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Sample records for inducible factor regulatory

  1. Ikkepsilon regulates viral-induced interferon regulatory factor-3 activation via a redox-sensitive pathway

    SciTech Connect

    Indukuri, Hemalatha; Castro, Shawn M.; Liao, S.-M.; Feeney, Lee Ann; Dorsch, Marion; Coyle, Anthony J.; Garofalo, Roberto P.; Brasier, Allan R.; Casola, Antonella . E-mail: ancasola@utmb.edu

    2006-09-15

    Respiratory syncytial virus (RSV)-induced chemokine gene expression occurs through the activation of a subset of transcription factors, including Interferon Regulatory Factor (IRF)-3. In this study, we have investigated the signaling pathway leading to RSV-induced IRF-3 activation and whether it is mediated by intracellular reactive oxygen species (ROS) generation. Our results show that RSV infection induces expression and catalytic activity of IKK{epsilon}, a noncanonical IKK-like kinase. Expression of a kinase-inactive IKK{epsilon} blocks RSV-induced IRF-3 serine phosphorylation, nuclear translocation and DNA-binding, leading to inhibition of RANTES gene transcription, mRNA expression and protein synthesis. Treatment of alveolar epithelial cells with antioxidants or with NAD(P)H oxidase inhibitors abrogates RSV-induced chemokine secretion, IRF-3 phosphorylation and IKK{epsilon} induction, indicating that ROS generation plays a fundamental role in the signaling pathway leading to IRF-3 activation, therefore, identifying a novel molecular target for the development of strategies aimed to modify the inflammatory response associated with RSV infection of the lung.

  2. Regulatory elements responsible for inducible expression of the granulocyte colony-stimulating factor gene in macrophages.

    PubMed Central

    Nishizawa, M; Nagata, S

    1990-01-01

    Granulocyte colony-stimulating factor (G-CSF) plays an essential role in granulopoiesis during bacterial infection. Macrophages produce G-CSF in response to bacterial endotoxins such as lipopolysaccharide (LPS). To elucidate the mechanism of the induction of G-CSF gene in macrophages or macrophage-monocytes, we have examined regulatory cis elements in the promoter of mouse G-CSF gene. Analyses of linker-scanning and internal deletion mutants of the G-CSF promoter by the chloramphenicol acetyltransferase assay have indicated that at least three regulatory elements are indispensable for the LPS-induced expression of the G-CSF gene in macrophages. When one of the three elements was reiterated and placed upstream of the TATA box of the G-CSF promoter, it mediated inducibility as a tissue-specific and orientation-independent enhancer. Although this element contains a conserved NF-kappa B-like binding site, the gel retardation assay and DNA footprint analysis with nuclear extracts from macrophage cell lines demonstrated that nuclear proteins bind to the DNA sequence downstream of the NF-kappa B-like element, but not to the conserved element itself. The DNA sequence of the binding site was found to have some similarities to the LPS-responsive element which was recently identified in the promoter of the mouse class II major histocompatibility gene. Images PMID:1691438

  3. S. macrurus myogenic regulatory factors induce mammalian skeletal muscle differentiation: Evidence for functional conservation of MRFs

    PubMed Central

    Kim, Hyun-Jung; Güth, Robert; Jonsson, Colleen B.; Unguez, Graciela A.

    2009-01-01

    The current-producing cells of the electric organ (EO), i.e., electrocytes, in Sternopygus macrurus derive from skeletal muscle fibers. Mature electrocytes are not contractile but they do retain some muscle proteins, are multinucleated, and receive cholinergic innervation. Electrocytes express the myogenic regulatory factors (MRFs) MyoD, myogenin, Myf5 and MRF4 despite their incomplete muscle phenotype. Although S. macrurus MRFs share functional domains that are highly conserved and their expression is confined to the myogenic lineage, their capability to induce the muscle phenotype has not been determined. To test the functional conservation of S. macrurus MRFs to transcriptionally activate skeletal muscle gene expression and induce the myogenic program, we transiently over-expressed S. macrurus MyoD (SmMyoD) and myogenin (SmMyoG) in mouse C3H/10T1/2 and NIH3T3 embryonic cells. RT-PCR and immunolabeling studies showed that SmMyoD and SmMyoG efficiently can convert these two cell lines into multinucleated myotubes that expressed differentiated muscle markers. The levels of myogenic induction by SmMyoD and SmMyoG were comparable to those obtained with mouse MRF homologs. Furthermore, SmMyoD and SmMyoG proteins were able to induce mouse MyoD and myogenin in C3H/10T1/2 cells. We conclude that S. macrurus MRFs are functionally conserved as they can transcriptionally activate skeletal muscle gene expression and induce the myogenic program in mammalian non-muscle cells. Hence, these data suggest that the partial muscle phenotype of electrocytes is not likely due to differences in the MRF-dependent transcriptional program between skeletal muscle and electric organ. PMID:19598116

  4. Interferon regulatory factor-8 modulates the development of tumour-induced CD11b+Gr-1+ myeloid cells.

    PubMed

    Stewart, Trina J; Greeneltch, Kristy M; Reid, Julia E; Liewehr, David J; Steinberg, Seth M; Liu, Kebin; Abrams, Scott I

    2009-09-01

    Tumour-induced myeloid-derived suppressor cells (MDSC) promote immune suppression and mediate tumour progression. However, the molecular basis for the generation of MDSC, which in mice co-express the CD11b(+) and Gr-1(+) cell surface markers remains unclear. Because CD11b(+)Gr-1(+) cells expand during progressive tumour growth, this suggests that tumour-induced events alter signalling pathways that affect normal myeloid cell development. Interferon regulatory factor-8 (IRF-8), a member of the IFN-gamma regulatory factor family, is essential for normal myelopoiesis. We therefore examined whether IRF-8 modulated tumour-induced CD11b(+)Gr-1(+) cell development or accumulation using both implantable (4T1) and transgenic (MMTV-PyMT) mouse models of mammary tumour growth. In the 4T1 model, both splenic and bone marrow-derived CD11b(+)Gr-1(+) cells of tumour-bearing mice displayed a marked reduction in IRF-8 expression compared to control populations. A causal link between IRF-8 expression and the emergence of tumour-induced CD11b(+)Gr-1(+) cells was explored in vivo using a double transgenic (dTg) mouse model designed to express transgenes for both IRF-8 and mammary carcinoma development. Despite the fact that tumour growth was unaffected, splenomegaly, as well as the frequencies and absolute numbers of CD11b(+)Gr-1(+) cells were significantly lower in dTg mice when compared with single transgenic tumour-bearing mice. Overall, these data reveal that IRF-8 plays an important role in tumour-induced development and/or accumulation of CD11b(+)Gr-1(+) cells, and establishes a molecular basis for the potential manipulation of these myeloid populations for cancer therapy. PMID:20196788

  5. TCDD Induces the Hypoxia-Inducible Factor (HIF)-1α Regulatory Pathway in Human Trophoblastic JAR Cells

    PubMed Central

    Liao, Tien-Ling; Chen, Su-Chee; Tzeng, Chii-Reuy; Kao, Shu-Huei

    2014-01-01

    The exposure to dioxin can compromise pregnancy outcomes and increase the risk of preterm births. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) has been demonstrated to induce placental hypoxia at the end of pregnancy in a rat model, and hypoxia has been suggested to be the cause of abnormal trophoblast differentiation and placental insufficiency syndromes. In this study, we demonstrate that the non-hypoxic stimulation of human trophoblastic cells by TCDD strongly increased hypoxia inducible factor-1 alpha (HIF-1α) stabilization. TCDD exposure induced the generation of reactive oxygen species (ROS) and nitric oxide. TCDD-induced HIF-1α stabilization and Akt phosphorylation was inhibited by pretreatment with wortmannin (a phosphatidylinositol 3-kinase (PI3K) inhibitor) or N-acetylcysteine (a ROS scavenger). The augmented HIF-1α stabilization by TCDD occurred via the ROS-dependent activation of the PI3K/Akt pathway. Additionally, a significant increase in invasion and metallomatrix protease-9 activity was found in TCDD-treated cells. The gene expression of vascular endothelial growth factor and placental growth factor was induced upon TCDD stimulation, whereas the protein levels of peroxisome proliferator-activated receptor γ (PPARγ), PPARγ coactivator-1α, mitochondrial transcription factor, and uncoupling protein 2 were decreased. Our results indicate that an activated HIF-1α pathway, elicited oxidative stress, and induced metabolic stress contribute to TCDD-induced trophoblastic toxicity. These findings may provide molecular insight into the TCDD-induced impairment of trophoblast function and placental development. PMID:25272228

  6. Altered expression of hypoxia-inducible factor-1α (HIF-1α) and its regulatory genes in gastric cancer tissues.

    PubMed

    Wang, Jihan; Ni, Zhaohui; Duan, Zipeng; Wang, Guoqing; Li, Fan

    2014-01-01

    Tissue hypoxia induces reprogramming of cell metabolism and may result in normal cell transformation and cancer progression. Hypoxia-inducible factor 1-alpha (HIF-1α), the key transcription factor, plays an important role in gastric cancer development and progression. This study aimed to investigate the underlying regulatory signaling pathway in gastric cancer using gastric cancer tissue specimens. The integration of gene expression profile and transcriptional regulatory element database (TRED) was pursued to identify HIF-1α ↔ NFκB1 → BRCA1 → STAT3 ← STAT1 gene pathways and their regulated genes. The data showed that there were 82 differentially expressed genes that could be regulated by these five transcription factors in gastric cancer tissues and these genes formed 95 regulation modes, among which seven genes (MMP1, TIMP1, TLR2, FCGR3A, IRF1, FAS, and TFF3) were hub molecules that are regulated at least by two of these five transcription factors simultaneously and were associated with hypoxia, inflammation, and immune disorder. Real-Time PCR and western blot showed increasing of HIF-1α in mRNA and protein levels as well as TIMP1, TFF3 in mRNA levels in gastric cancer tissues. The data are the first study to demonstrate HIF-1α-regulated transcription factors and their corresponding network genes in gastric cancer. Further study with a larger sample size and more functional experiments is needed to confirm these data and then translate into clinical biomarker discovery and treatment strategy for gastric cancer.

  7. Altered Expression of Hypoxia-Inducible Factor-1α (HIF-1α) and Its Regulatory Genes in Gastric Cancer Tissues

    PubMed Central

    Wang, Jihan; Ni, Zhaohui; Duan, Zipeng; Wang, Guoqing; Li, Fan

    2014-01-01

    Tissue hypoxia induces reprogramming of cell metabolism and may result in normal cell transformation and cancer progression. Hypoxia-inducible factor 1-alpha (HIF-1α), the key transcription factor, plays an important role in gastric cancer development and progression. This study aimed to investigate the underlying regulatory signaling pathway in gastric cancer using gastric cancer tissue specimens. The integration of gene expression profile and transcriptional regulatory element database (TRED) was pursued to identify HIF-1α ↔ NFκB1 → BRCA1 → STAT3 ← STAT1 gene pathways and their regulated genes. The data showed that there were 82 differentially expressed genes that could be regulated by these five transcription factors in gastric cancer tissues and these genes formed 95 regulation modes, among which seven genes (MMP1, TIMP1, TLR2, FCGR3A, IRF1, FAS, and TFF3) were hub molecules that are regulated at least by two of these five transcription factors simultaneously and were associated with hypoxia, inflammation, and immune disorder. Real-Time PCR and western blot showed increasing of HIF-1α in mRNA and protein levels as well as TIMP1, TFF3 in mRNA levels in gastric cancer tissues. The data are the first study to demonstrate HIF-1α-regulated transcription factors and their corresponding network genes in gastric cancer. Further study with a larger sample size and more functional experiments is needed to confirm these data and then translate into clinical biomarker discovery and treatment strategy for gastric cancer. PMID:24927122

  8. Interferon regulatory factor 3 is a key regulation factor for inducing the expression of SAMHD1 in antiviral innate immunity

    PubMed Central

    Yang, Shen; Zhan, Yuan; Zhou, Yanjun; Jiang, Yifeng; Zheng, Xuchen; Yu, Lingxue; Tong, Wu; Gao, Fei; Li, Liwei; Huang, Qinfeng; Ma, Zhiyong; Tong, Guangzhi

    2016-01-01

    SAMHD1 is a type I interferon (IFN) inducible host innate immunity restriction factor that inhibits an early step of the viral life cycle. The underlying mechanisms of SAMHD1 transcriptional regulation remains elusive. Here, we report that inducing SAMHD1 upregulation is part of an early intrinsic immune response via TLR3 and RIG-I/MDA5 agonists that ultimately induce the nuclear translocation of the interferon regulation factor 3 (IRF3) protein. Further studies show that IRF3 plays a major role in upregulating endogenous SAMHD1 expression in a mechanism that is independent of the classical IFN-induced JAK-STAT pathway. Both overexpression and activation of IRF3 enhanced the SAMHD1 promoter luciferase activity, and activated IRF3 was necessary for upregulating SAMHD1 expression in a type I IFN cascade. We also show that the SAMHD1 promoter is a direct target of IRF3 and an IRF3 binding site is sufficient to render this promoter responsive to stimulation. Collectively, these findings indicate that upregulation of endogenous SAMHD1 expression is attributed to the phosphorylation and nuclear translocation of IRF3 and we suggest that type I IFN induction and induced SAMHD1 expression are coordinated. PMID:27411355

  9. Regulatory factor X1-induced down-regulation of transforming growth factor β2 transcription in human neuroblastoma cells.

    PubMed

    Feng, Chenzhuo; Zuo, Zhiyi

    2012-06-29

    Regulatory factor X (RFX) proteins are transcription factors. Seven mammalian RFX proteins have been identified. RFX1 is the prototype RFX. However, its biological functions are not known. Here, RFX1 overexpression reduced fetal bovine serum-stimulated proliferation of SH-SY5Y cells, a human neuroblastoma cell line. This inhibition is associated with decreased transforming growth factor β2 (TGFβ2) and phospho-extracellular signal-regulated kinase (ERK). Exogenous TGFβ2 increased cell proliferation and phospho-ERK in cells overexpressing RFX1. An anti-TGFβ2 antibody and PD98059, an ERK activation inhibitor, inhibited SH-SY5Y cell proliferation. TGFβ2 promoter activity was decreased in cells overexpressing RFX1. Chromosome immunoprecipitation assay showed that RFX1 bound the TGFβ2 promoter. RFX1 down-regulation increased TGFβ2 in SH-SY5Y and HCN-1A cells, a normal human neuronal cell line. More importantly, TGFβ2 concentrations were negatively correlated with RFX1 levels in human medulloblastoma tissues with a R(2) of 0.464. These results suggest that RFX1 reduces cell proliferation through inhibiting the TGFβ2-ERK signaling pathway. RFX1 blocks TGFβ2 expression through its direct action on TGFβ2 transcription. This effect also appears in human brain tumor tissues. Because TGFβ is known to be involved in cancer development, our results provide initial evidence to suggest that RFX1 may play an important role in human tumor biology.

  10. A model for genetic and epigenetic regulatory networks identifies rare pathways for transcription factor induced pluripotency

    NASA Astrophysics Data System (ADS)

    Artyomov, Maxim; Meissner, Alex; Chakraborty, Arup

    2010-03-01

    Most cells in an organism have the same DNA. Yet, different cell types express different proteins and carry out different functions. This is because of epigenetic differences; i.e., DNA in different cell types is packaged distinctly, making it hard to express certain genes while facilitating the expression of others. During development, upon receipt of appropriate cues, pluripotent embryonic stem cells differentiate into diverse cell types that make up the organism (e.g., a human). There has long been an effort to make this process go backward -- i.e., reprogram a differentiated cell (e.g., a skin cell) to pluripotent status. Recently, this has been achieved by transfecting certain transcription factors into differentiated cells. This method does not use embryonic material and promises the development of patient-specific regenerative medicine, but it is inefficient. The mechanisms that make reprogramming rare, or even possible, are poorly understood. We have developed the first computational model of transcription factor-induced reprogramming. Results obtained from the model are consistent with diverse observations, and identify the rare pathways that allow reprogramming to occur. If validated, our model could be further developed to design optimal strategies for reprogramming and shed light on basic questions in biology.

  11. Transcription factors RUNX1 and RUNX3 in the induction and suppressive function of Foxp3+ inducible regulatory T cells

    PubMed Central

    Klunker, Sven; Chong, Mark M.W.; Mantel, Pierre-Yves; Palomares, Oscar; Bassin, Claudio; Ziegler, Mario; Rückert, Beate; Meiler, Flurina; Akdis, Mübeccel; Littman, Dan R.

    2009-01-01

    Forkhead box P3 (FOXP3)+CD4+CD25+ inducible regulatory T (iT reg) cells play an important role in immune tolerance and homeostasis. In this study, we show that the transforming growth factor-β (TGF-β) induces the expression of the Runt-related transcription factors RUNX1 and RUNX3 in CD4+ T cells. This induction seems to be a prerequisite for the binding of RUNX1 and RUNX3 to three putative RUNX binding sites in the FOXP3 promoter. Inactivation of the gene encoding RUNX cofactor core-binding factor-β (CBFβ) in mice and small interfering RNA (siRNA)-mediated suppression of RUNX1 and RUNX3 in human T cells resulted in reduced expression of Foxp3. The in vivo conversion of naive CD4+ T cells into Foxp3+ iT reg cells was significantly decreased in adoptively transferred CbfbF/F CD4-cre naive T cells into Rag2−/− mice. Both RUNX1 and RUNX3 siRNA silenced human T reg cells and CbfbF/F CD4-cre mouse T reg cells showed diminished suppressive function in vitro. Circulating human CD4+ CD25high CD127− T reg cells significantly expressed higher levels of RUNX3, FOXP3, and TGF-β mRNA compared with CD4+CD25− cells. Furthermore, FOXP3 and RUNX3 were colocalized in human tonsil T reg cells. These data demonstrate Runx transcription factors as a molecular link in TGF-β–induced Foxp3 expression in iT reg cell differentiation and function. PMID:19917773

  12. Interferon Regulatory Factor-1 Mediates Alveolar Macrophage Pyroptosis During LPS-Induced Acute Lung Injury in Mice

    PubMed Central

    Wu, Dongdong; Pan, Pinhua; Su, Xiaoli; Zhang, Lemeng; Qin, Qingwu; Tan, Hongyi; Huang, Li; Li, Yuanyuan

    2016-01-01

    ABSTRACT Previously, we demonstrated that pyroptosis in alveolar macrophages (AMs) plays an essential role in lipopolysaccharide (LPS)-induced acute lung injury. However, the underlying mechanism remains largely unclear. Here, we show that the absence of interferon regulatory factor 1 (IRF-1) in genetic knock-out mice strongly abrogates pyroptosis in AMs and alleviates the LPS-induced lung injury and systemic inflammation. Our study demonstrates that IRF-1 contributes to caspase-1 activation and apoptosis-associated speck-like protein containing a caspase activation and recruitment domain pyroptosome formation in AMs and leads to downstream inflammatory cytokine release, including that of IL-1β, IL-18, and HMGB1. The nuclear translocation of IRF-1 is linked to the presence of toll-like receptor 4 (TLR4). Our findings suggest that pyroptosis and the downstream inflammatory response in AMs induced by LPS is a process that is dependent on TLR4-mediated up-regulation of IRF-1. In summary, IRF-1 plays a key role in controlling caspase-1-dependent pyroptosis and inflammation. PMID:26939040

  13. Transcription factors GAF and HSF act at distinct regulatory steps to modulate stress-induced gene activation

    PubMed Central

    Fuda, Nicholas J.; Mahat, Dig B.; Core, Leighton J.; Guertin, Michael J.

    2016-01-01

    The coordinated regulation of gene expression at the transcriptional level is fundamental to development and homeostasis. Inducible systems are invaluable when studying transcription because the regulatory process can be triggered instantaneously, allowing the tracking of ordered mechanistic events. Here, we use precision run-on sequencing (PRO-seq) to examine the genome-wide heat shock (HS) response in Drosophila and the function of two key transcription factors on the immediate transcription activation or repression of all genes regulated by HS. We identify the primary HS response genes and the rate-limiting steps in the transcription cycle that GAGA-associated factor (GAF) and HS factor (HSF) regulate. We demonstrate that GAF acts upstream of promoter-proximally paused RNA polymerase II (Pol II) formation (likely at the step of chromatin opening) and that GAF-facilitated Pol II pausing is critical for HS activation. In contrast, HSF is dispensable for establishing or maintaining Pol II pausing but is critical for the release of paused Pol II into the gene body at a subset of highly activated genes. Additionally, HSF has no detectable role in the rapid HS repression of thousands of genes. PMID:27492368

  14. Suppression of preoptic sleep-regulatory neuronal activity during corticotropin-releasing factor-induced sleep disturbance.

    PubMed

    Gvilia, Irma; Suntsova, Natalia; Kumar, Sunil; McGinty, Dennis; Szymusiak, Ronald

    2015-11-01

    Corticotropin releasing factor (CRF) is implicated in sleep and arousal regulation. Exogenous CRF causes sleep suppression that is associated with activation of at least two important arousal systems: pontine noradrenergic and hypothalamic orexin/hypocretin neurons. It is not known whether CRF also impacts sleep-promoting neuronal systems. We hypothesized that CRF-mediated changes in wake and sleep involve decreased activity of hypothalamic sleep-regulatory neurons localized in the preoptic area. To test this hypothesis, we examined the effects of intracerebroventricular administration of CRF on sleep-wake measures and c-Fos expression in GABAergic neurons in the median preoptic nucleus (MnPN) and ventrolateral preoptic area (VLPO) in different experimental conditions. Administration of CRF (0.1 nmol) during baseline rest phase led to delayed sleep onset and decreases in total amount and mean duration of non-rapid eye movement (NREM) sleep. Administration of CRF during acute sleep deprivation (SD) resulted in suppression of recovery sleep and decreased c-Fos expression in MnPN/VLPO GABAergic neurons. Compared with vehicle controls, intracerebroventricular CRF potentiated disturbances of both NREM and REM sleep in rats exposed to a species-specific psychological stressor, the dirty cage of a male conspecific. The number of MnPN/VLPO GABAergic neurons expressing c-Fos was reduced in the CRF-treated group of dirty cage-exposed rats. These findings confirm the involvement of CRF in wake-sleep cycle regulation and suggest that increased CRF signaling in the brain 1) negatively affects homeostatic responses to sleep loss, 2) exacerbates stress-induced disturbances of sleep, and 3) suppresses the activity of sleep-regulatory neurons of the MnPN and VLPO. PMID:26333784

  15. MEK inhibition prevents tumour-shed transforming growth factor-β-induced T-regulatory cell augmentation in tumour milieu

    PubMed Central

    Hossain, Dewan M S; Panda, Abir K; Chakrabarty, Sreeparna; Bhattacharjee, Pushpak; Kajal, Kirti; Mohanty, Suchismita; Sarkar, Irene; Sarkar, Diptendra K; Kar, Santosh K; Sa, Gaurisankar

    2015-01-01

    Tumour progression is associated with immune-suppressive conditions that facilitate the escape of tumour cells from the regimen of immune cells, subsequently paralysing the host defence mechanisms. Induction of CD4+ CD25+ FoxP3+ T regulatory (Treg) cells has been implicated in the tumour immune escape mechanism, although the novel anti-cancer treatment strategies targeting Treg cells remain unknown. The focus of this study is to define the interaction between tumour and immune system, i.e. how immune tolerance starts and gradually leads to the induction of adaptive Treg cells in the tumour microenvironment. Our study identified hyperactivated mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) -signalling as a potential target for reversing Treg cell augmentation in breast cancer patients. In more mechanistic detail, pharmacological inhibitors of MEK/ERK signalling inhibited transforming growth factor-β (TGF-β) production in tumour cells that essentially blocked TGF-β-SMAD3/SMAD4-mediated induction of CD25/interleukin-2 receptor α on CD4+ T-cell surface. As a result high-affinity binding of interleukin-2 on those cells was prohibited, causing lack of Janus kinase 1 (JAK1)/JAK3-mediated signal transducer and activator of transcription 3 (STAT3)/STAT5 activation required for FoxP3 expression. Finally, for a more radical approach towards a safe MEK inhibitor, we validate the potential of multi-kinase inhibitor curcumin, especially the nano-curcumin made out of pure curcumin with greater bioavailability; in repealing tumour-shed TGF-β-induced Treg cell augmentation. PMID:25284464

  16. Distinctive Expression of Bcl-2 Factors in Regulatory T Cells Determines a Pharmacological Target to Induce Immunological Tolerance

    PubMed Central

    Gabriel, Sarah Sharon; Bon, Nina; Chen, Jin; Wekerle, Thomas; Bushell, Andrew; Fehr, Thomas; Cippà, Pietro Ernesto

    2016-01-01

    Distinctive molecular characteristics of functionally diverse lymphocyte populations may represent novel pharmacological targets for immunotherapy. The intrinsic apoptosis pathway is differently regulated among conventional and regulatory T cells (Tregs). Targeted pharmacological modulation of this pathway with a small molecule Bcl-2/Bcl-xL inhibitor (ABT-737) caused a selective depletion of effector T cells and a relative enrichment of Tregs in vivo. Treatment with ABT-737 resulted in a tolerogenic milieu, which was exploited to alleviate graft-versus-host disease, to prevent allograft rejection in a stringent fully MHC-mismatched skin transplantation model and to induce immunological tolerance in combination with bone marrow transplantation. This concept has the potential to find various applications for immunotherapy, since it allows pharmacologic exploitation of the immunomodulatory properties of Tregs without the need for cell manipulation ex vivo. PMID:26973650

  17. Nuclear factor kappaB/p49 is a negative regulatory factor in nerve growth factor-induced choline acetyltransferase promoter activity in PC12 cells.

    PubMed

    Toliver-Kinsky, T; Wood, T; Perez-Polo, J R

    2000-12-01

    Anovel nuclear factor kappaB (NF-kappaB) binding site has been identified within the promoter region of the mouse gene encoding choline acetyltransferase (ChAT), the enzyme that synthesizes acetylcholine and has been implicated in the cognitive deficits associated with aging and Alzheimer's disease. This binding site, which is located within the nerve growth factor (NGF)-responsive enhancer element, was recognized by the NF-kappaB protein p49 but not p65 or p50. p49 from both basal forebrain and PC12 nuclear extracts interacted with this specific sequence in electrophoretic mobility shift assays. Mutation of the NF-kappaB site caused an increase in NGF-induced promoter activation, whereas overexpression of p49 in NGF-differentiated PC12 cells caused a decrease in endogenous ChAT enzyme activity and a decrease in promoter activity that was specifically mediated through this NF-kappaB binding site. Treatment of PC12 cells with NGF resulted in a drastic reduction in nuclear p49 binding to the ChAT NF-kappaB site after 24 h, but nuclear p49 levels were not altered, suggesting that late NGF-mediated events prevent binding of p49 to the ChAT promoter by an unknown mechanism other than nuclear translocation. Decreased ChAT expression and increased NF-kappaB activity in the brain are associated with aging and Alzheimer's disease. These data indicate that p49 is a negative regulator of ChAT expression and suggest a possible mechanism for aging-associated declines in cholinergic function.

  18. cIRF-3, a new member of the interferon regulatory factor (IRF) family that is rapidly and transiently induced by dsRNA.

    PubMed Central

    Grant, C E; Vasa, M Z; Deeley, R G

    1995-01-01

    In mammals, some of the effects of interferon (IFN) on gene transcription are known to be mediated by a family of IFN-inducible DNA-binding proteins, the IFN regulatory factor (IRF) family, which includes both activators and repressors of transcription. Although IFN activities have been described in many vertebrates, little is known about regulation of IFN- or IFN-stimulated genes in species other than human and mouse. Here, we report the cloning of a chicken cDNA, cIRF-3, encoding a protein with a DNA-binding domain similar to that found in the mammalian IRF family of proteins. Similarity between cIRF-3 and the mammalian IRFs is comparable with that between known members of the family. It is most similar to the IRF proteins ICSBP and ISGF3 gamma but is equally divergent from both. Gel mobility shift assays indicate that cIRF-3 is capable of binding a known IFN-stimulated response element that is conserved between the mammalian and chicken Mx genes. Expression of the cIRF-3 gene can be induced to high levels by poly(I).poly(C). Induction is rapid and transient with no requirement for protein synthesis. Co-treatment of cells with cycloheximide results in superinduction of cIRF-3 mRNA. The structural and regulatory characteristics of cIRF-3 indicate that it is the first example of a non-mammalian IRF protein. Images PMID:7541908

  19. Apoptosis Induced by Mammalian Reovirus Is Beta Interferon (IFN) Independent and Enhanced by IFN Regulatory Factor 3- and NF-κB-Dependent Expression of Noxa

    PubMed Central

    Knowlton, Jonathan J.; Dermody, Terence S.

