
Inhibitors of topoisomerases do not block the passage of human lymphocyte chromosomes through mitosis
Authors: Sumner A. T. 
Published: April 2021

ABSTRACT
Cultured human lymphocytes have been treated with a number of topoisomerase inhibitors, to see whether topoisomerase II is involved in the process of chromosome segregation at anaphase. Results were assessed by examination of cytogenetical preparations of spread chromosomes Four effects were observed, although no inhibitor produced all four effects. These effects were: inhibition of entry into mitosis; chromosome breakage and rearrangement; inhibition of chromosome condensation; and inhibition of chromosome segregation. Evidence for the last was ambiguous. Although there was evidence that separation of chromatids was affected when cells were treated with colchicine as well as topoisomerase II inhibitors (most notably with nalidixic acid, which resulted in complete fusion of the chromatids), no evidence was obtained to show that, in the absence of colchicine, cells treated with inhibitors could not proceed through anaphase normally. The topoisomerase I inhibitor, camptothecin, differed from the topoisomerase II inhibitors in not showing any effect on chromosome condensation or any significant effect on segregation.
Effects of okadaic acid on mitotic HeLa cells
Authors: Ghosh Sibdas , Paweletz Neidhard , Schroeter Dieter 
Published: April 2021

ABSTRACT
Mitotic HeLa cells were treated with different concentrations of okadaic acid (OA), known to inhibit phosphatase 1 and 2A activities. The cytological effects on the course of mitosis were studied at the light microscopic, immunofluorescence and electron microscopic levels. At the lowest concentration used (1 nM), OA did not show any effect on mitosis, but at higher concentrations it showed pronounced effects. The mitotic chromosomes became scattered, the mitotic spindle became deranged and the cells failed to enter anaphase. At the electron microscopic level formation of isolated microtubules and regular trilaminar kinetochores were observed. An extensive growth of the endoplasmic reticulum could be noted in these cells. Decondensation of chromatin and nuclear envelope re-formation could be seen only after withdrawal of OA. A high frequency of multinucleate cells could be found after 24 h of recovery. Cells treated with 100 nM OA for 3 hours showed diplochromosomes in over 50% of mitotic cells after 24 h recovery. These were presumably formed due to the failure of sister chromatid separation in the earlier mitosis in the presence of OA. At the electron microscopic level the diplochromosomes showed a quadruplet structure. The role of phosphatase 1 in controlling some late mitotic events, i.e. sister chromatid separation, MPF-inactivation and nuclear envelope re-formation etc., is discussed.
Nuclear F-actin: a functional component of baculovirus-infected lepidopteran cells?
Authors: Volkman Loy E. , Talhouk Salma N. , Oppenheimer Daniel I. , Charlton Carol A. 
Published: April 2021

ABSTRACT
Cellular functions known to involve actin are thought to occur in the cytoplasm. Even though actin has frequently been found in the nucleus, systems well-suited for studying the function of such nuclear actin are rare. We observed filamentous (F) actin within nuclei of IPLB-Sf-21 cells infected with Autographa caUfornica M nuclear polyhedrosis virus (AcMNPV) as detected by laser confocal microscopy using fluorescent phalloidin probes. The nuclear F-actin co-localized with the major capsid protein of the virus during normal infections. Cytochalasin D, known to interfere with nucleocapsid morphogenesis, uncoupled the apparent co-localization of F-actin and the capsid protein and inhibited infectious progeny production. Inhibition was reversible throughout infection (even in the presence of a protein synthesis inhibitor) and the nuclear co-localization of F-actin and the capsid protein was re-established upon removal of the drug. These observations suggest that nuclear Factin plays a role in virus replication, and that AcMNPV-infected cells may constitute a useful system in which to expand our understanding of nuclear actin transport and function.
Tubulin in bovine retinal rod outer segments
Authors: Matesic Diane F. , Philp Nancy J. , Murray John M. , Liebman Paul A. 
Published: April 2021

