Endogenous endothelial cell nitric-oxide synthase modulates apoptosis in cultured breast cancer cells and is transcriptionally regulated by p53.
AuthorsMortensen K,Skouv J,Hougaard DM,Larsson LI
JournalThe Journal of biological chemistry
PubMed ID10608825
In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue.
AuthorsStrappe PM, Wang TH, McKenzie CA, Lowrie S, Simmonds P, Bell JE
JournalJ Virol Methods
PubMed ID9562406
A direct in situ polymerase chain reaction (IS-PCR) assay is described for the detection of HIV-1 proviral DNA in formalin fixed paraffin embedded brain tissue. Biotin-16-dUTP is incorporated during the PCR process and microwave pretreatment of tissue sections ensures that no non-specific incorporation into damaged or nicked genomic DNA occurs. ... More
SPACRCAN, a novel human interphotoreceptor matrix hyaluronan-binding proteoglycan synthesized by photoreceptors and pinealocytes.
AuthorsAcharya S, Foletta VC, Lee JW, Rayborn ME, Rodriguez IR, Young WS, Hollyfield JG
JournalJ Biol Chem
PubMed ID10702256
The interphotoreceptor matrix is a unique extracellular complex occupying the interface between photoreceptors and the retinal pigment epithelium in the fundus of the eye. Because of the putative supportive role in photoreceptor maintenance, it is likely that constituent molecules play key roles in photoreceptor function and may be targets for ... More
Detection of low copy numbers of HPV DNA by fluorescent in situ hybridization combined with confocal microscopy as an alternative to in situ polymerase chain reaction.
AuthorsLizard G, Chignol MC, Souchier C, Roignot P, Chardonnet Y, Schmitt D
JournalJ Virol Methods
PubMed ID9672129
'In genital lesions infected by human papillomavirus (HPV), histological criteria and HPV DNA typing are of prognostic value. Therefore, non-radioactive methods such as in situ hybridization are used extensively since they preserve the histological organization of the tissue, and allow the detection and characterization of HPV DNA. However, the sensitivity ... More
Ectopic expression of Drosophila ELAV and human HuD in Drosophila wing disc cells reveals functional distinctions and similarities.
AuthorsToba G, Qui J, Koushika SP, White K
JournalJ Cell Sci
PubMed ID12006625
'Drosophila ELAV and human HuD are two neuronal RNA binding proteins that show remarkable sequence homology, yet differ in their respective documented roles in post-transcriptional regulation. ELAV regulates neural-specific alternative splicing of specific transcripts, and HuD stabilizes specific mRNAs that are otherwise unstable due to AU-rich elements (AREs) in their ... More
Regulation of chemotactic and proadhesive responses to chemoattractant receptors by RGS (regulator of G-protein signaling) family members.
AuthorsBowman EP, Campbell JJ, Druey KM, Scheschonka A, Kehrl JH, Butcher EC
JournalJ Biol Chem
PubMed ID9774420
'Serpentine Galphai-linked receptors support rapid adhesion and directed migration of leukocytes and other cell types. The intracellular mechanisms mediating and regulating chemoattractant-directed adhesion and locomotion are only now beginning to be explored. RGS (for regulator of G-protein signaling) proteins are a recently described family that regulate Galphai-stimulated pathways by acting ... More
MAHRP-1, a novel Plasmodium falciparum histidine-rich protein, binds ferriprotoporphyrin IX and localizes to the Maurer's clefts.
AuthorsSpycher C, Klonis N, Spielmann T, Kump E, Steiger S, Tilley L, Beck HP
JournalJ Biol Chem
PubMed ID12815049
'Using a stage-specific cDNA library from Plasmodium falciparum we have identified a gene coding for a novel histidine-rich protein (MAHRP-1). The gene is exclusively transcribed during early erythrocyte stages and codes for a small transmembrane protein. The C-terminal region contains a polymorphic stretch of histidine-rich repeats. Fluorescence microscopy studies using ... More
Catalyzed reporter deposition, a novel method of signal amplification. II. Application to membrane immunoassays.
AuthorsBobrow MN, Shaughnessy KJ, Litt GJ
JournalJ Immunol Methods
PubMed ID1849153
'In a previous publication (Bobrow et al., J. Immunol. Methods (1989) 279-285), we described a novel signal amplification method, catalyzed reporter deposition (CARD), and its application to microplate immunoassays. The method utilizes the analyte-dependent reporter enzyme (ADRE) to catalyze the deposition of additional reporter onto the surface of a solid-phase ... More
Insect lipoprotein follows a transferrin-like recycling pathway that is mediated by the insect LDL receptor homologue.
AuthorsVan Hoof D, Rodenburg KW, Van der Horst DJ
JournalJ Cell Sci
PubMed ID12356906
'The lipoprotein of insects, high-density lipophorin (HDLp), is homologous to that of mammalian low-density lipoprotein (LDL) with respect to its apolipoprotein structure. Moreover, an endocytic receptor for HDLp has been identified (insect lipophorin receptor, iLR) that is homologus to the LDL receptor. We transfected LDL-receptor-expressing CHO cells with iLR cDNA ... More
Simultaneous visualization of G- and F-actin in endothelial cells.
AuthorsHaugland RP, You W, Paragas VB, Wells KS, DuBose DA
JournalJ Histochem Cytochem
PubMed ID8308251
'We developed site-specific fluorescent probes that permit simultaneous microscopic observation of G- and F-actin in bovine endothelial cells. G-actin distribution was visualized with fluorescein-deoxyribonuclease I (DNAse I). F-actin was labeled with phalloidin conjugated to the new long-wavelength fluorophore BODIPY 581/591 (581-nm excitation, 591-nm emission), which is spectrally similar to Texas ... More
Adenovirus-mediated overexpression of catalase in the cytosolic or mitochondrial compartment protects against cytochrome P450 2E1-dependent toxicity in HepG2 cells.
AuthorsBai J, Cederbaum AI
JournalJ Biol Chem
PubMed ID11071897
'Cytochrome P450 2E1 (CYP2E1) is an effective producer of reactive oxygen species such as superoxide radical and hydrogen peroxide, which may contribute to the development of alcohol liver disease or cytotoxicity. To investigate the protective role of catalase against CYP2E1-dependent cytotoxicity, E47 cells, a transfected HepG2 cell line overexpressing CYP2E1, ... More
Immunocytochemical localization of the NPY/PYY Y1 receptor in enteric neurons, endothelial cells, and endocrine-like cells of the rat intestinal tract.