    2012-01-01

    A variety of signal transduction pathways are activated in response to viral infection, which dampen viral replication and transmission. These mechanisms involve both the induction of type I interferons (IFNs), which evoke an antiviral state, and the triggering of apoptosis. Mammalian orthoreoviruses are double-stranded RNA viruses that elicit apoptosis in vitro and in vivo. The transcription factors interferon regulatory factor 3 (IRF-3) and nuclear factor kappa light-chain enhancer of activated B cells (NF-κB) are required for the expression of IFN-β and the efficient induction of apoptosis in reovirus-infected cells. However, it is not known whether IFN-β induction is required for apoptosis, nor have the genes induced by IRF-3 and NF-κB that are responsible for apoptosis been identified. To determine whether IFN-β is required for reovirus-induced apoptosis, we used type I IFN receptor-deficient cells, IFN-specific antibodies, and recombinant IFN-β. We found that IFN synthesis and signaling are dispensable for the apoptosis of reovirus-infected cells. These results indicate that the apoptotic response following reovirus infection is mediated directly by genes responsive to IRF-3 and NF-κB. Noxa is a proapoptotic BH3-domain-only protein of the Bcl-2 family that requires IRF-3 and NF-κB for efficient expression. We found that Noxa is strongly induced at late times (36 to 48 h) following reovirus infection in a manner dependent on IRF-3 and NF-κB. The level of apoptosis induced by reovirus is significantly diminished in cells lacking Noxa, indicating a key prodeath function for this molecule during reovirus infection. These results suggest that prolonged innate immune response signaling induces apoptosis by eliciting Noxa expression in reovirus-infected cells. PMID:22090144

  20. Transforming growth factor-β2 increases the capacity of retinal pigment epithelial cells to induce the generation of regulatory T cells.

    PubMed

    Yan, Feng; He, Jin; Tang, Li; Kong, Yi; Shi, Yuhua; Chen, Suihua; Huang, Zhenping

    2016-02-01

    The present study investigated the underlying mechanism of the induction of regulatory T cells (Tregs) by retinal pigment epithelial (RPE) cells and the characteristics of these Tregs. Human RPE cells were cultured in the presence or absence of transforming growth factor-β 2 (TGF-β2), and reverse-transcription quantitative PCR was performed to determine the mRNA expression of indoleamine 2,3-dioxygenase (IDO) and nuclear factor erythroid 2-related factor (Nrf2). Supernatants of RPE cell cultures were added to CD4+ T cells to induce Tregs. The RPE-induced Tregs were purified by two-step magnetic cell sorting. The natural Tregs were isolated from the peripheral blood mononuclear cells of healthy volunteers. Purified CD4+ CD25- T cells (2 x 10(5)/well) were cultured alone or with Tregs (various densities, natural or RPE-induced). The proliferation of CD4+ CD25- T cells was determined by 3H-thymidine incorporation. After 24 h of stimulation with TGF-β2, the mRNA expression of IDO in RPE cells was upregulated. The highest level of IDO mRNA expression was reached after 72 h of stimulation with TGF-β2. However, the Nrf2 mRNA expression was slightly decreased after 24 h of stimulation with TGF-β2 and significantly increased after 48-72 h of TGF-β2 stimulation. Increased levels of CD25 expression were observed on CD4+ T cells exposed to supernatants of RPE cell cultures treated with TGF-β2 and recombinant interleukin-2. The RPE-induced Tregs were more effective at suppressing the proliferation of CD4+ CD25- T cells compared with native Tregs. These findings suggested that IDO may be a signaling protein in RPE cells which is implicated in the induction of Tregs. RPE-induced Tregs have the potential to be applied for immunotherapy for ocular inflammatory diseases.

  1. Foot-and-mouth disease virus leader proteinase inhibits dsRNA-induced type I interferon transcription by decreasing interferon regulatory factor 3/7 in protein levels

    SciTech Connect

    Wang, Dang; Fang, Liurong; Luo, Rui; Ye, Rui; Fang, Ying; Xie, Lilan; Chen, Huanchun; Xiao, Shaobo

    2010-08-13

    Research highlights: {yields} FMDV L{sup pro} inhibits poly(I:C)-induced IFN-{alpha}1/{beta} mRNA expression. {yields} L{sup pro} inhibits MDA5-mediated activation of the IFN-{alpha}1/{beta} promoter. {yields} L{sup pro} significantly reduced the transcription of multiple IRF-responsive genes. {yields} L{sup pro} inhibits IFN-{alpha}1/{beta} promoter activation by decreasing IRF-3/7 in protein levels. {yields} The ability to process eIF-4G of L{sup pro} is not necessary to inhibit IFN-{alpha}1/{beta} activation. -- Abstract: The leader proteinase (L{sup pro}) of foot-and-mouth disease virus (FMDV) has been identified as an interferon-{beta} (IFN-{beta}) antagonist that disrupts the integrity of transcription factor nuclear factor {kappa}B (NF-{kappa}B). In this study, we showed that the reduction of double stranded RNA (dsRNA)-induced IFN-{alpha}1/{beta} expression caused by L{sup pro} was also associated with a decrease of interferon regulatory factor 3/7 (IRF-3/7) in protein levels, two critical transcription factors for activation of IFN-{alpha}/{beta}. Furthermore, overexpression of L{sup pro} significantly reduced the transcription of multiple IRF-responsive genes including 2',5'-OAS, ISG54, IP-10, and RANTES. Screening L{sup pro} mutants indicated that the ability to process eIF-4G of L{sup pro} is not required for suppressing dsRNA-induced activation of the IFN-{alpha}1/{beta} promoter and decreasing IRF-3/7 expression. Taken together, our results demonstrate that, in addition to disrupting NF-{kappa}B, L{sup pro} also decreases IRF-3/7 expression to suppress dsRNA-induced type I IFN production, suggesting multiple strategies used by FMDV to counteract the immune response to viral infection.

  2. Smad7 protein induces interferon regulatory factor 1-dependent transcriptional activation of caspase 8 to restore tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis.

    PubMed

    Hong, Suntaek; Kim, Hye-Youn; Kim, Jooyoung; Ha, Huyen Trang; Kim, Young-Mi; Bae, Eunjin; Kim, Tae Hyung; Lee, Kang Choon; Kim, Seong-Jin

    2013-02-01

    Smad7 has been known as a negative regulator for the transforming growth factor-β (TGF-β) signaling pathway through feedback regulation. However, Smad7 has been suspected to have other biological roles through the regulation of gene transcription. By screening differentially regulated genes, we found that the caspase 8 gene was highly up-regulated in Smad7-expressing cells. Smad7 was able to activate the caspase 8 promoter through recruitment of the interferon regulatory factor 1 (IRF1) transcription factor to the interferon-stimulated response element (ISRE) site. Interaction of Smad7 on the caspase 8 promoter was confirmed with electrophoretic mobility shift assay and chromatin immunoprecipitation experiment. Interestingly, Smad7 did not directly interact with the ISRE site, but it increased the binding activity of IRF1 with ISRE. These results support that Smad7 recruits IRF1 protein on the caspase 8 promoter and functions as a transcriptional coactivator. To confirm the biological significance of caspase 8 up-regulation, we tested tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL)-mediated cell death assay in breast cancer cells. Smad7 in apoptosis-resistant MCF7 cells markedly sensitized the cells to TRAIL-induced cell death by restoring the caspase cascade. Furthermore, restoration of caspase 8-mediated apoptosis pathway repressed the tumor growth in the xenograft model. In conclusion, we suggest a novel role for Smad7 as a transcriptional coactivator for caspase 8 through the interaction with IRF1 in regulation of the cell death pathway. PMID:23255602

  3. Lactobacilli Modulate Hypoxia-Inducible Factor (HIF)-1 Regulatory Pathway in Triple Negative Breast Cancer Cell Line

    PubMed Central

    Abedin-Do, Atieh; Mirfakhraie, Reza; Shirzad, Mahdieh; Ghafouri-Fard, Soudeh; Motevaseli, Elahe

    2016-01-01

    Objective Hypoxia-Inducible Factor (HIF)-1 plays an essential role in the body’s response to low oxygen concentrations and regulates expression of several genes implicated in homeostasis, vascularization, anaerobic metabolism as well as immunological responses. Increased levels of HIF-1α are associated with increased proliferation and more aggressive breast tumor development. Lactobacilli have been shown to exert anti-cancer effects on several malignancies including breast cancer. However, the exact mechanism of such effect is not clear yet. The aim of this study was to analyze the expression of selected genes from HIF pathway in a triple negative breast cancer cell line (expressing no estrogen and progesterone receptors as well as HER-2/Neu), MDA-MB-231, following treatment with two lactobacilli culture supernatants. Materials and Methods In this experimental study, we analyzed the expression of HIF-1α, SLC2A1, VHL, HSP90, XBP1 and SHARP1 genes from HIF pathway in MDA-MB-231 cells, before and after treatment with Lactobacillus crispatus and Lactobacillus rhamnosus culture supernatants (LCS and LRS, respectively) by means of quantitative reverse-transcription polymerase chain reaction (qRT-PCR). Results Both LRS and LCS had cytotoxic effects on MDA-MB-231 cells, while the former type was more cytotoxic. LRS dramatically down-regulated expression levels of the HIF-1α, HSP90 and SLC2A1 in the MDA-MB-231 cells. LCS had similar effect on the expression of HSP90, to what was observed in the LRS treatment. The expression level of tumor suppressor genes VHL and SHARP1 were also decreased in LCS treated cells. Conclusion Although both LCS and LRS had cytotoxic effects on the MDA-MB-231 cells, it is proposed that LRS could be more appropriate for pathway directed treatment modalities, as it did not decrease expression of tumor suppressor genes involved in HIF pathway. Down-regulation of HIF pathway mediated oncogenes by LRS suggests that the cytotoxic effects of this

  4. PDGF upregulates CLEC-2 to induce T regulatory cells

    PubMed Central

    Agrawal, Sudhanshu; Ganguly, Sreerupa; Hajian, Pega; Cao, Jia-Ning; Agrawal, Anshu

    2015-01-01

    The effect of platelet derived growth factor (PDGF) on immune cells is not elucidated. Here, we demonstrate PDGF inhibited the maturation of human DCs and induced IL-10 secretion. Culture of PDGF-DCs with T cells induced the polarization of T cells towards FoxP3 expressing T regulatory cells that secreted IL-10. Gene expression studies revealed that PDGF induced the expression of C-type lectin like receptor member 2, (CLEC-2) receptor on DCs. Furthermore, DCs transfected with CLEC-2 induced T regulatory cells in DC-T cell co-culture. CLEC-2 is naturally expressed on platelets. Therefore, to confirm whether CLEC-2 is responsible for inducing the T regulatory cells, T cells were cultured with either CLEC-2 expressing platelets or soluble CLEC-2. Both conditions resulted in the induction of regulatory T cells. The generation of T regulatory cells was probably due to the binding of CLEC-2 with its ligand podoplanin on T cells, since crosslinking of podoplanin on the T cells also resulted in the induction of T regulatory cells. These data demonstrate that PDGF upregulates the expression of CLEC-2 on cells to induce T regulatory cells. PMID:26416420

  5. PDGF upregulates CLEC-2 to induce T regulatory cells.

    PubMed

    Agrawal, Sudhanshu; Ganguly, Sreerupa; Hajian, Pega; Cao, Jia-Ning; Agrawal, Anshu

    2015-10-01

    The effect of platelet derived growth factor (PDGF) on immune cells is not elucidated. Here, we demonstrate PDGF inhibited the maturation of human DCs and induced IL-10 secretion. Culture of PDGF-DCs with T cells induced the polarization of T cells towards FoxP3 expressing T regulatory cells that secreted IL-10. Gene expression studies revealed that PDGF induced the expression of C-type lectin like receptor member 2, (CLEC-2) receptor on DCs. Furthermore, DCs transfected with CLEC-2 induced T regulatory cells in DC-T cell co-culture. CLEC-2 is naturally expressed on platelets. Therefore, to confirm whether CLEC-2 is responsible for inducing the T regulatory cells, T cells were cultured with either CLEC-2 expressing platelets or soluble CLEC-2. Both conditions resulted in the induction of regulatory T cells. The generation of T regulatory cells was probably due to the binding of CLEC-2 with its ligand podoplanin on T cells, since crosslinking of podoplanin on the T cells also resulted in the induction of T regulatory cells. These data demonstrate that PDGF upregulates the expression of CLEC-2 on cells to induce T regulatory cells.

  6. Intra-articular nuclear factor-κB blockade ameliorates collagen-induced arthritis in mice by eliciting regulatory T cells and macrophages

    PubMed Central

    Min, S-Y; Yan, M; Du, Y; Wu, T; Khobahy, E; Kwon, S-R; Taneja, V; Bashmakov, A; Nukala, S; Ye, Y; Orme, J; Sajitharan, D; Kim, H-Y; Mohan, C

    2013-01-01

    Nuclear factor (NF)-κB is a transcription factor implicated in the pathogenesis of autoimmune disorders such as rheumatoid arthritis (RA). Here we have examined the effect of intra-articular administration of the IKK inhibitor, NEMO-binding domain peptide (NBD), on the severity of collagen-induced arthritis (CIA). NBD peptides were injected intra-articularly into the knee joints of DBA/1J mice after the onset of disease. Collagen-injected mice given a scrambled peptide served as controls. Arthritis severity was determined by visual examination of paws. Intra-articular NBD injection reduced the arthritis score and ameliorated morphological signs of bone destruction compared to the controls. Serum levels of type-II collagen-specific immunoglobulin (Ig)G2a antibodies were lower in NBD-treated mice versus the control mice, whereas the levels of type-II collagen-specific IgG1 antibodies were increased by NBD treatment. NBD treatment diminished the proinflammatory cytokines interleukin (IL)-17 and interferon (IFN)-γ in serum, but increased the regulatory cytokine IL-10. NBD-treated CIA mice exhibited significantly higher percentages and numbers of forkhead box protein 3 (FoxP3+)CD4+CD25+ regulatory T cells than controls. Immunofluorescence analysis of NBD-treated mice revealed that FoxP3 and Ym1, a marker of alternatively activated macrophages, were juxtaposed to each other within draining inguinal lymph nodes. Intra-articular administration of NBD peptide is effective as an experimental therapy in a murine model of RA. Nevertheless, the intra-articular treatment modality is still associated with systemic effects on the immune system. PMID:23574318

  7. Nerve growth factor-induced derepression of peripherin gene expression is associated with alterations in proteins binding to a negative regulatory element.

    PubMed Central

    Thompson, M A; Lee, E; Lawe, D; Gizang-Ginsberg, E; Ziff, E B

    1992-01-01

    The peripherin gene, which encodes a neuronal-specific intermediate filament protein, is transcriptionally induced with a late time course when nerve growth factor (NGF) stimulates PC12 cells to differentiate into neurons. We have studied its transcriptional regulation in order to better understand the neuronal-specific end steps of the signal transduction pathway of NGF. By 5' deletion mapping of the peripherin promoter, we have localized two positive regulatory elements necessary for full induction by NGF: a distal positive element and a proximal constitutive element within 111 bp of the transcriptional start site. In addition, there is a negative regulatory element (NRE; -179 to -111), the deletion of which results in elevated basal expression of the gene. Methylation interference footprinting of the NRE defined a unique sequence, GGCAGGGCGCC, as the binding site for proteins present in nuclear extracts from both undifferentiated and differentiated PC12 cells. However, DNA mobility shift assays using an oligonucleotide probe containing the footprinted sequence demonstrate a prominent retarded complex in extracts from undifferentiated PC12 cells which migrates with slower mobility than do the complexes produced by using differentiated PC12 cell extract. Transfection experiments using peripherin-chloramphenicol acetyltransferase constructs in which the footprinted sequence has been mutated confirm that the NRE has a functional, though not exclusive, role in repressing peripherin expression in undifferentiated and nonneuronal cells. We propose a two-step model of activation of peripherin by NGF in which dissociation of a repressor from the protein complex at the NRE, coupled with a positive signal from the distal positive element, results in depression of the gene. Images PMID:1588954

  8. Interferon regulatory factor 4b (IRF4b) in Japanese flounder, Paralichthys olivaceus: Sequencing, ubiquitous tissue distribution and inducible expression by poly(I:C) and DNA virus.

    PubMed

    Liu, Dahai; Chen, Jinjing; Zhang, Haiyan; Hu, Mengzhu; Lou, Huimin; Liu, Qiuming; Zhang, Shicui; Hu, Guobin

    2016-09-01

    Interferon regulatory factor 4 (IRF4) in mammals is known to be critical in regulation of development and functions of lymphomyeloid cell lineages. Recent studies have demonstrated its involvement in immune responses to bacterial and viral challenges in teleosts. In this study, an IRF4 gene was cloned from Japanese flounder (Paralichthys olivaceus) and its expression in response to polyinosinic:polycytidylic acid [poly(I:C)] and lymphocystis disease virus (LCDV) stimulations was studied in vivo. The cloned gene spans over 5.9 kb, comprises eight exons and seven introns and encodes a putative protein of 456 amino acids. The deduced amino acid sequence possesses a conserved DNA-binding domain (DBD), an IRF-association domain (IAD) and a nuclear localization signal (NLS). Phylogenetic analysis clustered it into the teleost IRF4b clade and, thus, it was named Paralichthys olivaceus (Po)IRF4b. The constitutive expression of PoIRF4b transcripts was detectable in all examined organs, with highest levels found in lymphomyeloid-rich tissues. They were induced by both poly(I:C) and LCDV with a similar inducibility in immune or non-immune organs. Two waves of induced expression of PoIRF4b were observed with the two stimuli during a 7-day time course in the immune organs, with the early-phase induction being stronger. The maximum increases of PoIRF4b transcript levels ranged from 1.3 to 4.0-fold and appeared at day 1-5 post-injection depending on different organs and stimuli. In both stimulation cases, the strongest induction was detected in spleen and the weakest in muscle. These results indicate that PoIRF4b may participate in regulation of immune responses of flounders to both RNA and DNA virus infections. PMID:27084058

  9. Multiple myeloma oncogene 1 (MUM1)/interferon regulatory factor 4 (IRF4) upregulates monokine induced by interferon-gamma (MIG) gene expression in B-cell malignancy.

    PubMed

    Uranishi, M; Iida, S; Sanda, T; Ishida, T; Tajima, E; Ito, M; Komatsu, H; Inagaki, H; Ueda, R

    2005-08-01

    MUM1 (multiple myeloma oncogene 1)/IRF4 (interferon regulatory factor 4) is a transcription factor that is activated as a result of t(6;14)(p25;q32) in multiple myeloma. MUM1 expression is seen in various B-cell lymphomas and predicts an unfavorable outcome in some lymphoma subtypes. To elucidate its role in B-cell malignancies, we prepared MUM1-expressing Ba/F3 cells, which proliferated until higher cellular density than the parental cells, and performed cDNA microarray analysis to identify genes whose expression is regulated by MUM1. We found that the expression of four genes including FK506-binding protein 3 (FKBP3), the monokine induced by interferon-gamma(MIG), Fas apoptotic inhibitory molecule (Faim) and Zinc-finger protein 94 was altered in the MUM1-expressing cells. We then focused on MIG since its expression was immediately upregulated by MUM1. In reporter assays, MUM1 activated the MIG promoter in cooperation with PU.1, and the interaction between MUM1 and the MIG promoter sequence was confirmed. The expression of MIG was correlated with that of MUM1 in B-CLL cell lines, and treatment with neutralizing antibodies against MIG and its receptor, CXCR3, slightly inhibited the proliferation of two MUM1-expressing lines. These results suggest that MUM1 plays roles in the progression of B-cell lymphoma/leukemia by regulating the expression of various genes including MIG. Leukemia (2005) 19, 1471-1478. doi:10.1038/sj.leu.2403833; published online 16 June 2005.

  10. The regulatory effect of SC-236 (4-[5-(4-chlorophenyl)-3-(trifluoromethyl)-1-pyrazol-1-l] benzenesulfonamide) on stem cell factor induced migration of mast cells

    SciTech Connect

    Kim, Su-Jin; Jeong, Hyun-Ja; Park, Rae-Kil; Lee, Kang-Min; Kim, Hyung-Min; Um, Jae-Young; Hong, Seung-Heon . E-mail: jooklim@wonkwang.ac.kr

    2007-04-15

    SC-236 (4-[5-(4-chlorophenyl)-3-(trifluoromethyl)-1-pyrazol-1-]benzenesulfonamide; C{sub 16}H{sub 11}ClF{sub 3}N{sub 3}O{sub 2}S), is a highly selective cyclooxygenase (COX)-2 inhibitor. Recently, there have been reports that SC-236 protects against cartilage damage in addition to reducing inflammation and pain in osteoarthritis. However, the mechanism involved in the inflammatory allergic reaction has not been examined. Mast cells accumulation can be related to inflammatory conditions, including allergic rhinitis, asthma, and rheumatoid arthritis. The aim of the present study is to investigate the effects of SC-236 on stem cell factor (SCF)-induced migration, morphological alteration, and cytokine production of rat peritoneal mast cells (RPMCs). We observed that SCF significantly induced the migration and morphological alteration. The ability of SCF to enhance migration and morphological alteration was abolished by treatment with SC-236. In addition, production of tumor necrosis factor (TNF)-{alpha}, interleukin (IL)-1{beta}, and vascular endothelial growth factor (VEGF) production induced by SCF was significantly inhibited by treatment with SC-236. Previous work has demonstrated that SCF-induced migration and cytokine production of mast cells require p38 MAPK activation. We also showed that SC-236 suppresses the SCF-induced p38 MAPK activation in RPMCs. These data suggest that SC-236 inhibits migration and cytokine production through suppression of p38 MAPK activation. These results provided new insight into the pharmacological actions of SC-236 and its potential therapeutic role in the treatment of inflammatory allergic diseases.

  11. Regulatory effect of cytokine-induced neutrophil chemoattractant, epithelial neutrophil-activating peptide 78 and pyrrolidine dithiocarbamate on pulmonary neutrophil aggregation mediated by nuclear factor-κB in lipopolysaccharide-induced acute respiratory distress syndrome mice

    PubMed Central

    Wang, Hongman; Zhao, Jiping; Xue, Guansheng; Wang, Junfei; Wu, Jinxiang; Wang, Donghui; Dong, Liang

    2016-01-01

    In the present study, the regulatory effect of cytokine-induced neutrophil chemoattractant (CINC) and epithelial neutrophil-activating peptide 78 (ENA-78) on pulmonary neutrophil (PMN) accumulation in lipopolysaccharide (LPS)-induced acute respiratory distress syndrome (ARDS) mice, and the therapeutic effect of pyrrolidine dithiocarbamate (PDTC), was investigated. BALB/c mice were divided into control, LPS and PDTC + LPS groups using a random number table. The phosphorylation of nuclear factor-κB (NF-κB) was detected using a western blot, and the mRNA expression levels of CINC were evaluated using reverse transcription-quantitative polymerase chain reaction. The expression of NF-κB, CINC and ENA-78 was detected using immunohistochemistry. The production of interleukin (IL)-8 and IL-10 in serum and broncho-alveolar lavage fluid (BALF) was analyzed using an enzyme-linked immunosorbent assay. The total number of leukocytes and proportion of PMNs in BALF was also determined. Following injection with LPS (20 mg/kg), the expression levels of p-NF-κB, CINC and ENA-78 were increased in lung tissue, and the expression levels of IL-8, IL-10 and the number of PMNs increased in serum and BALF. However, in comparison with the LPS group, the degree of lung injury was reduced in ARDS mice that were treated with PDTC. In addition, the expression level of p-NF-κB and the production of chemokines in lung tissue decreased in ARDS mice that were treated with PDTC, and the number of PMNs in BALF also decreased. In conclusion, the results of the present study suggest that the LPS-induced phosphorylation of NF-κB may result in the synthesis and release of CINC and ENA-78, which induce the accumulation of PMNs in the lung. Therefore, PDTC may be used to reduce the production of chemokines and cytokines, thereby decreasing the activation of PMNs in lung tissue and reducing the damage of lung tissue in ARDS. PMID:27602092

  12. Regulatory effect of cytokine-induced neutrophil chemoattractant, epithelial neutrophil-activating peptide 78 and pyrrolidine dithiocarbamate on pulmonary neutrophil aggregation mediated by nuclear factor-κB in lipopolysaccharide-induced acute respiratory distress syndrome mice

    PubMed Central

    Wang, Hongman; Zhao, Jiping; Xue, Guansheng; Wang, Junfei; Wu, Jinxiang; Wang, Donghui; Dong, Liang

    2016-01-01

    In the present study, the regulatory effect of cytokine-induced neutrophil chemoattractant (CINC) and epithelial neutrophil-activating peptide 78 (ENA-78) on pulmonary neutrophil (PMN) accumulation in lipopolysaccharide (LPS)-induced acute respiratory distress syndrome (ARDS) mice, and the therapeutic effect of pyrrolidine dithiocarbamate (PDTC), was investigated. BALB/c mice were divided into control, LPS and PDTC + LPS groups using a random number table. The phosphorylation of nuclear factor-κB (NF-κB) was detected using a western blot, and the mRNA expression levels of CINC were evaluated using reverse transcription-quantitative polymerase chain reaction. The expression of NF-κB, CINC and ENA-78 was detected using immunohistochemistry. The production of interleukin (IL)-8 and IL-10 in serum and broncho-alveolar lavage fluid (BALF) was analyzed using an enzyme-linked immunosorbent assay. The total number of leukocytes and proportion of PMNs in BALF was also determined. Following injection with LPS (20 mg/kg), the expression levels of p-NF-κB, CINC and ENA-78 were increased in lung tissue, and the expression levels of IL-8, IL-10 and the number of PMNs increased in serum and BALF. However, in comparison with the LPS group, the degree of lung injury was reduced in ARDS mice that were treated with PDTC. In addition, the expression level of p-NF-κB and the production of chemokines in lung tissue decreased in ARDS mice that were treated with PDTC, and the number of PMNs in BALF also decreased. In conclusion, the results of the present study suggest that the LPS-induced phosphorylation of NF-κB may result in the synthesis and release of CINC and ENA-78, which induce the accumulation of PMNs in the lung. Therefore, PDTC may be used to reduce the production of chemokines and cytokines, thereby decreasing the activation of PMNs in lung tissue and reducing the damage of lung tissue in ARDS.

  13. Regulatory effect of hypoxia-inducible factor-1α on hCG-stimulated endothelin-2 expression in granulosa cells from the PMSG-treated rat ovary.

    PubMed

    Zhang, Jisen; Zhang, Zhenghong; Wu, Yanqing; Chen, Liyun; Luo, Qianping; Chen, Jiajie; Huang, Xiaohong; Cheng, Yong; Wang, Zhengchao

    2012-01-01

    Endothelin (ET)-2 plays a crucial role in ovarian ovulation in mammals. The present study was designed to test the hypothesis that hypoxia-inducible factor (HIF)-1α-mediated transcriptional activation contributes to the increased expression of ET-2 gene in response to hCG in rat ovarian granulosa cells (GCs) during gonadotropin-induced superovulation. By real-time RT-PCR analysis, ET-2 mRNA expression was found to significantly increase in cultured ovarian GCs after treatment with hCG, or even N-carbobenzoxyl-L-leucinyl-L-leucinyl-L-norvalinal (MG-132), while this increased ET-2 mRNA expression could also be blocked by ferrous ammonium sulfate (FAS) under human chorionic gonadotropin (hCG) treatment. Further analysis also found that these changes of ET-2 mRNA were consistent with HIF-1α expression or HIF-1 activity, and HIF-1α inhibitor echinomycin inhibited ovulation in rats. Taken together, these results indicate that ET-2 is transcriptionally activated by hCG through HIF-1α-mediated mechanism in GCs. This HIF-1α-induced transcriptional activation may be one of the important mechanisms mediating the increase of ET-2 expression in GCs during the gonadotropin-induced mammalian ovulatory process in vivo.

  14. Interferon regulatory factor-1 binds c-Cbl, enhances mitogen activated protein kinase signaling and promotes retinoic acid-induced differentiation of HL-60 human myelo-monoblastic leukemia cells.

    PubMed

    Shen, Miaoqing; Bunaciu, Rodica P; Congleton, Johanna; Jensen, Holly A; Sayam, Lavanya G; Varner, Jeffrey D; Yen, Andrew

    2011-12-01

    All-trans retinoic acid (RA) and interferons (IFNs) have efficacy in treating certain leukemias and lymphomas, respectively, motivating interest in their mechanism of action to improve therapy. Both RA and IFNs induce interferon regulatory factor-1 (IRF-1). We find that in HL-60 myeloblastic leukemia cells which undergo mitogen activated protien kinase (MAPK)-dependent myeloid differentiation in response to RA, IRF-1 propels differentiation. RA induces MAPK-dependent expression of IRF-1. IRF-1 binds c-Cbl, a MAPK related adaptor. Ectopic IRF-1 expression causes CD38 expression and activation of the Raf/MEK/ERK axis, and enhances RA-induced differentiation by augmenting CD38, CD11b, respiratory burst and G0 arrest. Ectopic IRF-1 expression also decreases the activity of aldehyde dehydrogenase 1, a stem cell marker, and enhances RA-induced ALDH1 down-regulation. Interestingly, expression of aryl hydrocarbon receptor (AhR), which is RA-induced and known to down-regulate Oct4 and drive RA-induced differentiation, also enhances IRF-1 expression. The data are consistent with a model whereby IRF-1 acts downstream of RA and AhR to enhance Raf/MEK/ERK activation and propel differentiation.

