scholarly journals Members of the syndecan family of heparan sulfate proteoglycans are expressed in distinct cell-, tissue-, and development-specific patterns.

1994 ◽  
Vol 5 (7) ◽  
pp. 797-805 ◽  
Author(s):  
C W Kim ◽  
O A Goldberger ◽  
R L Gallo ◽  
M Bernfield
Keyword(s):  
Heparan Sulfate ◽  
Family Member ◽  
Cultured Cells ◽  
Family Members ◽  
Cell Types ◽  
Heparan Sulfate Proteoglycans ◽  
Mrna Levels ◽  
Cell Tissue ◽  
Mouse Tissues ◽  
The Individual

The syndecans are a gene family of four transmembrane heparan sulfate proteoglycans that bind, via their HS chains, diverse components of the cellular microenvironment. To evaluate the expression of the individual syndecans, we prepared cDNA probes to compare mRNA levels in various adult mouse tissues and cultured mouse cells representing various epithelial, fibroblastic, endothelial, and neural cell types and B cells at various stages of differentiation. We also prepared antibody probes to assess whether the extracellular domains of the individual syndecans are shed into the conditioned media of cultured cells. Our results show that all cells and tissues studied, except B-stem cells, express at least one syndecan family member; most cells and tissues express multiple syndecans. However, each syndecan family member is expressed selectively in cell-, tissue-, and development-specific patterns. The extracellular domain of all syndecan family members is shed as an intact proteoglycan. Thus, most, if not all, cells acquire a distinctive repertoire of the four syndecan family members as they differentiate, resulting in selective patterns of expression that likely reflect distinct functions.

scholarly journals Distinctive populations of basement membrane and cell membrane heparan sulfate proteoglycans are produced by cultured cell lines.

1987 ◽  
Vol 105 (1) ◽  
pp. 529-539 ◽  
Author(s):  
J L Stow ◽  
M G Farquhar
Keyword(s):  
Extracellular Matrix ◽  
Cell Membrane ◽  
Basement Membrane ◽  
Heparan Sulfate ◽  
Cultured Cells ◽  
Cell Types ◽  
Heparan Sulfate Proteoglycans ◽  
Cell Layers ◽  
The Media ◽  
Membrane Type

We have investigated the nature and distribution of different populations of heparan sulfate proteoglycans (HSPGs) in several cell lines in culture. Clone 9 hepatocytes and NRK and CHO cells were biosynthetically labeled with 35SO4, and proteoglycans were isolated by DEAE-Sephacel chromatography. Heterogeneous populations of HSPGs and chondroitin/dermatan proteoglycans (CSPGs) were found in the media and cell layer extracts of all cultures. HSPGs were further purified from the media and cell layers and separated from CSPGs by ion exchange chromatography after chondroitinase ABC digestion. In all cell types, HSPGs were found both in the cell layers (20-70% of the total) as well as the medium. When the purified HSPG fractions were further separated by octyl-Sepharose chromatography, very little HSPG in the incubation media bound to the octyl-Sepharose, whereas 40-55% of that in the cell layers bound and could be eluted with 1% Triton X-100. This hydrophobic population most likely consists of membrane-intercalated HSPGs. Basement membrane-type HSPGs were identified by immunoprecipitation as a component (30-80%) of the unbound (nonhydrophobic) HSPG fraction. By immunofluorescence, basement membrane-type HSPGs were distributed in a reticular network in Clone 9 and NRK cell monolayers; by immunoelectron microscopy, these HSPGs were localized to irregular clumps of extracellular matrix located beneath and between cells. The cells did not produce a morphologically recognizable basement membrane layer under these culture conditions. When membrane-associated HSPGs were localized by immunoelectron microscopy, they were found in a continuous layer along the cell membrane of all cell types. The results demonstrate that two antigenically distinct populations of HSPG--an extracellular matrix and a membrane-intercalated population--are found at the surface of several different cultured cells lines; these populations can be distinguished from one another by differences in their distribution in the monolayers by immunocytochemistry and can be separated by hydrophobic chromatography; and basement membrane-type HSPGs are secreted and deposited in the extracellular matrix by cultured cells even though they do not produce a bona fide basement membrane-like layer.

