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Growth pattern of tumours in mice induced by murine Moloney sarcoma-virus and sarcoma-virus-transformed cells.

Weiland, F.; Weiland, E.; Mussgay, M.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /12/1979 Português
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Transplantation of a Moloney sarcoma-virus (MSV-M)-transformed producer cell line (Sac(+)) induced progressively or regressively growing tumours in mice. Progressive growth always occurred after transplantation of an MSV-M non-producer transformant (Sac(-)), whereas the MSV-M released from the producer cells (Sac virus) always induced tumours which regressed. In contrast to the non-producer, the producer transformant Sac(+) as well as Sac virus induced a strong immune response, detected in vitro by cell- and antibody-mediated cytotoxicity assays, and in vivo by transplantation immunity. Implantation of Sac(-) cells led to solid, under-vascularized tumours, consisting histologically of uniform densely packed tumour cells. Sac-virus-induced tumours, however, were very well vascularized and arose by proliferation of different connective-tissue cells. After transplantation of Sac(+) cells, tumours were found to consist of typical tumour cells morphologically similar to Sac(-) cells intermingled with proliferated connective-tissue cells. Cultivation of tumour fragments from Sac(+) and Sac(-) tumours was followed by outgrowth of transformed tumour cells with the properties of the originally implanted cells. Tumour explant cultures from Sac-virus-induced tumours did not lead to growth of stably transformed cells. Co-culture of mouse embryo fibroblasts (MEF) with Sac(+) cells resulted in overgrowth of the transformed cells. Infection of MEF with Sac virus led to transiently transformed cells. It is concluded that Sac(+) cell tumours will resist the strong immune defence mechanisms they induce and grow progressively...

TR4 Orphan Nuclear Receptor functions as an Apoptosis Modulator via Regulation of Bcl-2 gene Expression

Kim, Eungseok; Ma, Wen-Lung; Lin, Din-Lii; Inui, Shigeki; Chen, Yuh-Ling; Chang, Chawnshang
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
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While Bcl-2 plays an important role in cell apoptosis, its relationship to the orphan nuclear receptors remains unclear. Here we report that mouse embryonic fibroblast (MEF) cells prepared from TR4-deficient (TR4−/−) mice are more susceptible to UV-irradiation mediated apoptosis compared to TR4-Wildtype (TR4+/−) littermates. Substantial increasing TR4−/− MEF apoptosis to UV-irradiation was correlated to the down-regulation of Bcl-2 RNA and protein expression and collaterally increased caspase-3 activity. Furthermore, this TR4-induced Bcl-2 gene expression can be suppressed by co-transfection with TR4 coregulators, such as androgen receptor (AR) and receptor-interacting protein 140 (RIP140) in a dose-dependent manner. Together, our results demonstrate that TR4 might function as an apoptosis modulator through induction of Bcl-2 gene expression.

Altered Levels of STAT1 and STAT3 Influence the Neuronal Response to Interferon Gamma

Rose, R. Wesley; Vorobyeva, Anna G.; Skipworth, Jason D.; Nicolas, Emmanuelle; Rall, Glenn F.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
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As immune responses in the CNS are highly regulated, cell-specific differences in IFNγ signaling may be integral in dictating the outcome of host cell responses. In comparing the response of IFNγ-treated primary neurons to control MEF, we observed that neurons demonstrated lower basal expression of both STAT1 and STAT3, the primary signal transducers responsible for IFNγ signaling. Following IFNγ treatment of these cell populations, we noted muted and delayed STAT1 phosphorylation, no detectable STAT3 phosphorylation, and a 3-10-fold lower level of representative IFNγ-responsive gene transcripts. Moreover, in response to a brief pulse of IFNγ, a steady increase in STAT1 phosphorylation and IFNγ gene expression over 48 h was observed in neurons, as compared to rapid attenuation in MEF. These distinct response kinetics in IFNγ-stimulated neurons may reflect modifications in the IFNγ negative feedback loop, which may provide a mechanism for the cell-specific heterogeneity of responses to IFNγ.