ABSTRACT
Bovine rod outer segment (ROS) preparations contain a major 58 kDa protein doublet that was identified by immunoblot as tubulin. Quantification by gel densitometry showed that the total amount of tubulin was 5- to 10-fold higher than that attributable to the rod axoneme, suggesting additional role(s) for tubulin in photoreceptor cells. Approximately 20% of this non-axonemal tubulin (15% of total tubulin) is tightly associated with outer segment membranes. This fraction remains membrane-associated after extensive low- or high-salt washing, requiring detergents or protein denaturants for release from ROS membranes. Unlike
ROS soluble tubulin it associates tightly with liposomes upon detergent solubilization and reconstitution. The ROS membrane-associated tubulin is highly enriched in isolated ROS plasma membrane fractions compared to the total outer segment membrane pool and can be localized to the plasma membrane but not to disks by immunofluorescent staining, suggesting a possible role in the structure or electrophysiology of the rod outer segment plasma membrane.
Cell guidance by micropatterned adhesiveness in vitro
Authors: Clark Peter , Connolly Patricia , Moores Geoffrey R. 
Published: April 2021

ABSTRACT
Tracks of adhesiveness are believed to be involved in guiding morphogenetic cell migrations. Here, electronics microfabrication technology was used to manufacture patterns of alternating tracks of adhesive and non-adhesive substratum (untreated fused quartz and adjacent parallel tracks of hydrophobic treatment) of varying period (4, 6, 12, 24 and 50 μm). These experimental substrata were used to model, in vitro, possible differentially adhesive guidance cues. The effect of such patterned substrata was assessed using fibroblastic BHK cells and epithelial MDCK cells. Cells were oriented and elongated by these surfaces. Their responsiveness was dependent on cell type, cell-cell interactions, and the geometry of the patterns. Alignment of BHK cells increased with increasing pattern period.
Single MDCK cells aligned to all pattern periods, their elongation being period-dependent, whereas colonies were mainly unaffected. These in vitro data have important implications, which are discussed with regard to in vivo guidance cues. The ability of cells to bridge over non-adhesive regions will influence the effectiveness of linear cues, and will be important for guideposting. The geometry of patterns of differential adhesion is here shown to be an important factor in determining the precision with which local guidance of cells may be controlled.
The role of the cytoskeleton and intercellular junctions in the transcellular membrane protein polarity of bovine aortic endothelial cells in vitro
Authors: Stolz Donna Beer , Bannish Gregory , Jacobson Bruce S. 
Published: April 2021

ABSTRACTThis project examines the transcellular membrane protein polarity of bovine aortic endothelial cell (BAEC) monolayers in vitro with respect to the roles that intercellular junctions (as defined by comparing confluent and subconfluent monolayers) and the submembranous cytoskeleton play in controlling this phenomenon. Plasma membrane (PM) proteins obtained from apical (AP) and basolateral (BL) PM domains of confluent BAEC monolayers were isolated using the cationic colloidal silica technique and resolved by twodimensional gel electrophoresis (2-D PAGE). To facilitate the identification of domain-specific PM proteins, an isoelectric point/molecular weight database of the proteins from AP and BL PM domains was constructed. Domain-specific PM proteins were assessed for their interaction with the cytoskeleton by determining whether they co-isolated with a Triton X-100 detergentresistant cytoskeletal/extracellular matrix fraction. The maintenance of polarized PM protein segregation by intercellular junctional complexes was determined by comparing AP and BL protein patterns of confluent monolayers with patterns generated by subconfluent monolayers, which lack such junctional structures. Proteins isolated from AP and BL PM domains from both confluent states were immunoblotted with antibodies to angiotensin-converting enzyme (ACE) and collagen receptors (CR). ACE was restricted exclusively to the AP PM domain in the subconfluent condition, even though no apparent cytoskeletal interaction was observed. CRs, found to interact with the cytoskeleton in either confluence state, were predominantly segregated to the BL PM domain regardless of the presence or absence of cell-cell contact. Membrane proteins found by 2-D PAGE to be asymmetrically distributed in the absence of intercellular junctions were assessed for cytoskeletal interaction by their inability to be extracted by Triton X-100 from monolayers in the subconfluent state. Computer cross-referencing of 2-D PAGE peak lists and immunodetection generated from the above fractionation protocols identifies a set of four proteins associated with the cytoskeleton that remain segregated in the proper domain, and five proteins associated with the cytoskeleton that become equally distributed between AP and BL PM domains in the absence of intercellular junctions. Additionally, six proteins not associated with the cytoskeleton remain asymmetrically distributed to the AP domain in the subconfluent state. The data suggest that BAEC monolayers have unknown mechanisms, apart from intercellular junctions expressed at confluency or cytoskeletal binding, for maintaining transcellular PM protein polarity.