scholarly journals Role of heparan sulfate proteoglycans in the binding and uptake of apolipoprotein E-enriched remnant lipoproteins by cultured cells

1993 ◽  
Vol 268 (14) ◽  
pp. 10160-10167
Author(s):  
Z.S. Ji ◽  
W.J. Brecht ◽  
R.D. Miranda ◽  
M.M. Hussain ◽  
T.L. Innerarity ◽  
...  
Keyword(s):  
Apolipoprotein E ◽  
Heparan Sulfate ◽  
Cultured Cells ◽  
Heparan Sulfate Proteoglycans ◽  
Remnant Lipoproteins

scholarly journals The mammalian 43-kD acetylcholine receptor-associated protein (RAPsyn) is expressed in some nonmuscle cells.

1989 ◽  
Vol 108 (5) ◽  
pp. 1833-1840 ◽  
Author(s):  
L S Musil ◽  
D E Frail ◽  
J P Merlie
Keyword(s):  
Skeletal Muscle ◽  
Acetylcholine Receptor ◽  
Cell Lines ◽  
Neuromuscular Junctions ◽  
Cell Types ◽  
Cardiac Cells ◽  
Mrna Levels ◽  
Receptor Associated Protein ◽  
Mouse Tissues ◽  
Nonmuscle Cells

Torpedo electric organ and vertebrate neuromuscular junctions contain the receptor-associated protein of the synapse (RAPsyn) (previously referred to as the 43K protein), a nonactin, 43,000-Mr peripheral membrane protein associated with the cytoplasmic face of postsynaptic membranes at areas of high nicotinic acetylcholine receptor (AChR) density. Although not directly demonstrated, several lines of evidence suggest that RAPsyn is involved in the synthesis and/or maintenance of such AChR clusters. Microscopic and biochemical studies had previously indicated that RAPsyn expression is restricted to differentiated, AChR-synthesizing cells. Our recent finding that RAPsyn is also produced in undifferentiated myocytes (Frail, D.E., L.S. Musil, a. Bonanno, and J.P. Merlie, 1989. Neuron. 2:1077-1086) led to to examine whether RAPsyn is synthesized in cell types that never express AChR (i.e., cells of other than skeletal muscle origin). Various primary and established rodent cell lines were metabolically labeled with [35S]methionine, and extracts were immunoprecipitated with a monospecific anti-RAPsyn serum. Analysis of these immunoprecipitates by SDS-PAGE revealed detectable RAPsyn synthesis in some (notably fibroblast and Leydig tumor cell lines and primary cardiac cells) but not all (hepatocyte- and lymphocyte-derived) cell types. These results were further substantiated by peptide mapping studies of RAPsyn immunoprecipitated from different cells and quantitation of RAPsyn-encoding mRNA levels in mouse tissues. RAPsyn synthesized in both muscle and nonmuscle cells was shown to be tightly associated with membranes. These findings demonstrate that RAPsyn is not specific to skeletal muscle-derived cells and imply that it may function in a capacity either in addition to or instead of AChR clustering.

scholarly journals Analysis and identification of the Grem2 heparin/heparan sulfate-binding motif

2017 ◽  
Vol 474 (7) ◽  
pp. 1093-1107 ◽  
Author(s):  
Chandramohan Kattamuri ◽  
Kristof Nolan ◽  
Thomas B. Thompson
Keyword(s):  
Heparan Sulfate ◽  
Bone Morphogenetic Proteins ◽  
Family Members ◽  
Binding Motif ◽  
Heparin Binding ◽  
Surface Binding ◽  
Cell Surface Binding ◽  
Binding Epitope ◽  
Lysine Residues ◽  
The Individual