  15. The Pseudomonas aeruginosa PAO1 Two-Component Regulator CarSR Regulates Calcium Homeostasis and Calcium-Induced Virulence Factor Production through Its Regulatory Targets CarO and CarP

    PubMed Central

    Guragain, Manita; King, Michelle M.; Williamson, Kerry S.; Pérez-Osorio, Ailyn C.; Akiyama, Tatsuya; Khanam, Sharmily

    2016-01-01

    ABSTRACT Pseudomonas aeruginosa is an opportunistic human pathogen that causes severe, life-threatening infections in patients with cystic fibrosis (CF), endocarditis, wounds, or artificial implants. During CF pulmonary infections, P. aeruginosa often encounters environments where the levels of calcium (Ca2+) are elevated. Previously, we showed that P. aeruginosa responds to externally added Ca2+ through enhanced biofilm formation, increased production of several secreted virulence factors, and by developing a transient increase in the intracellular Ca2+ level, followed by its removal to the basal submicromolar level. However, the molecular mechanisms responsible for regulating Ca2+-induced virulence factor production and Ca2+ homeostasis are not known. Here, we characterized the genome-wide transcriptional response of P. aeruginosa to elevated [Ca2+] in both planktonic cultures and biofilms. Among the genes induced by CaCl2 in strain PAO1 was an operon containing the two-component regulator PA2656-PA2657 (here called carS and carR), while the closely related two-component regulators phoPQ and pmrAB were repressed by CaCl2 addition. To identify the regulatory targets of CarSR, we constructed a deletion mutant of carR and performed transcriptome analysis of the mutant strain at low and high [Ca2+]. Among the genes regulated by CarSR in response to CaCl2 are the predicted periplasmic OB-fold protein, PA0320 (here called carO), and the inner membrane-anchored five-bladed β-propeller protein, PA0327 (here called carP). Mutations in both carO and carP affected Ca2+ homeostasis, reducing the ability of P. aeruginosa to export excess Ca2+. In addition, a mutation in carP had a pleotropic effect in a Ca2+-dependent manner, altering swarming motility, pyocyanin production, and tobramycin sensitivity. Overall, the results indicate that the two-component system CarSR is responsible for sensing high levels of external Ca2+ and responding through its regulatory targets that

  16. The Protein Phosphatase 2A Regulatory Subunit B56γ Mediates Suppression of T Cell Receptor (TCR)-induced Nuclear Factor-κB (NF-κB) Activity*

    PubMed Central

    Breuer, Rebecca; Becker, Michael S.; Brechmann, Markus; Mock, Thomas; Arnold, Rüdiger; Krammer, Peter H.

    2014-01-01

    NF-κB is an important transcription factor in the immune system, and aberrant NF-κB activity contributes to malignant diseases and autoimmunity. In T cells, NF-κB is activated upon TCR stimulation, and signal transduction to NF-κB activation is triggered by a cascade of phosphorylation events. However, fine-tuning and termination of TCR signaling are only partially understood. Phosphatases oppose the role of kinases by removing phosphate moieties. The catalytic activity of the protein phosphatase PP2A has been implicated in the regulation of NF-κB. PP2A acts in trimeric complexes in which the catalytic subunit is promiscuous and the regulatory subunit confers substrate specificity. To understand and eventually target NF-κB-specific PP2A functions it is essential to define the regulatory PP2A subunit involved. So far, the regulatory PP2A subunit that mediates NF-κB suppression in T cells remained undefined. By performing a siRNA screen in Jurkat T cells harboring a NF-κB-responsive luciferase reporter, we identified the PP2A regulatory subunit B56γ as negative regulator of NF-κB in TCR signaling. B56γ was strongly up-regulated upon primary human T cell activation, and B56γ silencing induced increased IκB kinase (IKK) and IκBα phosphorylation upon TCR stimulation. B56γ silencing enhanced NF-κB activity, resulting in increased NF-κB target gene expression including the T cell cytokine IL-2. In addition, T cell proliferation was increased upon B56γ silencing. These data help to understand the physiology of PP2A function in T cells and the pathophysiology of diseases involving PP2A and NF-κB. PMID:24719332

  17. Growth factor-induced activation of a kinase activity which causes regulatory phosphorylation of p42/microtubule-associated protein kinase.

    PubMed Central

    L'Allemain, G; Her, J H; Wu, J; Sturgill, T W; Weber, M J

    1992-01-01

    p42/microtubule-associated protein kinase (p42mapk) is activated by tyrosine and threonine phosphorylation, and its regulatory phosphorylation is likely to be important in signalling pathways involved in growth control, secretion, and differentiation. Here we show that treatment of quiescent 3T3 cells with diverse agonists results in the appearance of an activity capable of causing the in vitro phosphorylation of p42mapk on the regulatory tyrosine and to a lesser extent on the regulatory threonine, resulting in enzymatic activation of the p42mapk. This p42mapk-activating activity is capable of phosphorylating a kinase-defective p42mapk mutant, thus confirming its activity as a kinase. Images PMID:1314951

  18. Ginsenoside F2 reduces hair loss by controlling apoptosis through the sterol regulatory element-binding protein cleavage activating protein and transforming growth factor-β pathways in a dihydrotestosterone-induced mouse model.

    PubMed

    Shin, Heon-Sub; Park, Sang-Yong; Hwang, Eun-Son; Lee, Don-Gil; Mavlonov, Gafurjon Turdalievich; Yi, Tae-Hoo

    2014-01-01

    This study was conducted to test whether ginsenoside F2 can reduce hair loss by influencing sterol regulatory element-binding protein (SREBP) cleavage-activating protein (SCAP) and the transforming growth factor beta (TGF-β) pathway of apoptosis in dihydrotestosterone (DHT)-treated hair cells and in a DHT-induced hair loss model in mice. Results for ginsenoside F2 were compared with finasteride. DHT inhibits proliferation of hair cells and induces androgenetic alopecia and was shown to activate an apoptosis signal pathway both in vitro and in vivo. The cell-based 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that the proliferation rates of DHT-treated human hair dermal papilla cells (HHDPCs) and HaCaTs increased by 48% in the ginsenoside F2-treated group and by 12% in the finasteride-treated group. Western blot analysis showed that ginsenoside F2 decreased expression of TGF-β2 related factors involved in hair loss. The present study suggested a hair loss related pathway by changing SCAP related apoptosis pathway, which has been known to control cholesterol metabolism. SCAP, sterol regulatory element-binding protein (SREBP) and caspase-12 expression in the ginsenoside F2-treated group were decreased compared to the DHT and finasteride-treated group. C57BL/6 mice were also prepared by injection with DHT and then treated with ginsenoside F2 or finasteride. Hair growth rate, density, thickness measurements and tissue histotological analysis in these groups suggested that ginsenoside F2 suppressed hair cell apoptosis and premature entry to catagen more effectively than finasteride. Our results indicated that ginsenoside F2 decreased the expression of TGF-β2 and SCAP proteins, which have been suggested to be involved in apoptosis and entry into catagen. This study provides evidence those factors in the SCAP pathway could be targets for hair loss prevention drugs.

  19. Regulatory role of NADPH oxidase in glycated LDL-induced upregulation of plasminogen activator inhibitor-1 and heat shock factor-1 in mouse embryo fibroblasts and diabetic mice.

    PubMed

    Zhao, Ruozhi; Le, Khuong; Moghadasian, Mohammed H; Shen, Garry X

    2013-08-01

    Cardiovascular disease is the predominant cause of death in diabetic patients. Fibroblasts are one of the major types of cells in the heart or vascular wall. Increased levels of glycated low-density lipoprotein (glyLDL) were detected in diabetic patients. Previous studies in our group demonstrated that oxidized LDL increased the amounts of NADPH oxidase (NOX), plasminogen activator inhibitor-1 (PAI-1), and heat shock factor-1 (HSF1) in fibroblasts. This study examined the expression of NOX, PAI-1, and HSF1 in glyLDL-treated wild-type or HSF1-deficient mouse embryo fibroblasts (MEFs) and in leptin receptor-knockout (db/db) diabetic mice. Treatment with physiologically relevant levels of glyLDL increased superoxide and H2O2 release and the levels of NOX4 and p22phox (an essential component of multiple NOX complexes) in wild-type or HSF1-deficient MEFs. The levels of HSF1 and PAI-1 were increased by glyLDL in wild-type MEFs, but not in HSF1-deficient MEFs. Diphenyleneiodonium (a nonspecific NOX inhibitor) or small interfering RNA for p22phox prevented glyLDL-induced increases in the levels of NOX4, HSF1, or PAI-1 in MEFs. The amounts of NOX4, HSF1, and PAI-1 were elevated in hearts of db/db diabetic mice compared to wild-type mice. The results suggest that glyLDL increased the abundance of NOX4 or p22phox via an HSF1-independent pathway, but that of PAI-1 via an HSF1-dependent manner. NOX4 plays a crucial role in glyLDL-induced expression of HSF1 and PAI-1 in mouse fibroblasts. Increased expression of NOX4, HSF1, and PAI-1 was detected in cardiovascular tissue of diabetic mice.

  20. Regulatory roles for NKT cell ligands in environmentally induced autoimmunity.

    PubMed

    Vas, Jaya; Mattner, Jochen; Richardson, Stewart; Ndonye, Rachel; Gaughan, John P; Howell, Amy; Monestier, Marc

    2008-11-15

    The development of autoimmune diseases is frequently linked to exposure to environmental factors such as chemicals, drugs, or infections. In the experimental model of metal-induced autoimmunity, administration of subtoxic doses of mercury (a common environmental pollutant) to genetically susceptible mice induces an autoimmune syndrome with rapid anti-nucleolar Ab production and immune system activation. Regulatory components of the innate immune system such as NKT cells and TLRs can also modulate the autoimmune process. We examined the interplay among environmental chemicals and NKT cells in the regulation of autoimmunity. Additionally, we studied NKT and TLR ligands in a tolerance model in which preadministration of a low dose of mercury in the steady state renders animals tolerant to metal-induced autoimmunity. We also studied the effect of Sphingomonas capsulata, a bacterial strain that carries both NKT cell and TLR ligands, on metal-induced autoimmunity. Overall, NKT cell activation by synthetic ligands enhanced the manifestations of metal-induced autoimmunity. Exposure to S. capsulata exacerbated autoimmunity elicited by mercury. Although the synthetic NKT cell ligands that we used are reportedly similar in their ability to activate NKT cells, they displayed pronounced differences when coinjected with environmental agents or TLR ligands. Individual NKT ligands differed in their ability to prevent or break tolerance induced by low-dose mercury treatment. Likewise, different NKT ligands either dramatically potentiated or inhibited the ability of TLR9 agonistic oligonucleotides to disrupt tolerance to mercury. Our data suggest that these differences could be mediated by the modification of cytokine profiles and regulatory T cell numbers.

  1. Transforming Growth Factor (TGF)-β1–producing Regulatory T Cells Induce Smad-mediated Interleukin 10 Secretion That Facilitates Coordinated Immunoregulatory Activity and Amelioration of TGF-β1–mediated Fibrosis

    PubMed Central

    Kitani, Atsushi; Fuss, Ivan; Nakamura, Kazuhiko; Kumaki, Fumiyuki; Usui, Takashi; Strober, Warren

    2003-01-01

    Interleukin (IL)-10 and transforming growth factor (TGF)-β1 are suppressor cytokines that frequently occur together during a regulatory T cell response. Here we used a one gene doxycycline (Dox)-inducible plasmid encoding TGF-β1 to analyze this association and test its utility. In initial studies, we showed that intranasal administration of this plasmid (along with Dox) led to the appearance of TGF-β1–producing cells (in spleen and lamina propria) and the almost concomitant appearance of IL-10–producing cells. Moreover, we showed that these cells exert Dox-regulated suppression of the T helper cell (Th)1-mediated inflammation in trinitrobenzene sulfonic acid colitis. In subsequent in vitro studies using retroviral TGF-β1 expression, we established that IL-10 production by Th1 cells occurs after exposure to TGF-β1 from either an endogenous or exogenous source. In addition, using a self-inactivating retrovirus luciferase reporter construct we showed that TGF-β1 induces Smad4, which then binds to and activates the IL-10 promoter. Furthermore, intranasal TGF-β1 plasmid administration ameliorates bleomycin-induced fibrosis in wild-type but not IL-10–deficient mice, strongly suggesting that the amelioration is IL-10 dependent and that IL-10 protects mice from TGF-β1–mediated fibrosis. Taken together, these findings suggest that the induction of IL-10 by TGF-β1 is not fortuitous, but instead fulfills important requirements of TGF-β1 function after its secretion by regulatory T cells. PMID:14557415

  2. Interferons and interferon regulatory factors in malaria.

    PubMed

    Gun, Sin Yee; Claser, Carla; Tan, Kevin Shyong Wei; Rénia, Laurent

    2014-01-01

    Malaria is one of the most serious infectious diseases in humans and responsible for approximately 500 million clinical cases and 500 thousand deaths annually. Acquired adaptive immune responses control parasite replication and infection-induced pathologies. Most infections are clinically silent which reflects on the ability of adaptive immune mechanisms to prevent the disease. However, a minority of these can become severe and life-threatening, manifesting a range of overlapping syndromes of complex origins which could be induced by uncontrolled immune responses. Major players of the innate and adaptive responses are interferons. Here, we review their roles and the signaling pathways involved in their production and protection against infection and induced immunopathologies.

  3. Transcription factor trapping by RNA in gene regulatory elements.

    PubMed

    Sigova, Alla A; Abraham, Brian J; Ji, Xiong; Molinie, Benoit; Hannett, Nancy M; Guo, Yang Eric; Jangi, Mohini; Giallourakis, Cosmas C; Sharp, Phillip A; Young, Richard A

    2015-11-20

    Transcription factors (TFs) bind specific sequences in promoter-proximal and -distal DNA elements to regulate gene transcription. RNA is transcribed from both of these DNA elements, and some DNA binding TFs bind RNA. Hence, RNA transcribed from regulatory elements may contribute to stable TF occupancy at these sites. We show that the ubiquitously expressed TF Yin-Yang 1 (YY1) binds to both gene regulatory elements and their associated RNA species across the entire genome. Reduced transcription of regulatory elements diminishes YY1 occupancy, whereas artificial tethering of RNA enhances YY1 occupancy at these elements. We propose that RNA makes a modest but important contribution to the maintenance of certain TFs at gene regulatory elements and suggest that transcription of regulatory elements produces a positive-feedback loop that contributes to the stability of gene expression programs.

  4. Circuitry and dynamics of human transcription factor regulatory networks

    PubMed Central

    Neph, Shane; Stergachis, Andrew B.; Reynolds, Alex; Sandstrom, Richard; Borenstein, Elhanan; Stamatoyannopoulos, John A.

    2012-01-01

    SUMMARY The combinatorial cross-regulation of hundreds of sequence-specific transcription factors defines a regulatory network that underlies cellular identity and function. Here we use genome-wide maps of in vivo DNaseI footprints to assemble an extensive core human regulatory network comprising connections among 475 sequence-specific transcription factors, and to analyze the dynamics of these connections across 41 diverse cell and tissue types. We find that human transcription factor networks are highly cell-selective and are driven by cohorts of factors that include regulators with previously unrecognized roles in control of cellular identity. Moreover, we identify many widely expressed factors that impact transcriptional regulatory networks in a cell-selective manner. Strikingly, in spite of their inherent diversity, all cell type regulatory networks independently converge on a common architecture that closely resembles the topology of living neuronal networks. Together, our results provide the first description of the circuitry, dynamics, and organizing principles of the human transcription factor regulatory network. PMID:22959076

  5. Biosynthesis of nitric oxide activates iron regulatory factor in macrophages.

    PubMed

    Drapier, J C; Hirling, H; Wietzerbin, J; Kaldy, P; Kühn, L C

    1993-09-01

    Biosynthesis of nitric oxide (NO) from L-arginine modulates activity of iron-dependent enzymes, including mitochondrial acontiase, an [Fe-S] protein. We examined the effect of NO on the activity of iron regulatory factor (IRF), a cytoplasmic protein which modulates both ferritin mRNA translation and transferrin receptor mRNA stability by binding to specific mRNA sequences called iron responsive elements (IREs). Murine macrophages were activated with interferon-gamma and lipopolysaccharide to induce NO synthase activity and cultured in the presence or absence of NG-substituted analogues of L-arginine which served as selective inhibitors of NO synthesis. Measurement of the nitrite concentration in the culture medium was taken as an index of NO production. Mitochondria-free cytosols were then prepared and aconitase activity as well as IRE binding activity and induction of IRE binding activity were correlated and depended on NO synthesis after IFN-gamma and/or LPS stimulation. Authentic NO gas as well as the NO-generating compound 3-morpholinosydnonimine (SIN-1) also conversely modulated aconitase and IRE binding activities of purified recombinant IRF. These results provide evidence that endogenously produced NO may modulate the post-transcriptional regulation of genes involved in iron homeostasis and support the hypothesis that the [Fe-S] cluster of IRF mediates iron-dependent regulation. PMID:7504626

  6. A role for interferon regulatory factor 4 in receptor editing.

    PubMed

    Pathak, Simanta; Ma, Shibin; Trinh, Long; Lu, Runqing

    2008-04-01

    Receptor editing is the primary means through which B cells revise antigen receptors and maintain central tolerance. Previous studies have demonstrated that interferon regulatory factor 4 (IRF-4) and IRF-8 promote immunoglobulin light-chain rearrangement and transcription at the pre-B stage. Here, the roles of IRF-4 and -8 in receptor editing were analyzed. Our results show that secondary rearrangement was impaired in IRF-4 but not IRF-8 mutant mice, suggesting that receptor editing is defective in the absence of IRF-4. The role of IRF-4 in receptor editing was further examined in B-cell-receptor (BCR) transgenic mice. Our results show that secondary rearrangement triggered by membrane-bound antigen was defective in the IRF-4-deficient mice. Our results further reveal that the defect in secondary rearrangement is more severe at the immunoglobulin lambda locus than at the kappa locus, indicating that IRF-4 is more critical for the lambda rearrangement. We provide evidence demonstrating that the expression of IRF-4 in immature B cells is rapidly induced by self-antigen and that the reconstitution of IRF-4 expression in the IRF-4 mutant immature B cells promotes secondary rearrangement. Thus, our studies identify IRF-4 as a nuclear effector of a BCR signaling pathway that promotes secondary rearrangement at the immature B-cell stage.

  7. Regulatory coding of lymphoid lineage choice by hematopoietic transcription factors

    NASA Technical Reports Server (NTRS)

    Warren, Luigi A.; Rothenberg, Ellen V.

    2003-01-01

    During lymphopoiesis, precursor cells negotiate a complex regulatory space, defined by the levels of several competing and cross-regulating transcription factors, before arriving at stable states of commitment to the B-, T- and NK-specific developmental programs. Recent perturbation experiments provide evidence that this space has three major axes, corresponding to the PU.1 versus GATA-1 balance, the intensity of Notch signaling through the CSL pathway, and the ratio of E-box transcription factors to their Id protein antagonists.

  8. The effect of hyperammonemia on myostatin and myogenic regulatory factor gene expression in broiler embryos

    PubMed Central

    Stern, R.A.; Ashwell, C.M.; Dasarathy, S.; Mozdziak, P.E.

    2015-01-01

    Myogenesis is facilitated by four myogenic regulatory factors and is significantly inhibited by myostatin. The objective of the current study was to examine embryonic gene regulation of myostatin/myogenic regulatory factors, and subsequent manipulations of protein synthesis, in broiler embryos under induced hyperammonemia. Broiler eggs were injected with ammonium acetate solution four times over 48 hours beginning on either embryonic day (ED) 15 or 17. Serum ammonia concentration was significantly higher (P < 0.05) in ammonium acetate injected embryos for both ED17 and ED19 collected samples when compared to sham-injected controls. Expression of mRNA, extracted from pectoralis major of experimental and control embryos, was measured using real-time quantitative PCR for myostatin, myogenic regulatory factors myogenic factor 5, myogenic determination factor 1, myogenin, myogenic regulatory factor 4, and paired box 7. A significantly lower (P < 0.01) myostatin expression was accompanied by a higher serum ammonia concentration in both ED17 and ED19 collected samples. Myogenic factor 5 expression was higher (P < 0.05) in ED17 collected samples administered ammonium acetate. In both ED17 and ED19 collected samples, myogenic regulatory factor 4 was lower (P ≤ 0.05) in ammonium acetate injected embryos. No significant difference was seen in myogenic determination factor 1, myogenin, or paired box 7 expression between treatment groups for either age of sample collection. Additionally, there was no significant difference in BrdU staining of histological samples taken from treated and control embryos. Myostatin protein levels were evaluated by Western blot analysis, and also showed lower myostatin expression (P < 0.05). Overall, it appears possible to inhibit myostatin expression through hyperammonemia, which is expected to have a positive effect on embryonic myogenesis and postnatal muscle growth. PMID:25689990

  9. Trypanosoma cruzi Induces Regulatory Dendritic Cells In Vitro▿

    PubMed Central

    Poncini, Carolina Verónica; Soto, Catalina Dirney Alba; Batalla, Estela; Solana, Maria Elisa; González Cappa, Stella Maris

    2008-01-01

    A main feature of acute infection with Trypanosoma cruzi is the presence of immunological disorders. A previous study demonstrated that acute infection with the virulent RA strain downregulates the expression of major histocompatibility complex class II (MHC-II) on antigen-presenting cells and impairs the T-cell stimulatory capacity of splenic dendritic cells (DC). In the present work, we assessed the ability of trypomastigotes (Tp) to modulate the differentiation stage and functionality of bone marrow-derived DC in vitro. We observed that the Tp stage of T. cruzi failed to activate DC, which preserved their low expression of MHC-II and costimulatory molecules, as well as their endocytic activity. We also show that Tp induced transforming growth factor β (TGF-β) secretion by DC and enhanced the gap between interleukin-10 (IL-10) and IL-12p70 production, showing a higher IL-10/IL-12p70 ratio upon lipopolysaccharide (LPS) treatment. In addition, we observed that Tp prevented DC full activation induced by LPS, thereby downregulating their MHC-II surface expression and inhibiting their capacity to stimulate lymphocyte proliferation. In vitro IL-10 neutralization during the differentiation process of DC with Tp+LPS showed a reversion of their inhibitory effect during mixed lymphocyte reaction. In contrast, only simultaneous neutralization of IL-10 and TGF-β, after DC differentiation, was involved in the partial restitution of lymphocyte proliferation. Since both TGF-β and IL-10 are immunosuppressive cytokines essential in the modulation of the immune response and important in the induction of tolerance, our results suggest for the first time that Tp are responsible for the generation of regulatory DC in vitro. PMID:18347042

  10. Putative Regulatory Factors Associated with Intramuscular Fat Content

    PubMed Central

    Cesar, Aline S. M.; Regitano, Luciana C. A.; Koltes, James E.; Fritz-Waters, Eric R.; Lanna, Dante P. D.; Gasparin, Gustavo; Mourão, Gerson B.; Oliveira, Priscila S. N.; Reecy, James M.; Coutinho, Luiz L.

    2015-01-01

    Intramuscular fat (IMF) content is related to insulin resistance, which is an important prediction factor for disorders, such as cardiovascular disease, obesity and type 2 diabetes in human. At the same time, it is an economically important trait, which influences the sensorial and nutritional value of meat. The deposition of IMF is influenced by many factors such as sex, age, nutrition, and genetics. In this study Nellore steers (Bos taurus indicus subspecies) were used to better understand the molecular mechanisms involved in IMF content. This was accomplished by identifying differentially expressed genes (DEG), biological pathways and putative regulatory factors. Animals included in this study had extreme genomic estimated breeding value (GEBV) for IMF. RNA-seq analysis, gene set enrichment analysis (GSEA) and co-expression network methods, such as partial correlation coefficient with information theory (PCIT), regulatory impact factor (RIF) and phenotypic impact factor (PIF) were utilized to better understand intramuscular adipogenesis. A total of 16,101 genes were analyzed in both groups (high (H) and low (L) GEBV) and 77 DEG (FDR 10%) were identified between the two groups. Pathway Studio software identified 13 significantly over-represented pathways, functional classes and small molecule signaling pathways within the DEG list. PCIT analyses identified genes with a difference in the number of gene-gene correlations between H and L group and detected putative regulatory factors involved in IMF content. Candidate genes identified by PCIT include: ANKRD26, HOXC5 and PPAPDC2. RIF and PIF analyses identified several candidate genes: GLI2 and IGF2 (RIF1), MPC1 and UBL5 (RIF2) and a host of small RNAs, including miR-1281 (PIF). These findings contribute to a better understanding of the molecular mechanisms that underlie fat content and energy balance in muscle and provide important information for the production of healthier beef for human consumption. PMID:26042666

  11. Putative regulatory factors associated with intramuscular fat content.

    PubMed

    Cesar, Aline S M; Regitano, Luciana C A; Koltes, James E; Fritz-Waters, Eric R; Lanna, Dante P D; Gasparin, Gustavo; Mourão, Gerson B; Oliveira, Priscila S N; Reecy, James M; Coutinho, Luiz L

    2015-01-01

    Intramuscular fat (IMF) content is related to insulin resistance, which is an important prediction factor for disorders, such as cardiovascular disease, obesity and type 2 diabetes in human. At the same time, it is an economically important trait, which influences the sensorial and nutritional value of meat. The deposition of IMF is influenced by many factors such as sex, age, nutrition, and genetics. In this study Nellore steers (Bos taurus indicus subspecies) were used to better understand the molecular mechanisms involved in IMF content. This was accomplished by identifying differentially expressed genes (DEG), biological pathways and putative regulatory factors. Animals included in this study had extreme genomic estimated breeding value (GEBV) for IMF. RNA-seq analysis, gene set enrichment analysis (GSEA) and co-expression network methods, such as partial correlation coefficient with information theory (PCIT), regulatory impact factor (RIF) and phenotypic impact factor (PIF) were utilized to better understand intramuscular adipogenesis. A total of 16,101 genes were analyzed in both groups (high (H) and low (L) GEBV) and 77 DEG (FDR 10%) were identified between the two groups. Pathway Studio software identified 13 significantly over-represented pathways, functional classes and small molecule signaling pathways within the DEG list. PCIT analyses identified genes with a difference in the number of gene-gene correlations between H and L group and detected putative regulatory factors involved in IMF content. Candidate genes identified by PCIT include: ANKRD26, HOXC5 and PPAPDC2. RIF and PIF analyses identified several candidate genes: GLI2 and IGF2 (RIF1), MPC1 and UBL5 (RIF2) and a host of small RNAs, including miR-1281 (PIF). These findings contribute to a better understanding of the molecular mechanisms that underlie fat content and energy balance in muscle and provide important information for the production of healthier beef for human consumption.

  12. Effect of regulatory cells induced by influenza virus during adoptive transfer

    SciTech Connect

    Volgarev, A.P.; Maiorova, L.P.; Smorodintsev, A.A.

    1987-06-01

    This paper examines the protective effect of humoral and cellular factors of immunity in experimental influenza infection in mice, with an analysis of the functional activity of the regulatory subpopulations of the lymphocytes, especially in the early stages of observation. The regulatory activity of the splenic lymphocytes was studied by assessing changes in the level of mitogen-induced proliferation of test cultures of lymphocytes from intact animals in the presence of lymphocytes obtained from infected mice at different times after infection. Tritium-labelled thymidine was used as the marker.

  13. Regulation of APC Activity by Phosphorylation and Regulatory Factors

    PubMed Central

    Kotani, Shuji; Tanaka, Hirofumi; Yasuda, Hideyo; Todokoro, Kazuo

    1999-01-01

    Ubiquitin-dependent proteolysis of Cut2/Pds1 and Cyclin B is required for sister chromatid separation and exit from mitosis, respectively. Anaphase-promoting complex/cyclosome (APC) specifically ubiquitinates Cut2/Pds1 at metaphase–anaphase transition, and ubiquitinates Cyclin B in late mitosis and G1 phase. However, the exact regulatory mechanism of substrate-specific activation of mammalian APC with the right timing remains to be elucidated. We found that not only the binding of the activators Cdc20 and Cdh1 and the inhibitor Mad2 to APC, but also the phosphorylation of Cdc20 and Cdh1 by Cdc2-Cyclin B and that of APC by Polo-like kinase and cAMP-dependent protein kinase, regulate APC activity. The cooperation of the phosphorylation/dephosphorylation and the regulatory factors in regulation of APC activity may thus control the precise progression of mitosis. PMID:10459014

  14. Dysregulated T helper cell differentiation in the absence of interferon regulatory factor 4.

    PubMed

    Lohoff, Michael; Mittrücker, Hans-Willi; Prechtl, Stefan; Bischof, Susi; Sommer, Frank; Kock, Sonja; Ferrick, David A; Duncan, Gordon S; Gessner, Andre; Mak, Tak W

    2002-09-01

    Certain IFN regulatory factor (IRF) transcription factors indirectly influence T helper (Th) cell differentiation by regulating the production of IL-12. Here, we show that IRF4 directly regulates Th cell differentiation in vitro and in vivo during murine leishmaniasis. In the absence of IRF4, IL-12-induced Th1 cell differentiation was compromised, while IL-4 failed to induce Th2 cell differentiation. Instead, IL-4 tended to induce Th1 cells, defined by production of IFN-gamma and TNF. Although early IL-4 signaling was normal in IRF4(-/-) Th cells, the protein GATA-3, a transcription factor critical for Th2 development, was not up-regulated following IL-4 treatment. Retroviral overexpression of GATA-3 rescued Th2 differentiation. Therefore, IRF4 deficiency manifests itself as severely dysregulated Th cell differentiation.

