bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dynamic chromatin: The regulatory domain organization of eukaryotic gene loci
Authors: Bonifer Constanze , Hecht Andreas , Saueressig Harald , Winter Diana M. , Sippel Albrecht E. 
Published: May 2005

AbstractIt is hypothesized that nuclear DNA is organized in topologically constrained loop domains defining basic units of higher order chromatin structure. Our studies are performed in order to investigate the functional relevance of this structural subdivision of eukaryotic chromatin for the control of gene expression. We used the chicken lysozyme gene locus as a model to examine the relation between chromatin structure and gene function. Several structural features of the lysozyme locus are known: the extension of the region of general DNAasel sensitivity of the active gene, the location of DNA‐sequences with high affinity for the nuclear matrix in vitro, and the position of DNAasel hypersensitive chromatin sites (DHSs). The pattern of DHSs changes depending on the transcriptional status of the gene. Functional studies demonstrated that DHSs mark the position of cis‐acting regulatory elements. Additionally, we discovered a novel cis‐activity of the border regions of the DNAasel sensitive domain (A‐elements). By eliminating the position effect on gene expression usually observed when genes are randomly integrated into the genome after transfection, A‐elements possibly serve as punctuation marks for a regulatory chromatin domain. Experiments using transgenic mice confirmed that the complete structurally defined lysozyme gene domain behaves as an independent regulatory unit, expressing the gene in a tissue specific and position independent manner. These expression features were lost in transgenic mice carrying a construct, in which the A‐elements as well as an upstream enhancer region were deleted, indicating the lack of a locus activation function on this construct. Experiments are designed in order to uncover possible hierarchical relationships between the different cis‐acting regulatory elements for stepwise gene activation during cell differentiation. We are aiming at the definition of the basic structural and functional requirements for position independent and high level gene expression. The result of these experiments will have important consequences for random gene transfer with predictable and reproducible expression of transgenes.
The nuclear matrix: A heuristic model for investigating genomic organization and function in the cell nucleus
Authors: Berezney Ronald 
Published: May 2005

AbstractDespite significant advances in deciphering the molecular events underlying genomic function, our understanding of these intergrated processes inside the functioning cell nucleus has, until recently, met with only very limited success. A major conundrum has been the “layers of complexity” characteristic of all cell structure and function. To understand how the cell nucleus functions, we must also understand how the cell nucleus is put together and functions as a whole. The value of this neo‐holistic approach is demonstrated by the enormous progress made in recent years in identifying a wide variety of nuclear functions associated with the nuclear matrix. In this article we summarize basic properties of in situ nuclear structure, isolated nuclear matrix systems, nuclear matrix‐associated functions, and DNA replication in particular. Emphasis is placed on identifying current problems and directions of research in this field and illustrating the intrinsic heuristic value of this global approach to genomic organization and function.
DNA and RNA within the nucleus: How much sequence‐specific spatial organization?
Authors: Carter Kenneth C. , Lawrence Jeanne Bentley 
Published: May 2005

AbstractSpatial organization of various nuclear components is often proposed as means by which nuclei more efficiently carry out their various tasks. Such functional compartmentalization may involve a sequence‐specific packaging and placement of DNA and RNA. Here we review recent insights, allowed primarily by advances in fluorescent in situ hybridization methodology, into the organization of nucleic acids within individual nuclei.
Influence of dexamethasone on the vitamin D–mediated regulation of osteocalcin gene expression
Authors: Schepmoes Grace , Breen Ellen , Owen Thomas A. , Aronow Michael A. , Stein Gary S. , Lian Jane B. 
Published: May 2005