Bone morphogenetic proteins (BMPs) are regulated by extracellular antagonists of the DAN (differential screening-selected gene aberrative in neuroblastoma) family. Similar to the BMP ligands, certain DAN family members have been shown to interact with heparin and heparan sulfate (HS). Structural studies of DAN family members Gremlin-1 and Gremlin-2 (Grem2) have revealed a dimeric growth factor-like fold where a series of lysine residues cluster along one face of the protein. In the present study, we used mutagenesis, heparin-binding measurements, and cell surface-binding analysis to identify lysine residues that are important for heparin/HS binding in Grem2. We determined that residues involved in heparin/HS binding, while not necessary for BMP antagonism, merge with the heparin/HS-binding epitope of BMP2. Furthermore, the Grem2–BMP2 complex has higher affinity for heparin than the individual proteins and this affinity is not abrogated when the heparin/HS-binding epitope of Grem2 is attenuated. Overall, the present study shows that the Grem2 heparin/HS and BMP-binding epitopes are unique and independent, where, interestingly, the Grem2–BMP2 complex exhibits a significant increase in binding affinity toward heparin moieties that appear to be partially independent of the Grem2 heparin/HS-binding epitope.

scholarly journals Isolation of an adhesion-mediating protein from chick neural retina adherons.

1985 ◽  
Vol 101 (3) ◽  
pp. 1071-1077 ◽  
Author(s):  
D Schubert ◽  
M LaCorbiere
Keyword(s):  
Cell Surface ◽  
Heparan Sulfate ◽  
Cultured Cells ◽  
Cell Types ◽  
Neural Retina ◽  
Cell Surface Receptor ◽  
Isolation And Characterization ◽  
Surface Receptor ◽  
Adhesive Interactions

Adherons are high molecular weight glycoprotein complexes which are released into the growth medium of cultured cells. They mediate the adhesive interactions of many cell types, including those of embryonic chick neural retina. The cell surface receptor for chick neural retina adherons has been purified, and shown to be a heparan sulfate proteoglycan (Schubert, D., and M. LaCorbiere, 1985, J. Cell Biol., 100:56-63). This paper describes the isolation and characterization of a protein in neural retina adherons which interacts specifically with the cell surface receptor. The 20,000-mol-wt protein, called retinal purpurin (RP), stimulates neural retina cell-substratum adhesion and prolongs the survival of neural retina cells in culture. The RP protein interacts with heparin and heparan sulfate, but not with other glycosaminoglycans. Monovalent antibodies against RP inhibit RP-cell adhesion as well as adheron-cell interactions. The RP protein is found in neural retina, but not in other tissues such as brain and muscle. These data suggest that RP plays a role in both the survival and adhesive interactions of neural retina cells.

Identification of the cells expressing cot proto-oncogene mRNA

1995 ◽  
Vol 108 (1) ◽  
pp. 97-103
Author(s):  
R. Ohara ◽  
S. Hirota ◽  
H. Onoue ◽  
S. Nomura ◽  
Y. Kitamura ◽  
...  
Keyword(s):  
Parotid Gland ◽  
Cultured Cells ◽  
Expression Patterns ◽  
Cell Types ◽  
Gene Expressions ◽  
Developmentally Regulated ◽  
Gastric Glands ◽  
Gastric Adenocarcinomas ◽  
Mouse Tissues ◽  
Glandular Cells

The cell types expressing cot proto-oncogene mRNA were identified by in situ hybridization (ISH) histochemistry. Among a variety of adult mouse tissues examined, four types of glandular cells expressing cot gene were identified: (1) granular duct cells in the submandibular and sublingual glands; (2) serous cells in the parotid gland; (3) peptic (chief) cells in gastric glands; and (4) goblet cells in colonic glands. Investigation of the developmentally regulated expression of cot mRNA using tissues of 14-day and 18-day embryos, newborn and weanling mice showed that cot gene is expressed only in morphologically differentiated and functionally activated cells of these four types. No other types of cells showing ISH signals were observed. Based on these results, cot gene expressions in cultured cells of colonic adenocarcinomas and gastric adenocarcinomas were examined. SW 480 and WiDr cells showed high expression of this gene and so should be useful for functional analysis of Cot kinase. The expression patterns of cot gene in tumor tissues of the parotid gland, and gastric and colonic glands were investigated. Two of the tissues overexpressed this gene markedly, suggesting that overproduction of Cot kinase may be one cause of their transformation.

