bluuebunny/crossref_metadata_2025_split huggingface dataset:
Disassembly of the Drosophila nuclear lamina in a homologous cell-free system
Authors: Maus Nancy , Stuurman Nico , Fisher Paul A. 
Published: April 2021

ABSTRACT
Stage 14 Drosophila oocytes are arrested in first meiotic metaphase. A cell-free extract of these oocytes catalyzes apparent disassembly of purified Drosophila nuclei as well as of nuclear lamin polymers formed in vitro from isolated interphase lamins. Biochemically, the oocyte extract catalyzes lamin solubilization and phosphorylation as well as characteristic changes in one- and two-dimensional gel mobility. A previously unidentified soluble lamin isoform is easily seen after in vitro disassembly. This isoform is detectable but present only in very small quantities in vivo and is apparently derived specifically from one of the two interphase lamin isoforms. Cell-free nuclear lamina disassembly is ATP-dependent and addition of calcium to extracts blocks disassembly as judged both morphologically and biochemically. This system will allow enzymological characterization of cell-free lamina disassembly as well as molecular analysis of specific Drosophila mutants.
Role of E-cadherin in the response of tumor cell aggregates to lymphatic, venous and arterial flow: measurement of cell-cell adhesion strength
Authors: Byers Stephen W. , Sommers Connie L. , Hoxter Becky , Mercurio Arthur M. , Tozeren Aydin 
Published: April 2021

ABSTRACT
Defects in the expression or function of the calcium dependent cell-cell adhesion molecule E-cadherin are common in invasive, metastatic carcinomas. In the present study the response of aggregates of breast epithelial cells and breast and colon carcinoma cells to forces imposed by laminar flow in a parallel plate flow channel was examined. Although E-cadherin negative tumor cells formed cell aggregates in the presence of calcium, these were significantly more likely than E-cadherin positive cell aggregates to disaggregate in response to low shear forces, such as those found in a lymphatic vessel or venule (&lt;3.5 dyn/cm2). E-cadherin positive normal breast epithelial cells and E-cadherin positive breast tumor cell aggregates could not be disaggregated when exposed to shear forces in excess of those found in arteries (&gt;100 dyn/cm2). E-cadherin negative cancer cells which had been transfected with E-cadherin exhibited large increases in adhesion strength only if the expressed protein was appropriately linked to the cytoskeleton. These results show that E-cadherin negative tumor cells, or cells in which the adhesion molecule is present but is inefficiently linked to the cytoskeleton, are far more likely than E-cadherin positive cells to detach from a tumor mass in response to low shear forces, such as those found in a lymphatic vessel or venule. Since a primary route of dissemination of many carcinoma cells is to the local lymph nodes these results point to a novel mechanism whereby defects in cell-cell adhesion could lead to carcinoma cell dissemination
The in vivo role of annexin VII (synexin): characterization of an annexin VII deficient Dictyostelium mutant indicates an involvement in Ca2+-regulated processes
Authors: Döring Volker , Veretout Francoise , Albrecht Richard , Mühlbauer Bettina , Schlatterer Christina , Schleicher Michael , Noegel Angelika A. 
Published: April 2021

ABSTRACT
Dictyostelium discoideum cells harbor two annexin VII isoforms of 47 and 51 kDa which are present throughout development. In immunofluorescence and cell fractionation studies annexin VII was found in the cytoplasm and on the plasma membrane. In gene disruption mutants lacking both annexin VII isoforms growth, pinocytosis, phagocytosis, chemotaxis and motility were not significantly impaired under routine laboratory conditions, and the cells were able to complete the developmental cycle on bacterial plates. On non-nutrient agar plates development was delayed by three to four hours and a significant number of aggregates was no longer able to form fruiting bodies. Exocytosis as determined by measuring extracellular cAMP phosphodiesterase, α-fucosidase and α-mannosidase activity was unaltered, the total amounts of these enzymes were however lower in the mutant than in the wild type. The mutant cells were markedly impaired when they were exposed to low Ca2+ concentrations by adding EGTA to the nutrient medium. Under these conditions growth, motility and chemotaxis were severely affected. The Ca2+ concentrations were similar in mutant and wild-type cells both under normal and Ca2+ limiting conditions; however, the distribution was altered under low Ca2+ conditions in SYN- cells. The data suggest that annexin VII is not required for membrane fusion events but rather contributes to proper Ca2+ homeostasis in the cell.
An antisense oligodeoxyribonucleotide to m-calpain mRNA inhibits myoblast fusion
Authors: Balcerzak D. , Poussard S. , Brustis J. J. , Elamrani N. , Soriano M. , Cottin P. , Ducastaing A. 
Published: April 2021