AbstractThe influence of dexamethasone on expression of the osteocalcin gene which encodes the most abundant non‐collagenous and only reported bone‐specific protein was examined in ROS 17/2.8 osteosarcoma cells which express a broad spectrum of genes related to bone formation. Consistent with previous reports, quantitation of cellular osteocalcin mRNA levels by Northern blot analysis, osteocalcin gene transcription by activity of the osteocalcin gene promoter fused to a chloramphenicol acetyl‐transferase (CAT) mRNA coding sequence following transfection into ROS 17/2.8 cells, and osteocalcin biosynthesis by radioimmunoassay indicate that dexamethasone in a concentration range of 10‐6to 10‐9M only modestly modifies basal levels of osteocalcin gene expression. However, dexamethasone significantly inhibits these parameters of the vitamin D–induced upregulation of osteocalcin gene expression in both proliferating and in confluent ROS 17/2.8 cells. In this study, we observed that the extent to which abrogation of the vitamin D response occurs is dependent on basal levels of osteocalcin gene expression as reflected by a complete inhibition of the vitamin D–induced upregulation in a ROS 17/2.8K subline with low basal expression and only a partial reduction of the vitamin D stimulation in a ROS 17/2.8C subline with eightfold higher levels of basal expression. This effect of glucocorticoid appears to be at the transcriptional and post‐transcriptional levels as demonstrated by a parallel decline in the cellular representation of osteocalcin mRNA, osteocalcin gene promoter activity, and osteocalcin biosynthesis. The complexity of the glucocorticoid effect on vitamin D–mediated transcriptional properties of the osteocalcin gene is indicated by persistence of sequence‐specific protein‐DNA interactions at two principal osteocalcin gene promoter regulatory elements, the osteocalcin (CCAAT) box which modulates basal level of transcription, and the vitamin D responsive element, where vitamin D–mediated enhancement of osteocalcin gene transcription is controlled.
Effect of monensin on ricin and fluid phase transport in polarized MDCK cells
Authors: Melby Elisabeth Løkken , Prydz Kristian , Olsnes Sjur , Sandvig Kirsten 
Published: May 2005

AbstractThe effect of monensin on endocytosis, transcytosis, recycling and transport to the Golgi apparatus in filter‐grown Madin‐Darby canine kidney (MDCK) cells was investigated using 125I‐labeled ricin as a marker for membrane transport, and horseradish peroxidase (HRP) as a marker for fluid phase transport. Monensin (10 μM) stimulated transcytosis of both markers about 3‐fold in the basolateral to apical direction. Transcytosis of HRP in the opposite direction, apical to basolateral, was reduced to approximately 50% of the control by monensin, whereas that of ricin was slightly increased. Recycling of markers endocytosed from the apical surface was reduced in the presence of monensin and there was an increased accumulation of both ricin and HRP in the cells. Transport of ricin to the Golgi apparatus increased to the same extent as the increase in intracellular accumulation. No change in recycling or accumulation was observed with monensin when the markers were added basolaterally, but transport of ricin to the Golgi apparatus increased almost 3‐fold. Our results indicate that basolateral to apical transcytosis is increased in the absence of low endosomal pH, and they suggest that apical to basolateral transcytosis of a membrane‐bound marker (ricin) is affected by monensin differently from that of a fluid phase marker (HRP).
Engineering T lymphocyte antigen specificity
Authors: Bolhuis Reinder L. H. , Sturm Els , Gratama Jan Willem , Braakman Eric 
Published: May 2005

AbstractTargeting of immune cells by bispecific antibodies has proven to be a powerful tool for the investigation of cellular cytotoxicity, lymphocyte activation and induction of cytokine production, as well as to represent an innovative form of immunotherapy for the treatment of cancer. The hallmark of this approach is the use of the specificity of monoclonal antibodies to join target and immune cells by virtue of the dual specificity of bispecific antibodies for the two entities. More precisely, the bispecific antibody has two different binding sites, which are capable of recognizing tumor associated antigens on the one hand and lymphocyte activation sites on the other. This process of crosslinking results in the activation of the lymphocyte and triggering of its lytic machinery, as well as lymphokine production. A major advantage of this therapeutic modality is, that use is made of the normal cellular immune defence system and therefore is only associated with minor toxicity. The distinct lymphocyte populations, which can be used for adoptive immunotherapy and the various bispecific antibody preparations, as well as the chimeric immunoglobulin/T cell receptor construction, are the major topics of this review.
Effect of cytokines and lipopolysaccharide on CD14 antigen expression in human monocytes and macrophages
Authors: Landmann R. , Ludwig C. , Obrist R. , Obrecht J. P. 
Published: May 2005