  15. Interaction of Trypanosoma cruzi adenylate cyclase with liver regulatory factors.

    PubMed Central

    Eisenschlos, C; Flawiá, M M; Torruella, M; Torres, H N

    1986-01-01

    Trypanosoma cruzi adenylate cyclase catalytic subunits may interact with regulatory factors from rat liver membranes, reconstituting heterologous systems which are catalytically active in assay mixtures containing MgATP. The systems show stimulatory responses to glucagon and guanosine 5'-[beta gamma-imido]triphosphate (p[NH]ppG) or fluoride. Reconstitution was obtained by three different methods: fusion of rat liver membranes (pretreated with N-ethylmaleimide) to T. cruzi membranes; interaction of detergent extracts of rat liver membranes with T. cruzi membranes; or interaction of purified preparations of T. cruzi adenylate cyclase and of liver membrane factors in phospholipid vesicles. The liver factors responsible for the guanine nucleotide effect were characterized as the NS protein. Data also indicate that reconstitution requires the presence of a membrane substrate. PMID:2947568

  16. Developmental Stage-Specific Hepatocytes Induce Maturation of HepG2 Cells by Rebuilding the Regulatory Circuit.

    PubMed

    Li, Yanning; Liu, Demei; Zong, Yanhong; Qi, Jinsheng; Li, Bin; Liu, Kun; Xiao, Hui

    2015-04-14

    On the basis of their characteristics, we presume that developmental stage-specific hepatocytes should have the ability to induce maturation of hepatoma cells. A regulatory circuit formed by hepatocyte nuclear factor (HNF)-4α, HNF-1α, HNF-6 and the upstream stimulatory factor (USF-1) play a key role in the maturation of embryonic hepatocytes; however, it is unclear whether the regulatory circuit mediates the embryonic induction of hepatoma cell maturation. In this study, 12.5-d to 15.5-d mouse embryonic hepatocytes or their medium were used to coculture or treat HepG2 cells, and the induced maturation was evaluated in vitro and in vivo. In the induced HepG2 cells, the components of the regulatory circuit were detected, their cross-regulation was evaluated and HNF-4α RNA interference was performed. We found that 13.5-d to 14.5-d embryonic hepatocytes could induce HepG2 cell maturation, demonstrated by morphological changes, increased maturation markers and decreased c-Myc and α-fetoprotein (AFP) in vitro. The majority of HepG2 tumors were eliminated by 13.5-d embryonic induction in vivo. All components of the regulatory circuit were upregulated and the binding of HNF-4α, HNF-1α, HNF-6 and USF-1 to their target sites was promoted to rebuild the regulatory circuit in the induced HepG2 cells. Moreover, RNA interference targeting HNF-4α, which is the core of the regulatory circuit, attenuated the induced maturation of HepG2 cells with downregulation of the regulatory circuit. These results revealed that developmental stage-specific hepatocytes could induce the maturation of HepG2 cells by rebuilding the regulatory circuit.

  17. Uncovering Transcriptional Regulatory Networks by Sparse Bayesian Factor Model

    NASA Astrophysics Data System (ADS)

    Meng, Jia; Zhang, Jianqiu(Michelle); Qi, Yuan(Alan); Chen, Yidong; Huang, Yufei

    2010-12-01

    The problem of uncovering transcriptional regulation by transcription factors (TFs) based on microarray data is considered. A novel Bayesian sparse correlated rectified factor model (BSCRFM) is proposed that models the unknown TF protein level activity, the correlated regulations between TFs, and the sparse nature of TF-regulated genes. The model admits prior knowledge from existing database regarding TF-regulated target genes based on a sparse prior and through a developed Gibbs sampling algorithm, a context-specific transcriptional regulatory network specific to the experimental condition of the microarray data can be obtained. The proposed model and the Gibbs sampling algorithm were evaluated on the simulated systems, and results demonstrated the validity and effectiveness of the proposed approach. The proposed model was then applied to the breast cancer microarray data of patients with Estrogen Receptor positive ([InlineEquation not available: see fulltext.]) status and Estrogen Receptor negative ([InlineEquation not available: see fulltext.]) status, respectively.

  18. Bayesian non-negative factor analysis for reconstructing transcription factor mediated regulatory networks

    PubMed Central

    2011-01-01

    Background Transcriptional regulation by transcription factor (TF) controls the time and abundance of mRNA transcription. Due to the limitation of current proteomics technologies, large scale measurements of protein level activities of TFs is usually infeasible, making computational reconstruction of transcriptional regulatory network a difficult task. Results We proposed here a novel Bayesian non-negative factor model for TF mediated regulatory networks. Particularly, the non-negative TF activities and sample clustering effect are modeled as the factors from a Dirichlet process mixture of rectified Gaussian distributions, and the sparse regulatory coefficients are modeled as the loadings from a sparse distribution that constrains its sparsity using knowledge from database; meantime, a Gibbs sampling solution was developed to infer the underlying network structure and the unknown TF activities simultaneously. The developed approach has been applied to simulated system and breast cancer gene expression data. Result shows that, the proposed method was able to systematically uncover TF mediated transcriptional regulatory network structure, the regulatory coefficients, the TF protein level activities and the sample clustering effect. The regulation target prediction result is highly coordinated with the prior knowledge, and sample clustering result shows superior performance over previous molecular based clustering method. Conclusions The results demonstrated the validity and effectiveness of the proposed approach in reconstructing transcriptional networks mediated by TFs through simulated systems and real data. PMID:22166063

  19. Clinical trials in "emerging markets": regulatory considerations and other factors.

    PubMed

    Singh, Romi; Wang, Ouhong

    2013-11-01

    Clinical studies are being placed in emerging markets as part of global drug development programs to access large pool of eligible patients and to benefit from a cost effective structure. However, over the last few years, the definition of "emerging markets" is being revisited, especially from a regulatory perspective. For purposes of this article, countries outside US, EU and the traditional "western countries" are discussed. Multiple factors are considered for placement of clinical studies such as adherence to Good Clinical Practice (GCP), medical infrastructure & standard of care, number of eligible patients, etc. This article also discusses other quantitative factors such as country's GDP, patent applications, healthcare expenditure, healthcare infrastructure, corruption, innovation, etc. These different factors and indexes are correlated to the number of clinical studies ongoing in the "emerging markets". R&D, healthcare expenditure, technology infrastructure, transparency, and level of innovation, show a significant correlation with the number of clinical trials being conducted in these countries. This is the first analysis of its kind to evaluate and correlate the various other factors to the number of clinical studies in a country. PMID:24070788

  20. Clinical trials in "emerging markets": regulatory considerations and other factors.

    PubMed

    Singh, Romi; Wang, Ouhong

    2013-11-01

    Clinical studies are being placed in emerging markets as part of global drug development programs to access large pool of eligible patients and to benefit from a cost effective structure. However, over the last few years, the definition of "emerging markets" is being revisited, especially from a regulatory perspective. For purposes of this article, countries outside US, EU and the traditional "western countries" are discussed. Multiple factors are considered for placement of clinical studies such as adherence to Good Clinical Practice (GCP), medical infrastructure & standard of care, number of eligible patients, etc. This article also discusses other quantitative factors such as country's GDP, patent applications, healthcare expenditure, healthcare infrastructure, corruption, innovation, etc. These different factors and indexes are correlated to the number of clinical studies ongoing in the "emerging markets". R&D, healthcare expenditure, technology infrastructure, transparency, and level of innovation, show a significant correlation with the number of clinical trials being conducted in these countries. This is the first analysis of its kind to evaluate and correlate the various other factors to the number of clinical studies in a country.

  1. Foxo transcription factors control regulatory T cell development and function

    PubMed Central

    Kerdiles, Yann M.; Stone, Erica L.; Beisner, Daniel L.; McGargill, Maureen A.; Ch'en, Irene L.; Stockmann, Christian; Katayama, Carol D.; Hedrick, Stephen M.

    2010-01-01

    SUMMARY Foxo transcription factors integrate extrinsic signals to regulate cell division, differentiation and survival, and specific functions of lymphoid and myeloid cells. Here we showed the absence of Foxo1 severely curtailed the development of Foxp3+ regulatory T (Treg) cells, and those that developed were nonfunctional in vivo. The loss of function included diminished CTLA-4 receptor expression as the Ctla4 gene was a direct target of Foxo1. T cell specific loss of Foxo1 resulted in exocrine pancreatitis, hind limb paralysis, multi-organ lymphocyte infiltration, anti-nuclear antibodies and expanded germinal centers. Foxo-mediated control over Treg cell specification was further revealed by the inability of TGF-β cytokine to suppress T-bet transcription factor in the absence of Foxo1, resulting in IFN-γ-secretion. In addition the absence of Foxo3 exacerbated the effects of the loss of Foxo1. Thus, Foxo transcription factors guide the contingencies of T cell differentiation and specific functions of effector cell populations. PMID:21167754

  2. Tumor suppressor properties of the splicing regulatory factor RBM10

    PubMed Central

    Hernández, Jordi; Bechara, Elias; Schlesinger, Doerte; Delgado, Javier; Serrano, Luis; Valcárcel, Juan

    2016-01-01

    ABSTRACT RBM10 is an RNA binding protein and alternative splicing regulator frequently mutated in lung adenocarcinomas. Recent results indicate that RBM10 inhibits proliferation of lung cancer cells by promoting skipping of exon 9 of the gene NUMB, a frequent alternative splicing change in lung cancer generating a negative regulator of Notch signaling. Complementing these observations, we show that knock down of RBM10 in human cancer cells enhances growth of mouse tumor xenografts, confirming that RBM10 acts as a tumor suppressor, while knock down of an oncogenic mutant version of RBM10 reduces xenograft tumor growth. A RBM10 mutation found in lung cancer cells, V354E, disrupts RBM10-mediated regulation of NUMB alternative splicing, inducing the cell proliferation-promoting isoform. We now show that 2 natural RBM10 isoforms that differ by the presence or absence of V354 in the second RNA Recognition Motif (RRM2), display similar regulatory effects on NUMB alternative splicing, suggesting that V354E actively disrupts RBM10 activity. Structural modeling localizes V354 in the outside surface of one α-helix opposite to the RNA binding surface of RBM10, and we show that the mutation does not compromise binding of the RRM2 domain to NUMB RNA regulatory sequences. We further show that other RBM10 mutations found in lung adenocarcinomas also compromise regulation of NUMB exon 9. Collectively, our previous and current results reveal that RBM10 is a tumor suppressor that represses Notch signaling and cell proliferation through the regulation of NUMB alternative splicing. PMID:26853560

  3. Transcriptional Regulatory Network Analysis of MYB Transcription Factor Family Genes in Rice

    PubMed Central

    Smita, Shuchi; Katiyar, Amit; Chinnusamy, Viswanathan; Pandey, Dev M.; Bansal, Kailash C.

    2015-01-01

    MYB transcription factor (TF) is one of the largest TF families and regulates defense responses to various stresses, hormone signaling as well as many metabolic and developmental processes in plants. Understanding these regulatory hierarchies of gene expression networks in response to developmental and environmental cues is a major challenge due to the complex interactions between the genetic elements. Correlation analyses are useful to unravel co-regulated gene pairs governing biological process as well as identification of new candidate hub genes in response to these complex processes. High throughput expression profiling data are highly useful for construction of co-expression networks. In the present study, we utilized transcriptome data for comprehensive regulatory network studies of MYB TFs by “top-down” and “guide-gene” approaches. More than 50% of OsMYBs were strongly correlated under 50 experimental conditions with 51 hub genes via “top-down” approach. Further, clusters were identified using Markov Clustering (MCL). To maximize the clustering performance, parameter evaluation of the MCL inflation score (I) was performed in terms of enriched GO categories by measuring F-score. Comparison of co-expressed cluster and clads analyzed from phylogenetic analysis signifies their evolutionarily conserved co-regulatory role. We utilized compendium of known interaction and biological role with Gene Ontology enrichment analysis to hypothesize function of coexpressed OsMYBs. In the other part, the transcriptional regulatory network analysis by “guide-gene” approach revealed 40 putative targets of 26 OsMYB TF hubs with high correlation value utilizing 815 microarray data. The putative targets with MYB-binding cis-elements enrichment in their promoter region, functional co-occurrence as well as nuclear localization supports our finding. Specially, enrichment of MYB binding regions involved in drought-inducibility implying their regulatory role in drought

  4. Transcription factor abundance controlled by an auto-regulatory mechanism involving a transcription start site switch

    PubMed Central

    Ngondo, Richard Patryk; Carbon, Philippe

    2014-01-01

    A transcriptional feedback loop is the simplest and most direct means for a transcription factor to provide an increased stability of gene expression. In this work performed in human cells, we reveal a new negative auto-regulatory mechanism involving an alternative transcription start site (TSS) usage. Using the activating transcription factor ZNF143 as a model, we show that the ZNF143 low-affinity binding sites, located downstream of its canonical TSS, play the role of protein sensors to induce the up- or down-regulation of ZNF143 gene expression. We uncovered that the TSS switch that mediates this regulation implies the differential expression of two transcripts with an opposite protein production ability due to their different 5′ untranslated regions. Moreover, our analysis of the ENCODE data suggests that this mechanism could be used by other transcription factors to rapidly respond to their own aberrant expression level. PMID:24234445

  5. Retinoic acid exerts dual regulatory actions on the expression and nuclear localization of interferon regulatory factor-1.

    PubMed

    Luo, Xin M; Ross, A Catharine

    2006-05-01

    Interferon regulatory factor-1 (IRF-1), a transcription factor and tumor suppressor involved in cell growth regulation and immune responses, has been shown to be induced by all-trans retinoic acid (ATRA). However, the factors controlling the cellular location and activity of IRF-1 are not well understood. In this study, we examined the expression of IRF-1 and its nuclear localization, DNA-binding activity, and target gene expression in human mammary epithelial MCF10A cells, a model of breast epithelial cell differentiation and carcinogenesis. Following initial treatment with ATRA, IRF-1 mRNA and protein were induced within 2 hrs, reached a peak (>30-fold induction) at 8 hrs, and declined afterwards. IRF-1 protein was predominantly cytoplasmic during this treatment. Although a second dose of ATRA or Am580 (a related retinoid selective for retinoic acid receptor-alpha [RARalpha]), given 16 hrs after the first dose, restimulated IRF-1 mRNA and protein levels to a similar level to that obtained by the first dose, IRF-1 was predominantly concentrated in the nucleus after restimulation. ATRA and Am580 also increased nuclear RARalpha, whereas retinoid X receptor-alpha (RXRalpha)--a dimerization partner for RARalpha, was localized to the nucleus upon second exposure to ATRA. However, ATRA and Am580 did not regulate the expression or activation of signal transducer and activator of transcription-1 (STAT-1), a transcription factor capable of inducing the expression of IRF-1, indicating an STAT-1-independent mechanism of regulation by ATRA and Am580. The increase in nuclear IRF-1 after retinoid restimulation was accompanied by enhanced binding to an IRF-E DNA response element, and elevated expression of an IRF-1 target gene, 2',5'-oligoadenylate synthetase-2. The dual effect of retinoids in increasing IRF-1 mRNA and protein and in augmenting the nuclear localization of IRF-1 protein may be essential for maximizing the tumor suppressor activity and the immunosurveillance

  6. Stress-induced Start Codon Fidelity Regulates Arsenite-inducible Regulatory Particle-associated Protein (AIRAP) Translation*

    PubMed Central

    Zach, Lolita; Braunstein, Ilana; Stanhill, Ariel

    2014-01-01

    Initial steps in protein synthesis are highly regulated processes as they define the reading frame of the translation machinery. Eukaryotic translation initiation is a process facilitated by numerous factors (eIFs), aimed to form a “scanning” mechanism toward the initiation codon. Translation initiation of the main open reading frame (ORF) in an mRNA transcript has been reported to be regulated by upstream open reading frames (uORFs) in a manner of re-initiation. This mode of regulation is governed by the phosphorylation status of eIF2α and controlled by cellular stresses. Another mode of translational initiation regulation is leaky scanning, and this regulatory process has not been extensively studied. We have identified arsenite-inducible regulatory particle-associated protein (AIRAP) transcript to be translationally induced during arsenite stress conditions. AIRAP transcript contains a single uORF in a poor-kozak context. AIRAP translation induction is governed by means of leaky scanning and not re-initiation. This induction of AIRAP is solely dependent on eIF1 and the uORF kozak context. We show that eIF1 is phosphorylated under specific conditions that induce protein misfolding and have biochemically characterized this site of phosphorylation. Our data indicate that leaky scanning like re-initiation is responsive to stress conditions and that leaky scanning can induce ORF translation by bypassing poor kozak context of a single uORF transcript. PMID:24898249

  7. Molecular characterization of interferon regulatory factor 1 in Bubalus bubalis.

    PubMed

    Stafuzza, N B; Borges, M M; Amaral-Trusty, M E J

    2015-01-01

    Interferon regulatory factor 1 (IRF1) is functionally diverse in the regulation of immune response and is considered to be an important candidate gene for studying disease susceptibility in mammals. In this paper, we characterized the whole sequence of the IRF1 gene in river buffalo (Bubalus bubalis) and compared genomic and the amino acid sequences between different species. The buffalo IRF1 gene was 7099 bp long and organized into 10 exons and nine introns. Its molecular structure showed exactly the same number of exons (10) and introns (nine) in bovids, mice, horses, humans, and chickens. However, rats did not have exon 5, but had the largest exon 4, which suggests that exon 5 was incorporated into exon 4. The coding and the amino acid sequences of the gene showed that identity varied from 73 to 99% at the coding sequence level and from 61 to 100% at the amino acid level when compared with other mammals and chickens. Comparative analysis of the gene sequence between two different buffalo breeds, Murrah and Mediterranean, revealed six potential SNPs that are primarily located in the 5' and 3'UTRs. PMID:26400319

  8. Altered oncomodules underlie chromatin regulatory factors driver mutations.

    PubMed

    Frigola, Joan; Iturbide, Ane; Lopez-Bigas, Nuria; Peiro, Sandra; Gonzalez-Perez, Abel

    2016-05-24

    Chromatin regulatory factors (CRFs), are known to be involved in tumorigenesis in several cancer types. Nevertheless, the molecular mechanisms through which driver alterations of CRFs cause tumorigenesis remain unknown. Here, we developed a CRFs Oncomodules Discovery approach, which mines several sources of cancer genomics and perturbaomics data. The approach prioritizes sets of genes significantly miss-regulated in primary tumors (oncomodules) bearing mutations of driver CRFs. We applied the approach to eleven TCGA tumor cohorts and uncovered oncomodules potentially associated to mutations of five driver CRFs in three cancer types. Our results revealed, for example, the potential involvement of the mTOR pathway in the development of tumors with loss-of-function mutations of MLL2 in head and neck squamous cell carcinomas. The experimental validation that MLL2 loss-of-function increases the sensitivity of cancer cell lines to mTOR inhibition lends further support to the validity of our approach. The potential oncogenic modules detected by our approach may guide experiments proposing ways to indirectly target driver mutations of CRFs. PMID:27095575

  9. Type I Interferons Induce T Regulatory 1 Responses and Restrict Humoral Immunity during Experimental Malaria

    PubMed Central

    Zander, Ryan A.; Guthmiller, Jenna J.; Graham, Amy C.; Burke, Bradly E.; Carr, Daniel J.J.

    2016-01-01

    CD4 T cell-dependent antibody responses are essential for limiting Plasmodium parasite replication and the severity of malaria; however, the factors that regulate humoral immunity during highly inflammatory, Th1-biased systemic infections are poorly understood. Using genetic and biochemical approaches, we show that Plasmodium infection-induced type I interferons limit T follicular helper accumulation and constrain anti-malarial humoral immunity. Mechanistically we show that CD4 T cell-intrinsic type I interferon signaling induces T-bet and Blimp-1 expression, thereby promoting T regulatory 1 responses. We further show that the secreted effector cytokines of T regulatory 1 cells, IL-10 and IFN-γ, collaborate to restrict T follicular helper accumulation, limit parasite-specific antibody responses, and diminish parasite control. This circuit of interferon-mediated Blimp-1 induction is also operational during chronic virus infection and can occur independently of IL-2 signaling. Thus, type I interferon-mediated induction of Blimp-1 and subsequent expansion of T regulatory 1 cells represent generalizable features of systemic, inflammatory Th1-biased viral and parasitic infections that are associated with suppression of humoral immunity. PMID:27732671

  10. Roles of Interferon Regulatory Factors in Chronic Myeloid Leukemia.

    PubMed

    Manzella, Livia; Tirrò, Elena; Pennisi, Maria Stella; Massimino, Michele; Stella, Stefania; Romano, Chiara; Vitale, Silvia Rita; Vigneri, Paolo

    2016-01-01

    The Interferon Regulatory Factor (IRF) family consists of multiple transcription factors involved in the regulation of a variety of biological processes. Originally identified as transcriptional regulators of the type I interferon system, IRFs play a pivotal role in adaptive immunity, cell growth, differentiation and tumorigenesis. Hence, understanding IRF biology has important implications in the host response to cancer development and progression. Many lines of evidence suggest that different IRFs are involved in the pathogenesis of Chronic Myeloid Leukemia (CML), a myeloproliferative disorder caused by the BCR-ABL oncoprotein. BCR-ABL displays constitutive tyrosine kinase activity that favors cell proliferation, inhibits apoptosis and allows cell survival even in the absence of proper adhesion to the extracellular matrix. Different BCR-ABL tyrosine kinase inhibitors are currently available for CML treatment. These drugs are able to generate eight year CML-specific overall survival rates >90%, only a minority of patients will achieve molecular responses compatible with drug discontinuation. Thus, there is an unmet need for additional therapeutic targets that may lead to the cure of most patients diagnosed with CML. A growing body of evidence has suggested a role for both IRF4 and IRF8 in the pathogenesis of CML. Furthermore, IRF1 is consistently deleted at one or both alleles in patients with leukemia and myelodysplasia. Finally, we have recently demonstrated that IRF5 is a target of BCR-ABL kinase activity and reduces CML cell proliferation. In this article, we provide an update on the current knowledge of the role of the IRFs in CML. PMID:26728039

  11. Chromatin properties of regulatory DNA probed by manipulation of transcription factors.

    PubMed

    Sharov, Alexei A; Nishiyama, Akira; Qian, Yong; Dudekula, Dawood B; Longo, Dan L; Schlessinger, David; Ko, Minoru S H

    2014-08-01

    Transcription factors (TFs) bind to DNA and regulate the transcription of nearby genes. However, only a small fraction of TF binding sites have such regulatory effects. Here we search for the predictors of functional binding sites by carrying out a systematic computational screening of a variety of contextual factors (histone modifications, nuclear lamin-bindings, and cofactor bindings). We used regression analysis to test if contextual factors are associated with upregulation or downregulation of neighboring genes following the induction or knockdown of the 9 TFs in mouse embryonic stem (ES) cells. Functional TF binding sites appeared to be either active (i.e., bound by P300, CHD7, mediator, cohesin, and SWI/SNF) or repressed (i.e., with H3K27me3 histone marks and bound by Polycomb factors). Active binding sites mediated the downregulation of nearby genes upon knocking down the activating TFs or inducing repressors. Repressed TF binding sites mediated the upregulation of nearby genes (e.g., poised developmental regulators) upon inducing TFs. In addition, repressed binding sites mediated repressive effects of TFs, identified by the downregulation of target genes after the induction of TFs or by the upregulation of target genes after the knockdown of TFs. The contextual factors associated with functions of DNA-bound TFs were used to improve the identification of candidate target genes regulated by TFs.

  12. Regulatory role of the respiratory supercomplex factors in Saccharomyces cerevisiae

    PubMed Central

    Rydström Lundin, Camilla; Ott, Martin; Ädelroth, Pia; Brzezinski, Peter

    2016-01-01

    The respiratory supercomplex factors (Rcf) 1 and 2 mediate supramolecular interactions between mitochondrial complexes III (ubiquinol-cytochrome c reductase; cyt. bc1) and IV (cytochrome c oxidase; CytcO). In addition, removal of these polypeptides results in decreased activity of CytcO, but not of cyt. bc1. In the present study, we have investigated the kinetics of ligand binding, the single-turnover reaction of CytcO with O2, and the linked cyt. bc1-CytcO quinol oxidation-oxygen-reduction activities in mitochondria in which Rcf1 or Rcf2 were removed genetically (strains rcf1Δ and rcf2Δ, respectively). The data show that in the rcf1Δ and rcf2Δ strains, in a significant fraction of the population, ligand binding occurs over a time scale that is ∼100-fold faster (τ ≅ 100 μs) than observed with the wild-type mitochondria (τ ≅ 10 ms), indicating structural changes. This effect is specific to removal of Rcf and not dissociation of the cyt. bc1–CytcO supercomplex. Furthermore, in the rcf1Δ and rcf2Δ strains, the single-turnover reaction of CytcO with O2 was incomplete. This observation indicates that the lower activity of CytcO is caused by a fraction of inactive CytcO rather than decreased CytcO activity of the entire population. Furthermore, the data suggest that the Rcf1 polypeptide mediates formation of an electron-transfer bridge from cyt. bc1 to CytcO via a tightly bound cyt. c. We discuss the significance of the proposed regulatory mechanism of Rcf1 and Rcf2 in the context of supramolecular interactions between cyt. bc1 and CytcO. PMID:27432958

  13. How tolerogenic dendritic cells induce regulatory T cells

    PubMed Central

    Maldonado, Roberto A.; von Andrian, Ulrich H.

    2010-01-01

    Since their discovery by Steinman and Cohn in 1973, dendritic cells (DCs) have become increasingly recognized for their crucial role as regulators of innate and adaptive immunity. DCs are exquisitely adept at acquiring, processing and presenting antigens to T cells. They also adjust the context (and hence the outcome) of antigen presentation in response to a plethora of environmental inputs that signal the occurence of pathogens or tissue damage. Such signals generally boost DC maturation, which promotes their migration from peripheral tissues into and within secondary lymphoid organs and their capacity to induce and regulate effector T cell responses. Conversely, more recent observations indicate that DCs are also crucial to ensure immunological peace. Indeed, DCs constantly present innocuous self and non-self antigens in a fashion that promotes tolerance, at least in part, through the control of regulatory T cells (Tregs). Tregs are specialized T cells that exert their immuno-suppressive function through a variety of mechanisms affecting both DCs and effector cells. Here, we review recent advances in our understanding of the relationship between tolerogenic DCs and Tregs. PMID:21056730

  14. Immune privilege induced by regulatory T cells in transplantation tolerance.

    PubMed

    Cobbold, Stephen P; Adams, Elizabeth; Graca, Luis; Daley, Stephen; Yates, Stephen; Paterson, Alison; Robertson, Nathan J; Nolan, Kathleen F; Fairchild, Paul J; Waldmann, Herman

    2006-10-01

    Immune privilege was originally believed to be associated with particular organs, such as the testes, brain, the anterior chamber of the eye, and the placenta, which need to be protected from any excessive inflammatory activity. It is now becoming clear, however, that immune privilege can be acquired locally in many different tissues in response to inflammation, but particularly due to the action of regulatory T cells (Tregs) induced by the deliberate therapeutic manipulation of the immune system toward tolerance. In this review, we consider the interplay between Tregs, dendritic cells, and the graft itself and the resulting local protective mechanisms that are coordinated to maintain the tolerant state. We discuss how both anti-inflammatory cytokines and negative costimulatory interactions can elicit a number of interrelated mechanisms to regulate both T-cell and antigen-presenting cell activity, for example, by catabolism of the amino acids tryptophan and arginine and the induction of hemoxygenase and carbon monoxide. The induction of local immune privilege has implications for the design of therapeutic regimens and the monitoring of the tolerant status of patients being weaned off immunosuppression. PMID:16972908

  15. Anti-Sigma Factors in E. coli: Common Regulatory Mechanisms Controlling Sigma Factors Availability

    PubMed Central

    Treviño-Quintanilla, Luis Gerardo; Freyre-González, Julio Augusto; Martínez-Flores, Irma

    2013-01-01

    In bacteria, transcriptional regulation is a key step in cellular gene expression. All bacteria contain a core RNA polymerase that is catalytically competent but requires an additional σ factor for specific promoter recognition and correct transcriptional initiation. The RNAP core is not able to selectively bind to a given σ factor. In contrast, different σ factors have different affinities for the RNAP core. As a consequence, the concentration of alternate σ factors requires strict regulation in order to properly control the delicate interplay among them, which favors the competence for the RNAP core. This control is archived by different σ/anti-σ controlling mechanisms that shape complex regulatory networks and cascades, and enable the response to sudden environmental cues, whose global understanding is a current challenge for systems biology. Although there have been a number of excellent studies on each of these σ/anti-σ post-transcriptional regulatory systems, no comprehensive comparison of these mechanisms in a single model organism has been conducted. Here, we survey all these systems in E. coli dissecting and analyzing their inner workings and highlightin their differences. Then, following an integral approach, we identify their commonalities and outline some of the principles exploited by the cell to effectively and globally reprogram the transcriptional machinery. These principles provide guidelines for developing biological synthetic circuits enabling an efficient and robust response to sudden stimuli. PMID:24396271

  16. Ubiquitination of Tumor Necrosis Factor Receptor-associated Factor 4 (TRAF4) by Smad Ubiquitination Regulatory Factor 1 (Smurf1) Regulates Motility of Breast Epithelial and Cancer Cells*

    PubMed Central

    Wang, Xiangchun; Jin, Chaoyang; Tang, Yi; Tang, Liu-Ya; Zhang, Ying E.

