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  • Arabidopsis TFL2/LHP1 specifically associates with genes marked by trimethylation of histone H3 lysine 27. 17542647

    TERMINAL FLOWER 2/LIKE HETEROCHROMATIN PROTEIN 1 (TFL2/LHP1) is the only Arabidopsis protein with overall sequence similarity to the HETEROCHROMATIN PROTEIN 1 (HP1) family of metazoans and S. pombe. TFL2/LHP1 represses transcription of numerous genes, including the flowering-time genes FLOWERING LOCUS T (FT) and FLOWERING LOCUS C (FLC), as well as the floral organ identity genes AGAMOUS (AG) and APETALA 3 (AP3). These genes are also regulated by proteins of the Polycomb repressive complex 2 (PRC2), and it has been proposed that TFL2/LHP1 represents a potential stabilizing factor of PRC2 activity. Here we show by chromatin immunoprecipitation and hybridization to an Arabidopsis Chromosome 4 tiling array (ChIP-chip) that TFL2/LHP1 associates with hundreds of small domains, almost all of which correspond to genes located within euchromatin. We investigated the chromatin marks to which TFL2/LHP1 binds and show that, in vitro, TFL2/LHP1 binds to histone H3 di- or tri-methylated at lysine 9 (H3K9me2 or H3K9me3), the marks recognized by HP1, and to histone H3 trimethylated at lysine 27 (H3K27me3), the mark deposited by PRC2. However, in vivo TFL2/LHP1 association with chromatin occurs almost exclusively and co-extensively with domains marked by H3K27me3, but not H3K9me2 or -3. Moreover, the distribution of H3K27me3 is unaffected in lhp1 mutant plants, indicating that unlike PRC2 components, TFL2/LHP1 is not involved in the deposition of this mark. Rather, our data suggest that TFL2/LHP1 recognizes specifically H3K27me3 in vivo as part of a mechanism that represses the expression of many genes targeted by PRC2.
    Tipo de documento:
    Referencia
    Referencia del producto:
    Múltiplo
    Nombre del producto:
    Múltiplo

  • Integrin-driven actin polymerization consolidates long-term potentiation. 16567651

    Long-term potentiation (LTP), like memory, becomes progressively more resistant to disruption with time after its formation. Here we show that threshold conditions for inducing LTP cause a rapid, long-lasting increase in polymerized filamentous actin in dendritic spines of adult hippocampus. Two independent manipulations that reverse LTP disrupted this effect when applied shortly after induction but not 30 min later. Function-blocking antibodies to beta1 family integrins selectively eliminated both actin polymerization and stabilization of LTP. We propose that the initial stages of consolidation involve integrin-driven events common to cells engaged in activities that require rapid morphological changes.
    Tipo de documento:
    Referencia
    Referencia del producto:
    Múltiplo
    Nombre del producto:
    Múltiplo

  • Multiple amino acid changes at the first glycosylation motif in NS1 protein of West Nile virus are necessary for complete attenuation for mouse neuroinvasiveness. 21945257

    West Nile virus (WNV), like all members of the Japanese encephalitis (JE) serogroup except JE virus, contains three N-linked glycosylation (N-X-S/T) sites in the NS1 protein at asparagine residues NS1(130), NS1(175) and NS1(207). Previously we showed that the ablation of these glycosylation sites in WNV, by substitution of asparagine for alanine, attenuated mouse neuroinvasiveness; however, full attenuation was not achieved and the virus retained a neurovirulence phenotype. Sequence of viral RNA extracted from mouse brains revealed a reversion at the NS1(130) site in some mice that succumbed to the attenuated NS1(130A/175A/207A) strain. Here, we further attenuated WNV by mutating the asparagine to serine or glutamine in addition to mutating other residues in the NS1(130-132) glycosylation motif. These mutants proved to further attenuate WNV for both neuroinvasiveness and neurovirulence in mice. NS1(130-132QQA/175A/207A), the most attenuated mutant virus, showed modest changes in infectivity titers versus the parental strain, was not temperature sensitive, and did not show reversion in mice. Mutant virus was completely attenuated for neuroinvasiveness after intraperitoneal inoculation with >1,000,000 PFU, and mice were protected against lethal challenge. Overall, we showed that changing the asparagine of the NS1(130) glycosylation motif to a serine or glutamine attenuated WNV further than the asparagine to alanine substitution. Further, mutating all three of the amino acids of the NS1(130-132) glycosylation motif (NTT-QQA) along with NS1(175) and NS1(207) asparagine to alanine mutations gave the most stable and attenuated strain.
    Tipo de documento:
    Referencia
    Referencia del producto:
    MAB8152
    Nombre del producto:
    Anti-West Nile Virus/Kunjin Antibody, NS1, clone 3.1112G