Interferon enhances the susceptibility of virus-infected fibroblasts to cytotoxic T cells

Fonte: The Rockefeller University Press Publicador: The Rockefeller University Press
Tipo: Artigo de Revista Científica
Publicado em 01/01/1985 Português
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Interferon (IFN) pretreatment of low-passage mouse embryonic fibroblasts (MEF) infected with lymphocytic choriomeningitis virus or vaccinia virus rendered these cells two to three times more susceptible to lysis by H-2 restricted, virus-specific cytotoxic T lymphocytes (CTL) than control, virus-infected MEF. The increased sensitivity to lysis correlated with increased expression of surface H-2 antigens, but not viral antigens. Continuous cell lines already highly sensitive to CTL-mediated lysis and already expressing high levels of surface H-2 antigens were unaffected by IFN pretreatment. These results suggest that IFN treatment, by increasing surface H-2 levels, may result in increased association of surface H-2 and virus antigens, leading to enhanced recognition and lysis by virus-specific CTL.

Continuous replication of Friend virus complex (spleen focus-forming virus-lymphatic leukemia-inducing virus) in mouse embryo fibroblasts. Retention of leukemogenicity and loss of immunosuppressive properties

Fonte: The Rockefeller University Press Publicador: The Rockefeller University Press
Tipo: Artigo de Revista Científica
Publicado em 01/10/1975 Português
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Exposure of NIH Swiss mouse embryo fibroblasts (MEF) to infectious Friend virus (FV) complex [containing defective spleen focus-forming virus (SFFV) and endogenous NB-tropic leukemia-inducing helper virus (LLV-F)] resulted in the productive infection of these cells by both SFFV and LLV-F. Stocks of SFFV derived after extensive growth in this Swiss MEF cell culture system are fully leukemogenic in adult mice for the induction of erythroleukemia and spleen foci. In addition, in vitro- derived LLV-F, when isolated free of SFFV, is fully leukemogenic for the induction of lymphatic leukemia when inoculated into susceptible newborn BALB/c mice. The host range of in vitro-derived FV complex (i.e., FV-TC) for focus formation in vivo is NB-tropic. Unlike in vivo- derived FV complex, FV-TC does not suppress the responsiveness of murine thymocytes to concanavalin A (Con A) in vitro. Rather, FV-TC acts as a mitogen to nonspecifically stimulate the proliferation of BALB/c thymocytes. The mitogenicity of in vitro-derived FV complex is directly associated with the presence of type-C virus particles, is a heat-labile and UV-sensitive property of the virus, and may be primarily due to LLV since equivalent amounts of LLV with or without SFFV present are equally mitogenic. One in vivo passage of FV-TC resulted in the total loss of this mitogenic property with the reappearance of full immunosuppressive properties. This result demonstrates a clear association between in vivo growth of FV and its ability to suppress mouse thymocytes...

Negative Regulation of the Sapk/Jnk Signaling Pathway by Presenilin 1

Kim, Jin Woo; Chang, Tong-Shin; Lee, Ji Eun; Huh, Sung-Ho; Yeon, Seung Woo; Yang, Wan Seok; Joe, Cheol O.; Mook-Jung, Inhee; Tanzi, Rudolph E.; Kim, Tae-Wan; Choi, Eui-Ju
Fonte: The Rockefeller University Press Publicador: The Rockefeller University Press
Tipo: Artigo de Revista Científica
Publicado em 30/04/2001 Português
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Presenilin 1 (PS1) plays a pivotal role in Notch signaling and the intracellular metabolism of the amyloid β-protein. To understand intracellular signaling events downstream of PS1, we investigated in this study the action of PS1 on mitogen-activated protein kinase pathways. Overexpressed PS1 suppressed the stress-induced stimulation of stress-activated protein kinase (SAPK)/c-Jun NH2-terminal kinase (JNK) in human embryonic kidney 293 cells. Interestingly, two functionally inactive PS1 mutants, PS1(D257A) and PS1(D385A), failed to inhibit UV-stimulated SAPK/JNK. Furthermore, H2O2- or UV-stimulated SAPK activity was higher in mouse embryonic fibroblast (MEF) cells from PS1-null mice than in MEF cells from PS+/+ mice. MEFPS1(−/−) cells were more sensitive to the H2O2-induced apoptosis than MEFPS1(+/+) cells. Ectopic expression of PS1 in MEFPS1(−/−) cells suppressed H2O2-stimulated SAPK/JNK activity and apoptotic cell death. Together, our data suggest that PS1 inhibits the stress-activated signaling by suppressing the SAPK/JNK pathway.