    2013-01-01

    Smad ubiquitin regulatory factors (Smurfs) are HECT-domain ubiquitin E3 ligases that regulate diverse cellular processes, including normal and tumor cell migration. However, the underlying mechanism of the Smurfs' role in cell migration is not fully understood. Here we show that Smurf1 induces ubiquitination of tumor necrosis factor receptor-associated factor 4 (TRAF4) at K190. Using the K190R mutant of TRAF4, we demonstrate that Smurf1-induced ubiquitination is required for proper localization of TRAF4 to tight junctions in confluent epithelial cells. We further show that TRAF4 is essential for the migration of both normal mammary epithelial and breast cancer cells. The ability of TRAF4 to promote cell migration is also dependent on Smurf1-mediated ubiquitination, which is associated with Rac1 activation by TRAF4. These results reveal a new regulatory circuit for cell migration, consisting of Smurf1-mediated ubiquitination of TRAF4 and Rac1 activation. PMID:23760265

  17. Identification and functional analysis of interferon regulatory factor 3 in Lateolabrax maculatus.

    PubMed

    Chen, Xiao-Wu; Wei, Qun; Wang, Zhi-Peng; Wang, Chun-Lei; Bi, Yan-Hui; Gu, Yi-Feng

    2016-10-01

    The interferon (IFN) regulatory factor 3 (IRF3) is a member of the IFN regulatory transcription factor family, which binds to the IFN-stimulated response element (ISRE) within the promoter of IFN genes and IFN-stimulated genes. In this study, the IRF3 cDNA of sea perch Lateolabrax maculatus (SpIRF3) was identified, which contained 1781 bp with an open reading frame of 1398 bp that coded a 465 amino acid protein. The SpIRF3 protein shared conserved characterizations with its homologues and displayed the conserved DNA-binding domain, IRF association domain, serine-rich C-terminal domain, and tryptophan residue cluster. Phylogenetic analysis illustrated that SpIRF3 belonged to the IRF3 subfamily. Subcellular localization analysis showed that SpIRF3 mainly resided in the cytoplasm without stimuli but translocated into nuclei in the presence of poly I:C. Real-time PCR data indicated that SpIRF3 was transcriptionally up-regulated by poly I:C stimulation in various organs. Moreover, reporter assay revealed that SpIRF3 functioned as a modulator in triggering the IFN response by inducing the activity of IFN and ISRE-containing promoter. These data revealed that SpIRF3 was a potential molecule in the IFN immune defense system against viral infection. PMID:27181713

  18. Transcriptional up-regulation of inhibitory PAS domain protein gene expression by hypoxia-inducible factor 1 (HIF-1): a negative feedback regulatory circuit in HIF-1-mediated signaling in hypoxic cells.

    PubMed

    Makino, Yuichi; Uenishi, Rie; Okamoto, Kensaku; Isoe, Tsubasa; Hosono, Osamu; Tanaka, Hirotoshi; Kanopka, Arvydas; Poellinger, Lorenz; Haneda, Masakazu; Morimoto, Chikao

    2007-05-11

    The inhibitory PAS (Per/Arnt/Sim) domain protein (IPAS), a dominant negative regulator of hypoxia-inducible transcription factors (HIFs), is potentially implicated in negative regulation of angiogenesis in such tissues as the avascular cornea of the eye. We have previously shown IPAS mRNA expression is up-regulated in hypoxic tissues, which at least in part involves hypoxia-dependent alternative splicing of the transcripts from the IPAS/HIF-3alpha locus. In the present study, we demonstrate that a hypoxia-driven transcriptional mechanism also plays a role in augmentation of IPAS gene expression. Isolation and analyses of the promoter region flanking to the first exon of IPAS gene revealed a functional hypoxia response element at position -834 to -799, whereas the sequence upstream of the HIF-3alpha first exon scarcely responded to hypoxic stimuli. A transient transfection experiment demonstrated that HIF-1alpha mediates IPAS promoter activation via the functional hypoxia response element under hypoxic conditions and that a constitutively active form of HIF-1alpha is sufficient for induction of the promoter in normoxic cells. Moreover, chromatin immunoprecipitation and electrophoretic mobility shift assays showed binding of the HIF-1 complex to the element in a hypoxia-dependent manner. Taken together, HIF-1 directly up-regulates IPAS gene expression through a mechanism distinct from RNA splicing, providing a further level of negative feedback gene regulation in adaptive responses to hypoxic/ischemic conditions. PMID:17355974

  19. Development of disease-resistant rice using regulatory components of induced disease resistance

    PubMed Central

    Takatsuji, Hiroshi

    2014-01-01

    Infectious diseases cause huge crop losses annually. In response to pathogen attacks, plants activate defense systems that are mediated through various signaling pathways. The salicylic acid (SA) signaling pathway is the most powerful of these pathways. Several regulatory components of the SA signaling pathway have been identified, and are potential targets for genetic manipulation of plants’ disease resistance. However, the resistance associated with these regulatory components is often accompanied by fitness costs; that is, negative effects on plant growth and crop yield. Chemical defense inducers, such as benzothiadiazole and probenazole, act on the SA pathway and induce strong resistance to various pathogens without major fitness costs, owing to their ‘priming effect.’ Studies on how benzothiadiazole induces disease resistance in rice have identified WRKY45, a key transcription factor in the branched SA pathway, and OsNPR1/NH1. Rice plants overexpressing WRKY45 were extremely resistant to rice blast disease caused by the fungus Magnaporthe oryzae and bacterial leaf blight disease caused by Xanthomonas oryzae pv. oryzae (Xoo), the two major rice diseases. Disease resistance is often accompanied by fitness costs; however, WRKY45 overexpression imposed relatively small fitness costs on rice because of its priming effect. This priming effect was similar to that of chemical defense inducers, although the fitness costs were amplified by some environmental factors. WRKY45 is degraded by the ubiquitin–proteasome system, and the dual role of this degradation partly explains the priming effect. The synergistic interaction between SA and cytokinin signaling that activates WRKY45 also likely contributes to the priming effect. With a main focus on these studies, I review the current knowledge of SA-pathway-dependent defense in rice by comparing it with that in Arabidopsis, and discuss potential strategies to develop disease-resistant rice using signaling components

  20. Antiviral factors and type I/III interferon expression associated with regulatory factors in the oral epithelial cells from HIV-1-serodiscordant couples

    PubMed Central

    Cervantes, Cesar A. C.; Oliveira, Luanda M. S.; Manfrere, Kelly C. G.; Lima, Josenilson F.; Pereira, Natalli Z.; Duarte, Alberto J. S.; Sato, Maria N.

    2016-01-01

    Individuals who remain HIV-seronegative despite repeated unprotected exposure to the virus are defined as exposed seronegative (ESN) individuals. Innate and adaptive immunity, as well as genetic factors, provide ESNs with important advantages that allow for low infection susceptibility. The majority of HIV-1-infected individuals undergo antiretroviral therapy, which can decrease the level of HIV-1 exposure in ESNs. We analyzed type I interferon (IFN)-related antiviral and regulatory factors in peripheral blood mononuclear cells (PBMCs) and oral epithelial cells from serodiscordant couples. Our findings revealed that ESNs did not induce the expression of antiviral factors (APOBEC-3G, TRIM5-α, SAMDH1, STING, TBk1) or regulatory factors (Trex, Foxo3, Socs3, IL-10) in PBMCs, unlike their HIV-1-infected partners. In contrast, ESNs upregulated APOBEC-3G and type I/III IFNs (IFNs-α,-β/-λ) in oral mucosal epithelial cells similar to their HIV-infected partners. The serodiscordant groups exhibited an increased expression of type I IFN-induced regulators, such as Trex and Foxo3, in oral epithelial cells. TLR7, TLR8 and TLR9 were expressed in oral epithelial cells of both ESNs and HIV-1-infected subjects. These findings revealed evidence of antiviral factors, type I/III interferon and regulatory factor expression only in the oral mucosal compartment of ESNs, while HIV-1-infected partners systemically and oral mucosal expressed the antiviral profile. PMID:27168019

  1. Antiviral Activity of Porcine Interferon Regulatory Factor 1 against Swine Viruses in Cell Culture.

    PubMed

    Li, Yongtao; Chang, Hongtao; Yang, Xia; Zhao, Yongxiang; Chen, Lu; Wang, Xinwei; Liu, Hongying; Wang, Chuanqing; Zhao, Jun

    2015-11-17

    Interferon regulatory factor 1 (IRF1), as an important transcription factor, is abundantly induced upon virus infections and participates in host antiviral immune responses. However, the roles of porcine IRF1 (poIRF1) in host antiviral defense remain poorly understood. In this study, we determined that poIRF1 was upregulated upon infection with viruses and distributed in nucleus in porcine PK-15 cells. Subsequently, we tested the antiviral activities of poIRF1 against several swine viruses in cells. Overexpression of poIRF1 can efficiently suppress the replication of viruses, and knockdown of poIRF1 promotes moderately viral replication. Interestingly, overexpression of poIRF1 enhances dsRNA-induced IFN-β and IFN-stimulated response element (ISRE) promoter activation, whereas knockdown of poIRF1 cannot significantly affect the activation of IFN-β promoter induced by RNA viruses. This study suggests that poIRF1 plays a significant role in cellular antiviral response against swine viruses, but might be dispensable for IFN-β induction triggered by RNA viruses in PK-15 cells. Given these results, poIRF1 plays potential roles in cellular antiviral responses against swine viruses.

  2. Multiple signaling pathways leading to the activation of interferon regulatory factor 3.

    PubMed

    Servant, Marc J; Grandvaux, Nathalie; Hiscott, John

    2002-09-01

    Virus infection of susceptible cells activates multiple signaling pathways that orchestrate the activation of genes, such as cytokines, involved in the antiviral and innate immune response. Among the kinases induced are the mitogen-activated protein (MAP) kinases, Jun-amino terminal kinases (JNK) and p38, the IkappaB kinase (IKK) and DNA-PK. In addition, virus infection also activates an uncharacterized VAK responsible for the C-terminal phosphorylation and subsequent activation of interferon regulatory factor 3 (IRF-3). Virus-mediated activation of IRF-3 through VAK is dependent on viral entry and transcription, since replication deficient virus failed to induce IRF-3 activity. The pathways leading to VAK activation are not well characterized, but IRF-3 appears to represent a novel cellular detection pathway that recognizes viral nucleocapsid (N) structure. Recently, the range of inducers responsible for IRF-3 activation has increased. In addition to virus infection, recognition of bacterial infection mediated through lipopolysaccharide by Toll-like receptor 4 has also been reported. Furthermore, MAP kinase kinase kinase (MAP KKK)-related pathways and DNA-PK induce N-terminal phosphorylation of IRF-3. This review summarizes recent observations in the identification of novel signaling pathways leading to IRF-3 activation.

  3. Identification of Neurodegenerative Factors Using Translatome-Regulatory Network Analysis

    PubMed Central

    Brichta, Lars; Shin, William; Jackson-Lewis, Vernice; Blesa, Javier; Yap, Ee-Lynn; Walker, Zachary; Zhang, Jack; Roussarie, Jean-Pierre; Alvarez, Mariano J.; Califano, Andrea; Przedborski, Serge; Greengard, Paul

    2016-01-01

    For degenerative disorders of the central nervous system, the major obstacle to therapeutic advancement has been the challenge of identifying the key molecular mechanisms underlying neuronal loss. We developed a combinatorial approach including translational profiling and brain regulatory network analysis to search for key determinants of neuronal survival or death. Following the generation of transgenic mice for cell type-specific profiling of midbrain dopaminergic neurons, we established and compared translatome libraries reflecting the molecular signature of these cells at baseline or under degenerative stress. Analysis of these libraries by interrogating a context-specific brain regulatory network led to the identification of a repertoire of intrinsic upstream regulators that drive the dopaminergic stress response. The altered activity of these regulators was not associated with changes in their expression levels. This strategy can be generalized for the elucidation of novel molecular determinants involved in the degeneration of other classes of neurons. PMID:26214373

  4. Ex Vivo Induced Regulatory Human/Murine Mesenchymal Stem Cells as Immune Modulators.

    PubMed

    Hinden, Liad; Shainer, Reut; Almogi-Hazan, Osnat; Or, Reuven

    2015-07-01

    Over the past decade there has been a growing interest in using mesenchymal stem cells (MSCs) as an immune-regulatory agent for prevention and treatment of various immune disorders including graft-versus-host disease (GVHD), transplanted organ rejection, and autoimmune diseases. However, the high diversity in the results from clinical trials using MSCs for such disorders emphasizes the need for MSCs to be "professionalized" ex vivo to a more defined regulatory phenotype before administering to patients. To this aim, we have established an ex vivo immunomodulatory triple combination treatment (TCT) for MSCs, using IFNγ, TGFβ, and kynurenine. We show that pretreated MSCs acquire an immunomodulatory phenotype, have improved regulatory functions, and upregulate the expression of inducible nitric oxide synthase, indoleamine 2,3-dioxygenase, cyclooxygenase-2 (COX2), heme oxygenase 1, leukemia inhibitory factor (LIF), and programmed death ligand 1. We define the pathway of kynurenine induced aryl hydrocarbon receptor activation in MSCs and how it contributes to the upregulation of COX2 expression and IL-6 downregulation. The combination of reduced IL-6 secretion with enhanced LIF expression leads to the inhibition of Th17 differentiation in coculture of TCT MSCs and lymphocytes. To test the immunomodulatory function of TCT MSCs in vivo, we used the cells as GVHD prophylaxis in a GVHD mouse model. TCT MSCs administration significantly decreased GVHD score and improved mouse survival. Importantly, single administration could attenuate disease symptoms for more than 3 weeks. Based on these results, we suggest considering TCT MSCs as an improved cell therapy for systemic diseases with an underlying inflammatory and immunologic etiology. Stem Cells 2015;33:2256-2267.

  5. VEGFA/VEGFR2-targeted therapies prevent the VEGFA-induced proliferation of regulatory T cells in cancer

    PubMed Central

    Terme, Magali; Tartour, Eric; Taieb, Julien

    2013-01-01

    Some of the anti-angiogenic agents currently used to treat solid malignancies have effects on tumor endothelial cells as well as on immune cells. We have recently demonstrated that targeting the vascular endothelial growth factor A (VEGFA)/VEGF receptor 2 (VEGFR2) signaling pathway reduces the proportion of regulatory T cells (Treg) in a mouse model of colorectal cancer (CRC) and in metastatic CRC patients as it inhibits tumor-induced Treg proliferation. PMID:24083078

  6. MicroRNA and transcription factor mediated regulatory network for ovarian cancer: regulatory network of ovarian cancer.

    PubMed

    Ying, Huanchun; Lv, Jing; Ying, Tianshu; Li, Jun; Yang, Qing; Ma, Yuan

    2013-10-01

    A better understanding on the regulatory interactions of microRNA (miRNA) target genes and transcription factor (TF) target genes in ovarian cancer may be conducive for developing early diagnosis strategy. Thus, gene expression data and miRNA expression data were downloaded from The Cancer Genome Atlas in this study. Differentially expressed genes and miRNAs were selected out with t test, and Gene Ontology enrichment analysis was performed with DAVID tools. Regulatory interactions were retrieved from miRTarBase, TRED, and TRANSFAC, and then networks for miRNA target genes and TF target genes were constructed to globally present the mechanisms. As a result, a total of 1,939 differentially expressed genes were identified, and they were enriched in 28 functions, among which cell cycle was affected to the most degree. Besides, 213 differentially expressed miRNAs were identified. Two regulatory networks for miRNA target genes and TF target genes were established and then both were combined, in which E2F transcription factor 1, cyclin-dependent kinase inhibitor 1A, cyclin E1, and miR-16 were the hub genes. These genes may be potential biomarkers for ovarian cancer.

  7. Effect of Adoptive Transfer or Depletion of Regulatory T Cells on Triptolide-induced Liver Injury

    PubMed Central

    Wang, Xinzhi; Sun, Lixin; Zhang, Luyong; Jiang, Zhenzhou

    2016-01-01

    Objective: The aim of this study is to clarify the role of regulatory T cell (Treg) in triptolide (TP)-induced hepatotoxicity. Methods: Female C57BL/6 mice received either adoptive transfer of Tregs or depletion of Tregs, then underwent TP administration and were sacrificed 24 h after TP administration. Liver injury was determined according to alanine transaminase (ALT) and aspartate transaminase (AST) levels in serum and histopathological change in liver tissue. Hepatic frequencies of Treg cells and the mRNA expression levels of transcription factor Forkhead box P3 and retinoid orphan nuclear receptor γt (RORγt), interleukin-10 (IL-10), suppressor of cytokine signaling (SOCS), and Notch/Notch ligand were investigated. Results: During TP-induced liver injury, hepatic Treg and IL-10 decreased, while T helper 17 cells cell-transcription factor RORγt, SOCS and Notch signaling increased, accompanied with liver inflammation. Adoptive transfer of Tregs ameliorated the severity of TP-induced liver injury, accompanied with increased levels of hepatic Treg and IL-10. Adoptive transfer of Tregs remarkably inhibited the expression of RORγt, SOCS3, Notch1, and Notch3. On the contrary, depletion of Treg cells in TP-administered mice resulted in a notable increase of RORγt, SOCS1, SOCS3, and Notch3, while the Treg and IL-10 of liver decreased. Consistent with the exacerbation of liver injury, higher serum levels of ALT and AST were detected in Treg-depleted mice. Conclusion: These results showed that adoptive transfer or depletion of Tregs attenuated or aggravated TP-induced liver injury, suggesting that Tregs could play important roles in the progression of liver injury. SOCS proteins and Notch signaling affected Tregs, which may contribute to the pathogenesis of TP-induced hepatotoxicity. PMID:27148057

  8. Mangiferin inhibits macrophage classical activation via downregulating interferon regulatory factor 5 expression.

    PubMed

    Wei, Zhiquan; Yan, Li; Chen, Yixin; Bao, Chuanhong; Deng, Jing; Deng, Jiagang

    2016-08-01

    Mangiferin is a natural polyphenol and the predominant effective component of Mangifera indica Linn. leaves. For hundreds of years, Mangifera indica Linn. leaf has been used as an ingredient in numerous traditional Chinese medicine preparations for the treatment of bronchitis. However, the pharmacological mechanism of mangiferin in the treatment of bronchitis remains to be elucidated. Macrophage classical activation is important role in the process of bronchial airway inflammation, and interferon regulatory factor 5 (IRF5) has been identified as a key regulatory factor for macrophage classical activation. The present study used the THP‑1 human monocyte cell line to investigate whether mangiferin inhibits macrophage classical activation via suppressing IRF5 expression in vitro. THP‑1 cells were differentiated to macrophages by phorbol 12‑myristate 13‑acetate. Macrophages were polarized to M1 macrophages following stimulation with lipopolysaccharide (LPS)/interferon‑γ (IFN‑γ). Flow cytometric analysis was conducted to detect the M1 macrophages. Reverse transcription‑quantitative polymerase chain reaction was used to investigate cellular IRF5 gene expression. Levels of proinflammatory cytokines and IRF5 were assessed following cell culture and cellular homogenization using enzyme‑linked immunosorbent assay. IRF5 protein and nuclei co‑localization was performed in macrophages with laser scanning confocal microscope immunofluorescence analysis. The results of the present study demonstrated that mangiferin significantly inhibits LPS/IFN‑γ stimulation‑induced classical activation of macrophages in vitro and markedly decreases proinflammatory cytokine release. In addition, cellular IRF5 expression was markedly downregulated. These results suggest that the inhibitory effect of mangiferin on classical activation of macrophages may be exerted via downregulation of cellular IRF5 expression levels. PMID:27277156

  9. Mangiferin inhibits macrophage classical activation via downregulating interferon regulatory factor 5 expression

    PubMed Central

    Wei, Zhiquan; Yan, Li; Chen, Yixin; Bao, Chuanhong; Deng, Jing; Deng, Jiagang

    2016-01-01

    Mangiferin is a natural polyphenol and the predominant effective component of Mangifera indica Linn. leaves. For hundreds of years, Mangifera indica Linn. leaf has been used as an ingredient in numerous traditional Chinese medicine preparations for the treatment of bronchitis. However, the pharmacological mechanism of mangiferin in the treatment of bronchitis remains to be elucidated. Macrophage classical activation is important role in the process of bronchial airway inflammation, and interferon regulatory factor 5 (IRF5) has been identified as a key regulatory factor for macrophage classical activation. The present study used the THP-1 human monocyte cell line to investigate whether mangiferin inhibits macrophage classical activation via suppressing IRF5 expression in vitro. THP-1 cells were differentiated to macrophages by phorbol 12-myristate 13-acetate. Macrophages were polarized to M1 macrophages following stimulation with lipopolysaccharide (LPS)/interferon-γ (IFN-γ). Flow cytometric analysis was conducted to detect the M1 macrophages. Reverse transcription-quantitative polymerase chain reaction was used to investigate cellular IRF5 gene expression. Levels of proinflammatory cytokines and IRF5 were assessed following cell culture and cellular homogenization using enzyme-linked immunosorbent assay. IRF5 protein and nuclei co-localization was performed in macrophages with laser scanning confocal microscope immunofluorescence analysis. The results of the present study demonstrated that mangiferin significantly inhibits LPS/IFN-γ stimulation-induced classical activation of macrophages in vitro and markedly decreases proinflammatory cytokine release. In addition, cellular IRF5 expression was markedly downregulated. These results suggest that the inhibitory effect of mangiferin on classical activation of macrophages may be exerted via downregulation of cellular IRF5 expression levels. PMID:27277156

  10. Integrated Response to Inducers by Communication between a Catabolic Pathway and Its Regulatory System▿

    PubMed Central

    Martínez-Pérez, Olga; López-Sánchez, Aroa; Reyes-Ramírez, Francisca; Floriano, Belén; Santero, Eduardo

    2007-01-01

    Efficient gene regulation of metabolic pathways implies that the profile of molecules inducing the pathway matches that of the molecules that are metabolized. Gratuitous induction, a well-known phenomenon in catabolic pathways, is the consequence of differences in the substrate and inducer profiles. This phenomenon is particularly evident in pathways for biodegradation of organic contaminants that can be induced by a variety of molecules similar to the real substrates. Analysis of the regulation of tetralin biodegradation genes in mutant strains with mutations that affect each component of the initial dioxygenase enzymatic complex indicated that the response of the regulatory system to potential inducers is altered differently depending on the mutated component. Based on the expression phenotypes of a number of single or double mutants, we propose a model that represents an unprecedented way of communication between a catabolic pathway and its regulatory system to prevent efficient induction by a molecule that is not a real substrate. This communication allows a better fit of the substrate and inducer profiles, thus minimizing gratuitous induction, without a requirement for optimal coevolution to match the specificity of catabolic enzymes and their regulatory systems. Modulation of the regulatory system in this way not only provides a more appropriate response to potential inducers recognized by the regulatory system but also may properly adjust the levels of gene expression to the substrate availability. PMID:17351041

  11. Role of cellular caspases, nuclear factor-kappa B and interferon regulatory factors in Bluetongue virus infection and cell fate

    PubMed Central

    2010-01-01

    Background Bluetongue virus (BTV) infection causes haemorrhagic disease in ruminants and induces cell death. The pathogenesis in animals and in cell culture has been linked to BTV-induced apoptosis. Results In this report, we investigated BTV-induced apoptosis in cell culture in depth and show that both extrinsic (caspase-8 activation) and intrinsic (caspase-9 activation) pathways play roles in BTV apoptosis. Further, by using chemical inhibitors and knock-out cell lines, we show that these pathways act independently of each other in BTV infected cells. In addition to activation of caspase-8, -9 and executioner caspase-3, we also identified that BTV infection causes the activation of caspase-7, which results in the cleavage of poly (ADP-ribose) polymerase (PARP). BTV-induced cell death appears to be due to apoptosis rather than necrosis, as the HMBG-1 was not translocated from the nucleus. We also examined if NF-κB response is related to BTV-induced apoptosis as in reovirus. Our data suggests that NF-κB response is not linked to the induction of apoptosis. It is controlled by the degradation of only IκBα but not IκBβ, resulting in a rapid transient response during BTV infection. This was supported using an NF-κB dependent luciferase reporter gene assay, which demonstrated early response, that appeared to be suppressed by the late stage of BTV replication. Furthermore, virus titres were higher in the presence of NF-κB inhibitor (SN50), indicating that NF-κB has a role in initiating an antiviral environment. In addition, we show that BTV infection induces the translocation of interferon regulatory factors (IRF-3 and IRF-7) into the nucleus. The induction of IRF responses, when measured by IRF dependent luciferase reporter gene assay, revealed that the IRF responses, like NF-κB response, were also at early stage of infection and mirrored the timing of NF-κB induction. Conclusion BTV triggers a wide range of caspase activities resulting in cell apoptosis

  12. Environmental Factors Inducing Human Cancers

    PubMed Central

    Parsa, N

    2012-01-01

    Background An explosion of research has been done in discovering how human health is affected by environmental factors. I will discuss the impacts of environmental cancer causing factors and how they continue to cause multiple disruptions in cellular networking. Some risk factors may not cause cancer. Other factors initiate consecutive genetic mutations that would eventually alter the normal pathway of cellular proliferations and differentiation. Genetic mutations in four groups of genes; (Oncogenes, Tumor suppressor genes, Apoptosis genes and DNA repairing genes) play a vital role in altering the normal cell division. In recent years, molecular genetics have greatly increased our understanding of the basic mechanisms in cancer development and utilizing these molecular techniques for cancer screening, diagnosis, prognosis and therapies. Inhibition of carcinogenic exposures wherever possible should be the goal of cancer prevention programs to reduce exposures from all environmental carcinogens. PMID:23304670

  13. Effect of heparin on bovine epithelial lens cell proliferation induced by heparin affin regulatory peptide.

    PubMed

    Delbé, J; Vacherot, F; Laaroubi, K; Barritault, D; Courty, J

    1995-07-01

    HARP (heparin affin regulatory peptide) is an 18 kDa heparin binding protein, also known as HB-GAM or pleiotrophin (PTN) which has been primarily isolated from brain and uterus, and displays neurite outgrowth, angiogenic and mitogenic activities. Previously, we have expressed the human cDNA encoding human HARP in NIH 3T3 cells. Purified recombinant HARP displayed mitogenic activity for endothelial cells. Its NH2-terminal sequence indicates that the HARP molecule possesses a three amino acid extension from the signal peptide more than the NH2-terminal described. For HB-GAM or PTN, these three amino acids may be essential for the stability and the mitogenic activity of this growth factor. In an attempt to further study the mode of action of this growth factor, we have investigated the mitogenic effect of HARP on various cell types. In contrast to FGF-2, HARP failed to induce stimulation of DNA synthesis on a CCL39 cell line. However, we found that in quiescent bovine epithelial lens (BEL) cells, the stimulation of DNA synthesis induced by HARP is dose-dependent (EC50: 2.5 ng/ml) and maximal stimulation is as potent as that induced by FGF-2 (EC50: 25 pg/ml). Interestingly, when BEL cells were allowed to quiesce in the presence of serum, the stimulation induced by HARP is considerably less potent. In this highly responsive cell system, heparin could potentiate the mitogenic activity of HARP at very low doses (0.1-1 microgram/ml) and inhibit this activity at concentrations of 10 micrograms/ml. In contrast to its protective effect on FGF-1 and -2, heparin was unable to preserve HARP from tryptic and chymotryptic degradations.

  14. Inflammation-inducing Factors of Mycoplasma pneumoniae

    PubMed Central

    Shimizu, Takashi

    2016-01-01

    Mycoplasma pneumoniae, which causes mycoplasmal pneumonia in human, mainly causes pneumonia in children, although it occasionally causes disease in infants and geriatrics. Some pathogenic factors produced by M. pneumoniae, such as hydrogen peroxide and Community-Acquired Respiratory Distress Syndrome (CARDS) toxin have been well studied. However, these factors alone cannot explain this predilection. The low incidence rate of mycoplasmal pneumonia in infants and geriatrics implies that the strong inflammatory responses induced by M. pneumoniae coordinate with the pathogenic factors to induce pneumonia. However, M. pneumoniae lacks a cell wall and does not possess an inflammation-inducing endotoxin, such as lipopolysaccharide (LPS). In M. pneumoniae, lipoproteins were identified as an inflammation-inducing factor. Lipoproteins induce inflammatory responses through Toll-like receptors (TLR) 2. Because Mycoplasma species lack a cell wall and lipoproteins anchored in the membrane are exposed, lipoproteins and TLR2 have been thought to be important for the pathogenesis of M. pneumoniae. However, recent reports suggest that M. pneumoniae also induces inflammatory responses also in a TLR2-independent manner. TLR4 and autophagy are involved in this TLR2-independent inflammation. In addition, the CARDS toxin or M. pneumoniae cytadherence induces inflammatory responses through an intracellular receptor protein complex called the inflammasome. In this review, the inflammation-inducing factors of M. pneumoniae are summarized. PMID:27065977

  15. Hypoxia-Inducible Factor as an Angiogenic Master Switch

    PubMed Central

    Hashimoto, Takuya; Shibasaki, Futoshi

    2015-01-01

    Hypoxia-inducible factors (HIFs) regulate the transcription of genes that mediate the response to hypoxia. HIFs are constantly expressed and degraded under normoxia, but stabilized under hypoxia. HIFs have been widely studied in physiological and pathological conditions and have been shown to contribute to the pathogenesis of various vascular diseases. In clinical settings, the HIF pathway has been studied for its role in inhibiting carcinogenesis. HIFs might also play a protective role in the pathology of ischemic diseases. Clinical trials of therapeutic angiogenesis after the administration of a single growth factor have yielded unsatisfactory or controversial results, possibly because the coordinated activity of different HIF-induced factors is necessary to induce mature vessel formation. Thus, manipulation of HIF activity to simultaneously induce a spectrum of angiogenic factors offers a superior strategy for therapeutic angiogenesis. Because HIF-2α plays an essential role in vascular remodeling, manipulation of HIF-2α is a promising approach to the treatment of ischemic diseases caused by arterial obstruction, where insufficient development of collateral vessels impedes effective therapy. Eukaryotic initiation factor 3 subunit e (eIF3e)/INT6 interacts specifically with HIF-2α and induces the proteasome inhibitor-sensitive degradation of HIF-2α, independent of hypoxia and von Hippel-Lindau protein. Treatment with eIF3e/INT6 siRNA stabilizes HIF-2α activity even under normoxic conditions and induces the expression of several angiogenic factors, at levels sufficient to produce functional arteries and veins in vivo. We have demonstrated that administration of eIF3e/INT6 siRNA to ischemic limbs or cold-injured brains reduces ischemic damage in animal models. This review summarizes the current understanding of the relationship between HIFs and vascular diseases. We also discuss novel oxygen-independent regulatory proteins that bind HIF-α and the implications

  16. Transcription factor binding at enhancers: shaping a genomic regulatory landscape in flux

    PubMed Central

    Palstra, Robert-Jan; Grosveld, Frank

    2012-01-01

    The mammalian genome is packed tightly in the nucleus of the cell. This packing is primarily facilitated by histone proteins and results in an ordered organization of the genome in chromosome territories that can be roughly divided in heterochromatic and euchromatic domains. On top of this organization several distinct gene regulatory elements on the same chromosome or other chromosomes are thought to dynamically communicate via chromatin looping. Advances in genome-wide technologies have revealed the existence of a plethora of these regulatory elements in various eukaryotic genomes. These regulatory elements are defined by particular in vitro assays as promoters, enhancers, insulators, and boundary elements. However, recent studies indicate that the in vivo distinction between these elements is often less strict. Regulatory elements are bound by a mixture of common and lineage-specific transcription factors which mediate the long-range interactions between these elements. Inappropriate modulation of the binding of these transcription factors can alter the interactions between regulatory elements, which in turn leads to aberrant gene expression with disease as an ultimate consequence. Here we discuss the bi-modal behavior of regulatory elements that act in cis (with a focus on enhancers), how their activity is modulated by transcription factor binding and the effect this has on gene regulation. PMID:23060900

  17. Enhancement of alkaloid production in opium and California poppy by transactivation using heterologous regulatory factors.