AuthorsJackerott M, Larsson LI
JournalJ Histochem Cytochem
PubMed ID9389767
'Neuropeptide Y (NPY) and peptide YY (PYY) are structurally related peptides that are considered to mediate inhibitory actions on gastrointestinal motility, secretion, and blood flow. Several receptor subtypes for these peptides have been identified and the Y1, Y2, Y4/PP1, Y5, and Y5/PP2/Y2b receptors have been cloned. In this article we ... More
Reduction of connexin43 expression and dye-coupling during neuronal differentiation of human NTera2/clone D1 cells.
AuthorsBani-Yaghoub M, Bechberger JF, Naus CC
JournalJ Neurosci Res
PubMed ID9211986
'Gap junctions are plasma membrane specializations that allow direct communication among adjoining cells. We used a human pluripotential teratocarcinoma cell line, NTera-2/clone D1 (NT2/D1), as a model to study gap junctions in CNS neurons and their neuronal precursors. These cells were differentiated following retinoic acid (RA) treatment for 4 weeks ... More
Expression and partial characterization of kinesin-related proteins in differentiating and adult skeletal muscle.
AuthorsGinkel LM, Wordeman L
JournalMol Biol Cell
PubMed ID11102514
'Using pan-kinesin antibodies to screen a differentiating C2C12 cell library, we identified the kinesin proteins KIF3A, KIF3B, and conventional kinesin heavy chain to be present in differentiating skeletal muscle. We compared the expression and subcellular localization characteristics of these kinesins in myogenic cells to others previously identified in muscle, neuronal, ... More
Nuclear import of insulin-like growth factor-binding protein-3 and -5 is mediated by the importin beta subunit.
AuthorsSchedlich LJ, Le Page SL, Firth SM, Briggs LJ, Jans DA, Baxter RC
JournalJ Biol Chem
PubMed ID10811646
'Although insulin-like growth factor-binding protein (IGFBP)-3 and IGFBP-5 are known to modulate cell growth by reversibly sequestering extracellular insulin-like growth factors, several reports have suggested that IGFBP-3, and possibly also IGFBP-5, have important insulin-like growth factor-independent effects on cell growth. These effects may be related to the putative nuclear actions ... More
[Tetrazolium methods for the histochemical investigation of hydrolases (author's transl)]
AuthorsGossrau R
JournalHistochemistry
PubMed ID103869
'Using freeze-dried or sections from fresh-frozen or aldehyde-fixed material nitro BT (NBT), tetranito BT (TNBT), distyryl nitro BT (DS-NBT), thiocarbamyl nitro BT (TC-NBT) or benzothiazolylstyrylphthalhydrazidyl tetrazolium chloride (BSPT) were tested as auxiliary reagents for the localization of glycosidases, phosphatases and non-specific esterases with indoxyl substrates in rat tissues. By means ... More
Use of methacrylate De-embedding protocols for in situ hybridization on semithin plastic sections with multiple detection strategies.
AuthorsWarren KC, Coyne KJ, Waite JH, Cary SC
JournalJ Histochem Cytochem
PubMed ID9446821
'New plastic resins are gradually replacing traditional paraffin-embedding in situ hybridization (ISH) strategies. One unique resin that has not been fully investigated or exploited with respect to light microscopic ISH is a methacrylate mixture. The advantage of this resin is its ability to be removed from tissues postsectioning, dramatically increasing ... More
Apoptogenic ganglioside GD3 directly induces the mitochondrial permeability transition.
AuthorsKristal BS, Brown AM
JournalJ Biol Chem
PubMed ID10438487
'Early events in apoptotic cascades initiated by ceramides or by activation of the surface receptor CD95 (Fas/APO-1) include the formation of ganglioside GD3. GD3 appears to be both necessary and sufficient to propagate this lipid-mediated apoptotic pathway. Later events common to many apoptotic pathways include induction of the mitochondrial permeability ... More
Identification of deoxyribonuclease II as an endonuclease involved in apoptosis.
AuthorsBarry MA, Eastman A
JournalArch Biochem Biophys
PubMed ID8424678
'Cell death occurs by apoptosis during programmed deletion of cells and following exposure to cytotoxic agents. Central to the mechanism of apoptosis is internucleosomal DNA digestion by an endogenous endonuclease which is thought to mediate cell death. An axiom of apoptosis is that the endonuclease involved is a Ca2+/Mg(2+)-dependent endonuclease. ... More
Close proximity of a cytoplasmic loop of subunit a with c subunits of the ATP synthase from Escherichia coli.
AuthorsZhang D, Vik SB
JournalJ Biol Chem
PubMed ID12525480
'Interactions between subunit a and the c subunits of the Escherichia coli ATP synthase are thought to control proton translocation through the F(o) sector. In this study cysteine substitution mutagenesis was used to define the cytoplasmic ends of the first three transmembrane spans of subunit a, as judged by accessibility ... More
Assessment of transcriptional gene activity in situ by application of HOPE-fixed, paraffin-embedded tissues.
'We report the use of HOPE-fixation (HOPE = Hepes-Glutamic acid buffer mediated Organic solvent Protection Effect) for specimens utilized for in situ hybridization targeting mRNA. For this purpose, an optimized protocol was developed and repeatedly tested on HOPE-fixed lung specimens. We observed that neither pretreatment, permeabilizing the cells, nor prehybridization ... More
Cytosolic calmodulin is increased in SK-N-SH human neuroblastoma cells due to release of calcium from intracellular stores.
AuthorsMcGinnis KM, Shariat-Madar Z, Gnegy ME
JournalJ Neurochem
PubMed ID9422356
'Muscarinic receptor stimulation elicits a redistribution of calmodulin (CaM) from the membrane fraction to cytosol in the human neuroblastoma cell line SK-N-SH. Increasing the intracellular Ca2+ concentration with ionomycin also elevates cytosolic CaM. The aim of this study was to investigate the roles of extracellular and intracellular Ca2+ pools in ... More
An alpha-actinin binding site of zyxin is essential for subcellular zyxin localization and alpha-actinin recruitment.
AuthorsReinhard M, Zumbrunn J, Jaquemar D, Kuhn M, Walter U, Trueb B
JournalJ Biol Chem
PubMed ID10224105
'The LIM domain protein zyxin is a component of adherens type junctions, stress fibers, and highly dynamic membrane areas and appears to be involved in microfilament organization. Chicken zyxin and its human counterpart display less than 60% sequence identity, raising concern about their functional identity. Here, we demonstrate that human ... More
Phosphoinositide 3-kinase is involved in the tumor-specific activation of human breast cancer cell Na(+)/H(+) exchange, motility, and invasion induced by serum deprivation.