  • Ethanol withdrawal influences survival and morphology of developing rat hippocampal neurons in vitro. 14745305

    BACKGROUND: Previous studies in this laboratory have shown that, like their counterparts in vivo, fetal rat hippocampal pyramidal neurons in culture develop abnormally small dendritic arbors when exposed to ethanol. This study asked whether ethanol's inhibitory effects on dendritic development differ when the duration of ethanol exposure and timing of withdrawal are varied to correspond with early versus later stages of development and whether ethanol withdrawal influences survival of these neurons. METHODS: We compared neurons exposed continuously for 6 or 14 days to ethanol (70 mM) with neurons transferred from ethanol-containing medium to control medium either 1 day after adding ethanol (before dendrites elongated) or 6 days after adding ethanol (after dendrites began elongating). We then performed morphometric and cell density analyses at 6 and 14 days using digital images of neurons immunostained with microtubule-associated protein 2 (MAP2) to visualize dendrites. RESULTS: Continuous exposure to ethanol decreased the length and number of dendrites formed but had no effect on neuron survival compared with controls without ethanol. Dendritic length was less inhibited when ethanol was withdrawn after 1 day, but the number of dendrites per cell was unchanged compared with neurons continuously exposed to ethanol. Withdrawal from ethanol at 1 day slightly enhanced the survival of neurons assessed at 14 days compared with neurons in control medium and with neurons exposed continuously to ethanol. In contrast, withdrawal from ethanol at 6 days severely decreased the number of neurons at 14 days. CONCLUSIONS: These results suggest that dendrites can achieve normal length when ethanol exposure is limited to only 1 day and withdrawal occurs before dendrites begin elongating. However, a persistent reduction in dendrite number results in smaller overall dendritic arbor size. Although continuous exposure to ethanol has little effect on neuron survival in these cultures, and exposure limited to 1 day followed by withdrawal can be neuroprotective against cell death associated with increased time in culture, longer exposure before withdrawal can trigger cell death.
    Tipo de documento:
    Referencia
    Referencia del producto:
    MAB3418
    Nombre del producto:
    Anti-MAP2 Antibody, clone AP20

  • Exposure to bioaerosols during the growth season in an organic greenhouse tomato production using Supresivit(R) (Trichoderma harzianum) and Mycostop(R) (Streptomyces gris ... 20622135

    In working environments, especially in confined spaces like greenhouses, elevated concentrations of airborne microorganisms may become a problem for workers' health. Additionally, the use of microbial pest control agents may increase exposure to microorganisms. The aim of this study was to investigate tomato growers' exposure to naturally occurring bioaerosol components (dust, bacteria, fungi, actinomycetes, (1-->3)-beta-D-glucans and endotoxin) and microbial pest control agents applied by drip irrigation. Airborne dust was collected with filter samplers and analyzed for microorganisms by plate counts and total counts in microscope. Analysis of (1-->3)-beta-D-glucan and endotoxin content were performed by kinetic, chromatic Limulus Amoebocyte Lysate tests. The fungal strain (Trichoderma harzianum) from the biocontrol product Supresivit(R) was identified by PCR analysis. Measurements were performed on the day of drip irrigation and one week, one month and three months after the irrigation. T. harzianum from Supresivit(R) could only be detected on the day of treatment. Streptomyces griseoviridis, an applied microbial pest control agent, was not detected in the air during this investigation. We found that bioaerosol exposure increases during the growth season and that exposure to fungi, bacteria, and endotoxin can reach levels during the harvest period, that may cause respiratory symptoms in growers. The collected data indicates that MPCAs applied by drip irrigation do not become airborne later in the season.
    Tipo de documento:
    Referencia
    Referencia del producto:
    2100
    Nombre del producto:
    Protein-Concentrate Kit (Micro)

  • Mesodiencephalic dopaminergic neuronal differentiation does not involve GLI2A-mediated SHH-signaling and is under the direct influence of canonical WNT signaling. 24865218