    PubMed

    Apuya, Nestor R; Park, Joon-Hyun; Zhang, Liping; Ahyow, Maurice; Davidow, Patricia; Van Fleet, Jennifer; Rarang, Joel C; Hippley, Matthew; Johnson, Thomas W; Yoo, Hye-Dong; Trieu, Anthony; Krueger, Shannon; Wu, Chuan-yin; Lu, Yu-ping; Flavell, Richard B; Bobzin, Steven C

    2008-02-01

    Genes encoding regulatory factors isolated from Arabidopsis, soybean and corn have been screened to identify those that modulate the expression of genes encoding for enzymes involved in the biosynthesis of morphinan alkaloids in opium poppy (Papaver somniferum) and benzophenanthridine alkaloids in California poppy (Eschscholzia californica). In opium poppy, the over-expression of selected regulatory factors increased the levels of PsCOR (codeinone reductase), Ps4'OMT (S-adenosyl-l-methionine:3'-hydroxy-N-methylcoclaurine 4'-O-methyltransferase) and Ps6OMT [(R,S)-norcoclaurine 6-O-methyltransferase] transcripts by 10- to more than 100-fold. These transcriptional activations translated into an enhancement of alkaloid production in opium poppy of up to at least 10-fold. In California poppy, the transactivation effect of regulatory factor WRKY1 resulted in an increase of up to 60-fold in the level of EcCYP80B1 [(S)-N-methylcoclaurine 3'-hydroxylase] and EcBBE (berberine bridge enzyme) transcripts. As a result, the accumulations of selected alkaloid intermediates were enhanced up to 30-fold. The transactivation effects of other regulatory factors led to the accumulation of the same intermediates. These regulatory factors also led to the production of new alkaloids in California poppy callus culture.

  18. Enhancement of alkaloid production in opium and California poppy by transactivation using heterologous regulatory factors.

    PubMed

    Apuya, Nestor R; Park, Joon-Hyun; Zhang, Liping; Ahyow, Maurice; Davidow, Patricia; Van Fleet, Jennifer; Rarang, Joel C; Hippley, Matthew; Johnson, Thomas W; Yoo, Hye-Dong; Trieu, Anthony; Krueger, Shannon; Wu, Chuan-yin; Lu, Yu-ping; Flavell, Richard B; Bobzin, Steven C

    2008-02-01

    Genes encoding regulatory factors isolated from Arabidopsis, soybean and corn have been screened to identify those that modulate the expression of genes encoding for enzymes involved in the biosynthesis of morphinan alkaloids in opium poppy (Papaver somniferum) and benzophenanthridine alkaloids in California poppy (Eschscholzia californica). In opium poppy, the over-expression of selected regulatory factors increased the levels of PsCOR (codeinone reductase), Ps4'OMT (S-adenosyl-l-methionine:3'-hydroxy-N-methylcoclaurine 4'-O-methyltransferase) and Ps6OMT [(R,S)-norcoclaurine 6-O-methyltransferase] transcripts by 10- to more than 100-fold. These transcriptional activations translated into an enhancement of alkaloid production in opium poppy of up to at least 10-fold. In California poppy, the transactivation effect of regulatory factor WRKY1 resulted in an increase of up to 60-fold in the level of EcCYP80B1 [(S)-N-methylcoclaurine 3'-hydroxylase] and EcBBE (berberine bridge enzyme) transcripts. As a result, the accumulations of selected alkaloid intermediates were enhanced up to 30-fold. The transactivation effects of other regulatory factors led to the accumulation of the same intermediates. These regulatory factors also led to the production of new alkaloids in California poppy callus culture. PMID:17961129

  19. Gene Regulatory Network Inference of Immunoresponsive Gene 1 (IRG1) Identifies Interferon Regulatory Factor 1 (IRF1) as Its Transcriptional Regulator in Mammalian Macrophages

    PubMed Central

    Tallam, Aravind; Perumal, Thaneer M.; Antony, Paul M.; Jäger, Christian; Fritz, Joëlle V.; Vallar, Laurent; Balling, Rudi; del Sol, Antonio; Michelucci, Alessandro

    2016-01-01

    Immunoresponsive gene 1 (IRG1) is one of the highest induced genes in macrophages under pro-inflammatory conditions. Its function has been recently described: it codes for immune-responsive gene 1 protein/cis-aconitic acid decarboxylase (IRG1/CAD), an enzyme catalysing the production of itaconic acid from cis-aconitic acid, a tricarboxylic acid (TCA) cycle intermediate. Itaconic acid possesses specific antimicrobial properties inhibiting isocitrate lyase, the first enzyme of the glyoxylate shunt, an anaplerotic pathway that bypasses the TCA cycle and enables bacteria to survive on limited carbon conditions. To elucidate the mechanisms underlying itaconic acid production through IRG1 induction in macrophages, we examined the transcriptional regulation of IRG1. To this end, we studied IRG1 expression in human immune cells under different inflammatory stimuli, such as TNFα and IFNγ, in addition to lipopolysaccharides. Under these conditions, as previously shown in mouse macrophages, IRG1/CAD accumulates in mitochondria. Furthermore, using literature information and transcription factor prediction models, we re-constructed raw gene regulatory networks (GRNs) for IRG1 in mouse and human macrophages. We further implemented a contextualization algorithm that relies on genome-wide gene expression data to infer putative cell type-specific gene regulatory interactions in mouse and human macrophages, which allowed us to predict potential transcriptional regulators of IRG1. Among the computationally identified regulators, siRNA-mediated gene silencing of interferon regulatory factor 1 (IRF1) in macrophages significantly decreased the expression of IRG1/CAD at the gene and protein level, which correlated with a reduced production of itaconic acid. Using a synergistic approach of both computational and experimental methods, we here shed more light on the transcriptional machinery of IRG1 expression and could pave the way to therapeutic approaches targeting itaconic acid levels

  20. Reconstruction and topological characterization of the sigma factor regulatory network of Mycobacterium tuberculosis.

    PubMed

    Chauhan, Rinki; Ravi, Janani; Datta, Pratik; Chen, Tianlong; Schnappinger, Dirk; Bassler, Kevin E; Balázsi, Gábor; Gennaro, Maria Laura

    2016-03-31

    Accessory sigma factors, which reprogram RNA polymerase to transcribe specific gene sets, activate bacterial adaptive responses to noxious environments. Here we reconstruct the complete sigma factor regulatory network of the human pathogen Mycobacterium tuberculosis by an integrated approach. The approach combines identification of direct regulatory interactions between M. tuberculosis sigma factors in an E. coli model system, validation of selected links in M. tuberculosis, and extensive literature review. The resulting network comprises 41 direct interactions among all 13 sigma factors. Analysis of network topology reveals (i) a three-tiered hierarchy initiating at master regulators, (ii) high connectivity and (iii) distinct communities containing multiple sigma factors. These topological features are likely associated with multi-layer signal processing and specialized stress responses involving multiple sigma factors. Moreover, the identification of overrepresented network motifs, such as autoregulation and coregulation of sigma and anti-sigma factor pairs, provides structural information that is relevant for studies of network dynamics.

  1. Reconstruction and topological characterization of the sigma factor regulatory network of Mycobacterium tuberculosis

    PubMed Central

    Chauhan, Rinki; Ravi, Janani; Datta, Pratik; Chen, Tianlong; Schnappinger, Dirk; Bassler, Kevin E.; Balázsi, Gábor; Gennaro, Maria Laura

    2016-01-01

    Accessory sigma factors, which reprogram RNA polymerase to transcribe specific gene sets, activate bacterial adaptive responses to noxious environments. Here we reconstruct the complete sigma factor regulatory network of the human pathogen Mycobacterium tuberculosis by an integrated approach. The approach combines identification of direct regulatory interactions between M. tuberculosis sigma factors in an E. coli model system, validation of selected links in M. tuberculosis, and extensive literature review. The resulting network comprises 41 direct interactions among all 13 sigma factors. Analysis of network topology reveals (i) a three-tiered hierarchy initiating at master regulators, (ii) high connectivity and (iii) distinct communities containing multiple sigma factors. These topological features are likely associated with multi-layer signal processing and specialized stress responses involving multiple sigma factors. Moreover, the identification of overrepresented network motifs, such as autoregulation and coregulation of sigma and anti-sigma factor pairs, provides structural information that is relevant for studies of network dynamics. PMID:27029515

  2. Interferon-Regulatory Factor 5-Dependent Signaling Restricts Orthobunyavirus Dissemination to the Central Nervous System

    PubMed Central

    Proenca-Modena, Jose Luiz; Hyde, Jennifer L.; Sesti-Costa, Renata; Lucas, Tiffany; Pinto, Amelia K.; Richner, Justin M.; Gorman, Matthew J.; Lazear, Helen M.

    2015-01-01

    ABSTRACT Interferon (IFN)-regulatory factor 5 (IRF-5) is a transcription factor that induces inflammatory responses after engagement and signaling by pattern recognition receptors. To define the role of IRF-5 during bunyavirus infection, we evaluated Oropouche virus (OROV) and La Crosse virus (LACV) pathogenesis and immune responses in primary cells and in mice with gene deletions in Irf3, Irf5, and Irf7 or in Irf5 alone. Deletion of Irf3, Irf5, and Irf7 together resulted in uncontrolled viral replication in the liver and spleen, hypercytokinemia, extensive liver injury, and an early-death phenotype. Remarkably, deletion of Irf5 alone resulted in meningoencephalitis and death on a more protracted timeline, 1 to 2 weeks after initial OROV or LACV infection. The clinical signs in OROV-infected Irf5−/− mice were associated with abundant viral antigen and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL)-positive cells in several regions of the brain. Circulating dendritic cell (DC) subsets in Irf5−/− mice had higher levels of OROV RNA in vivo yet produced lower levels of type I IFN than wild-type (WT) cells. This result was supported by data obtained in vitro, since a deficiency of IRF-5 resulted in enhanced OROV infection and diminished type I IFN production in bone marrow-derived DCs. Collectively, these results indicate a key role for IRF-5 in modulating the host antiviral response in peripheral organs that controls bunyavirus neuroinvasion in mice. IMPORTANCE Oropouche virus (OROV) and La Crosse virus (LACV) are orthobunyaviruses that are transmitted by insects and cause meningitis and encephalitis in subsets of individuals in the Americas. Recently, we demonstrated that components of the type I interferon (IFN) induction pathway, particularly the regulatory transcription factors IRF-3 and IRF-7, have key protective roles during OROV infection. However, the lethality in Irf3−/− Irf7−/− (DKO) mice infected with OROV

  3. Duplication of a promiscuous transcription factor drives the emergence of a new regulatory network.

    PubMed

    Pougach, Ksenia; Voet, Arnout; Kondrashov, Fyodor A; Voordeckers, Karin; Christiaens, Joaquin F; Baying, Bianka; Benes, Vladimir; Sakai, Ryo; Aerts, Jan; Zhu, Bo; Van Dijck, Patrick; Verstrepen, Kevin J

    2014-01-01

    The emergence of new genes throughout evolution requires rewiring and extension of regulatory networks. However, the molecular details of how the transcriptional regulation of new gene copies evolves remain largely unexplored. Here we show how duplication of a transcription factor gene allowed the emergence of two independent regulatory circuits. Interestingly, the ancestral transcription factor was promiscuous and could bind different motifs in its target promoters. After duplication, one paralogue evolved increased binding specificity so that it only binds one type of motif, whereas the other copy evolved a decreased activity so that it only activates promoters that contain multiple binding sites. Interestingly, only a few mutations in both the DNA-binding domains and in the promoter binding sites were required to gradually disentangle the two networks. These results reveal how duplication of a promiscuous transcription factor followed by concerted cis and trans mutations allows expansion of a regulatory network.

  4. Duplication of a promiscuous transcription factor drives the emergence of a new regulatory network

    PubMed Central

    Pougach, Ksenia; Voet, Arnout; Kondrashov, Fyodor A.; Voordeckers, Karin; Christiaens, Joaquin F.; Baying, Bianka; Benes, Vladimir; Sakai, Ryo; Aerts, Jan; Zhu, Bo; Van Dijck, Patrick; Verstrepen, Kevin J.

    2014-01-01

    The emergence of new genes throughout evolution requires rewiring and extension of regulatory networks. However, the molecular details of how the transcriptional regulation of new gene copies evolves remain largely unexplored. Here we show how duplication of a transcription factor gene allowed the emergence of two independent regulatory circuits. Interestingly, the ancestral transcription factor was promiscuous and could bind different motifs in its target promoters. After duplication, one paralogue evolved increased binding specificity so that it only binds one type of motif, whereas the other copy evolved a decreased activity so that it only activates promoters that contain multiple binding sites. Interestingly, only a few mutations in both the DNA-binding domains and in the promoter binding sites were required to gradually disentangle the two networks. These results reveal how duplication of a promiscuous transcription factor followed by concerted cis and trans mutations allows expansion of a regulatory network. PMID:25204769

  5. Modulation of interferon regulatory factor 5 activities by the Kaposi sarcoma-associated herpesvirus-encoded viral interferon regulatory factor 3 contributes to immune evasion and lytic induction.

    PubMed

    Bi, Xiaohui; Yang, Lisong; Mancl, Margo E; Barnes, Betsy J

    2011-04-01

    Multiple Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded proteins with potential roles in KSHV-associated neoplasms have been identified. KSHV encodes 4 genes with homology to transcription factors of the interferon (IFN) regulatory factor (IRF) family. Viral IRF3 (vIRF3) is expressed in latently KSHV-infected primary effusion lymphoma (PEL) cells and was recently shown to be essential for the survival of PEL cells. The focus of this study was to determine the mechanism(s) of vIRF3 oncogenic activity contributing to KSHV-associated lymphoma. We report that vIRF3 interacts with the amino-terminal DNA binding domain of human IRF5, leading to a complex manipulation of IRF5 function. vIRF3 associated with both exogenous and endogenous IRF5, thereby inhibiting IRF5-mediated IFN promoter activation and the synthesis of biologically active type I IFNs by blocking its binding to endogenous IFNA promoters. The function of this interaction was not limited to the IFN system as IRF5-mediated cell growth regulation was significantly altered by overexpression of vIRF3 in B cells. vIRF3 prevented IRF5-mediated growth inhibition and G2/M cell cycle arrest. Important, IRF5 was upregulated by the protein kinase C agonist 12-O-tetradecanoyl-phorbol-13-acetate in BCBL1 PEL cells and interaction with vIRF3 was observed at the endogenous p21 promoter in response to 12-O-tetradecanoyl-phorbol-13-acetate, suggesting that these 2 proteins cooperate in the regulation of lytic cycle-induced G1 arrest, which is an important early step for the reactivation of KSHV. In conclusion, cellular IRF5 and vIRF3 interact, leading to the functional modulation of IRF5-mediated type I IFN expression and cell cycle regulation. These findings support an important role for vIRF3 in immune evasion and cell proliferation that likely contribute to the survival of PEL cells.

  6. Decreased Transcription Factor Binding Levels Nearby Primate Pseudogenes Suggest Regulatory Degeneration

    PubMed Central

    Douglas, Gavin M.; Wilson, Michael D.; Moses, Alan M.

    2016-01-01

    Characteristics of pseudogene degeneration at the coding level are well-known, such as a shift toward neutral rates of nonsynonymous substitutions and gain of frameshift mutations. In contrast, degeneration of pseudogene transcriptional regulation is not well understood. Here, we test two predictions of regulatory degeneration along a pseudogenized lineage: 1) Decreased transcription factor (TF) binding and 2) accelerated evolution in putative cis-regulatory regions. We find evidence for decreased TF binding levels nearby two primate pseudogenes compared with functional liver genes. However, the majority of TF-bound sequences nearby pseudogenes do not show evidence for lineage-specific accelerated rates of evolution. We conclude that decreases in TF binding level could be a marker for regulatory degeneration, while sequence degeneration in primate cis-regulatory modules may be obscured by background rates of TF binding site turnover. PMID:26882985

  7. Tumor Suppressor Interferon-Regulatory Factor 1 Counteracts the Germinal Center Reaction Driven by a Cancer-Associated Gammaherpesvirus

    PubMed Central

    Mboko, Wadzanai P.; Olteanu, Horatiu; Ray, Avijit; Xin, Gang; Darrah, Eric J.; Kumar, Suresh N.; Kulinski, Joseph M.; Cui, Weiguo; Dittel, Bonnie N.; Gauld, Stephen B.

    2015-01-01

    ABSTRACT Gammaherpesviruses are ubiquitous pathogens that are associated with the development of B cell lymphomas. Gammaherpesviruses employ multiple mechanisms to transiently stimulate a broad, polyclonal germinal center reaction, an inherently mutagenic stage of B cell differentiation that is thought to be the primary target of malignant transformation in virus-driven lymphomagenesis. We found that this gammaherpesvirus-driven germinal center expansion was exaggerated and lost its transient nature in the absence of interferon-regulatory factor 1 (IRF-1), a transcription factor with antiviral and tumor suppressor functions. Uncontrolled and persistent expansion of germinal center B cells led to pathological changes in the spleens of chronically infected IRF-1-deficient animals. Additionally, we found decreased IRF-1 expression in cases of human posttransplant lymphoproliferative disorder, a malignant condition associated with gammaherpesvirus infection. The results of our study define an unappreciated role for IRF-1 in B cell biology and provide insight into the potential mechanism of gammaherpesvirus-driven lymphomagenesis. IMPORTANCE Gammaherpesviruses establish lifelong infection in most adults and are associated with B cell lymphomas. While the infection is asymptomatic in many hosts, it is critical to identify individuals who may be at an increased risk of virus-induced cancer. Such identification is currently impossible, as the host risk factors that predispose individuals toward viral lymphomagenesis are poorly understood. The current study identifies interferon-regulatory factor 1 (IRF-1) to be one of such candidate host factors. Specifically, we found that IRF-1 enforces long-term suppression of an inherently mutagenic stage of B cell differentiation that gammaherpesviruses are thought to target for transformation. Correspondingly, in the absence of IRF-1, chronic gammaherpesvirus infection induced pathological changes in the spleens of infected animals

  8. Activation of Vago by interferon regulatory factor (IRF) suggests an interferon system-like antiviral mechanism in shrimp.

    PubMed

    Li, Chaozheng; Li, Haoyang; Chen, Yixiao; Chen, Yonggui; Wang, Sheng; Weng, Shao-Ping; Xu, Xiaopeng; He, Jianguo

    2015-01-01

    There is a debate on whether invertebrates possess an antiviral immunity similar to the interferon (IFN) system of vertebrates. The Vago gene from arthropods encodes a viral-activated secreted peptide that restricts virus infection through activating the JAK-STAT pathway and is considered to be a cytokine functionally similar to IFN. In this study, the first crustacean IFN regulatory factor (IRF)-like gene was identified in Pacific white shrimp, Litopenaeus vannamei. The L. vannamei IRF showed similar protein nature to mammalian IRFs and could be activated during virus infection. As a transcriptional regulatory factor, L. vannamei IRF could activate the IFN-stimulated response element (ISRE)-containing promoter to regulate the expression of mammalian type I IFNs and initiate an antiviral state in mammalian cells. More importantly, IRF could bind the 5'-untranslated region of L. vannamei Vago4 gene and activate its transcription, suggesting that shrimp Vago may be induced in a similar manner to that of IFNs and supporting the opinion that Vago might function as an IFN-like molecule in invertebrates. These suggested that shrimp might possess an IRF-Vago-JAK/STAT regulatory axis, which is similar to the IRF-IFN-JAK/STAT axis of vertebrates, indicating that invertebrates might possess an IFN system-like antiviral mechanism.

  9. Activation of Vago by interferon regulatory factor (IRF) suggests an interferon system-like antiviral mechanism in shrimp

    PubMed Central

    Li, Chaozheng; Li, Haoyang; Chen, Yixiao; Chen, Yonggui; Wang, Sheng; Weng, Shao-Ping; Xu, Xiaopeng; He, Jianguo

    2015-01-01

    There is a debate on whether invertebrates possess an antiviral immunity similar to the interferon (IFN) system of vertebrates. The Vago gene from arthropods encodes a viral-activated secreted peptide that restricts virus infection through activating the JAK-STAT pathway and is considered to be a cytokine functionally similar to IFN. In this study, the first crustacean IFN regulatory factor (IRF)-like gene was identified in Pacific white shrimp, Litopenaeus vannamei. The L. vannamei IRF showed similar protein nature to mammalian IRFs and could be activated during virus infection. As a transcriptional regulatory factor, L. vannamei IRF could activate the IFN-stimulated response element (ISRE)-containing promoter to regulate the expression of mammalian type I IFNs and initiate an antiviral state in mammalian cells. More importantly, IRF could bind the 5′-untranslated region of L. vannamei Vago4 gene and activate its transcription, suggesting that shrimp Vago may be induced in a similar manner to that of IFNs and supporting the opinion that Vago might function as an IFN-like molecule in invertebrates. These suggested that shrimp might possess an IRF-Vago-JAK/STAT regulatory axis, which is similar to the IRF-IFN-JAK/STAT axis of vertebrates, indicating that invertebrates might possess an IFN system-like antiviral mechanism. PMID:26459861

  10. Tissue factor activation: is disulfide bond switching a regulatory mechanism?

    PubMed Central

    Ghosh, Samit; Mandal, Samir K.

    2007-01-01

    A majority of tissue factor (TF) on cell surfaces exists in a cryptic form (ie, coagulation function inactive) but retains its functionality in cell signaling. Recent studies have suggested that cryptic TF contains unpaired cysteine thiols and that activation involves the formation of the disulfide bond Cys186-Cys 209 and that protein disulfide isomerase (PDI) regulates TF coagulant and signaling activities by targeting this disulfide bond. This study was carried out to investigate the validity of this novel concept. Although treatment of MDA 231 tumor cells, fibroblasts, and stimulated endothelial cells with the oxidizing agent HgCl2 markedly increased the cell-surface TF coagulant activity, the increase is associated with increased anionic phospholipids at the cell surface. Annexin V, which binds to anionic phospholipids, attenuated the increased TF coagulant activity. It is noteworthy that treatment of cells with reducing agents also increased the cell surface TF activity. No evidence was found for either detectable expression of PDI at the cell surface or association of TF with PDI. Furthermore, reduction of PDI with the gene silencing had no effect on either TF coagulant or cell signaling functions. Overall, the present data undermine the recently proposed hypothesis that PDI-mediated disulfide exchange plays a role in regulating TF procoagulant and cell signaling functions. PMID:17726162

  11. Zinc regulates a key transcriptional pathway for epileptogenesis via metal-regulatory transcription factor 1

    PubMed Central

    van Loo, Karen M. J.; Schaub, Christina; Pitsch, Julika; Kulbida, Rebecca; Opitz, Thoralf; Ekstein, Dana; Dalal, Adam; Urbach, Horst; Beck, Heinz; Yaari, Yoel; Schoch, Susanne; Becker, Albert J.

    2015-01-01

    Temporal lobe epilepsy (TLE) is the most common focal seizure disorder in adults. In many patients, transient brain insults, including status epilepticus (SE), are followed by a latent period of epileptogenesis, preceding the emergence of clinical seizures. In experimental animals, transcriptional upregulation of CaV3.2 T-type Ca2+-channels, resulting in an increased propensity for burst discharges of hippocampal neurons, is an important trigger for epileptogenesis. Here we provide evidence that the metal-regulatory transcription factor 1 (MTF1) mediates the increase of CaV3.2 mRNA and intrinsic excitability consequent to a rise in intracellular Zn2+ that is associated with SE. Adeno-associated viral (rAAV) transfer of MTF1 into murine hippocampi leads to increased CaV3.2 mRNA. Conversely, rAAV-mediated expression of a dominant-negative MTF1 abolishes SE-induced CaV3.2 mRNA upregulation and attenuates epileptogenesis. Finally, data from resected human hippocampi surgically treated for pharmacoresistant TLE support the Zn2+-MTF1-CaV3.2 cascade, thus providing new vistas for preventing and treating TLE. PMID:26498180

  12. Interferon Regulatory Factor 6 Has a Protective Role in the Host Response to Endotoxic Shock

    PubMed Central

    Volk, Paige; Moreland, Jessica G.; Dunnwald, Martine

    2016-01-01

    Interferon Regulatory Factor (IRF) 6, a member of the IRF family, is essential for epidermal and orofacial embryonic development. Irf6 is strongly expressed in keratinocytes, in which it regulates epidermal proliferation, differentiation, and migration. A recent role for Irf6 in Toll-like receptor 2-dependent chemokine gene expression was also reported in an epithelial cell line. However, a function for Irf6 in innate immune cells was not previously reported. In the present study, we investigated the expression and function of Irf6 in bone marrow-derived neutrophils and macrophages. We show here, using a conditional knockout of Irf6 in lysosymeM expressing cells, that Irf6 is required for resistance to LPS-induced endotoxic shock. In addition, Irf6-deficient bone marrow-derived neutrophils exhibited increased chemotactic index and velocity compared with wild-type cells in vitro. TLR4-specific KC and IL6 secretions were upregulated in Irf6-deficient bone marrow-derived macrophages in vitro. These cells also exhibited an increased level of phosphorylated IkBa. Collectively, our findings suggest a role for Irf6 in the resistance to endotoxic shock due to NFk-B-mediated alteration of cytokine production. PMID:27035130

  13. A novel tumor necrosis factor-responsive transcription factor which recognizes a regulatory element in hemopoietic growth factor genes

    SciTech Connect

    Shannon, M.F.; Pell, L.M.; Kuczek, E.S.; Occhiodoro, F.S.; Dunn, S.M.; Vadas, M.A. ); Lenardo, M.J. )

    1990-06-01

    A conserved DNA sequence element, termed cytokine 1 (CK-1), is found in the promoter regions of many hemopoietic growth factor (HGF) genes. Mutational analyses and modification interference experiments show that this sequence specifically binds a nuclear transcription factor, NF-GMa, which is a protein with a molecular mass of 43 kilodaltons. It interacts with different affinities with the CK-1-like sequence from a number of HGF genes, including granulocyte macrophage colony-stimulating factor (GM-CSF), granulocyte (G)-CSF, interleukin 3 (IL-3), and IL-5. The authors show that the level of NF-GMa binding is induced in embryonic fibroblasts by tumor necrosis factor {alpha} (TNF-{alpha}) treatment and that the CK-1 sequence from the G-CSF gene is a TNF-{alpha}-responsive enhancer in these cells.

  14. Systematic genetic analysis of transcription factors to map the fission yeast transcription-regulatory network.

    PubMed

    Chua, Gordon

    2013-12-01

    Mapping transcriptional-regulatory networks requires the identification of target genes, binding specificities and signalling pathways of transcription factors. However, the characterization of each transcription factor sufficiently for deciphering such networks remains laborious. The recent availability of overexpression and deletion strains for almost all of the transcription factor genes in the fission yeast Schizosaccharomyces pombe provides a valuable resource to better investigate transcription factors using systematic genetics. In the present paper, I review and discuss the utility of these strain collections combined with transcriptome profiling and genome-wide chromatin immunoprecipitation to identify the target genes of transcription factors.