AuthorsReshkin SJ, Bellizzi A, Albarani V, Guerra L, Tommasino M, Paradiso A, Casavola V
JournalJ Biol Chem
PubMed ID10681510
'Whereas the tumor acidic extracellular pH plays a crucial role in the invasive process, the mechanism(s) behind this acidification, especially in low nutrient conditions, are unclear. The regulation of the Na(+)/H(+) exchanger (NHE) and invasion by serum deprivation were studied in a series of breast epithelial cell lines representing progression ... More
Analysis of CMF1 reveals a bone morphogenetic protein-independent component of the cardiomyogenic pathway.
AuthorsPabón-Peña LM, Goodwin RL, Cise LJ, Bader D
JournalJ Biol Chem
PubMed ID10747923
'Disruption of the CMF1 function in anterior mesoderm inhibits cardiac myogenesis in avian embryos. In the present study, we show that CMF1 is a member of an emerging family of proteins that includes centromeric protein-F, mitosin, and LEK1. These proteins are characterized by their large size (350 kDa), dynamic subcellular ... More
Immunocytochemistry of endothelial nitric oxide synthase in the rat brain: a light and electron microscopical study using the tyramide signal amplification technique.
AuthorsStanarius A, Töpel I, Schulz S, Noack H, Wolf G
JournalActa Histochem
PubMed ID9429601
'There are many inconsistencies in the literature about the cellular and subcellular distribution of the endothelial isoform of nitric oxide synthase (eNOS) in the brain. We have re-investigated its localization by light and electron microscopical (LM, EM) immunocytochemistry and the NADPH-diaphorase reaction. Using bovine aortic tissue as a positive control ... More
Cocaine- and amphetamine-regulated transcript in the rat vagus nerve: A putative mediator of cholecystokinin-induced satiety.
AuthorsBroberger C, Holmberg K, Kuhar MJ, Hökfelt T
JournalProc Natl Acad Sci U S A
PubMed ID10557351
'Cocaine- and amphetamine-regulated transcript (CART) is widely expressed in the central nervous system. Recent studies have pointed to a role for CART-derived peptides in inhibiting feeding behavior. Although these actions have generally been attributed to hypothalamic CART, it remains to be determined whether additional CART pathways exist that link signals ... More
The synthesis, kinetic characterization and application of a novel biotinylated affinity label for cathepsin B.
AuthorsWalker B, Cullen BM, Kay G, Halliday IM, McGinty A, Nelson J
JournalBiochem J
PubMed ID1575690
'In this study we report on the synthesis, kinetic characterization and application of a novel biotinylated and active-site-directed inactivator of cathepsin B. Thus the peptidyldiazomethane biotinyl-Phe-Ala-diazomethane has been synthesized by a combination of solid-phase and solution methodologies and has been shown to be a very efficient inactivator of bovine and ... More
Subtypes Y1 and Y2 of the neuropeptide Y receptor are respectively expressed in pro-opiomelanocortin- and neuropeptide-Y-containing neurons of the rat hypothalamic arcuate nucleus.
AuthorsBroberger C, Landry M, Wong H, Walsh JN, Hökfelt T
JournalNeuroendocrinology
PubMed ID9430445
'The arcuate nucleus of the hypothalamus houses a number of neurochemically different cell populations. Among these, a dense cluster of small neuropeptide-Y (NPY)-expressing neurons is located in its ventromedial subdivision and a pro-opiomelanocortin (POMC)-expressing neuron population in its ventrolateral part. Furthermore, both neuropeptide Y Y1 and Y2 receptors (Y1-Rs and ... More
Incorporation of monoclonal antibodies into cells by osmotic permeabilization. Effect on cellular metabolism.
AuthorsChakrabarti R, Pfeiffer NE, Wylie DE, Schuster SM
JournalJ Biol Chem
PubMed ID2566602
'Incorporation of asparagine synthetase-specific monoclonal antibodies into L5178Y D10/R (L-asparaginase-resistant) murine lymphoma cells by osmotic lysis of pinocytic vesicles was used to evaluate the potential of the technique for macromolecular incorporation for metabolic studies. Nonspecific effects of the incorporation procedure included temporary inhibition of protein and DNA synthesis by 80-85% ... More
Apoptosis of T cells mediated by galectin-1.
AuthorsPerillo NL, Pace KE, Seilhamer JJ, Baum LG
JournalNature
PubMed ID7501023
'Galectin-1, a member of the family of beta-galactoside binding proteins, has growth regulatory and immunomodulatory activities. We report here that galectin-1, expressed by stromal cells in human thymus and lymph nodes, is present at sites of cell death by apoptosis during normal T-cell development and maturation. Galectin-1 induced apoptosis of ... More
Herbimycin A inhibits both dephosphorylation and translocation of cofilin induced by opsonized zymosan in macrophagelike U937 cells.
'We previously reported that a 21-kDa phosphoprotein may play an important role in superoxide production through dephosphorylation by neutrophillike differentiated HL-60 cells (Suzuki et al., 1995, Biochim Biophys Acta 1266: 261-267). The phosphoprotein was identified as cofilin, an actin-binding protein, and the activation-induced changes in its intracellular distribution have been ... More
Distinct roles for the yeast phosphatidylinositol 4-kinases, Stt4p and Pik1p, in secretion, cell growth, and organelle membrane dynamics.
AuthorsAudhya A, Foti M, Emr SD
JournalMol Biol Cell
PubMed ID10930462
'The yeast Saccharomyces cerevisiae possesses two genes that encode phosphatidylinositol (PtdIns) 4-kinases, STT4 and PIK1. Both gene products phosphorylate PtdIns at the D-4 position of the inositol ring to generate PtdIns(4)P, which plays an essential role in yeast viability because deletion of either STT4 or PIK1 is lethal. Furthermore, although ... More
The action of chromium(III) in fixation of animal tissues.
AuthorsKiernan JA
JournalHistochem J
PubMed ID2416714
'Chromic salts have been studied as fixatives of mammalian tissues for light microscopy, and the binding of the metal has been examined histochemically. Tissues bind chromium(III) from aqueous solutions less acid than pH 2.5; the metal attaches mainly to collagen and basement membranes. Solutions containing chromium(III) as the only active ... More
Fluorogenic and chromogenic substrates used in bacterial diagnostics.
AuthorsManafi M, Kneifel W, Bascomb S
JournalMicrobiol Rev
PubMed ID1943991
'Methods based on the application of chromogenic and fluorogenic substrates enable specific and rapid detection of a variety of bacterial enzymatic activities. By using these techniques, enzymatic reactions can be examined simultaneously or individually, either directly on the isolation plate or in cell suspensions. For this purpose, various testing principles ... More
The use of alkaline phosphatase-conjugated anti-immunoglobulin with immunoblots for determining the specificity of monoclonal antibodies to protein mixtures.