scholarly journals The Microtubule Plus-End Proteins EB1 and Dynactin Have Differential Effects on Microtubule Polymerization

2003 ◽  
Vol 14 (4) ◽  
pp. 1405-1417 ◽  
Author(s):  
Lee A. Ligon ◽  
Spencer S. Shelly ◽  
Mariko Tokito ◽  
Erika L.F. Holzbaur
Keyword(s):  
Cultured Cells ◽  
Cell Types ◽  
Microtubule Dynamics ◽  
In Vitro Assays ◽  
Nucleation Effect ◽  
Microtubule Polymerization ◽  
The Individual ◽  
Different Cell Types ◽  
Dynactin Complex

Several microtubule-binding proteins including EB1, dynactin, APC, and CLIP-170 localize to the plus-ends of growing microtubules. Although these proteins can bind to microtubules independently, evidence for interactions among them has led to the hypothesis of a plus-end complex. Here we clarify the interaction between EB1 and dynactin and show that EB1 binds directly to the N-terminus of the p150Glued subunit. One function of a plus-end complex may be to regulate microtubule dynamics. Overexpression of either EB1 or p150Glued in cultured cells bundles microtubules, suggesting that each may enhance microtubule stability. The morphology of these bundles, however, differs dramatically, indicating that EB1 and dynactin may act in different ways. Disruption of the dynactin complex augments the bundling effect of EB1, suggesting that dynactin may regulate the effect of EB1 on microtubules. In vitro assays were performed to elucidate the effects of EB1 and p150Glued on microtubule polymerization, and they show that p150Gluedhas a potent microtubule nucleation effect, whereas EB1 has a potent elongation effect. Overall microtubule dynamics may result from a balance between the individual effects of plus-end proteins. Differences in the expression and regulation of plus-end proteins in different cell types may underlie previously noted differences in microtubule dynamics.

scholarly journals Characterization of Palladin, a Novel Protein Localized to Stress Fibers and Cell Adhesions

2000 ◽  
Vol 150 (3) ◽  
pp. 643-656 ◽  
Author(s):  
Mana M. Parast ◽  
Carol A. Otey
Keyword(s):  
Cultured Cells ◽  
Focal Adhesions ◽  
Cell Types ◽  
Cell Junctions ◽  
Stress Fibers ◽  
Western Blots ◽  
Embryonic Tissues ◽  
Mouse Tissues ◽  
Antisense Constructs

Here, we describe the identification of a novel phosphoprotein named palladin, which colocalizes with α-actinin in the stress fibers, focal adhesions, cell–cell junctions, and embryonic Z-lines. Palladin is expressed as a 90–92-kD doublet in fibroblasts and coimmunoprecipitates in a complex with α-actinin in fibroblast lysates. A cDNA encoding palladin was isolated by screening a mouse embryo library with mAbs. Palladin has a proline-rich region in the NH2-terminal half of the molecule and three tandem Ig C2 domains in the COOH-terminal half. In Northern and Western blots of chick and mouse tissues, multiple isoforms of palladin were detected. Palladin expression is ubiquitous in embryonic tissues, and is downregulated in certain adult tissues in the mouse. To probe the function of palladin in cultured cells, the Rcho-1 trophoblast model was used. Palladin expression was observed to increase in Rcho-1 cells when they began to assemble stress fibers. Antisense constructs were used to attenuate expression of palladin in Rcho-1 cells and fibroblasts, and disruption of the cytoskeleton was observed in both cell types. At longer times after antisense treatment, fibroblasts became fully rounded. These results suggest that palladin is required for the normal organization of the actin cytoskeleton and focal adhesions.

scholarly journals Tissue-Specific and Inducer-Specific Differential Induction of ISG56 and ISG54 in Mice