  15. BIP induces mice CD19(hi) regulatory B cells producing IL-10 and highly expressing PD-L1, FasL.

    PubMed

    Tang, Youfa; Jiang, Qing; Ou, Yanghui; Zhang, Fan; Qing, Kai; Sun, Yuanli; Lu, Wenjie; Zhu, Huifen; Gong, Feili; Lei, Ping; Shen, Guanxin

    2016-01-01

    Many studies have shown that B cells possess a regulatory function in mouse models of autoimmune diseases. Regulatory B cells can modulate immune response through many types of molecular mechanisms, including the production of IL-10 and the expression of PD-1 Ligand and Fas Ligand, but the microenvironmental factors and mechanisms that induce regulatory B cells have not been fully identified. BIP (binding immunoglobulin protein), a member of the heat shock protein 70 family, is a type of evolutionarily highly conserved protein. In this article, we have found that IL-10(+), PD-L1(hi) and FasL(hi) B cells are discrete cell populations, but enriched in CD19(hi) cells. BIP can induce IL-10-producing splenic B cells, IL-10 secretion and B cells highly expressing PD-L1 and FasL. CD40 signaling acts in synergy with BIP to induce regulatory B cells. BIP increased surface CD19 molecule expression intensity and IL-10(+), PD-L1(hi) and FasL(hi) B cells induced by BIP share the CD19(hi) phenotype. Furthermore, B cells treated with BIP and anti-CD40 can lead to suppression of T cell proliferation and the effect is partially IL-10-dependent and mainly BIP-induced. Taken together, our findings identify a novel function of BIP in the induction of regulatory B cells and add a new reason for the therapy of autoimmune disorders or other inflammatory conditions.

  16. Insights into the Interferon Regulatory Factor Activation from the Crystal Structure of Dimeric IRF5

    SciTech Connect

    Chen, W.; Lam, S; Srinath, H; Jiang, Z; Correia, J; Schiffer, C; Fitzgerald, K; Lin, K; Royer, Jr., W

    2008-01-01

    The interferon regulatory factors (IRFs) are involved in the innate immune response and are activated by phosphorylation. The structure of a pseudophosphorylated IRF5 activation domain now reveals structural changes in the activated form that would turn an autoinhibitory region into a dimerization interface. In vivo analysis supports the relevance of such a dimer to transcriptional activation.

  17. Uncovering a Dual Regulatory Role for Caspases During Endoplasmic Reticulum Stress-induced Cell Death

    PubMed Central

    Anania, Veronica G.; Yu, Kebing; Gnad, Florian; Pferdehirt, Rebecca R.; Li, Han; Ma, Taylur P.; Jeon, Diana; Fortelny, Nikolaus; Forrest, William; Ashkenazi, Avi; Overall, Christopher M.; Lill, Jennie R.

    2016-01-01

    Many diseases are associated with endoplasmic reticulum (ER) stress, which results from an accumulation of misfolded proteins. This triggers an adaptive response called the “unfolded protein response” (UPR), and prolonged exposure to ER stress leads to cell death. Caspases are reported to play a critical role in ER stress-induced cell death but the underlying mechanisms by which they exert their effect continue to remain elusive. To understand the role caspases play during ER stress, a systems level approach integrating analysis of the transcriptome, proteome, and proteolytic substrate profile was employed. This quantitative analysis revealed transcriptional profiles for most human genes, provided information on protein abundance for 4476 proteins, and identified 445 caspase substrates. Based on these data sets many caspase substrates were shown to be downregulated at the protein level during ER stress suggesting caspase activity inhibits their cellular function. Additionally, RNA sequencing revealed a role for caspases in regulation of ER stress-induced transcriptional pathways and gene set enrichment analysis showed expression of multiple gene targets of essential transcription factors to be upregulated during ER stress upon inhibition of caspases. Furthermore, these transcription factors were degraded in a caspase-dependent manner during ER stress. These results indicate that caspases play a dual role in regulating the cellular response to ER stress through both post-translational and transcriptional regulatory mechanisms. Moreover, this study provides unique insight into progression of the unfolded protein response into cell death, which may help identify therapeutic strategies to treat ER stress-related diseases. PMID:27125827

  18. Hypoxia-Inducible Factor-1α and Autoimmune Lupus, Arthritis.

    PubMed

    Yang, Zu-Cheng; Liu, Yi

    2016-06-01

    Hypoxia elicits an orchestrated response in cells, tissues, and entire organisms to survive a hypoxic challenge. On a molecular level, this response can be controlled by oxygen-dependent stabilization of the transcription factor hypoxia-inducible factor (HIF)-1α. Recently, studies have shown that HIF-1α plays an important role in the development and function of T helper (Th) cells, regulatory T (Treg) cells, and dendritic cells (DCs). Because these cells are critical in the pathogenesis of autoimmune diseases, such as systemic lupus erythematosus and rheumatoid arthritis, the roles of HIF-1α in these autoimmune disorders cannot be neglected. In this review, we discuss recent findings on the important roles of HIF-1α in immune cells and the possible pathologic roles of HIF-1α in autoimmune diseases. The obtained information may lead to deeper insights into the roles of HIF-1α in these disorders. PMID:27032396

  19. Immune regulatory and neuroprotective properties of preimplantation factor: From newborn to adult.

    PubMed

    Barnea, E R; Almogi-Hazan, O; Or, R; Mueller, M; Ria, F; Weiss, L; Paidas, M J

    2015-12-01

    Embryonic-maternal interaction from the earliest stages of gestation has a key, sustained role in neurologic development, persisting into adulthood. Early adverse events may be detrimental in adulthood. Protective factors present during gestation could significantly impact post-natal therapy. The role of PreImplantation Factor (PIF) within this context is herein examined. Secreted by viable early embryos, PIF establishes effective embryonic-maternal communication and exerts essential trophic and protective roles by reducing oxidative stress and protein misfolding and by blunting the nocive let-7 microRNA related pathway. PIF's effects on systemic immunity lead to comprehensive immune modulation, not immune suppression. We examine PIF's role in protecting embryos from adverse maternal environment, which can lead to neurological disorders that may only manifest post-nataly: Synthetic PIF successfully translates endogenous PIF features in both pregnant and non-pregnant clinically relevant models. Specifically PIF has neuroprotective effects in neonatal prematurity. In adult relapsing-remitting neuroinflammation, PIF reverses advanced paralysis while promoting neurogenesis. PIF reversed Mycobacterium smegmatis induced brain infection. In graft-vs.-host disease, PIF reduced skin ulceration, liver inflammation and colon ulceration while maintaining beneficial anti-cancer, graft-vs.-leukemia effect. Clinical-grade PIF has high-safety profile even at supraphysiological doses. The FDA awarded Fast-Track designation, and university-sponsored clinical trials for autoimmune disorder are ongoing. Altogether, PIF properties point to its determining regulatory role in immunity, inflammation and transplant acceptance. Specific plans for using PIF for the treatment of complex neurological disorders (ie. traumatic brain injury, progressive paralysis), including neuroprotection from newborn to adult, are presented.

  20. Immune regulatory and neuroprotective properties of preimplantation factor: From newborn to adult.

    PubMed

    Barnea, E R; Almogi-Hazan, O; Or, R; Mueller, M; Ria, F; Weiss, L; Paidas, M J

    2015-12-01

    Embryonic-maternal interaction from the earliest stages of gestation has a key, sustained role in neurologic development, persisting into adulthood. Early adverse events may be detrimental in adulthood. Protective factors present during gestation could significantly impact post-natal therapy. The role of PreImplantation Factor (PIF) within this context is herein examined. Secreted by viable early embryos, PIF establishes effective embryonic-maternal communication and exerts essential trophic and protective roles by reducing oxidative stress and protein misfolding and by blunting the nocive let-7 microRNA related pathway. PIF's effects on systemic immunity lead to comprehensive immune modulation, not immune suppression. We examine PIF's role in protecting embryos from adverse maternal environment, which can lead to neurological disorders that may only manifest post-nataly: Synthetic PIF successfully translates endogenous PIF features in both pregnant and non-pregnant clinically relevant models. Specifically PIF has neuroprotective effects in neonatal prematurity. In adult relapsing-remitting neuroinflammation, PIF reverses advanced paralysis while promoting neurogenesis. PIF reversed Mycobacterium smegmatis induced brain infection. In graft-vs.-host disease, PIF reduced skin ulceration, liver inflammation and colon ulceration while maintaining beneficial anti-cancer, graft-vs.-leukemia effect. Clinical-grade PIF has high-safety profile even at supraphysiological doses. The FDA awarded Fast-Track designation, and university-sponsored clinical trials for autoimmune disorder are ongoing. Altogether, PIF properties point to its determining regulatory role in immunity, inflammation and transplant acceptance. Specific plans for using PIF for the treatment of complex neurological disorders (ie. traumatic brain injury, progressive paralysis), including neuroprotection from newborn to adult, are presented. PMID:26546485

  1. Regulatory roles for Tiam1, a guanine nucleotide exchange factor for Rac1, in glucose-stimulated insulin secretion in pancreatic beta-cells.

    PubMed

    Veluthakal, Rajakrishnan; Madathilparambil, Suresh Vasu; McDonald, Phillip; Olson, Lawrence Karl; Kowluru, Anjaneyulu

    2009-01-01

    Using various biochemical, pharmacological and molecular biological approaches, we have recently reported regulatory roles for Rac1, a small G-protein, in glucose-stimulated insulin secretion (GSIS). However, little is understood with respect to localization of, and regulation by, specific regulatory factors of Rac1 in GSIS. Herein, we investigated regulatory roles for Tiam1, a specific nucleotide exchange factor (GEF) for Rac1, in GSIS in pancreatic beta-cells. Western blot analysis indicated that Tiam1 is predominantly cytosolic in distribution. NSC23766, a specific inhibitor of Tiam1-mediated activation of Rac1, markedly attenuated glucose-induced, but not KCl-induced insulin secretion in INS 832/13 cells and normal rat islets. Further, NSC23766 significantly reduced glucose-induced activation (i.e. GTP-bound form) and membrane association of Rac1 in INS 832/13 cells and rat islets. Moreover, siRNA-mediated knock-down of Tiam1 markedly inhibited glucose-induced membrane trafficking and activation of Rac1 in INS 832/13 cells. Interestingly, however, in contrast to the inhibitory effects of NSC23766, Tiam1 gene depletion potentiated GSIS in these cells; such a potentiation of GSIS was sensitive to extracellular calcium. Together, our studies present the first evidence for a regulatory role for Tiam1/Rac1-sensitive signaling step in GSIS. They also provide evidence for the existence of a potential Rac1/Tiam1-independent, but calcium-sensitive component for GSIS in these cells.

  2. Regulatory Lymphocytes Are Key Factors in MHC-Independent Resistance to EAE

    PubMed Central

    Marín, Nieves; Mecha, Miriam; Espejo, Carmen; Mestre, Leyre; Eixarch, Herena; Montalban, Xavier; Álvarez-Cermeño, José C.; Guaza, Carmen; Villar, Luisa M.

    2014-01-01

    Background and Objectives. Resistant and susceptible mouse strains to experimental autoimmune encephalomyelitis (EAE), an inducible demyelinating experimental disease serving as animal model for multiple sclerosis, have been described. We aimed to explore MHC-independent mechanisms inducing resistance to EAE. Methods. For EAE induction, female C57BL/6 (susceptible strain) and CD1 (resistant outbred strain showing heterogeneous MHC antigens) mice were immunized with the 35–55 peptide of myelin oligodendrocyte glycoprotein (MOG35−55). We studied T cell proliferation, regulatory and effector cell subpopulations, intracellular and serum cytokine patterns, and titers of anti-MOG serum antibodies. Results. Upon immunization with MOG35−55, T lymphocytes from susceptible mice but not that of resistant strain were capable of proliferating when stimulated with MOG35−55. Accordingly, resistant mice experienced a rise in regulatory B cells (P = 0.001) and, to a lower extent, in regulatory T cells (P = 0.02) compared with C57BL/6 susceptible mice. As a consequence, MOG35−55-immunized C57BL/6 mice showed higher percentages of CD4+ T cells producing both IFN-gamma (P = 0.02) and IL-17 (P = 0.009) and higher serum levels of IL-17 (P = 0.04) than resistant mice. Conclusions. Expansion of regulatory B and T cells contributes to the induction of resistance to EAE by an MHC-independent mechanism. PMID:24868560

  3. Regulatory Role of Cannabinoid Receptor 1 in Stress-Induced Excitotoxicity and Neuroinflammation

    PubMed Central

    Zoppi, Silvia; Pérez Nievas, Beatriz G; Madrigal, José L M; Manzanares, Jorge; Leza, Juan C; García-Bueno, Borja

    2011-01-01

    Exposure to stress elicits excitoxicity and neuroinflammation in the brain, contributing to cell death and damage in stress-related neurological and neuropsychiatric diseases. The endocannabinoid system is present in stress-responsive neural circuits and has been proposed as an endogenous neuroprotective system activated in some neuropathological scenarios to restore homeostasis. To elucidate the possible regulatory role of cannabinoid receptor 1 (CB1) in stress-induced excitotoxicity and neuroinflammation, both genetic and pharmacological approaches were used alternatively: (1) wild-type (WT) and CB1 knockout mice (CB1-KO) were exposed to immobilization/acoustic stress (2 h/day for 4 days) and (2) to specifically activate CB1, the selective CB1 agonist Arachidonyl-2′-chloroethylamide (ACEA) (2.5 mg/kg) was intraperitoneally administered daily to some groups of animals. Stress exposure increased CB1 mRNA and protein expression in the prefrontal cortex of WT mice in a mechanism related to N-methyl--aspartate glutamate receptor activation. Daily ACEA pretreatment prevented stress-induced: (1) upregulation of CB1 mRNA and protein, (2) decrease in glutamate uptake and glutamate astroglial transporter excitatory amino acid transporter 2 expression, (3) increase in consecutive proinflammatory molecules, such as cytokines (tumor necrosis factor-α and MCP-1), nuclear factor kappa B, and enzymatic sources, such as inducible nitric oxide synthase (NOS-2) and cyclooxygenase-2 (COX-2), (4) increase in lipid peroxidation; although having no effect on plasma corticosterone. Interestingly, a possible related mechanism could be the positive ACEA modulation of the antiinflammatory pathway deoxyprostaglandin/peroxisome proliferator-activated receptor γ (15d-PGJ2/PPARγ). Conversely, KO animal experiments indicated that a lack of CB1 produces hypothalamic/pituitary/adrenal (HPA) axis dysregulation and exacerbates stress-induced excitotoxic/neuroinflammatory responses. These

  4. From System-Wide Differential Gene Expression to Perturbed Regulatory Factors: A Combinatorial Approach.

    PubMed

    Mahajan, Gaurang; Mande, Shekhar C

    2015-01-01

    High-throughput experiments such as microarrays and deep sequencing provide large scale information on the pattern of gene expression, which undergoes extensive remodeling as the cell dynamically responds to varying environmental cues or has its function disrupted under pathological conditions. An important initial step in the systematic analysis and interpretation of genome-scale expression alteration involves identification of a set of perturbed transcriptional regulators whose differential activity can provide a proximate hypothesis to account for these transcriptomic changes. In the present work, we propose an unbiased and logically natural approach to transcription factor enrichment. It involves overlaying a list of experimentally determined differentially expressed genes on a background regulatory network coming from e.g. literature curation or computational motif scanning, and identifying that subset of regulators whose aggregated target set best discriminates between the altered and the unaffected genes. In other words, our methodology entails testing of all possible regulatory subnetworks, rather than just the target sets of individual regulators as is followed in most standard approaches. We have proposed an iterative search method to efficiently find such a combination, and benchmarked it on E. coli microarray and regulatory network data available in the public domain. Comparative analysis carried out on artificially generated differential expression profiles, as well as empirical factor overexpression data for M. tuberculosis, shows that our methodology provides marked improvement in accuracy of regulatory inference relative to the standard method that involves evaluating factor enrichment in an individual manner. PMID:26562430

  5. From System-Wide Differential Gene Expression to Perturbed Regulatory Factors: A Combinatorial Approach.

    PubMed

    Mahajan, Gaurang; Mande, Shekhar C

    2015-01-01

    High-throughput experiments such as microarrays and deep sequencing provide large scale information on the pattern of gene expression, which undergoes extensive remodeling as the cell dynamically responds to varying environmental cues or has its function disrupted under pathological conditions. An important initial step in the systematic analysis and interpretation of genome-scale expression alteration involves identification of a set of perturbed transcriptional regulators whose differential activity can provide a proximate hypothesis to account for these transcriptomic changes. In the present work, we propose an unbiased and logically natural approach to transcription factor enrichment. It involves overlaying a list of experimentally determined differentially expressed genes on a background regulatory network coming from e.g. literature curation or computational motif scanning, and identifying that subset of regulators whose aggregated target set best discriminates between the altered and the unaffected genes. In other words, our methodology entails testing of all possible regulatory subnetworks, rather than just the target sets of individual regulators as is followed in most standard approaches. We have proposed an iterative search method to efficiently find such a combination, and benchmarked it on E. coli microarray and regulatory network data available in the public domain. Comparative analysis carried out on artificially generated differential expression profiles, as well as empirical factor overexpression data for M. tuberculosis, shows that our methodology provides marked improvement in accuracy of regulatory inference relative to the standard method that involves evaluating factor enrichment in an individual manner.

  6. metagene Profiles Analyses Reveal Regulatory Element’s Factor-Specific Recruitment Patterns

    PubMed Central

    Samb, Rawane; Lemaçon, Audrey; Bilodeau, Steve; Droit, Arnaud

    2016-01-01

    ChIP-Sequencing (ChIP-Seq) provides a vast amount of information regarding the localization of proteins across the genome. The aggregation of ChIP-Seq enrichment signal in a metagene plot is an approach commonly used to summarize data complexity and to obtain a high level visual representation of the general occupancy pattern of a protein. Here we present the R package metagene, the graphical interface Imetagene and the companion package similaRpeak. Together, they provide a framework to integrate, summarize and compare the ChIP-Seq enrichment signal from complex experimental designs. Those packages identify and quantify similarities or dissimilarities in patterns between large numbers of ChIP-Seq profiles. We used metagene to investigate the differential occupancy of regulatory factors at noncoding regulatory regions (promoters and enhancers) in relation to transcriptional activity in GM12878 B-lymphocytes. The relationships between occupancy patterns and transcriptional activity suggest two different mechanisms of action for transcriptional control: i) a “gradient effect” where the regulatory factor occupancy levels follow transcription and ii) a “threshold effect” where the regulatory factor occupancy levels max out prior to reaching maximal transcription. metagene, Imetagene and similaRpeak are implemented in R under the Artistic license 2.0 and are available on Bioconductor. PMID:27538250

  7. metagene Profiles Analyses Reveal Regulatory Element's Factor-Specific Recruitment Patterns.

    PubMed

    Joly Beauparlant, Charles; Lamaze, Fabien C; Deschênes, Astrid; Samb, Rawane; Lemaçon, Audrey; Belleau, Pascal; Bilodeau, Steve; Droit, Arnaud

    2016-08-01

    ChIP-Sequencing (ChIP-Seq) provides a vast amount of information regarding the localization of proteins across the genome. The aggregation of ChIP-Seq enrichment signal in a metagene plot is an approach commonly used to summarize data complexity and to obtain a high level visual representation of the general occupancy pattern of a protein. Here we present the R package metagene, the graphical interface Imetagene and the companion package similaRpeak. Together, they provide a framework to integrate, summarize and compare the ChIP-Seq enrichment signal from complex experimental designs. Those packages identify and quantify similarities or dissimilarities in patterns between large numbers of ChIP-Seq profiles. We used metagene to investigate the differential occupancy of regulatory factors at noncoding regulatory regions (promoters and enhancers) in relation to transcriptional activity in GM12878 B-lymphocytes. The relationships between occupancy patterns and transcriptional activity suggest two different mechanisms of action for transcriptional control: i) a "gradient effect" where the regulatory factor occupancy levels follow transcription and ii) a "threshold effect" where the regulatory factor occupancy levels max out prior to reaching maximal transcription. metagene, Imetagene and similaRpeak are implemented in R under the Artistic license 2.0 and are available on Bioconductor. PMID:27538250

  8. Generation and immunosuppressive functions of p53-induced human adaptive regulatory T cells

    PubMed Central

    Mandapathil, Magis; Visus, Carmen; Finn, Olivera J; Lang, Stephan; Whiteside, Theresa L

    2013-01-01

    Inducible regulatory T cells (iTregs, also called Tr1 cells) are generated in the periphery (circulation or tissue) of cancer patients upon the encounter of naïve CD4+ T cells with tumor-associated antigens. As p53 is often inactivated by genetic or epigenetic events during oncogenesis, p53-induced Tr1 cells might play a key role in establishing immunosuppressive networks in cancer patients. Tr1 cells were generated by co-culturing circulating CD4+CD25− T cells with autologous immature dendritic cells pulsed with a wild-type (WT) p53-derived peptide or an unrelated peptide derived from mucin 1 (MUC1). The Tr1 phenotype and the specificity for p53 of these cells were confirmed by multicolor flow cytometry. Moreover, the Tr1 cell-mediated suppression of T-cell proliferation was evaluated by CFSE-based flow cytometry, while their ability to alter the T-cell cytokine profile by ELISA and Luminex assays. The capacity of p53-induced Tr1 cells to suppress the generation and function of cytotoxic T lymphcoytes (CTLs) was assessed by flow cytometry and ELISPOT. Of note, low doses of the p53-derived peptide (p53low) induced greater numbers of Tr1 cells than the same peptide employed at high doses (p53high). Moreover, Tr1/p53low cells not secreted higher levels of interleukin-10 and transforming growth factor β1, but also mediated more robust suppressive effects on CTL proliferation than Tr1/p53high cells. Tr1/p53low cells, Tr1/p53high cells, as well as Tr1 cells generated with low doses of an unrelated MUC1-derived peptide were equally effective in suppressing the expansion and antitumor activity of p53-reactive CTLs. p53low induced the expansion of highly suppressive p53-reactive Tr1 cells. However, the capacity of these Tr1 cells to suppress the generation and function of p53-reactive CTLs was independent of their antigen-specificity. PMID:24073385

  9. [ASSESSMENT OF EXTREME FACTORS OF SHIFT WORK IN ARCTIC CONDITIONS BY WORKERS WITH DIFFERENT REGULATORY PROCESSES].

    PubMed

    Korneeva, Ya A; Simonova, N N

    2016-01-01

    A man working on a shift basis in the Arctic, every day is under the influence of various extreme factors which are inevitable for oil and gas indudtry. To adapt to shift work employees use various resources of the individual. The purpose of research is the determination of personal resources of shift workers to overcome the adverse factors of the environment in the Arctic. The study involved 191 builder of main gas pipelines, working in shifts in the Tyumen region (the length of the shift 52 days of arrival) at the age of 23 to 59 (mean age 34.9 ± 8.1) years. Methods: psychological testing, questioning, observation, descriptive statistics, discriminant step by step analysis. There was revealed the correlation between the subjective assessment of the majority of adverse climatic factors in the regulatory process "assessment of results"; production factors--regulatory processes such as flexibility, autonomy, simulation, and the general level of self-regulation; social factors are more associated with the severity of such regulatory processes, flexibility and evaluation of results. PMID:27430072

  10. Scientific and regulatory standards for assessing product performance using the similarity factor, f2.

    PubMed

    Stevens, Ruth E; Gray, Vivian; Dorantes, Angelica; Gold, Lynn; Pham, Loan

    2015-03-01

    The similarity factor, f2, measures the sameness of dissolution profiles. The following commentary is an overview of discussions and presentations from a group of industry and US regulatory experts that have integrated the science and regulatory research and practice for assessing product performance, particularly for modified-release (MR) dosage forms, using f2. For a drug development sponsor or applicant with an orally complex dosage formulation, it is critical to understand dissolution methods and the similarity factor and how and/or when to apply it in their NDA, ANDA, or PMA submission. As part of any regulatory submission, it is critical to justify that the product performance has not been impacted by any change in the manufacturing process and/or the delayed and/or prolonged drug release characteristics compared to a similar conventional or another orally complex dosage form. The purposes of this document are (1) to provide a description of appropriate dissolution methods, how is the f2 calculated and how it can be used to justify product performance similarity, or not; (2) to provide an overview of alternative methods available for dissolution profile comparisons, and (3) to illustrate how applying these concepts in a focused way supports approval of submissions and regulatory dossiers and aligns them with on-going science and regulatory initiatives. A case study will be used as an example to demonstrate how dissolution testing and the f2 calculation results can impact regulatory outcomes from an NDA (505(b)(1)), NDA (505(b)(2)), ANDA (505(j)), supplemental NDAs/ANDAs, or PMA perspective.

  11. KLF2 is a rate-limiting transcription factor that can be targeted to enhance regulatory T-cell production

    PubMed Central

    Pabbisetty, Sudheer K.; Rabacal, Whitney; Maseda, Damian; Cendron, Delphine; Collins, Patrick L.; Hoek, Kristen L.; Parekh, Vrajesh V.; Aune, Thomas M.; Sebzda, Eric

    2014-01-01

    Regulatory T cells (Tregs) are a specialized subset of CD4+ T cells that maintain self-tolerance by functionally suppressing autoreactive lymphocytes. The Treg compartment is composed of thymus-derived Tregs (tTregs) and peripheral Tregs (pTregs) that are generated in secondary lymphoid organs after exposure to antigen and specific cytokines, such as TGF-β. With regard to this latter lineage, pTregs [and their ex vivo generated counterparts, induced Tregs (iTregs)] offer particular therapeutic potential because these cells can be raised against specific antigens to limit autoimmunity. We now report that transcription factor Krüppel-like factor 2 (KLF2) is necessary for the generation of iTregs but not tTregs. Moreover, drugs that limit KLF2 proteolysis during T-cell activation enhance iTreg development. To the authors’ knowledge, this study identifies the first transcription factor to distinguish between i/pTreg and tTreg ontogeny and demonstrates that KLF2 is a therapeutic target for the production of regulatory T cells. PMID:24979767

  12. Characterizing the interplay betwen mulitple levels of organization within bacterial sigma factor regulatory networks

    SciTech Connect

    Yu, Qiu; Nagarajan, Harish; Embree, Mallory; Shieu, Wendy; Abate, Elisa; Juarez, Katy; Cho, Byung-Kwan; Elkins, James G; Nevin, Kelly P.; Barrett, Christian; Lovley, Derek; Palsson, Bernhard O.; Zengler, Karsten

    2013-01-01

    Bacteria contain multiple sigma factors, each targeting diverse, but often overlapping sets of promoters, thereby forming a complex network. The layout and deployment of such a sigma factor network directly impacts global transcriptional regulation and ultimately dictates the phenotype. Here we integrate multi-omic data sets to determine the topology, the operational, and functional states of the sigma factor network in Geobacter sulfurreducens, revealing a unique network topology of interacting sigma factors. Analysis of the operational state of the sigma factor network shows a highly modular structure with sN being the major regulator of energy metabolism. Surprisingly, the functional state of the network during the two most divergent growth conditions is nearly static, with sigma factor binding profiles almost invariant to environmental stimuli. This first comprehensive elucidation of the interplay between different levels of the sigma factor network organization is fundamental to characterize transcriptional regulatory mechanisms in bacteria.

  13. Histone deacetylase inhibitor trichostatin A enhances myogenesis by coordinating muscle regulatory factors and myogenic repressors

    SciTech Connect

    Hagiwara, Hiroki; Saito, Fumiaki; Masaki, Toshihiro; Ikeda, Miki; Nakamura-Ohkuma, Ayami; Shimizu, Teruo; Matsumura, Kiichiro

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer We investigated the effect of TSA, one of most potent HDACIs, on myogenesis using the C2C12 skeletal muscle cell line. Black-Right-Pointing-Pointer TSA enhances the expression of myosin heavy chain without affecting DAPC expression. Black-Right-Pointing-Pointer TSA enhances the expression of the early MRFs, Myf5 and MEF2, and suppresses the late MRF, myogenin, after 24 h treatment. Black-Right-Pointing-Pointer TSA enhances the expression of the myogenic repressors, Ids, which inhibit myogenic differentiation. Black-Right-Pointing-Pointer TSA promotes myogenesis by coordinating the expression of MRFs and myogenic repressors. -- Abstract: Histone deacetylase inhibitors (HDACIs) are known to promote skeletal muscle formation. However, their mechanisms that include effects on the expression of major muscle components such as the dystrophin-associated proteins complex (DAPC) or myogenic regulatory factors (MRFs) remain unknown. In this study, we investigated the effects of HDACIs on skeletal muscle formation using the C2C12 cell culture system. C2C12 myoblasts were exposed to trichostatin A (TSA), one of the most potent HDACIs, and differentiation was subsequently induced. We found that TSA enhances the expression of myosin heavy chain without affecting DAPC expression. In addition, TSA increases the expression of the early MRFs, Myf5 and MEF2, whereas it suppresses the expression of the late MRF, myogenin. Interestingly, TSA also enhances the expression of Id1, Id2, and Id3 (Ids). Ids are myogenic repressors that inhibit myogenic differentiation. These findings suggest that TSA promotes gene expression in proliferation and suppresses it in the differentiation stage of muscle formation. Taken together, our data demonstrate that TSA enhances myogenesis by coordinating the expression of MRFs and myogenic repressors.