ABSTRACT
Previous studies have led to the hypothesis of a possible role for m-calpain (EC 3.4.22.17) in myoblast fusion in culture in vitro. To support this hypothesis, an antisense strategy has been used with cultured primary rat myoblasts. Using an appropriate antisense oligodeoxyribonucleotide to mcalpain mRNA, an inhibition of myoblast fusion has been observed, the maximum being obtained when the cell culture was treated with 30 µM of oligomer. Synthesis of m-calpain was decreased by 48% while high concentrations of antisense oligonucleotide do not significantly affect myoblast proliferation. The specificity of m-calpain intervention during fusion has also been confirmed using antisense oligonucleotides to µ-calpain and p94 mRNAs, respectively.
A preadipose 3T3 cell variant highly sensitive to adipogenic factors and to human growth hormone
Authors: Salazar-Olivo L. A. , Castro-Muñozledo F. , Kuri-Harcuch W. 
Published: April 2021

ABSTRACT
We describe a new Swiss 3T3 preadipose clone, 3T3-F442A/C4, which shows higher sensitivity to serum adipogenic factors and to human growth hormone as compared to other 3T3 preadipose clones. The 3T3-F442A/C4 clone exhibited several characteristics different from the parental 3T3-F442A cells, mainly a high extent of adipose conversion under culture conditions that are nonadipogenic for the parental cells. The 3T3-F442A/C4 cells are not committed to undergo adipose differentiation, since they do not differentiate into adipocytes under serum-free or low-serum culture conditions, unless adipogenic factors or growth hormone are added into the culture medium. The 3T3-F442A/C4 cells showed 1.5-to 3.6-fold higher sensitivity to serum adipogenic factors and 5-to 6-fold higher sensitivity to human growth hormone as compared to the 3T3-F442A cells. The 3T3-F442A/C4 variant clone also differed from the parental clone by having a shorter population doubling time, an increased saturation density, and lower activity levels in some biochemical markers of adipose differentiation. On the other hand, the new variant clone has a similar proportion of cells susceptible to become adipocytes, and a similar response to insulin as compared to the parental cells. Our results show that the 3T3-F442A/C4 cells represent a new 3T3 preadipose clone that could be useful as a bioassay to evaluate growth hormone activity, as well as to purify and characterize hormones, adipogenic factors, and those compounds that affect mammalian adipogenesis.
Rapid sequestration of DPP IV/CD26 and other cell surface proteins in an autophagic-like compartment in Caco-2 cells treated with forskolin
Authors: Baricault Laurent , Fransen Jack A. M. , Garcia Martine , Sapin Catherine , Codogno Patrice , Ginsel Leo A. , Trugnan1 Germain 
Published: April 2021

ABSTRACT
The enterocytic differentiation of Caco-2 cells, a human colon adenocarcinoma cell line, is accompanied by the transcriptionally regulated expression of a subset of proteins and their correct sorting towards the cell surface. In the present work we have explored the possibility that post-translational events may interfere with this process by investigating the short term effects of a potent adenylyl cyclase activator, forskolin, on cell surface expression of dipeptidyl peptidase IV. Previous works have shown that this protein is targeted towards the apical domain through either a direct or an indirect route. Domain specific biochemical experiments demonstrate that cell surface expression of neosynthesized dipeptidyl peptidase IV rapidly decreases after a 1 hour forskolin treatment. Both initial basolateral and apical dipeptidyl peptidase IV membrane delivery were altered by forskolin treatment. Decrease of dipeptidyl peptidase IV cell surface expression was not restricted to this protein, since membrane expression of ‘525’ antigen, a basolateral protein and of sucrase-isomaltase, an apically targeted hydrolase, which unlike dipeptidyl peptidase IV mainly follows a direct route to the brush border membrane, also decreases. In addition endocytosis of proteins from the apical and from the basolateral domain was essentially unchanged, suggesting that forskolin’s target may be located on the exocytic pathway. Confocal laser scanning microscopy and immuno-electron microscopy studies demonstrate that, within 5 minutes of forskolin treatment, the cell surface proteins studied accumulate in intracellular vesicles which were co-labeled with a polyclonal antibody raised against Lamp-1, a lysosomal membrane marker. Electron microscopy studies show that these vesicles display an autophagic-like morphology. Finally, biochemical experiments indicate that dibutyryl cAMP does not mimick the forskolin effect, thus suggesting that it is a cAMP-independent phenomenon.
Mitochondrial genome repositioning during the differentiation of the african trypanosome between life cycle forms is microtubule mediated
Authors: Matthews Keith R. , Sherwin Trevor , Gull Keith 
Published: April 2021