    Sonic Hedgehog (SHH) and WNT proteins are key regulators in many developmental processes, like embryonic patterning and brain development. In the brain, SHH is expressed in a gradient starting in the floor plate (FP) progressing ventrally in the midbrain, where it is thought to be involved in the development and specification of mesodiencephalic dopaminergic (mdDA) neurons. GLI2A-mediated SHH-signaling induces the expression of Gli1, which is inhibited when cells start expressing SHH themselves. To determine whether mdDA neurons receive GLI2A-mediated SHH-signaling during differentiation, we used a BAC-transgenic mouse model expressing eGFP under the control of the Gli1 promoter. This mouse-model allowed for mapping of GLI2A-mediated SHH-signaling temporal and spatial in the mouse midbrain. Since mdDA neurons are born from E10.5, peaking at E11.0-E12.0, we examined Gli1-eGFP embryos at E11.5, E12.5, and E13.5, indicating whether Gli1 was induced before or during mdDA development and differentiation. Our data indicate that GLI2A-mediated SHH-signaling is not involved in mdDA neuronal differentiation. However, it appears to be involved in the differentiation of neurons which make up a subset of the red nucleus (RN). In order to detect whether mdDA neuronal differentiation may be under the control of canonical WNT-signaling, we used a transgenic mouse-line expressing LacZ under the influence of stable β-catenin. Here, we show that TH+ neurons of the midbrain receive canonical WNT-signaling during differentiation. Therefore, we suggest that early SHH-signaling is indirectly involved in mdDA development through early patterning of the midbrain area, whereas canonical WNT-signaling is directly involved in the differentiation of the mdDA neuronal population.
    Tipo de documento:
    Referencia
    Referencia del producto:
    Múltiplo
    Nombre del producto:
    Múltiplo

  • Circulating mouse Flk1+/c-Kit+/CD45- cells function as endothelial progenitors cells (EPCs) and stimulate the growth of human tumor xenografts. 25047738

    Endothelial progenitor cells (EPCs) have been demonstrated to have stem-cell like as well as mature endothelial functions. However, controversy remains as to their origins, immunophenotypic markings, and contribution to the tumor vascular network and tumor survival.Flow cytometric analysis and sorting was used to isolate Flk-1+/c-Kit+/CD45- cells. Matrigel and methycellulose assays, flow cytometry, and gene array analyses were performed to characterize several murine EPC cell populations. Human tumor xenografts were used to evaluate the impact of EPCs on tumor growth and vascular development.Flk-1+/c-Kit+/CD45- cells were present at low levels in most murine organs with the highest levels in adipose, aorta/vena cava, and lung tissues. Flk-1+/c-Kit+/CD45- cells demonstrated stem cell qualities through colony forming assays and mature endothelial function by expression of CD31, uptake of acLDL, and vascular structure formation in matrigel. High passage EPCs grown in vitro became more differentiated and lost stem-cell markers. EPCs were found to have hemangioblastic properties as demonstrated by the ability to rescue mice given whole body radiation. Systemic injection of EPCs increased the growth of human xenograft tumors and vessel density.Flk-1+/C-Kit+/CD45- cells function as endothelial progenitor cells. EPCs are resident in most murine tissue types and localize to human tumor xenografts. Furthermore, the EPC population demonstrates stem-cell and mature endothelial functions and promoted the growth of tumors through enhanced vascular network formation. Given the involvement of EPCs in tumor development, this unique host-derived population may be an additional target to consider for anti-neoplastic therapy.
    Tipo de documento:
    Referencia
    Referencia del producto:
    AB3209
    Nombre del producto:
    Anti-Oct-4 Antibody

  • Non-integrating gamma-retroviral vectors as a versatile tool for transient zinc-finger nuclease delivery. 24722320

    Designer nucleases, like zinc-finger nucleases (ZFNs), represent valuable tools for targeted genome editing. Here, we took advantage of the gamma-retroviral life cycle and produced vectors to transfer ZFNs in the form of protein, mRNA and episomal DNA. Transfer efficacy and ZFN activity were assessed in quantitative proof-of-concept experiments in a human cell line and in mouse embryonic stem cells. We demonstrate that retrovirus-mediated protein transfer (RPT), retrovirus-mediated mRNA transfer (RMT), and retrovirus-mediated episome transfer (RET) represent powerful methodologies for transient protein delivery or protein expression. Furthermore, we describe complementary strategies to augment ZFN activity after gamma-retroviral transduction, including serial transduction, proteasome inhibition, and hypothermia. Depending on vector dose and target cell type, gene disruption frequencies of up to 15% were achieved with RPT and RMT, and greater than 50% gene knockout after RET. In summary, non-integrating gamma-retroviral vectors represent a versatile tool to transiently deliver ZFNs to human and mouse cells.
    Tipo de documento:
    Referencia
    Referencia del producto:
    MAB3580
    Nombre del producto:
    Anti-Green Fluorescent Protein Antibody