2007 ◽  
Vol 81 (16) ◽  
pp. 8656-8665 ◽  
Author(s):  
Fulvia Terenzi ◽  
Christine White ◽  
Srabani Pal ◽  
Bryan R. G. Williams ◽  
Ganes C. Sen
Keyword(s):  
Cultured Cells ◽  
Cell Types ◽  
Type I Interferons ◽  
Specific Cell ◽  
Type I ◽  
Mrna Levels ◽  
Tissue Specific ◽  
Interferon Stimulated Genes ◽  
Differential Pattern

ABSTRACT The interferon-stimulated genes (ISGs) ISG56 and ISG54 are strongly induced in cultured cells by type I interferons (IFNs), viruses, and double-stranded RNA (dsRNA), which activate their transcription by various signaling pathways. Here we studied the stimulus-dependent induction of both genes in vivo. dsRNA, which is generated during virus infection, induced the expression of both genes in all organs examined. Induction was not seen in STAT1-deficient mice, indicating that dsRNA-induced gene expression requires endogenous IFN. We further examined the regulation of these ISGs in several organs from mice injected with dsRNA or IFN-β. Both ISG56 and ISG54 were widely expressed and at comparable levels. However, in organs isolated from mice injected with IFN-α the expression of ISG54 was reduced and more restricted in distribution compared with the expression level and distribution of ISG56. When we began to study specific cell types, splenic B cells showed ISG54 but not ISG56 expression in response to all agonists. Finally, in livers isolated from mice infected with vesicular stomatitis virus, the expression of ISG56, but not ISG54, was induced; this difference was observed at both protein and mRNA levels. These studies have revealed unexpected complexity in IFN-stimulated gene induction in vivo. For the first time we showed that the two closely related genes are expressed in a tissue-specific and inducer-specific manner. Furthermore, our findings provide the first evidence of a differential pattern of expression of ISG54 and ISG56 genes by IFN-α and IFN-β.

scholarly journals The genetic association of the transcription factor NPAT with glycemic response to metformin involves regulation of fuel selection

PLoS ONE ◽  
2021 ◽  
Vol 16 (7) ◽  
pp. e0253533
Author(s):  
Changwei Chen ◽  
Jennifer R. Gallagher ◽  
Jamie Tarlton ◽  
Lidy van Aalten ◽  
Susan E. Bray ◽  
...  
Keyword(s):  
Ataxia Telangiectasia ◽  
Treatment Success ◽  
Cell Types ◽  
Underlying Mechanism ◽  
Metabolic Phenotype ◽  
Poor Responders ◽  
Mrna Levels ◽  
Mouse Tissues ◽  
First Choice ◽  
Primary Mouse

The biguanide, metformin, is the first-choice therapeutic agent for type-2 diabetes, although the mechanisms that underpin metformin clinical efficacy remain the subject of much debate, partly due to the considerable variation in patient response to metformin. Identification of poor responders by genotype could avoid unnecessary treatment and provide clues to the underlying mechanism of action. GWAS identified SNPs associated with metformin treatment success at a locus containing the NPAT (nuclear protein, ataxia-telangiectasia locus) and ATM (ataxia-telangiectasia mutated) genes. This implies that gene sequence dictates a subsequent biological function to influence metformin action. Hence, we modified expression of NPAT in immortalized cell lines, primary mouse hepatocytes and mouse tissues, and analysed the outcomes on metformin action using confocal microscopy, immunoblotting and immunocytochemistry. In addition, we characterised the metabolic phenotype of npat heterozygous knockout mice and established the metformin response following development of insulin resistance. NPAT protein was localised in the nucleus at discrete loci in several cell types, but over-expression or depletion of NPAT in immortalised cell models did not change cellular responses to biguanides. In contrast, metformin regulation of respiratory exchange ratio (RER) was completely lost in animals lacking one allele of npat. There was also a reduction in metformin correction of impaired glucose tolerance, however no other metabolic abnormalities, or response to metformin, were found in the npat heterozygous mice. In summary, we provide methodological advancements for the detection of NPAT, demonstrate that minor reductions in NPAT mRNA levels (20–40%) influence metformin regulation of RER, and propose that the association between NPAT SNPs and metformin response observed in GWAS, could be due to loss of metformin modification of cellular fuel usage.

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