Interplay Between Bax, Reactive Oxygen Species Production And Cardiolipin Oxidation During Apoptosis

Jiang, Jianfei; Huang, Zhentai; Zhao, Qing; Feng, Weihong; Belikova, Natalia A.; Kagan, Valerian E.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
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Bax/Bak activation and cardiolipin peroxidation are essential for cytochrome c release during apoptosis, yet, the link between them remains elusive. We report that sequence of events after exposure of mouse embryonic fibroblast (MEF) cells to actinomycin D followed the order: Bax translocation -> superoxide production -> cardiolipin peroxidation. Genetic ablation of Bax/Bak inhibited actinomycin D induced superoxide production and cardiolipin peroxidation. Rotenone caused robust superoxide generation but did not trigger cardiolipin peroxidation in Bax/Bak double knockout MEF cells. This suggests that, in addition to participating in ROS generation, Bax/Bak play another specific role in cardiolipin oxidation. In isolated mitochondria, recombinant Bax enhanced succinate induced cardiolipin oxidation and cytochrome c release. Mitochondrial peroxidase activity, likely involved in cardiolipin peroxidation, was enhanced upon incubation with recombinant Bax. Thus cardiolipin peroxidation may be causatively and time-dependently related to Bax/Bak effects on ROS generation and peroxidase activation of cytochrome c.

Angular-Dependent Metal-Enhanced Fluorescence from Silver Island Films

Aslan, Kadir; Malyn, Stuart N.; Geddes, Chris D.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em 03/03/2008 Português
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In this letter we report the observation of angular-dependent Metal Enhanced Fluorescence (MEF) from fluorophores deposited onto silver island films (SiFs). When illuminated with laser light (473 nm) at angles of 45 and 90 degrees from the surface, SiFs scattered light at wide observation angles biased by the direction of the incident light. We observed angular-dependent MEF (10-fold) from FITC-HSA immobilized onto the SiFs, again slightly biased with respect to the direction of the incident light. We also measured the photostability of FITC from the back of the glass substrate at angles of 225 and 340 degrees.

A Novel Protein, Luman/CREB3 Reruitment Factor, Inhibits Luman Activation of the Unfolded Protein Response▿

Audas, Timothy E.; Li, Yu; Liang, Genqing; Lu, Rui
Fonte: American Society for Microbiology (ASM) Publicador: American Society for Microbiology (ASM)
Tipo: Artigo de Revista Científica
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Luman/CREB3 (also called LZIP) is an endoplasmic reticulum (ER)-bound cellular transcription factor. It has been implicated in the mammalian unfolded protein response (UPR), as well as herpes simplex virus reactivation from latency in sensory neurons. Here, we report the identification of a novel Luman recruitment factor (LRF). Like Luman, LRF is a UPR-responsive basic-region leucine zipper protein that is prone to proteasomal degradation. Being a highly unstable protein, LRF interacts with Luman through the leucine zipper region and promotes Luman degradation. LRF was found to recruit the nuclear form of Luman to discrete nuclear foci, which overlap with the nuclear receptor coactivator GRIP1 bodies, and repress the transactivation activity of Luman. Compared to LRF+/+ mouse embryonic fibroblast (MEF) cells, the levels of CHOP, EDEM, and Herp were elevated in LRF−/− MEF cells. We propose that LRF is a negative regulator of the UPR. For Luman, it may represent another level of regulation following Luman proteolytic cleavage on the ER and nuclear translocation. In addition to inducing rapid Luman turnover, LRF may repress the transactivation potential of Luman by sequestering it in the LRF nuclear bodies away from key cofactors (such as HCF-1) that are required for transcriptional activation.

Human ES cells: Starting Culture from Frozen Cells

Trish, Erin; Dimos, John; Eggan, Kevin
Fonte: MyJove Corporation Publicador: MyJove Corporation
Tipo: Artigo de Revista Científica
Publicado em 09/11/2006 Português
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Here we demonstrate how our lab begins a HuES human embryonic stem cell line culture from a frozen stock. First, a one to two day old ten cm plate of approximately one (to two) million irradiated mouse embryonic fibroblast feeder cells is rinsed with HuES media to remove residual serum and cell debris, and then HuES media added and left to equilibrate in the cell culture incubator. A frozen vial of cells from long term liquid nitrogen storage or a -80C freezer is sourced and quickly submerged in a 37C water bath for quick thawing. Cells in freezing media are then removed from the vial and placed in a large volume of HuES media. The large volume of HuES media facilitates removal of excess serum and DMSO, which can cause HuES human embryonic stem cells to differentiate. Cells are gently spun out of suspension, and then re-suspended in a small volume of fresh HuES media that is then used to seed the MEF plate. It is considered important to seed the MEF plate by gently adding the HuES cells in a drop wise fashion to evenly disperse them throughout the plate. The newly established HuES culture plate is returned to the incubator for 48 hrs before media is replaced, then is fed every 24 hours thereafter.