AuthorsEy PL, Ashman LK
JournalMethods Enzymol
PubMed ID3523128
Non-radioactive labelling and immunoprecipitation analysis of leukocyte surface proteins using different methods of protein biotinylation.
AuthorsKähne T, Ansorge S
JournalJ Immunol Methods
PubMed ID8308295
'The biotinylation of surface proteins and the detection of immunoprecipitated protein(s) after transfer to nitrocellulose using chemiluminescence methods is a highly sensitive alternative to hazardous radioactive labelling procedures. Ligation of proteins using the common biotin-NHS-ester (N-hydroxysuccinimido-biotin) is often associated with a decrease in immunoreactivity. Here a new non-radioactive method of ... More
Visualization of antigenic proteins on Western blots.
AuthorsKnecht DA, Dimond RL
JournalAnal Biochem
PubMed ID6201085
'A new technique for the detection of antibodies bound to proteins blotted onto nitrocellulose paper was developed. The method is rapid, sensitive, and does not require radioactive probes. Proteins transferred to nitrocellulose paper are first reacted with primary antibody followed by reaction with an alkaline phosphatase conjugated second antibody. The ... More
Identification of protein kinase C isozymes responsible for the phosphorylation of photoreceptor-specific RGS9-1 at Ser475.
AuthorsSokal I, Hu G, Liang Y, Mao M, Wensel TG, Palczewski K,
JournalJ Biol Chem
PubMed ID12499365
'Inactivation of the visual G-protein transducin by GTP hydrolysis is regulated by the GTPase-accelerating protein (GAP) RGS9-1. Regulation of RGS9-1 itself is poorly understood, but we found previously that it is subject to a light- and Ca(2+)-sensitive phosphorylation on Ser(475). Because there are much higher RGS9-1 levels in cones than ... More
High-resolution analyses of two different classes of tumor cells in situ tagged with alternative histochemical marker genes.
AuthorsLin WC, Pretlow TP, Pretlow TG, Culp LA
JournalAm J Pathol
PubMed ID1466397
'To evaluate interactions of two different tumor cell classes during the establishment of micrometastases at the single-cell level, two different BALB/c 3T3 tumor cell derivatives were established that harbor different histochemical marker genes: bacterial lacZ in a EJ-Harvey ras transformant (abbreviated LZEJ cells) and human placental alkaline phosphatase (ALP) gene ... More
Three subunit a isoforms of mouse vacuolar H(+)-ATPase. Preferential expression of the a3 isoform during osteoclast differentiation.
AuthorsToyomura T, Oka T, Yamaguchi C, Wada Y, Futai M
JournalJ Biol Chem
PubMed ID10722719
'Vacuolar H(+)-ATPase (V-ATPase) is a multi-subunit enzyme with a membrane peripheral catalytic (V(1)) and an intrinsic (V(o)) sector. We have identified three cDNA clones coding for isoforms of mouse V(o) subunit a (a1, a2, and a3). They exhibit 48-52% identity with each other and high similarity to subunit a of ... More
Monitoring CD8 T cell responses to NY-ESO-1: correlation of humoral and cellular immune responses.
AuthorsJäger E, Nagata Y, Gnjatic S, Wada H, Stockert E, Karbach J, Dunbar PR, Lee SY, Jungbluth A, Jäger D, Arand M, Ritter G, Cerundolo V, Dupont B, Chen YT, Old LJ, Knuth A
JournalProc Natl Acad Sci U S A
PubMed ID10781081
'NY-ESO-1, a member of the cancer-testis family of antigens, is expressed in a subset of a broad range of different human tumor types. Patients with advanced NY-ESO-1-expressing tumors frequently develop humoral immunity to NY-ESO-1, and three HLA A2-restricted peptides were defined previously as targets for cytotoxic CD8(+) T cells in ... More
Plasticity of retrovirus-labelled myotubes in the newt limb regeneration blastema.
AuthorsKumar A, Velloso CP, Imokawa Y, Brockes JP
JournalDev Biol
PubMed ID10656757
'Two important indices of myogenic differentiation are the formation of syncytial myotubes and the postmitotic arrest from the cell cycle, both of which occur after fusion of mononucleate cells. We show here that these indices are reversed in the environment of the urodele limb regeneration blastema. In order to introduce ... More
The thrombin receptor is present in myoblasts and its expression is repressed upon fusion.
AuthorsSuidan HS, Niclou SP, Dreessen J, Beltraminelli N, Monard D
JournalJ Biol Chem
PubMed ID8910573
'Cultured myoblasts derived from limb muscle of newborn rats express thrombin receptor immunoreactivity on their surface. Receptor expression is repressed upon myoblast fusion. This is due at least in part to a decrease in the amount of the thrombin receptor mRNA. Addition of thrombin triggers calcium transients only in mono- ... More
The biotin-(strept)avidin system: principles and applications in biotechnology.
AuthorsDiamandis EP, Christopoulos TK
JournalClin Chem
PubMed ID2032315
'The biotin-(strept)avidin system has been used for many years in a variety of different applications. Here we present a general overview of the system, describe its components and advantages, and show how the system is used in various applications, with emphasis on immunological and nucleic acid hybridization assays. This system ... More
Embryonic gene expression resolved at the cellular level by fluorescence in situ hybridization.
AuthorsParatore C, Suter U, Sommer L
JournalHistochem Cell Biol
PubMed ID10429965
'Tyramide signal amplification has successfully been applied to enhance detection limits of both immunological reactions and in situ hybridization methods. The technique uses short-range deposition of activated tyramide mediated by horseradish peroxidase. We have adapted this method to fluorescence in situ hybridization on embryonic tissue sections using fluorophore-labeled tyramide. The ... More
Chicken oviductal ecto-ATP-diphosphohydrolase. Purification and characterization.
AuthorsStrobel RS, Nagy AK, Knowles AF, Buegel J, Rosenberg MD
JournalJ Biol Chem
PubMed ID8663133
'An ecto-ATP diphosphohydrolase (ATPDase) was purified to homogeneity from vesiculosomes shed from chicken oviduct. First, the ecto-ATPDase-enriched vesiculosomes were concentrated by filtration, differential centrifugation, and exclusion chromatography. Next, the nonionic detergent, Nonidet P-40, was used to extract the ecto-ATPDase from vesiculosomal membranes, and the solubilized enzyme was further purified by ... More
Sensitive two-color whole-mount in situ hybridizations using digoxygenin- and dinitrophenol-labeled RNA probes.
AuthorsLong S, Rebagliati M,
JournalBiotechniques
PubMed ID11911652
Biotinylated DNA probes: sensitivity and applications.