AbstractThe 52 kD myeloid membrane glycoprotein CD14 represents the receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein (LBP); it is involved in LPS induced tumor necrosis factor‐alpha production. Expression of CD14 increases in monocytes differentiating into macrophages, and it is reduced by rIFNg in monocytes in vitro. In the present study CD14 membrane antigen expression was investigated in cultures of human mononuclear leucocytes (PBL), in elutriated, purified monocytes, and in blood monocyte derived Teflon cultured macrophages. Cells were incubated for 15 or 45 h with rIL‐1, rIL‐2, rIL‐3, rIL‐4, rIL‐5, rIL‐6, rTNFa, rGM‐CSF, rM‐CSF, rTGFb1, rIFNa, lipopolysaccharide (LPS), and, as a control, rIFNg. The monoclonal antibodies Leu‐M3 and MEM 18 were used for labelling of CD14 antigen by indirect immunofluorescence and FACS analysis of scatter gated monocytes or macrophages. IFNg concentrations were determined in PBL culture supernatants by ELISA. rIFNa and rIL‐2 reduced CD14 in 15 and 45 h PBL cultures, an effect mediated by endogenous IFNg, since it was abolished by simultaneous addition of an anti‐IFNg antibody. rIFNa and rIL‐2 were ineffective in purified monocytes or macrophages. rIL‐4 strongly reduced CD14 in PBL and purified monocytes after 45 h, whereas in macrophages the decrease was weak, although measurable after 15 h. The other cytokines investigated did not change CD14 antigen expression. Cycloheximide alone reduced CD14, but when added in combination with rIFNg the effect on CD14 downregulation was more pronounced. The effect of rIFNg on CD14 in PBL cultures was dose‐dependently inhibited by rIL‐4 and this inhibition is probably due to an IL‐4 mediated blockade of IFNg secretion. LPS at a low dose increased CD14, at a high dose it produced a variable decrease of CD14 in PBL, which was probably due to LPS induced IFNg secretion. LPS strongly enhanced CD14 in 45 h cultures of purified monocytes. The results, showing that CD14 antigen expression is upregulated by LPS and downregulated by rIFNg and rIL‐4, suggest that the LPS‐LBP receptor is involved in the feedback response of IFNg and IL‐4 to LPS stimulation.
Protein‐Tyrosine phosphatases and the regulation of insulin action
Authors: Goldstein Barry J. 
Published: May 2005

AbstractProtein‐tyrosine phosphatases (PTPases) play an important role in the regulation of insulin action by dephosphorylating the active (autophosphorylated) form of the insulin receptor and attenuating its tyrosine kinase activity. PTPases can also modulate post‐receptor signalling by catalyzing the dephosphorylation of cellular substrates of the insulin receptor kinase. Dramatic advances have recently been made in our understanding of PTPases as an extensive family of transmembrane and intracellular proteins that are involved in a number of pathways of cellular signal transduction. Identification of the PTPase(s) which act on various components of the insulin action cascade will not only enhance our understanding of insulin signalling but will also clarify the potential involvement of PTPases in the pathophysiology of insulin‐resistant disease states. This brief review provides a summary of reversible tyrosine phosphorlyation events in insulin action and available data on candidate PTPases in liver and skeletal muscle that may be involved in the regulation of insulin action.
Insulin/IGF‐1 hybrid receptors: Implications for the dominant‐negative phenotype in syndromes of insulin resistance
Authors: Frattali Anne L. , Treadway Judith L. , Pessin Jeffrey E. 
Published: May 2005