Sulforhodamine Adsorbed Langmuir-Blodgett Layers on Silver Island Films: Effect of Probe Distance on the Metal-Enhanced Fluorescence

Ray, Krishanu; Badugu, Ramachandram; Lakowicz, Joseph R.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em //2007 Português
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Metal-Enhanced Fluorescence (MEF) has become an important method in biomedical sensing. In this paper, we present the distance-dependent MEF of sulforhodamine B (SRB) monolayer on silver island films (SIFs). SRB is electrostatically incorporated into the Langmuir-Blodgett (LB) layers of octadecylamine (ODA) deposited on glass and SIFs substrates. The distances between SRB and SIFs or glass surfaces are controlled by depositing a varied number of inert stearic acid (SA) spacer layers. SRB is incorporated into positively charged LB layers of ODA by immersing the ODA deposited substrates into aqueous solution of SRB. Dye incorporated ODA layers with 10 nm separation distance from the SIFs surface show maximum metal-enhanced fluorescence intensity; ~7-fold increase in intensity as compared to that from the glass surface. The corresponding enhancement factor is reduced with increasing or decreasing the probe distance from the SIFs surface. Additionally, SRB on SIF surfaces show reduced lifetimes. We observed the shortest lifetime from the SRB with 5 nm distance from the SIF surfaces and the lifetime increased consistently with increasing the distances between the fluorophore and the SIFs surface. These observed spectral changes, increase in fluorescence intensity and decreased fluorescence lifetimes...

Comprehensive Dissection of PDGF-PDGFR Signaling Pathways in PDGFR Genetically Defined Cells

Wu, Erxi; Palmer, Nathan; Tian, Ze; Moseman, Annie P.; Galdzicki, Michal; Wang, Xuetao; Berger, Bonnie; Zhang, Hongbing; Kohane, Isaac S.
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 24/11/2008 Português
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Despite the growing understanding of PDGF signaling, studies of PDGF function have encountered two major obstacles: the functional redundancy of PDGFRα and PDGFRβ in vitro and their distinct roles in vivo. Here we used wild-type mouse embryonic fibroblasts (MEF), MEF null for either PDGFRα, β, or both to dissect PDGF-PDGFR signaling pathways. These four PDGFR genetically defined cells provided us a platform to study the relative contributions of the pathways triggered by the two PDGF receptors. They were treated with PDGF-BB and analyzed for differential gene expression, in vitro proliferation and differential response to pharmacological effects. No genes were differentially expressed in the double null cells, suggesting minimal receptor-independent signaling. Protean differentiation and proliferation pathways are commonly regulated by PDGFRα, PDGFRβ and PDGFRα/β while each receptor is also responsible for regulating unique signaling pathways. Furthermore, some signaling is solely modulated through heterodimeric PDGFRα/β.

From MEFs to Matrigel 2: Splitting hESCs from MEFs onto Matrigel

Khvorostov, Ivan; Zhang, Jin; Teitell, Michael
Fonte: MyJove Corporation Publicador: MyJove Corporation
Tipo: Artigo de Revista Científica
Publicado em 09/06/2008 Português
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This video demonstrates how to grow human embryonic stem cells (hESCs) on mouse embryonic fibroblast (MEF) feeder cells, how to passage hESCs from MEF plates to feeder cell-free Matrigel plates.

Nrf2 protects human bladder urothelial cells from arsenite and monomethylarsonous acid toxicity