AuthorsNeumann R, Rudloff P, Eggers HJ
JournalNaturwissenschaften
PubMed ID3762729
Image acquisition in 2-D electrophoresis.
AuthorsPatton WF, Lim MJ, Shepro D
JournalMethods Mol Biol
PubMed ID10027261
A tetrazolium method for non-specific alkaline phosphatase.
AuthorsMcGadey J
JournalHistochemie
PubMed ID5507043
Double in situ hybridization on mouse embryos for detection of overlapping regions of gene expression.
AuthorsBueno D, Skinner J, Abud H, Heath JK
JournalTrends Genet
PubMed ID8909126
Staining for enzymatic activity after gel electrophoresis, I.
AuthorsGabriel O, Gersten DM
JournalAnal Biochem
PubMed ID1381872
Stability of nitroblue tetrazolium-based alkaline phosphatase substrates.
AuthorsSmejkal GB, Kaul CA
JournalJ Histochem Cytochem
PubMed ID11511689
This report demonstrates the stability of NBT substrate after multiple exposures to alkaline phosphatase. Perhaps more important than the ability to reuse substrates, the report provides some insight into the mechanisms by which tetrazoliums are reduced and evidence for the formation of an intermediary product, i.e., a half-formazan that is ... More
Western blots using stained protein gels.
AuthorsThompson D, Larson G
JournalBiotechniques
PubMed ID1381193
A general method is described for the electrophoretic transfer of proteins from stained gels to membranes and subsequent Western detection of specific proteins on the stained membranes. Proteins are separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and the gels are stained using either of two different methods followed by ... More
A synaptic localization domain in the synaptic cleft protein laminin beta 2 (s-laminin)
AuthorsMartin PT, Ettinger AJ, Sanes JR
JournalScience
PubMed ID7618109
The basal lamina that ensheaths skeletal muscle fibers traverses the synaptic cleft at the neuromuscular junction. Synaptic and extrasynaptic portions of the basal lamina contain different laminin beta chains: beta 2 (or s) at synapses and beta 1 (or B1) extrasynaptically. Laminin beta 2 is also confined to synapselike patches ... More
A monoclonal antibody specific to a song system nuclear antigen in estrildine finches.
AuthorsAkutagawa E, Konishi M
JournalNeuron
PubMed ID11545714
This paper describes a monoclonal antibody that recognizes a molecule whose expression is mostly restricted to some of the forebrain areas that control singing behavior in adult estrildine species studied, including the zebra, Bengalese, and spice finches. When the song system displays extreme sexual dimorphism, as in these species, antibody ... More
Interaction of tau with the neural plasma membrane mediated by tau's amino-terminal projection domain.
AuthorsBrandt R, Léger J, Lee G
JournalJ Cell Biol
PubMed ID8522593
The neuronal microtubule-associated protein tau is required for the development of cell polarity in cultured neurons. Using PC12 cells that stably express tau and tau amino-terminal fragments, we report that tau interacts with the neural plasma membrane through its amino-terminal projection domain. In differentiated PC12 transfectants, tau is found in ... More
Specific in situ hybridization of Haemobartonella felis with a DNA probe and tyramide signal amplification.
AuthorsBerent LM, Messick JB, Cooper SK, Cusick PK
JournalVet Pathol
PubMed ID10643980
Haemobartonella felis is an epierythrocytic bacterium suspected to be the causative agent of feline infectious anemia. Previous studies with a polymerase chain reaction assay have identified a mycoplasmal 16S rRNA gene sequence that coincides with clinical disease and the presence of organisms in the blood. Tissues from a cat experimentally ... More
Two histochemical marker genes, Drosophila alcohol dehydrogenase (ADH) and human placental alkaline phosphatase (ALP), were cloned into the recombinant retroviral vectors pLJ and pgag beta-actin. The resulting vectors were transfected into retroviral producer cell lines, psi CRE and psi CRIP, and stable recombinant retrovirus producers were isolated. Recombinant virus was ... More
Monitoring gene expression in Chinese hamster ovary cells using secreted apoaequorin.
AuthorsInouye S, Tsuji FI
JournalAnal Biochem
PubMed ID1621948
A luminescence method for monitoring gene expression in Chinese hamster ovary cells using apoaequorin as a secreted reporter enzyme is described. In this method, the cell is not disrupted prior to assay as in the earlier aequorin procedure and in the firefly method. The apoaequorin secretion vector is constructed by ... More
Nerve growth factor protects oligodendrocytes from tumor necrosis factor-alpha-induced injury through Akt-mediated signaling mechanisms.
Tumor necrosis factor-alpha is thought to be one of the most important inflammatory cytokines associated with the demyelinating disease multiple sclerosis. We determined whether neurotrophins could protect oligodendrocytes from tumor necrosis factor-alpha-mediated cytotoxicity. Among the neurotrophins tested, nerve growth factor was most effective at preventing cell death. Nerve growth factor ... More
Accurate detection of both glycoproteins and total proteins on blots: control of side reactions occurring after periodate oxidation of proteins.
AuthorsBouchez-Mahiout I, Doyen C, Laurière M
JournalElectrophoresis
PubMed ID10424463
The detection of glycoproteins based on the periodate oxidation of their carbohydrate moiety, and their conjugation to digoxigenin hydrazide directly on blots, leads to high background staining, especially with polyvinylidene difluoride (PVDF) membranes. The addition of Tween-20 to all incubation solutions, except at the oxidation step, strongly reduced the background ... More
The bHLH transcription factor Olig2 promotes oligodendrocyte differentiation in collaboration with Nkx2.2.
AuthorsZhou Q, Choi G, Anderson DJ
JournalNeuron
PubMed ID11567617
Olig2, a basic helix-loop-helix (bHLH) transcription factor, is expressed in a restricted domain of the spinal cord ventricular zone that sequentially generates motoneurons and oligodendrocytes. Just prior to oligo-dendrocyte precursor formation, the domains of Olig2 and Nkx2.2 expression switch from being mutually exclusive to overlapping, and Neurogenins1 and 2 are ... More
Direct extracellular contact between integrin alpha(3)beta(1) and TM4SF protein CD151.
AuthorsYauch RL, Kazarov AR, Desai B, Lee RT, Hemler ME
JournalJ Biol Chem
PubMed ID10734060
Previously we established that the alpha(3)beta(1) integrin shows stable, specific, and stoichiometric association with the TM4SF (tetraspannin) protein CD151. Here we used a membrane impermeable cross-linking agent to show a direct association between extracellular domains of alpha(3)beta(1) and CD151. The alpha(3)beta(1)-CD151 association site was then mapped using chimeric alpha(6)/alpha(3) integrins ... More
Novel chemical method for the preparation of nucleic acids for nonisotopic hybridization.