AbstractClassical insulin and IGF‐1 receptors are α2β2 heterotetrameric complexes synthesized from two identical αβ half‐receptor precursors [1,2]. Recent data strongly suggests, however, that nonidentical αβ half‐receptor precursors can assemble to generate hybrid holoreceptor species both in vivo and in vitro [3–6,41]. This review focuses primarily on two types of hybrid receptors. The first type is an insulin/IGF‐1 hybrid receptor generated by the association of an αβ insulin half‐receptor with an αβ IGF‐1 half‐receptor. The second type is one formed from a wildtype (kinase‐active) insulin or IGF‐1 αβ half‐receptor and a mutant (kinase‐inactive) insulin αβ half‐receptor. Although the functional properties of insulin/IGF‐1 hybrid receptors have not yet been completely defined, wildtype/mutant hybrid receptors are essentially substrate kinase inactive [6]. These data indicate that the mutant αβ half‐receptor exerts a transdominant inhibition upon the wildtype αβ half‐receptor within the α2β2 holoreceptor complex. This defect in substrate kinase activity may contribute to the molecular defect underlying some syndromes of severe insulin resistance and diabetes. Heterozygous individuals expressing both wildtype and mutant tyrosine kinase‐defective insulin receptor precursors demonstrate varying degrees of insulin resistance and diabetes [7–11]. In addition, cell lines which express both endogenous wildtype and transfected kinase‐defective insulin receptors display markedly decreased insulin and IGF‐1 sensitivity and responsiveness [12–14]. Formation of hybrid receptors which results in premature termination of insulin signal transduction may be one mechanism underlying the observation that kinase‐inactive receptors inhibit the function of native receptors.
Acute and chronic signals controlling glucose transport in skeletal muscle
Authors: Klip Amira , Marette André 
Published: May 2005

AbstractGlucose transport into muscle cells occurs through facilitated diffusion mediated primarily by the GLUT1 and GLUT4 glucose transporters. These transporter proteins are controlled by acute and chronic exposure to insulin, glucose, muscle contraction, and hypoxia. We propose that acute responses occur through recruitment of pre‐formed glucose transporters from an intracellular storage site to the plasma membrane. In contrast, chronic control is achieved by changes in transporter biosynthesis and protein stability. Using subcellular fractionation of rat skeletal muscle, recruitment of GLUT4 glucose transporters to the plasma membrane is demonstrated by acute exposure to insulin in vivo. The intracellular pool appears to arise from a unique organelle depleted of transverse tubule, plasma membrane, or sarcoplasmic reticulum markers. In diabetic rats, GLUT4 content in the plasma membranes and in the intracellular pool is reduced, and incomplete insulin‐dependent GLUT4 recruitment is observed, possibly through a defective incorporation of transporters to the plasma membrane. The lower content of GLUT4 transporters in the muscle plasma membranes is reversed by restoration of normoglycemia with phlorizin treatment.In some muscle cells in culture, GLUT1 is the only transporter expressed yet they respond to insulin, suggesting that this transporter can also be regulated by acute mechanisms. In the L6 muscle cell line, GLUT1 transporter content diminishes during myogenesis and GLUT4 appears after cell fusion, reaching a molar ratio of about 1:1 in the plasma membrane. Prolonged exposure to high glucose diminishes the amount of GLUT1 protein in the plasma membrane by both endocytosis and reduced biosynthesis, and lowers GLUT4 protein content in the absence of changes in GLUT4 mRNA possibly through increased protein degradation. These studies suggest that the relative contribution of each transporter to transport activity, and the mechanisms by which glucose exerts control of the glucose transporters, will be key subjects of future investigations.
Acetyl‐CoA carboxylase in reuber hepatoma cells: Variation in enzyme activity, insulin regulation, and cellular lipid content
Authors: Bianchi Antonella , Evans Joseph L. , Nordlund Ann‐Charlotte , Watts Thomas D. , Witters Lee A. 
Published: May 2005