  • Ouabain induces cell proliferation through calcium-dependent phosphorylation of Akt (protein kinase B) in opossum kidney proximal tubule cells. 16807298

    Cardiotonic glycosides, like ouabain, inhibit Na(+)-K(+)-ATPase. Recent evidence suggests that low molar concentrations of ouabain alter cell growth. Studies were conducted to examine the effect of ouabain on Akt phosphorylation and rate of cell proliferation in opossum kidney (OK) proximal tubule cells. Cells exposed to 10 nM ouabain displayed increased Akt Ser(473) phosphorylation, as evidenced by an increase in phospho-Akt Ser(473) band density. Ouabain-stimulated Akt Ser(473) phosphorylation was inhibited by pretreatment with phosphatidylinositol 3-kinase (PI3K) inhibitors (LY294002 and wortmannin), a PLC inhibitor (edelfosine), and an Akt inhibitor. Moreover, ouabain-mediated Akt Ser(473) phosphorylation was suppressed by reduction of extracellular calcium (EGTA) or when intracellular calcium was buffered by BAPTA-AM. An inhibitor of calcium store release (TMB-8) and an inhibitor of calcium entry via store-operated calcium channels (SKF96365) also suppressed ouabain-mediated Akt Ser(473) phosphorylation. In fura-2 AM-loaded cells, 10 nM ouabain increased capacitative calcium entry (CCE). Ouabain at 10 nM did not significantly alter baseline cytoplasmic calcium concentration in control cells. However, treatment with 10 nM ouabain caused a significantly higher ATP-mediated calcium store release. After 24 h, 10 nM ouabain increased the rate of cell proliferation. The Akt inhibitor, BAPTA-AM, SKF96365, and cyclopiazonic acid suppressed the increase in the rate of cell proliferation caused by 10 nM ouabain. Ouabain at 10 nM caused a detectable increase in (86)Rb uptake but did not significantly alter Na(+)-K(+)-ATPase (ouabain-sensitive pNPPase) activity in crude membranes or cell sodium content. Taken together, the results point to a role for CCE and Akt phosphorylation, in response to low concentrations of ouabain, that increase the rate of cell proliferation without inhibiting Na(+)-K(+)-ATPase-mediated ion transport.
    Tipo de documento:
    Referencia
    Referencia del producto:
    AB9094
    Nombre del producto:
    Anti-Sodium Pump (Na+ pump (Na+-K+-ATPase)) α 2 Isoform Antibody

  • GABA and glutamate immunoreactivity in tentacles of the sea anemone Phymactis papillosa (LESSON 1830). 20309875

    Sea anemones have a structurally simple nervous system that controls behaviors like feeding, locomotion, aggression, and defense. Specific chemical and tactile stimuli are transduced by ectodermal sensory cells and transmitted via a neural network to cnidocytes and epithelio-muscular cells, but the nature of the neurotransmitters operating in these processes is still under discussion. Previous studies demonstrated an important role of peptidergic transmission in cnidarians, but during the last decade the contribution of conventional neurotransmitters became increasingly evident. Here, we used immunohistochemistry on light and electron microscopical preparations to investigate the localization of glutamate and GABA in tentacle cross-sections of the sea anemone Phymactis papillosa. Our results demonstrate strong glutamate immunoreactivity in the nerve plexus, while GABA labeling was most prominent in the underlying epithelio-muscular layer. Immunoreactivity for both molecules was also found in glandular epithelial cells, and putative sensory cells were GABA positive. Under electron microscopy, both glutamate and GABA immunogold labeling was found in putative neural processes within the neural plexus. These data support a function of glutamate and GABA as signaling molecules in the nervous system of sea anemones.
    Tipo de documento:
    Referencia
    Referencia del producto:
    AB133
    Nombre del producto:
    Anti-Glutamate Antibody