AuthorsViscidi RP, Connelly CJ, Yolken RH
JournalJ Clin Microbiol
PubMed ID2939103
A novel chemical method was used to prepare biotin-labeled nucleic acids for nonisotopic hybridization. The method involves the transamination of unpaired cytosine residues in polynucleotides with sodium bisulfite and ethylenediamine. Primary amino groups on the cytosine derivatives are then reacted with biotinyl-e-aminocaproic acid N-hydroxysuccinimide ester. Biotinylated probes hybridized with 1 ... More
Complex contribution of the 3'-untranslated region to the expressional regulation of the human inducible nitric-oxide synthase gene. Involvement of the RNA-binding protein HuR.
AuthorsRodriguez-Pascual F, Hausding M, Ihrig-Biedert I, Furneaux H, Levy AP, Förstermann U, Kleinert H
JournalJ Biol Chem
PubMed ID10859327
Cytokine stimulation of human DLD-1 cells resulted in a marked expression of nitric-oxide synthase (NOS) II mRNA and protein accompanied by only a moderate increase in transcriptional activity. Also, there was a basal transcription of the NOS II gene, which did not result in measurable NOS II expression. The 3'-untranslated ... More
Identification of a 40-kDa cell surface sialoglycoprotein with the characteristics of a major influenza C virus receptor in a Madin-Darby canine kidney cell line.
AuthorsZimmer G, Klenk HD, Herrler G
JournalJ Biol Chem
PubMed ID7629082
Infection of cells by influenza C virus is known to be initiated by virus attachment to cell surface glycoconjugates containing N-acetyl-9-O-acetylneuraminic acid. Using an in vitro virus binding assay, we have detected this carbohydrate on several glycoproteins of Madin-Darby canine kidney cells (type I), a polarized epithelial cell line permissive ... More
Rapid and sensitive colorimetric method for visualizing biotin-labeled DNA probes hybridized to DNA or RNA immobilized on nitrocellulose: Bio-blots.
AuthorsLeary JJ, Brigati DJ, Ward DC
JournalProc Natl Acad Sci U S A
PubMed ID6191328
Biotin-labelled DNA probes, prepared by nick-translation in the presence of biotinylated analogs of TTP, are hybridized to DNA or RNA immobilized on nitrocellulose filters. After removal of residual probe, the filters are incubated for 2--5 min with a preformed complex made with avidin-DH (or streptavidin) and biotinylated polymers of intestinal ... More
Brefeldin A-dependent membrane tubule formation reconstituted in vitro is driven by a cell cycle-regulated microtubule motor.
AuthorsRobertson AM, Allan VJ
JournalMol Biol Cell
PubMed ID10712511
Treatment of cultured cells with brefeldin A (BFA) induces the formation of extensive membrane tubules from the Golgi apparatus, trans-Golgi network, and early endosomes in a microtubule-dependent manner. We have reconstituted this transport process in vitro using Xenopus egg cytosol and a rat liver Golgi-enriched membrane fraction. The presence of ... More
Characterization of Yeast Yea4p, a uridine diphosphate-N-acetylglucosamine transporter localized in the endoplasmic reticulum and required for chitin synthesis.
AuthorsRoy SK, Chiba Y, Takeuchi M, Jigami Y
JournalJ Biol Chem
PubMed ID10788474
Chitin is an essential cell wall component, synthesis of which is regulated throughout the cell cycle in the yeast Saccharomyces cerevisiae. We cloned an S. cerevisiae gene, YEA4, whose product is homologous to the Kluyveromyces lactis uridine diphosphate-N-acetylglucosamine (UDP-GlcNAc) transporter. An epitope-tagged Yea4p localized mainly in the 10,000 x g ... More
Comparison of in-situ hybridization, direct and indirect in-situ PCR as well as tyramide signal amplification for the detection of HPV.
AuthorsWiedorn KH, Kühl H, Galle J, Caselitz J, Vollmer E
JournalHistochem Cell Biol
PubMed ID10090569
One hundred paraffin-embedded cervical biopsy specimens were tested for the presence of human papilloma virus (HPV) by in situ hybridization (ISH), and by direct and indirect in situ PCR (IS-PCR) in order to evaluate the efficiency of the different in situ methods in detecting HPV infection. ISH was performed using ... More
Identification of basic residues involved in drug export function of human multidrug resistance-associated protein 2.
AuthorsRyu S, Kawabe T, Nada S, Yamaguchi A
JournalJ Biol Chem
PubMed ID10978330
Multidrurg resistance-associated protein 2 (MRP2)/canalicular multispecific organic anion transporter (cMOAT) is involved in the ATP-dependent export of organic anions across the bile canalicular membrane. To identify functional amino acid residues that play essential roles in the substrate transport, each of 13 basic residues around transmembrane regions (TMs) 6-17 were replaced ... More
Channel-forming activity of immunoaffinity-purified connexin32 in single phospholipid membranes.
AuthorsRhee SK, Bevans CG, Harris AL
JournalBiochemistry
PubMed ID8703927
Connexin32, a member of the family of proteins that forms gap junction channels between cells, was immunoaffinity-purified from rat liver using a monoclonal antibody, under nondenaturing conditions and reconstituted into unilamellar phospholipid liposomes and bilayers. Gel-filtration studies indicate that the connexin32 is purified predominantly in structures of a size consistent ... More
A luminescent ruthenium complex for ultrasensitive detection of proteins immobilized on membrane supports.
SYPRO Ruby protein blot stain provides a sensitive, gentle, fluorescence-based method for detecting proteins on nitrocellulose or polyvinylidene difluoride (PVDF) membranes. SYPRO Ruby dye is a permanent stain composed of ruthenium as part of an organic complex that interacts noncovalently with proteins. Stained proteins can be excited by ultraviolet light ... More
Cytosolic targeting domains of gamma and delta calmodulin-dependent protein kinase II.
AuthorsCaran N, Johnson LD, Jenkins KJ, Tombes RM
JournalJ Biol Chem
PubMed ID11535587
Ca(2+)/calmodulin-dependent protein kinase II (CaMK-II) isozyme variability is the result of alternative usage of variable domain sequences. Isozyme expression is cell type-specific to transduce the appropriate Ca(2+) signals. We have determined the subcellular targeting domain of delta(E) CaMK-II, an isozyme that induces neurite outgrowth, and of a structurally similar isozyme, ... More
Heat shock and oxidative stress-induced exposure of hydrophobic protein domains as common signal in the induction of hsp68.