ABSTRACT
The cell cycle of the African trypanosome requires a precise orchestration of nuclear and mitochondrial genome (kinetoplast) positioning to ensure faithful segregation during division. The controls underlying these events must be subject to modulation, however, as the respective positioning of these organelles changes during the parasite’s complex life cycle. We have studied mitochondrial DNA repositioning during differentiation between the trypanosome’s bloodstream and procyclic form. We have found that repositioning occurs simultaneously with the DNA replication phase of the cell cycle of the differentiating parasite. Furthermore, we demonstrate, at the cell and individual microtubule level, that this organelle repositioning is achieved via microtubule-dependent processes. Our results have implications for the control of cell differentiation and division in African trypanosomes.
Polar organization of gamma-tubulin in acentriolar mitotic spindles of Drosophila melanogastercells
Authors: Debec Alain , Détraves Claire , Montmory Colette , Géraud Gérard , Wright Michel 
Published: April 2021

ABSTRACT
The spindle pole localization of gamma-tubulin was compared in wild type and acentriolar cultured Drosophila cells using polyclonal antibodies specifically raised against the carboxy terminal amino acid sequence of Drosophila gamma-tubulin-1 (-KSEDSRSVTSAGS). During inter phase, γ-tubulin was present in the centrosome of wild type cells and accumulated around this organelle in a cell cycle dependent manner. In contrast, no such structure was observed in acentriolar cells. Wild type mitoses were homogeneously composed of biconical spindles, with two centrosome-associated γ-tubulin spots at the poles. The mitotic apparatuses observed in the acentriolar cells were heterogeneous; multipolar mitoses, bipolar mitoses with a barrel-shaped spindle and bipolar mitoses with biconical spindles were observed. In acentriolar cells, γ-tubulin accu mulation at mitotic poles was dependent on spindle micro tubule integrity. Most acentriolar spindles presented a dispersed γ-tubulin labeling at the poles. Only well polarized and biconical acentriolar spindles showed a strong γ-tubulin polar spot. Finally, acentriolar mitotic poles were not organized around true centrosomes. In contrast to wild type cells, in acentriolar cells the Bx63 centrosome-associated antigen was absent and the γ-tubulin containing material dispersed readily following micro tubule disassembly. These observations confirm that γ-tubulin plays an essential role in the nucleation of micro tubules even in the absence of mitotic polar organelles. In addition the data suggest that the mechanisms involved in the bipolarization of wild type and acentriolar mitoses are different, and that centrioles play a role in the spatial organization of the nucleating material containing γ-tubulin.
An antagonist of integrin αvβ3 prevents maturation of blood vessels during embryonic neovascularization
Authors: Drake Christopher J. , Cheresh David A. , Little Charles D. 
Published: April 2021

ABSTRACT
Experimental data in this study demonstrate that integrin αvβ3 is fundamentally involved in the maturation of blood vessels during embryonic neovascularization (vasculogenesis). Integrin αvβ3 was specifically expressed on the surface of angioblasts during vessel development in quail embryos and vitronectin, a ligand for αvβ3, localized to the basal surface of these cells. More importantly, microinjec tion of the anti-αvβ3 monoclonal antibody, LM609, disrupted the normal pattern of vascular development. After exposure to LM609 the angioblasts in experimental embryos appeared as clusters of rounded cells lacking normal cellular protrusions. This led to disruption of lumen formation and abnormal vessel patterning. These findings demonstrate that during vasculogenesis ligation of integrin αvβ3 on the surface of primordial endothelial cells is critical for the differentiation and maturation of blood vessels. Similar studies on chicken chorioallantoic membrane showed that LM609 blocks angiogenesis. Together the two studies suggest that integrin αvβ3 plays a role in neovascularization of tissues.
Giardia gene predicts a 183 kDa nucleotide-binding head-stalk protein
Authors: Marshall Jonathan , Holberton David V. 
Published: April 2021