AbstractRuber hapatoma cells are useful cultured lines for the study of insulin action, lipid and lipoprotein metabolism, and the regulation of acetyl‐CoA carboxylase (ACC), the rate‐limiting enzyme of faty acid biosynthesis. During investigations in deifferent clonal lines of these of these cells, we have uncovered marked intercellular variability in the activity, enzyme content, and insulin regulation of ACC paralleled by differences in cellular neutral lipid (triglyceride) content. Two contrasting clonal liners, Fao and H356A‐1, have been studied in detail. Several features distinguish these two lines, including differences in ACC activity and enzyme kinetics, the content of the two major hepatic ACC isozymes (M, 280,000 and 265,000 Da) and their heteroisozymic complex, the extent of ACC phosphorylation, and the ability of ACC to be activated on stimulation by insulin and insulinomimetic agonists. As studied by Nile Red staining and fluorscence‐activated cell sorting, these two lines also display marked differences in neutral lipid content, which correlates with both basal levels of ACC activity and inhobition of ACC by the fatty acid analog, 5‐(tetradecyloxy)‐2‐furoic and (TOFA). Thsese results emphasize the importance of characterization of any particular clonal line of Reuber cells for studies of enzyme regulation, substrate metabolism, and hormone action. With respect to ACC, studies in contrasting colonal lines of Ruber cells could provide valuable clues to understanding both the complex mechanisms of intracellular ACC regulation in the absence and presence of hormones and its regulatory role (s) in overla hepatic lipid metabolism.
mRNA association with the cytoskeletal framework likely represents a physiological binding event
Authors: Biegel Diane , Pachter Joel S. 
Published: May 2005

AbstractA multitude of studies has indicated that the vast majority of mRNA and polyribosomes is associated with the detergent‐resistant cytoskeletal faramework (CSK). However, the nature and purpose of this association remain unclear. To begin unraveling the factors which may medicate this phenomenon, we examined the extent of association of four mRNAs (ubulin, vimentin, actin, and histone mRNA) with the CSKs NIH T3 cells over a wide range of salt concentrations. Results indicate that the vast majority ( > 90%) of each of these mRNAs remains associated with the CSK after detergent extration of cells in low ionic strength buffer (25 mM NaCI). This association is manifest under conditions that cause the complete depolymerization of microtubules but that leave microfilaments and intermediate filaments intact. Even after extensive washing in buffer of approximately physiological ionic strength (150 mM NaCI), 75–85% of these mRNAs still remain associated with the CSK. However, at least 50% of each of these mRNAs can be eluted from the CSK by washing with buffer containing 250 mM NaCI. Not all the mRNAs, though, display the same elution profile. This suggests that different binding sites and/or different binding affinities may exist for different mRNAs. Surprisingly, close to 50% of the polyribosome population remains bound to the CSK despite washing in as much as 1.0 M NaCI. These adherent polyribosomes appear to be of the same size as those that are eluted, alllaying the possibility that they are retained by the CSK nor physically trapped within the meshwork of cytoskeletal filaments. Instead, mRNA appears to be bound to the CSK via interactions that are, at least partially, ionic in nature and strong enough to exist intracellulary. Moreover, the binding sites for these mRNAs on the CSK are not associated with microtubules, but may be associated with wither the microflament or intermedicate filament network.
Identification of a core motif that is recognized by three members of the HMG class of transcriptional regulators: IRE‐ABP, SRY, and TCF‐1α
Authors: Alexander‐Bridges M. , Ercolani Louis , Kong X. F. , Nasrin Nargis 
Published: May 2005

AbstractInsulin induces glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) gene transcription in part by regulating one or more proteins that bind a cis‐acting element, IRE‐A. We have recently cloned a protein, IRE‐ABP, that binds the IRE‐A element. IRE‐ABP is a member of the HMG class of transcriptional regulators and is 67% identical within its HMG box domain to the candidate gene for the testis‐determining factor, SRY. IRE‐ABP and SRY share binding specificity for the IRE‐A motif. This sequence is also highly conserved with a core motif, 5′‐Py‐ctttg(a/t)‐3′, contained in T‐cell specific genes that have high affinity for TCF‐1α, another member of the HMG class of transcriptional regulators. Thus, diverse members of the HMG family interact with similar nucleotide sequences to regulate expression of genes that initiate and maintain the differentiated phenotype. We have found this core motif in the upstream region of many genes that are positively and negatively regulated by insulin. These observations suggest that IRE‐ABP or a related family member may coordinate the expression of these genes. The HMG family of proteins has diverse functions ranging from the regulation of differentiation and mating type in yeast to the regulation of tissue‐and species‐specific gene expression in mammals. Insulin regulates GAPDH gene transcription in a tissue‐specfic manner. We propose that members of the IRE‐ABP family play an important role in controlling tissue specificity of the insulin response.