AuthorsGosslau A, Ruoff P, Mohsenzadeh S, Hobohm U, Rensing L
JournalJ Biol Chem
PubMed ID11042186
The hypothesis of a common signal for heat shock (HS) and oxidative stress (OS) was analyzed in C6 cells with regard to the induction of heat shock proteins (Hsps). The synthesis rate and level of the strictly inducible Hsp68 was significantly higher after HS (44 degrees C) compared with OS ... More
ETL, a novel seven-transmembrane receptor that is developmentally regulated in the heart. ETL is a member of the secretin family and belongs to the epidermal growth factor-seven-transmembrane subfamily.
AuthorsNechiporuk T, Urness LD, Keating MT
JournalJ Biol Chem
PubMed ID11050079
Using differential display of rat fetal and postnatal cardiomyocytes, we have identified a novel seven-transmembrane receptor, ETL. The cDNA-predicted amino acid sequence of ETL indicated that it encodes a 738-aa protein composed of a large extracellular domain with epidermal growth factor (EGF)-like repeats, a seven-transmembrane domain, and a short cytoplasmic ... More
5-Lipoxygenase interacts with coactosin-like protein.
We have recently identified coactosin-like protein (CLP) in a yeast two-hybrid screen using 5-lipoxygenase (5LO) as a bait. In this report, we demonstrate a direct interaction between 5LO and CLP. 5LO associated with CLP, which was expressed as a glutathione S-transferase fusion protein, in a dose-dependent manner. Coimmunoprecipitation experiments using ... More
Histamine treatment induces rearrangements of orthogonal arrays of particles (OAPs) in human AQP4-expressing gastric cells.
AuthorsCarmosino M, Procino G, Nicchia GP, Mannucci R, Verbavatz JM, Gobin R, Svelto M, Valenti G
JournalJ Cell Biol
PubMed ID11564760
To test the involvement of the water channel aquaporin (AQP)-4 in gastric acid physiology, the human gastric cell line (HGT)-1 was stably transfected with rat AQP4. AQP4 was immunolocalized to the basolateral membrane of transfected HGT-1 cells, like in native parietal cells. Expression of AQP4 in transfected cells increased the ... More
Characterization of palladin, a novel protein localized to stress fibers and cell adhesions.
AuthorsParast MM, Otey CA
JournalJ Cell Biol
PubMed ID10931874
Here, we describe the identification of a novel phosphoprotein named palladin, which colocalizes with alpha-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 alpha-actinin in fibroblast lysates. A cDNA encoding palladin was ... More
Identification of an active site arginine in rat choline acetyltransferase by alanine scanning mutagenesis.
AuthorsWu D, Hersh LB
JournalJ Biol Chem
PubMed ID7493935
Kinetic as well as chemical modification studies have implicated the presence of an active site arginine in choline acetyltransferase, whose function is to stabilize coenzyme binding by interacting with the 3'-phosphate of the coenzyme A substrate. In order to identify this residue seven conserved arginines in rat choline acetyltransferase were ... More
Arabidopsis eIF3e (INT-6) associates with both eIF3c and the COP9 signalosome subunit CSN7.
AuthorsYahalom A, Kim TH, Winter E, Karniol B, von Arnim AG, Chamovitz DA
JournalJ Biol Chem
PubMed ID11029466
The Arabidopsis COP9 signalosome is a multisubunit repressor of photomorphogenesis that is conserved among eukaryotes. This complex may have a general role in development. As a step in dissecting the biochemical mode of action of the COP9 signalosome, we determined the sequence of proteins that copurify with this complex. Here ... More
Yop1p, the yeast homolog of the polyposis locus protein 1, interacts with Yip1p and negatively regulates cell growth.
AuthorsCalero M, Whittaker GR, Collins RN
JournalJ Biol Chem
PubMed ID11278413
Rab proteins are small GTPases that are essential elements of the protein transport machinery of eukaryotic cells. Each round of membrane transport requires a cycle of Rab protein nucleotide binding and hydrolysis. We have recently characterized a protein, Yip1p, which appears to play a role in Rab-mediated membrane transport in ... More
Epithelial cell-specific laminin 5 is required for survival of early thymocytes.
AuthorsKim MG, Lee G, Lee SK, Lolkema M, Yim J, Hong SH, Schwartz RH
JournalJ Immunol
PubMed ID10861052
The gene LamC2 encoding the gamma2 chain of laminin 5, an epithelial cell-specific extracellular matrix protein, was identified in a PCR-based subtracted cDNA library from mouse thymic stromal cells. The mRNA existed in two alternative forms (5.1 and 2.4 kb). The full-length message was highly expressed in SCID thymus and ... More
Identification and isolation of a 45-kDa calcium-dependent lactoferrin receptor from rat hepatocytes.
AuthorsBennatt DJ, McAbee DD
JournalBiochemistry
PubMed ID9204883
Isolated rat hepatocytes bind, internalize, and degrade bovine lactoferrin (Lf) via high-affinity Ca2+-dependent sites [<10(6) sites/cell; McAbee et al., (1993) Biochemistry 32, 13749-13760]. In this study, we identified a 45-kDa Ca2+-dependent Lf binding protein on rat hepatocytes by three independent approaches. First, dithiobis(sulfosuccimidylproprionate) (DTSSP) cross-linked 125I-Lf to a 45-kDa adduct ... More
Enzyme-based antigen localization and quantitation in cell and tissue samples (Midwestern assay).
AuthorsRoth KA, Brenner JW, Selznick LA, Gokden M, Lorenz RG
JournalJ Histochem Cytochem
PubMed ID9389766
Quantitation of antigen concentration in cell and tissue samples typically requires antigen extraction, which precludes antigen localization in the same sample. Similarly, antigen immunolocalization in fixed cells or tissue sections provides limited information about antigen concentration. We have developed a rapid and sensitive assay for simultaneous antigen localization and quantitation ... More
Functional reconstitution and characterization of recombinant human alpha 1-glycine receptors.
AuthorsCascio M, Shenkel S, Grodzicki RL, Sigworth FJ, Fox RO
JournalJ Biol Chem
PubMed ID11145968
By utilizing a baculoviral expression system described previously (Cascio, M., Schoppa, N. E., Grodzicki, R. L., Sigworth, F. J., and Fox, R. O. (1993) J. Biol. Chem. 268, 22135-22142), functional recombinant homomeric human alpha(1)-glycine receptors (GlyR) were overexpressed in insect cell culture, solubilized, purified, and reconstituted into lipid vesicles via ... More
A recombinant retrovirus encoding alkaline phosphatase confirms clonal boundary assignment in lineage analysis of murine retina.