ABSTRACT
Previously described extended proteins from the cytoskele ton of Giardia lamblia (β-giardin, median body protein) have been found to be segmented coiled coils with regular structural repeat patterns in their amino acid sequences. Screening a λZAPII library derived from Giardia genomic DNA with an antibody directed against a 34×103Mr giardin isoform selected a gene encoding a much larger polypep tide chain (HPSR2), the sequence of which was determined by chromosome walking the open reading frame. The complete gene has been cloned and expressed as a recom binant protein of 183×103Mr. The predicted amino acid sequence of the protein has identifiable features suggesting that it might be a motor protein with an amino-terminal hydrolytic domain attached to a long coiled coil stalk. The presumed head domain is 211 residues and contains a P-loop sequence conserved in purine nucleotide-binding proteins. The remaining 1409 amino acids mainly make up a region of heptad repeats such as in myosin or the kinesin stalk, ending in a small (67 amino acids) carboxy-terminal domain. Fourier analysis of the predicted stalk shows the presence of a strong physical repeat created by regular heptad phase changes dividing the coil into segments of 25 residues. This structure most closely resembles the smaller microtubule-associated median body protein which has segments of 24 residues.
Active cyclin B-cdc2 kinase does not inhibit DNA replication and cannot drive prematurely fertilized sea urchin eggs into mitosis
Authors: Genevière-Garrigues Anne-Marie , Barakat Abdelhamid , Dorée Marcel , Moreau Jean-Luc , Picard André 
Published: April 2021

ABSTRACT
Feedback mechanisms preventing M phase occurrence before S phase completion are assumed to depend on inhi bition of cyclin B-cdc2 kinase activation by unreplicated DNA. In sea urchin, fertilization stimulates protein synthesis and releases eggs from G1 arrest. We found that in the one-cell sea urchin embryo cyclin B-cdc2 kinase undergoes partial activation before S phase, reaching in S phase a level that is sufficient for G2-M phase transition. S phase entry is not inhibited by this level of cyclin B-dependent kinase activity. Inhibition of DNA replication by aphidicolin suppresses nuclear envelope breakdown, yet it does not prevent the microtubule array from being converted from its interphasic to its mitotic state. Moreover, mitotic cytoplasmic events occur at the same time in control and aphidicolin-treated embryos. Thus unreplicated DNA only prevents mitotic nuclear, not cyto plasmic, events from occurring prematurely. These results together show that the inhibition of cyclin B-cdc2 kinase activation is probably not the only mechanism that prevents mitotic nuclear events from occurring as long as DNA replication has not been completed. In contrast, cyto plasmic mitotic events seem to be controlled by a timing mechanism independent of DNA replication, set up at fer tilization, that prevents premature opening of a window for mitotic events.
Factors affecting the timing and imprinting of replication on a mammalian chromosome
Authors: Bickmore Wendy A. , Carothers Andrew D. 
Published: April 2021

ABSTRACT
Fluorescence in situ hybridisation has been used to follow replication of the short arm of human chromosome 11 using chromosome anomalies to distinguish the maternally- and paternally-derived homologues. The temporal difference in replication timing within and between chromosomes has been estimated by combining S phase detection with dual colour fluorescence in situ hybridisation. Proximal regions of 11p, including the WT1 gene, tend to replicate earlier on the maternally-derived chromosome than on the paternally-derived homologue. More distal parts of 11p (including the IGF2 gene) have the opposite imprint. The average difference in replication timing between homologous loci in the population of cells is small compared to the differences between loci along a single chromosome. The imprint is not strictly adhered to since many nuclei have hybridisation patterns opposite to the trend within the population. The nature of the imprinting signal has been investigated. Absolute replication time, but not the imprint, was affected by azacytidine, an inhibitor of DNA methylation. The replication imprint was modified by treatments that inhibit histone deacetylation. We suggest that replication imprinting reflects differences in chromatin structure between homologues.
The roles of nucleolar structure and function in the subcellular location of the HIV-1 Rev protein
Authors: Dundr Miroslav , Leno Gregory H. , Hammarskjöld Marie-Louise , Rekosh David , Helga-Maria C. , Olson Mark O. J. 
Published: April 2021