Wang, Xiao-Jun; Sun, Zheng; Chen, Weimin; Eblin, Kylee E.; Gandolfi, A. Jay; Zhang, Donna D.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
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Arsenic is widely spread in our living environment and imposes a big challenge on human health worldwide. Arsenic damages biological systems through multiple mechanisms including the generation of reactive oxygen species. The transcription factor Nrf2 regulates the cellular antioxidant response that protects cells from various insults. In this study, the protective role of Nrf2 in arsenic toxicity was investigated in a human bladder urothelial cell line, UROtsa. Using an UROtsa cell line stably infected with Nrf2-siRNA, we clearly demonstrate that compromised Nrf2 expression sensitized the cells to As(III)- and MMA(III)-induced toxicity. On the other hand, the activation of the Nrf2 pathway by tert-butylhydroquinone (tBHQ) and sulforaphane (SF), the known Nrf2-inducers, rendered UROtsa cells more resistant to As(III)- and MMA(III). Furthermore, the wild type mouse embryo fibroblast (WT-MEF) cells were protected from As(III)- and MMA(III)-induced toxicity following Nrf2 activation by tBHQ or SF whereas neither tBHQ nor SF conferred protection in the Nrf2−/−-MEF cells, demonstrating that tBHQ- or SF-mediated protection against As(III)- and MMA(III)-induced toxicity depends on Nrf2 activation. These results, obtained by both loss of function and gain of function analyses...

ATF4 is Necessary and Sufficient for ER Stress-induced Upregulation of REDD1 Expression

Whitney, Michael L.; Jefferson, Leonard S.; Kimball, Scot R.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
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In response to a variety of cell stresses, e.g. endoplasmic reticulum (ER) stress, expression of REDD1 (regulated in development and DNA damage responses) is transcriptionally upregulated. However, the mechanism through which ER stress acts to upregulate REDD1 expression is unknown. In the present study, REDD1 expression was found to be upregulated by ER stress in several cell lines. However, in MEF cells lacking the eIF2α kinase PERK, ER stress failed to upregulate REDD1 expression, demonstrating that phosphorylation of eIF2α was necessary for the effect. Moreover, ER stress led to upregulated expression of the transcription factor ATF4, but in MEF cells lacking ATF4, REDD1 mRNA expression was not increased by ER stress. In contrast, exogenous expression of ATF4 was sufficient to induce REDD1 expression. Overall, the results suggest that REDD1 expression is upregulated during ER stress through a mechanism involving activation of PERK, phosphorylation of eIF2α, and increased ATF4 expression.

The ubiquitously expressed bZIP inhibitor, JDP2, suppresses the transcription of its homologue immediate early gene counterpart, ATF3

Weidenfeld-Baranboim, Keren; Hasin, Tal; Darlyuk, Ilona; Heinrich, Ronit; Elhanani, Ofer; Pan, Jianzhi; Yokoyama, Kazunari K.; Aronheim, Ami
Fonte: Oxford University Press Publicador: Oxford University Press
Tipo: Artigo de Revista Científica
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JDP2 is a ubiquitously expressed bZIP repressor protein. JDP2 binds TPA response element and cyclic AMP response element located within various promoters. JDP2 displays a high degree of homology to the immediate early gene ATF3. ATF3 plays a crucial role in the cellular adaptive response to multiple stress insults as well as growth stimuli. We have identified ATF3 as a potential target gene for JDP2 repression. JDP2 regulates the ATF3 promoter potentially through binding to both the consensus ATF/CRE site and a non-consensus ATF3 auto-repression DNA-binding element. Expression of ATF3 protein in wild-type mouse embryo fibroblast (MEF) cells is below the detectable levels, whereas, JDP2 disrupted MEF cells display noticeable level of ATF3 protein. Following either serum or ER stress stimulation, ATF3 expression is potentiated in JDP2-KO fibroblast cells as compared with wild-type cells. Mice with either JDP2 over-expression or JDP2 disruption display undetectable level of ATF3 protein. However, ATF3 induction in response to either growth or stress signals is dependent on JDP2 expression level. ATF3 induction is attenuated in JDP2 over-expressing mice whereas is potentiated in JDP2-KO mice as compared with the corresponding wild-type mice. Collectively...

Metal-Enhanced Fluorescence from Nanoparticulate Zinc Films

Aslan, Kadir; Previte, Michael J.R.; Zhang, Yongxia; Geddes, Chris D.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em 27/11/2008 Português
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A detailed study of metal-enhanced fluorescence (MEF) from fluorophores in the blue-to- red spectral region placed in close proximity to thermally evaporated zinc nanostructured films is reported. The zinc nanostructured films were deposited onto glass microscope slides as individual particles and were 1–10 nm in height and 20–100 nm in width, as characterized by Atomic Force Microscopy. The surface plasmon resonance peak of the zinc nanostructured films was ≈ 400 nm. Finite-difference time-domain calculations for single and multiple nanostructures organized in a staggered fashion on a solid support predict, as expected, that the electric fields are concentrated both around and between the nanostructures. Additionally, Mie scattering calculations show that the absorption and scattering components of the extinction spectrum are dominant in the UV and visible spectral ranges, respectively. Enhanced fluorescence emission accompanied by no significant changes in excited state lifetimes of fluorophores with emission wavelengths in the visible blue-to-red spectral range near-to zinc nanostructured films were observed, implying that MEF from zinc nanostructured films is mostly due to an electric field enhancement effect.