AuthorsFields-Berry SC, Halliday AL, Cepko CL
JournalProc Natl Acad Sci U S A
PubMed ID1731342
Recombinant retroviruses encoding the histochemically detectable enzyme beta-galactosidase have been used to investigate lineage in the vertebrate nervous system. Identification of the descendants of individual progenitors is straightforward when progeny cells are arranged in a reproducible, clustered pattern, but difficulties in interpretation arise when progeny migrate extensively and/or in an ... More
Localization of the cyclic ADP-ribose-dependent calcium signaling pathway in hepatocyte nucleus.
AuthorsKhoo KM, Han MK, Park JB, Chae SW, Kim UH, Lee HC, Bay BH, Chang CF
JournalJ Biol Chem
PubMed ID10818108
CD38 is a type II transmembrane glycoprotein found on both hematopoietic and non-hematopoietic cells. It is known for its involvement in the metabolism of cyclic ADP-ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate, two nucleotides with calcium mobilizing activity independent of inositol trisphosphate. It is generally believed that CD38 is ... More
Alternative splicing of the first F3 domain from chicken collagen XIV affects cell adhesion and heparin binding.
AuthorsImhof M, Trueb B
JournalJ Biol Chem
PubMed ID11098058
The N terminus of chicken collagen XIV is subject to alternative splicing. The longer isoform contains a fibronectin type III (F3) domain at its N terminus, whereas the shorter isoform is lacking this domain. Alternative splicing of the F3 domain is developmentally regulated. At early embryonic stages, both isoforms are ... More
Quantitative transcript imaging in normal and heat-shocked Drosophila embryos by using high-density oligonucleotide arrays.
AuthorsLeemans R, Egger B, Loop T, Kammermeier L, He H, Hartmann B, Certa U, Hirth F, Reichert H
JournalProc Natl Acad Sci U S A
PubMed ID11035778
Embryonic development in Drosophila is characterized by an early phase during which a cellular blastoderm is formed and gastrulation takes place, and by a later postgastrulation phase in which key morphogenetic processes such as segmentation and organogenesis occur. We have focused on this later phase in embryogenesis with the goal ... More
IL-4 plays a crucial role in regulating oxidative damage in the liver during schistosomiasis.
AuthorsLa Flamme AC, Patton EA, Bauman B, Pearce EJ
JournalJ Immunol
PubMed ID11160238
Liver enlargement and hepatocyte proliferation, normal responses in wild-type (WT) mice infected with the parasitic helminth Schistosoma mansoni, were found to be severely impaired in infected IL-4(-/-) mice. Compared with WT mice, increased levels of O(2)(-), NO, and the more highly reactive ONOO(-) were detected in the liver and produced ... More
Selective degradation of oxidized calmodulin by the 20 S proteasome.
AuthorsFerrington DA, Sun H, Murray KK, Costa J, Williams TD, Bigelow DJ, Squier TC
JournalJ Biol Chem
PubMed ID11010965
We have investigated the mechanisms that target oxidized calmodulin for degradation by the proteasome. After methionine oxidation within calmodulin, rates of degradation by the 20 S proteasome are substantially enhanced. Mass spectrometry was used to identify the time course of the proteolytic fragments released from the proteasome. Oxidized calmodulin is ... More
Dual targeting property of the N-terminal signal sequence of P4501A1. Targeting of heterologous proteins to endoplasmic reticulum and mitochondria.
Recent studies from our laboratory showed that the beta-naphthoflavone-inducible cytochrome P4501A1 is targeted to both the endoplasmic reticulum (ER) and mitochondria. In the present study, we have further investigated the ability of the N-terminal signal sequence (residues 1-44) of P4501A1 to target heterologous proteins, dihydrofolate reductase, and the mature portion ... More
Novel E-cadherin-mediated adhesion in peripheral nerve: Schwann cell architecture is stabilized by autotypic adherens junctions.
AuthorsFannon AM, Sherman DL, Ilyina-Gragerova G, Brophy PJ, Friedrich VL, Colman DR
JournalJ Cell Biol
PubMed ID7698985
Previous studies (Blank, W. F., M. B. Bunge, and R. P. Bunge. 1974. Brain Res. 67:503-518) showed that Schwann cell paranodal membranes were disrupted in calcium free medium suggesting that cadherin mediated mechanisms may operate to maintain the integrity of the paranodal membrane complex. Using antibodies against the fifth extracellular ... More
An optimized method for in situ hybridization with signal amplification that allows the detection of rare mRNAs.
AuthorsYang H, Wanner IB, Roper SD, Chaudhari N
JournalJ Histochem Cytochem
PubMed ID10082745
In situ hybridization (ISH) using nonradioactive probes enables mRNAs to be detected with improved cell resolution but compromised sensitivity compared to ISH with radiolabeled probes. To detect rare mRNAs, we optimized several parameters for ISH using digoxygenin (DIG)-labeled probes, and adapted tyramide signal amplification (TSA) in combination with alkaline phosphatase ... More
Mixed lineage kinase 2 interacts with clathrin and influences clathrin-coated vesicle trafficking.
AuthorsAkbarzadeh S, Ji H, Frecklington D, Marmy-Conus N, Mok YF, Bowes L, Devereux L, Linsenmeyer M, Simpson RJ, Dorow DS
JournalJ Biol Chem
PubMed ID12105200
Mixed lineage kinase 2 (MLK2) is a protein kinase that signals in the stress-activated Jun N-terminal kinase signal transduction pathway. We used immunoprecipitation and mass spectrometric analysis to identify MLK2-binding proteins in cell lines with inducible expression of green fluorescent protein-tagged MLK2. Here we report the identification of clathrin as ... More
Molecular and biochemical characterization of a novel oxysterol-binding protein (OSBP2) highly expressed in retina.
AuthorsMoreira EF, Jaworski C, Li A, Rodriguez IR
JournalJ Biol Chem
PubMed ID11278871
We are interested in understanding the possible function(s) of the oxysterol-binding proteins in mediating oxysterol cytotoxicity in the retina. In this study we describe the cloning, localization, and biological activity of a novel oxysterol-binding protein (OSBP2), and complete the molecular characterization of the previously known OSBP1. Both OSBP genes contain ... More
A high-resolution, fluorescence-based method for localization of endogenous alkaline phosphatase activity.
AuthorsCox WG, Singer VL
JournalJ Histochem Cytochem
PubMed ID10544217
We describe a high-resolution, fluorescence-based method for localizing endogenous alkaline phosphatase in tissues and cultured cells. This method utilizes ELF (Enzyme-Labeled Fluorescence)-97 phosphate, which yields an intensely fluorescent yellow-green precipitate at the site of enzymatic activity. We compared zebrafish intestine, ovary, and kidney cryosections stained for endogenous alkaline phosphatase using ... More