ABSTRACT
The human immunodeficiency virus 1 (HIV-1) Rev transactivator protein plays a critical role in the regulation of expression of structural proteins by controlling the pathway of mRNA transport. The Rev protein is located predominantly in the nucleoli of HIV-1 infected or Rev-expressing cells. Previous studies demonstrated that the Rev protein forms a specific complex in vitro with protein B23 which is suggested to be a nucleolar receptor and/or carrier for the Rev protein. To study the role of the nucleolus and nucleolar proteins in Rev function, transfected COS-7 or transformed CMT3 cells expressing the Rev protein were examined for subcellular locations of Rev and other proteins using indirect immunofluorescence and immunoelectron microscopy. One day after transfection the Rev protein was found in most cells only in the nucleolar dense fibrillar and granular components where it colocalized with protein B23. These were designated class 1 cells. In a second class of cells Rev and B23 accumulated in the nucleoplasm as well as in nucleoli. Treatment of class 1 cells with actinomycin D (AMD) under conditions that blocked only RNA polymerase I transcription caused Rev to completely redistribute from nucleoli to the cytoplasm. Simultaneously, protein B23 was partially released from nucleoli, mostly into the nucleoplasm, with detectable amounts in the cytoplasm. In cells recovering from AMD treatment in the presence of cycloheximide Rev and B23 showed coincident relocation to nucleoli. Class 2 cells were resistant to AMD-induced Rev redistribution. Selective inhibition of RNA polymerase II transcription by α-amanitin or by DRB did not cause Rev to be released into the cytoplasm suggesting that active preribosomal RNA transcription is required for the nucleolar location of Rev. However, treatment with either of the latter two drugs at higher doses and for longer times caused partial disruption of nucleoli accompanied by translocation of the Rev protein to the cytoplasm. These results suggest that the nucleolar location of Rev depends on continuous preribosomal RNA transcription and a substantially intact nucleolar structure.
cAMP increasing agents prevent the stimulation of heat-shock protein 70 (HSP70) gene expression by cadmium chloride in human myeloid cell lines
Authors: Vilaboa Nuria E. , Calle Consuelo , Pérez Concepción , Blas Elena de , García-Bermejo Laura , Aller Patricio 
Published: April 2021

ABSTRACT
Treatment of U-937 human promonocytic cells with the cAMP increasing agents isoproterenol plus theophylline decreased the basal level of heat-shock protein 70 (HSP70) mRNA. In addition, the cAMP increasing agents attenuated the increase in HSP70 mRNA and protein levels produced by cadmium chloride in U-937 and other human myeloid cell lines, reduced the capacity of cadmium treatment to generate stress-tolerance, and attenuated the cadmium-produced stimulation of heat-shock factor (HSF) binding activity. By contrast, isoproterenol plus theophylline failed to attenuate the stimulation of HSP70 gene expression and HSF binding activity caused by heat-shock. Isoproterenol plus theophylline did not prevent the uptake of cadmium into the cells, and increased to a similar extent the intracellular cAMP levels in cadmium- and heat-treated cells. The cAMP increasing agents reduced the induction by cadmium of the HSP27 stress gene, but failed to attenuate other cadmium-elicited stress reactions such as the inhibition of total protein synthesis. It is concluded that cAMP does not inhibit the stress response as a whole, but it interferes with some step of the pathway by which cadmium specifically stimulates HSF binding activity and as a consequence HSP70 gene expression, in human myeloid cell lines.
The signal recognition particle and related small cytoplasmic ribonucleoprotein particles
Authors: Bovia Fabrice , Strub Katharina 
Published: April 2021

ABSTRACT
Recently, a number of novel small cytoplasmic ribonucleoprotein particles have been identified that comprise RNA and protein subunits related to the signal recognition particle (SRP). Here we discuss the latest results on the structure and functions of SRP together with the structures and putative functions of the novel SRP-related ribonucleoprotein particles.