Microwave-Accelerated and Metal-Enhanced Fluorescence Myoglobin Detection on Silvered Surfaces: Potential Application to Myocardial Infarction Diagnosis

Aslan, Kadir; Geddes, Chris D.
Fonte: Kluwer Academic Publishers-Plenum Publishers Publicador: Kluwer Academic Publishers-Plenum Publishers
Tipo: Artigo de Revista Científica
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In this short paper, we describe a novel approach to both significantly accelerate and optically amplify fluorescence-based immunoassays. Our approach utilizes metal-enhanced fluorescence (MEF) to intrinsically optically amplify fluorescence signatures, which, when combined with the use of low-power microwaves to kinetically accelerate assays, provides for both ultrafast and ultrabright immunoassays. Surprisingly, the use of low-power microwaves and silver nanostructures provides for localized heating, concentrating the effect to the particles themselves as compared to the generic heating of the high dielectric assay fluid. We have subsequently applied our microwave-accelerated MEF approach to the detection of myoglobin, where its rapid quantification is paramount for the clinical assessment of an acute myocardial infarction.

Genome-Wide uH2A Localization Analysis Highlights Bmi1-Dependent Deposition of the Mark at Repressed Genes

Kallin, Eric M.; Cao, Ru; Jothi, Raja; Xia, Kai; Cui, Kairong; Zhao, Keji; Zhang, Yi
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
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Polycomb group (PcG) proteins control organism development by regulating the expression of developmental genes. Transcriptional regulation by PcG proteins is achieved, at least partly, through the PRC2-mediated methylation on lysine 27 of histone H3 (H3K27) and PRC1-mediated ubiquitylation on lysine 119 of histone H2A (uH2A). As an integral component of PRC1, Bmi1 has been demonstrated to be critical for H2A ubiquitylation. Although recent studies have revealed the genome-wide binding patterns of some of the PRC1 and PRC2 components, as well as the H3K27me3 mark, there have been no reports describing genome-wide localization of uH2A. Using the recently developed ChIP-Seq technology, here, we report genome-wide localization of the Bmi1-dependent uH2A mark in MEF cells. Gene promoter averaging analysis indicates a peak of uH2A just inside the transcription start site (TSS) of well-annotated genes. This peak is enriched at promoters containing the H3K27me3 mark and represents the least expressed genes in WT MEF cells. In addition, peak finding reveals regions of local uH2A enrichment throughout the mouse genome, including almost 700 gene promoters. Genes with promoter peaks of uH2A exhibit lower-level expression when compared to genes that do not contain promoter peaks of uH2A. Moreover...

Human DNA polymerase β polymorphism, Arg137Gln, impairs its polymerase activity and interaction with PCNA and the cellular base excision repair capacity

Guo, Zhigang; Zheng, Li; Dai, Huifang; Zhou, Mian; Xu, Hong; Shen, Binghui
Fonte: Oxford University Press Publicador: Oxford University Press
Tipo: Artigo de Revista Científica
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DNA polymerase β (Pol β) is a key enzyme in DNA base excision repair, and an important factor for maintaining genome integrity and stability. More than 30% of human tumors characterized to date express DNA Pol β variants, many of which result from a single nucleotide residue substitution. However, in most cases, their precise functional deficiency and relationship to cancer susceptibility are still unknown. In the current work, we show that a polymorphism encoding an arginine to glutamine substitution, R137Q, has lower polymerase activity. The substitution also affects the interaction between Pol β and proliferating cell nuclear antigen (PCNA). These defects impair the DNA repair capacity of Pol β in reconstitution assays, as well as in cellular extracts. Expression of wild-type Pol β in pol β−/− mouse embryonic fibroblast (MEF) cells restored cellular resistance to DNA damaging reagents such as methyl methanesulfonate (MMS) and N-methyl-N-nitrosourea (MNU), while expression of R137Q in pol β−/− MEF cells failed to do so. These data indicate that polymorphisms in base excision repair genes may contribute to the onset and development of cancers.