  14. A direct regulatory interaction between chaperonin TRiC and stress-responsive transcription factor HSF1.

    PubMed

    Neef, Daniel W; Jaeger, Alex M; Gomez-Pastor, Rocio; Willmund, Felix; Frydman, Judith; Thiele, Dennis J

    2014-11-01

    Heat shock transcription factor 1 (HSF1) is an evolutionarily conserved transcription factor that protects cells from protein-misfolding-induced stress and apoptosis. The mechanisms by which cytosolic protein misfolding leads to HSF1 activation have not been elucidated. Here, we demonstrate that HSF1 is directly regulated by TRiC/CCT, a central ATP-dependent chaperonin complex that folds cytosolic proteins. A small-molecule activator of HSF1, HSF1A, protects cells from stress-induced apoptosis, binds TRiC subunits in vivo and in vitro, and inhibits TRiC activity without perturbation of ATP hydrolysis. Genetic inactivation or depletion of the TRiC complex results in human HSF1 activation, and HSF1A inhibits the direct interaction between purified TRiC and HSF1 in vitro. These results demonstrate a direct regulatory interaction between the cytosolic chaperone machine and a critical transcription factor that protects cells from proteotoxicity, providing a mechanistic basis for signaling perturbations in protein folding to a stress-protective transcription factor. PMID:25437552

  15. Pervasive variation of transcription factor orthologs contributes to regulatory network evolution.

    PubMed

    Nadimpalli, Shilpa; Persikov, Anton V; Singh, Mona

    2015-03-01

    Differences in transcriptional regulatory networks underlie much of the phenotypic variation observed across organisms. Changes to cis-regulatory elements are widely believed to be the predominant means by which regulatory networks evolve, yet examples of regulatory network divergence due to transcription factor (TF) variation have also been observed. To systematically ascertain the extent to which TFs contribute to regulatory divergence, we analyzed the evolution of the largest class of metazoan TFs, Cys2-His2 zinc finger (C2H2-ZF) TFs, across 12 Drosophila species spanning ~45 million years of evolution. Remarkably, we uncovered that a significant fraction of all C2H2-ZF 1-to-1 orthologs in flies exhibit variations that can affect their DNA-binding specificities. In addition to loss and recruitment of C2H2-ZF domains, we found diverging DNA-contacting residues in ~44% of domains shared between D. melanogaster and the other fly species. These diverging DNA-contacting residues, found in ~70% of the D. melanogaster C2H2-ZF genes in our analysis and corresponding to ~26% of all annotated D. melanogaster TFs, show evidence of functional constraint: they tend to be conserved across phylogenetic clades and evolve slower than other diverging residues. These same variations were rarely found as polymorphisms within a population of D. melanogaster flies, indicating their rapid fixation. The predicted specificities of these dynamic domains gradually change across phylogenetic distances, suggesting stepwise evolutionary trajectories for TF divergence. Further, whereas proteins with conserved C2H2-ZF domains are enriched in developmental functions, those with varying domains exhibit no functional enrichments. Our work suggests that a subset of highly dynamic and largely unstudied TFs are a likely source of regulatory variation in Drosophila and other metazoans.

  16. Identification of tissue-specific cis-regulatory modules based on interactions between transcription factors

    PubMed Central

    Yu, Xueping; Lin, Jimmy; Zack, Donald J; Qian, Jiang

    2007-01-01

    Background Evolutionary conservation has been used successfully to help identify cis-acting DNA regions that are important in regulating tissue-specific gene expression. Motivated by increasing evidence that some DNA regulatory regions are not evolutionary conserved, we have developed an approach for cis-regulatory region identification that does not rely upon evolutionary sequence conservation. Results The conservation-independent approach is based on an empirical potential energy between interacting transcription factors (TFs). In this analysis, the potential energy is defined as a function of the number of TF interactions in a genomic region and the strength of the interactions. By identifying sets of interacting TFs, the analysis locates regions enriched with the binding sites of these interacting TFs. We applied this approach to 30 human tissues and identified 6232 putative cis-regulatory modules (CRMs) regulating 2130 tissue-specific genes. Interestingly, some genes appear to be regulated by different CRMs in different tissues. Known regulatory regions are highly enriched in our predicted CRMs. In addition, DNase I hypersensitive sites, which tend to be associated with active regulatory regions, significantly overlap with the predicted CRMs, but not with more conserved regions. We also find that conserved and non-conserved CRMs regulate distinct gene groups. Conserved CRMs control more essential genes and genes involved in fundamental cellular activities such as transcription. In contrast, non-conserved CRMs, in general, regulate more non-essential genes, such as genes related to neural activity. Conclusion These results demonstrate that identifying relevant sets of binding motifs can help in the mapping of DNA regulatory regions, and suggest that non-conserved CRMs play an important role in gene regulation. PMID:17996093

  17. Ethylene Response Factors: A Key Regulatory Hub in Hormone and Stress Signaling.

    PubMed

    Müller, Maren; Munné-Bosch, Sergi

    2015-09-01

    Ethylene is essential for many developmental processes and a key mediator of biotic and abiotic stress responses in plants. The ethylene signaling and response pathway includes Ethylene Response Factors (ERFs), which belong to the transcription factor family APETALA2/ERF. It is well known that ERFs regulate molecular response to pathogen attack by binding to sequences containing AGCCGCC motifs (the GCC box), a cis-acting element. However, recent studies suggest that several ERFs also bind to dehydration-responsive elements and act as a key regulatory hub in plant responses to abiotic stresses. Here, we review some of the recent advances in our understanding of the ethylene signaling and response pathway, with emphasis on ERFs and their role in hormone cross talk and redox signaling under abiotic stresses. We conclude that ERFs act as a key regulatory hub, integrating ethylene, abscisic acid, jasmonate, and redox signaling in the plant response to a number of abiotic stresses.

  18. Dietary antigens limit mucosal immunity by inducing regulatory T cells in the small intestine.

    PubMed

    Kim, Kwang Soon; Hong, Sung-Wook; Han, Daehee; Yi, Jaeu; Jung, Jisun; Yang, Bo-Gie; Lee, Jun Young; Lee, Minji; Surh, Charles D

    2016-02-19

    Dietary antigens are normally rendered nonimmunogenic through a poorly understood "oral tolerance" mechanism that involves immunosuppressive regulatory T (Treg) cells, especially Treg cells induced from conventional T cells in the periphery (pTreg cells). Although orally introducing nominal protein antigens is known to induce such pTreg cells, whether a typical diet induces a population of pTreg cells under normal conditions thus far has been unknown. By using germ-free mice raised and bred on an elemental diet devoid of dietary antigens, we demonstrated that under normal conditions, the vast majority of the small intestinal pTreg cells are induced by dietary antigens from solid foods. Moreover, these pTreg cells have a limited life span, are distinguishable from microbiota-induced pTreg cells, and repress underlying strong immunity to ingested protein antigens.

  19. Mining 3D genome structure populations identifies major factors governing the stability of regulatory communities

    PubMed Central

    Dai, Chao; Li, Wenyuan; Tjong, Harianto; Hao, Shengli; Zhou, Yonggang; Li, Qingjiao; Chen, Lin; Zhu, Bing; Alber, Frank; Jasmine Zhou, Xianghong

    2016-01-01

    Three-dimensional (3D) genome structures vary from cell to cell even in an isogenic sample. Unlike protein structures, genome structures are highly plastic, posing a significant challenge for structure-function mapping. Here we report an approach to comprehensively identify 3D chromatin clusters that each occurs frequently across a population of genome structures, either deconvoluted from ensemble-averaged Hi-C data or from a collection of single-cell Hi-C data. Applying our method to a population of genome structures (at the macrodomain resolution) of lymphoblastoid cells, we identify an atlas of stable inter-chromosomal chromatin clusters. A large number of these clusters are enriched in binding of specific regulatory factors and are therefore defined as ‘Regulatory Communities.' We reveal two major factors, centromere clustering and transcription factor binding, which significantly stabilize such communities. Finally, we show that the regulatory communities differ substantially from cell to cell, indicating that expression variability could be impacted by genome structures. PMID:27240697

  20. Interaction between Major Nitrogen Regulatory Protein NIT2 and Pathway-Specific Regulatory Factor NIT4 Is Required for Their Synergistic Activation of Gene Expression in Neurospora crassa

    PubMed Central

    Feng, Bo; Marzluf, George A.

    1998-01-01

    In Neurospora crassa, the major nitrogen regulatory protein, NIT2, a member of the GATA family of transcription factors, controls positively the expression of numerous genes which specify nitrogen catabolic enzymes. Expression of the highly regulated structural gene nit-3, which encodes nitrate reductase, is dependent upon a synergistic interaction of NIT2 with a pathway-specific control protein, NIT4, a member of the GAL4 family of fungal regulatory factors. The NIT2 and NIT4 proteins both bind at specific recognition elements in the nit-3 promoter, but, in addition, we show that a direct protein-protein interaction between NIT2 and NIT4 is essential for optimal expression of the nit-3 structural gene. Neurospora possesses at least five different GATA factors which control different areas of cellular function, but which have a similar DNA binding specificity. Significantly, only NIT2, of the several Neurospora GATA factors examined, interacts with NIT4. We propose that protein-protein interactions of the individual GATA factors with additional pathway-specific regulatory factors determine each of their specific regulatory functions. PMID:9632783

  1. Uncovering MicroRNA and Transcription Factor Mediated Regulatory Networks in Glioblastoma.

    PubMed

    Sun, Jingchun; Gong, Xue; Purow, Benjamin; Zhao, Zhongming

    2012-01-01

    Glioblastoma multiforme (GBM) is the most common and lethal brain tumor in humans. Recent studies revealed that patterns of microRNA (miRNA) expression in GBM tissue samples are different from those in normal brain tissues, suggesting that a number of miRNAs play critical roles in the pathogenesis of GBM. However, little is yet known about which miRNAs play central roles in the pathology of GBM and their regulatory mechanisms of action. To address this issue, in this study, we systematically explored the main regulation format (feed-forward loops, FFLs) consisting of miRNAs, transcription factors (TFs) and their impacting GBM-related genes, and developed a computational approach to construct a miRNA-TF regulatory network. First, we compiled GBM-related miRNAs, GBM-related genes, and known human TFs. We then identified 1,128 3-node FFLs and 805 4-node FFLs with statistical significance. By merging these FFLs together, we constructed a comprehensive GBM-specific miRNA-TF mediated regulatory network. Then, from the network, we extracted a composite GBM-specific regulatory network. To illustrate the GBM-specific regulatory network is promising for identification of critical miRNA components, we specifically examined a Notch signaling pathway subnetwork. Our follow up topological and functional analyses of the subnetwork revealed that six miRNAs (miR-124, miR-137, miR-219-5p, miR-34a, miR-9, and miR-92b) might play important roles in GBM, including some results that are supported by previous studies. In this study, we have developed a computational framework to construct a miRNA-TF regulatory network and generated the first miRNA-TF regulatory network for GBM, providing a valuable resource for further understanding the complex regulatory mechanisms in GBM. The observation of critical miRNAs in the Notch signaling pathway, with partial verification from previous studies, demonstrates that our network-based approach is promising for the identification of new and important

  2. Modulation of Mitochondrial Antiviral Signaling by Human Herpesvirus 8 Interferon Regulatory Factor 1

    PubMed Central

    Hwang, Keun Young

    2015-01-01

    ABSTRACT Mitochondrial lipid raft-like microdomains, experimentally also termed mitochondrial detergent-resistant membrane fractions (mDRM), play a role as platforms for recruiting signaling molecules involved in antiviral responses such as apoptosis and innate immunity. Viruses can modulate mitochondrial functions for their own survival and replication. However, viral regulation of the antiviral responses via mDRM remains incompletely understood. Here, we report that human herpesvirus 8 (HHV-8) gene product viral interferon regulatory factor 1 (vIRF-1) is targeted to mDRM during virus replication and negatively regulates the mitochondrial antiviral signaling protein (MAVS)-mediated antiviral responses. The N-terminal region of vIRF-1 interacts directly with membrane lipids, including cardiolipin. In addition, a GxRP motif within the N terminus of vIRF-1, conserved in the mDRM-targeting region of mitochondrial proteins, including PTEN-induced putative kinase 1 (PINK1) and MAVS, was found to be important for vIRF-1 association with mitochondria. Furthermore, MAVS, which has the potential to promote vIRF-1 targeting to mDRM possibly by inducing cardiolipin exposure on the outer membrane of mitochondria, interacts with vIRF-1, which, in turn, inhibits MAVS-mediated antiviral signaling. Consistent with these results, vIRF-1 targeting to mDRM contributes to promotion of HHV-8 productive replication and inhibition of associated apoptosis. Combined, our results suggest novel molecular mechanisms for negative-feedback regulation of MAVS by vIRF-1 during virus replication. IMPORTANCE Successful virus replication is in large part achieved by the ability of viruses to counteract apoptosis and innate immune responses elicited by infection of host cells. Recently, mitochondria have emerged to play a central role in antiviral signaling. In particular, mitochondrial lipid raft-like microdomains appear to function as platforms in cell apoptosis signaling. However, viral regulation

  3. Regulatory Mechanisms of the Molecular Pathways in Fibrosis Induced by MicroRNAs

    PubMed Central

    Yang, Cui; Zheng, Si-Dao; Wu, Hong-Jin; Chen, Shao-Jun

    2016-01-01

    Objective: MicroRNAs (miRNAs or miRs) play critical roles in the fibrotic process in different organs. We summarized the latest research progress on the roles and mechanisms of miRNAs in the regulation of the molecular signaling pathways involved in fibrosis. Data Sources: Papers published in English from January 2010 to August 2015 were selected from the PubMed and Web of Science databases using the search terms “microRNA”, “miR”, “transforming growth factor β”, “tgf β”, “mitogen-activated protein kinase”, “mapk”, “integrin”, “p38”, “c-Jun NH2-terminal kinase”, “jnk”, “extracellular signal-regulated kinase”, “erk”, and “fibrosis”. Study Selection: Articles were obtained and reviewed to analyze the regulatory effects of miRNAs on molecular signaling pathways involved in the fibrosis. Results: Recent evidence has shown that miRNAs are involved in regulating fibrosis by targeting different substrates in the molecular processes that drive fibrosis, such as immune cell sensitization, effector cell activation, and extracellular matrix remodeling. Moreover, several important molecular signaling pathways involve in fibrosis, such as the transforming growth factor-beta (TGF-β) pathway, mitogen-activated protein kinase (MAPK) pathways, and the integrin pathway are regulated by miRNAs. Third, regulation of the fibrotic pathways induced by miRNAs is found in many other tissues in addition to the heart, lung, liver, and kidney. Interestingly, the actions of many drugs on the human body are also induced by miRNAs. It is encouraging that the fibrotic process can be blocked or reversed by targeting specific miRNAs and their signaling pathways, thereby protecting the structures and functions of different organs. Conclusions: miRNAs not only regulate molecular signaling pathways in fibrosis but also serve as potential targets of novel therapeutic interventions for fibrosing diseases. PMID:27647197

  4. Comparative analysis of the transcription-factor gene regulatory networks of E. coli and S. cerevisiae

    PubMed Central

    Guzmán-Vargas, Lev; Santillán, Moisés

    2008-01-01

    Background The regulatory interactions between transcription factors (TF) and regulated genes (RG) in a species genome can be lumped together in a single directed graph. The TF's and RG's conform the nodes of this graph, while links are drawn whenever a transcription factor regulates a gene's expression. Projections onto TF nodes can be constructed by linking every two nodes regulating a common gene. Similarly, projections onto RG nodes can be made by linking every two regulated genes sharing at least one common regulator. Recent studies of the connectivity pattern in the transcription-factor regulatory network of many organisms have revealed some interesting properties. However, the differences between TF and RG nodes have not been widely explored. Results After analysing the RG and TF projections of the transcription-factor gene regulatory networks of Escherichia coli and Saccharomyces cerevisiae, we found several common characteristic as well as some noticeable differences. To better understand these differences, we compared the properties of the E. coli and S. cerevisiae RG- and TF-projected networks with those of the corresponding projections built from randomized versions of the original bipartite networks. These last results indicate that the observed differences are mostly due to the very different ratios of TF to RG counts of the E. coli and S. cerevisiae bipartite networks, rather than to their having different connectivity patterns. Conclusion Since E. coli is a prokaryotic organism while S. cerevisiae is eukaryotic, there are important differences between them concerning processing of mRNA before translation, DNA packing, amount of junk DNA, and gene regulation. From the results in this paper we conclude that the most important effect such differences have had on the development of the corresponding transcription-factor gene regulatory networks is their very different ratios of TF to RG numbers. This ratio is more than three times larger in S

  5. Erythropoietin is a hypoxia inducible factor-induced protective molecule in experimental autoimmune neuritis.

    PubMed

    Luo, Bangwei; Jiang, Man; Yang, Xiaofeng; Zhang, Zhiyuan; Xiong, Jian; Schluesener, Hermann J; Zhang, Zhiren; Wu, Yuzhang

    2013-08-01

    Experimental autoimmune neuritis (EAN), an autoantigen-specific T-cell-mediated disease model for human demyelinating inflammatory disease of the peripheral nervous system, is characterized by self-limitation. Here we investigated the regulation and contribution of erythropoietin (EPO) in EAN self-limitation. In EAN sciatic nerves, hypoxia, and protein and mRNA levels of hypoxia-inducible factor 1α (HIF-1α), HIF-2α, EPO and EPO receptor (EPOR) were induced in parallel at disease peak phase but reduced at recovery periods. Further, the deactivation of HIF reduced EAN-induced EPO/EPOR upregulation in EAN, suggesting the central contribution of HIF to EPO/EPOR induction. The deactivation of EPOR signalling exacerbated EAN progression, implying that endogenous EPO contributed to EAN recovery. Exogenous EPO treatment greatly improved EAN recovery. In addition, EPO was shown to promote Schwann cell survival and myelin production. In EAN, EPO treatment inhibited lymphocyte proliferation and altered helper T cell differentiation by inducing increase of Foxp3(+)/CD4(+) regulatory T cells and decrease of IFN-γ(+)/CD4(+) Th1 cells. Furthermore, EPO inhibited inflammatory macrophage activation and promoted its phagocytic activity. In summary, our data demonstrated that EPO was induced in EAN by HIF and contributed to EAN recovery, and endogenous and exogenous EPO could effectively suppress EAN by attenuating inflammation and exerting direct cell protection, indicating that EPO contributes to the self-recovery of EAN and could be a potent candidate for treatment of autoimmune neuropathies. PMID:23603807

  6. Novel Foxp3− IL-10− Regulatory T-cells Induced by B-Cells Alleviate Intestinal Inflammation in Vivo

    PubMed Central

    Shao, Tzu-Yu; Hsu, Ling-Hui; Chien, Chien-Hui; Chiang, Bor-Luen

    2016-01-01

    Recent studies have revealed various Foxp3− regulatory T (Treg) cell subsets effectively protect mice from colitis. In the present study, we demonstrated that B cells induced a particular subset of regulatory T (Treg-of-B) cells, expressing programmed cell death 1 (PD-1), inducible costimulator (ICOS), lymphocyte-activation gene 3 (LAG3), glucocorticoid-induced tumor necrosis factor receptor (GITR), and OX-40, did not express Foxp3. Treg-of-B cells produced abundant levels of IL-10 and low levels of IL-4 and TGF-β. Adoptive transfer of Treg-of-B cells protected mice from CD4+CD45RBhi T-cell-induced colitis, including infiltration of leukocytes, depletion of goblet cells, epithelial hyperplasia, and inhibition of Th1 and Th17 cytokines. These features were similar to IL-10-producing type 1 regulatory T (Tr1) cells; however, IL-10-deficient Treg-of-B cells maintained their suppressive function in vitro as well as in vivo, while the regulation of Tr1 cells depended on IL-10. In conclusion, Treg-of-B cells protected against experimental colitis through an IL-10-independent mechanism. We reported a novel subpopulation of regulatory T cells was different from conventional Foxp3+ Treg and IL-10-producing Tr1 cells. PMID:27581189

  7. Transcription Factors OVOL1 and OVOL2 Induce the Mesenchymal to Epithelial Transition in Human Cancer

    PubMed Central

    Roca, Hernan; Hernandez, James; Weidner, Savannah; McEachin, Richard C.; Fuller, David; Sud, Sudha; Schumann, Taibriana; Wilkinson, John E.; Zaslavsky, Alexander; Li, Hangwen; Maher, Christopher A.; Daignault-Newton, Stephanie; Healy, Patrick N.; Pienta, Kenneth J.

    2013-01-01

    Cell plasticity regulated by the balance between the mesenchymal to epithelial transition (MET) and the opposite program, EMT, is critical in the metastatic cascade. Several transcription factors (TFs) are known to regulate EMT, though the mechanisms of MET remain unclear. We demonstrate a novel function of two TFs, OVOL1 and OVOL2, as critical inducers of MET in human cancers. Our findings indicate that the OVOL-TFs control MET through a regulatory feedback loop with EMT-inducing TF ZEB1, and the regulation of mRNA splicing by inducing Epithelial Splicing Regulatory Protein 1 (ESRP1). Using mouse prostate tumor models we show that expression of OVOL-TFs in mesenchymal prostate cancer cells attenuates their metastatic potential. The role of OVOL-TFs as inducers of MET is further supported by expression analyses in 917 cancer cell lines, suggesting their role as crucial regulators of epithelial-mesenchymal cell plasticity in cancer. PMID:24124593

  8. Regulatory T cells in human and angiotensin II-induced mouse abdominal aortic aneurysms

    PubMed Central

    Zhou, Yi; Wu, Wenxue; Lindholt, Jes S.; Sukhova, Galina K.; Libby, Peter; Yu, Xueqing; Shi, Guo-Ping

    2015-01-01

    Aims Regulatory T cells (Tregs) protect mice from angiotensin II (Ang-II)-induced abdominal aortic aneurysms (AAA). This study tested whether AAA patients are Treg-insufficient and the Treg molecular mechanisms that control AAA pathogenesis. Methods and results ELISA determined the Foxp3 concentration in blood cell lysates from 485 AAA patients and 204 age- and sex-matched controls. AAA patients exhibited lower blood cell Foxp3 expression than controls (P < 0.0001). Pearson's correlation test demonstrated a significant but negative correlation between Foxp3 and AAA annual expansion rate before (r = –0.147, P = 0.007) and after (r = –0.153, P = 0.006) adjustment for AAA risk factors. AAA in apolipoprotein E-deficient (Apoe–/–) mice that received different doses of Ang-II exhibited a negative correlation of lesion Foxp3+ Treg numbers with AAA size (r = –0.883, P < 0.0001). Adoptive transfer of Tregs from wild-type (WT) and IL10-deficient (Il10–/–) mice increased AAA lesion Treg content, but only WT mice Tregs reduced AAA size, AAA incidence, blood pressure, lesion macrophage and CD4+ and CD8+ T-cell accumulation, and angiogenesis with concurrent increase of lesion collagen content. Both AAA lesion immunostaining and plasma ELISA demonstrated that adoptive transfer of WT Tregs, but not Il10–/– Tregs, reduced the expression of MCP-1. In vitro cell culture and aortic ring assay demonstrated that only Tregs from WT mice, but not those from Il10–/– mice, reduced macrophage MCP-1 secretion, macrophage and vascular cell protease expression and activity, and aortic ring microvessel formation. Conclusion This study supports a protective role of Tregs in human and experimental AAA by releasing IL10 to suppress inflammatory cell chemotaxis, arterial wall remodelling, and angiogenesis. PMID:25824145

  9. Evolution of DNA specificity in a transcription factor family produced a new gene regulatory module.

    PubMed

    McKeown, Alesia N; Bridgham, Jamie T; Anderson, Dave W; Murphy, Michael N; Ortlund, Eric A; Thornton, Joseph W

    2014-09-25

    Complex gene regulatory networks require transcription factors (TFs) to bind distinct DNA sequences. To understand how novel TF specificity evolves, we combined phylogenetic, biochemical, and biophysical approaches to interrogate how DNA recognition diversified in the steroid hormone receptor (SR) family. After duplication of the ancestral SR, three mutations in one copy radically weakened binding to the ancestral estrogen response element (ERE) and improved binding to a new set of DNA sequences (steroid response elements, SREs). They did so by establishing unfavorable interactions with ERE and abolishing unfavorable interactions with SRE; also required were numerous permissive substitutions, which nonspecifically improved cooperativity and affinity of DNA binding. Our findings indicate that negative determinants of binding play key roles in TFs' DNA selectivity and-with our prior work on the evolution of SR ligand specificity during the same interval-show how a specific new gene regulatory module evolved without interfering with the integrity of the ancestral module. PMID:25259920

  10. Using RSAT to scan genome sequences for transcription factor binding sites and cis-regulatory modules.

    PubMed

    Turatsinze, Jean-Valery; Thomas-Chollier, Morgane; Defrance, Matthieu; van Helden, Jacques

    2008-01-01

    This protocol shows how to detect putative cis-regulatory elements and regions enriched in such elements with the regulatory sequence analysis tools (RSAT) web server (http://rsat.ulb.ac.be/rsat/). The approach applies to known transcription factors, whose binding specificity is represented by position-specific scoring matrices, using the program matrix-scan. The detection of individual binding sites is known to return many false predictions. However, results can be strongly improved by estimating P value, and by searching for combinations of sites (homotypic and heterotypic models). We illustrate the detection of sites and enriched regions with a study case, the upstream sequence of the Drosophila melanogaster gene even-skipped. This protocol is also tested on random control sequences to evaluate the reliability of the predictions. Each task requires a few minutes of computation time on the server. The complete protocol can be executed in about one hour.

  11. Hypoxia-Inducible Factor in Thyroid Carcinoma

    PubMed Central

    Burrows, Natalie; Babur, Muhammad; Resch, Julia; Williams, Kaye J.; Brabant, Georg

    2011-01-01

    Intratumoural hypoxia (low oxygen tension) is associated with aggressive disease and poor prognosis. Hypoxia-inducible factor-1 is a transcription factor activated by hypoxia that regulates the expression of genes that promote tumour cell survival, progression, metastasis, and resistance to chemo/radiotherapy. In addition to hypoxia, HIF-1 can be activated by growth factor-signalling pathways such as the mitogen-activated protein kinases- (MAPK-) and phosphatidylinositol-3-OH kinases- (PI3K-) signalling cascades. Mutations in these pathways are common in thyroid carcinoma and lead to enhanced HIF-1 expression and activity. Here, we summarise current data that highlights the potential role of both hypoxia and MAPK/PI3K-induced HIF-1 signalling in thyroid carcinoma progression, metastatic characteristics, and the potential role of HIF-1 in thyroid carcinoma response to radiotherapy. Direct or indirect targeting of HIF-1 using an MAPK or PI3K inhibitor in combination with radiotherapy may be a new potential therapeutic target to improve the therapeutic response of thyroid carcinoma to radiotherapy and reduce metastatic burden. PMID:21765994

  12. Development of a doxycycline-inducible lentiviral plasmid with an instant regulatory feature.

    PubMed

    Yang, Tian; Burrows, Christopher; Park, Jeong Hyeon

    2014-03-01

    Lentiviruses provide highly efficient gene delivery vehicles in both dividing and non-dividing cells. Inducible gene expression systems often employ a specific cell line that constitutively expresses a regulatory protein for transgene expression. As one of such inducible expression systems the Tet-On system uses a cell line expressing reverse tetracycline-responsive transcriptional activator (rtTA). The rtTA protein binds to the tetracycline-responsive element (TRE) in the promoter and activates transcription of a transgene in a doxycycline-dependent manner. To establish a universal and instant regulatory system without generating Tet-On cell lines, the cDNAs of rtTA and a testing target gene (PPM1B) were cloned in the bi-directional TRE-containing promoters. Here, we examined whether a basal leaky expression of rtTA allows instantly inducible expression of both rtTA itself and the target gene, PPM1B in a single plasmid using the two mini-CMV promoters. Transient transfection of the lentiviral plasmids into human embryonic kidney HEK293T cells showed a significant induction of PPM1B expression in response to doxycycline, suggesting that these lentiviral plasmids can be used as an instantly inducible mammalian expression vector. However, the expression of rtTA by lentiviral transduction shows a minimal expression without a consistent response to doxycycline, suggesting that the utility of these lentiviral vectors is limited. A potential solution to overcome lentiviral transgene inactivation is proposed. PMID:24727543

  13. A Minimal Regulatory Network of Extrinsic and Intrinsic Factors Recovers Observed Patterns of CD4+ T Cell Differentiation and Plasticity

    PubMed Central

    Martinez-Sanchez, Mariana Esther; Mendoza, Luis; Villarreal, Carlos; Alvarez-Buylla, Elena R.

    2015-01-01

    CD4+ T cells orchestrate the adaptive immune response in vertebrates. While both experimental and modeling work has been conducted to understand the molecular genetic mechanisms involved in CD4+ T cell responses and fate attainment, the dynamic role of intrinsic (produced by CD4+ T lymphocytes) versus extrinsic (produced by other cells) components remains unclear, and the mechanistic and dynamic understanding of the plastic responses of these cells remains incomplete. In this work, we studied a regulatory network for the core transcription factors involved in CD4+ T cell-fate attainment. We first show that this core is not sufficient to recover common CD4+ T phenotypes. We thus postulate a minimal Boolean regulatory network model derived from a larger and more comprehensive network that is based on experimental data. The minimal network integrates transcriptional regulation, signaling pathways and the micro-environment. This network model recovers reported configurations of most of the characterized cell types (Th0, Th1, Th2, Th17, Tfh, Th9, iTreg, and Foxp3-independent T regulatory cells). This transcriptional-signaling regulatory network is robust and recovers mutant configurations that have been reported experimentally. Additionally, this model recovers many of the plasticity patterns documented for different T CD4+ cell types, as summarized in a cell-fate map. We tested the effects of various micro-environments and transient perturbations on such transitions among CD4+ T cell types. Interestingly, most cell-fate transitions were induced by transient activations, with the opposite behavior associated with transient inhibitions. Finally, we used a novel methodology was used to establish that T-bet, TGF-β and suppressors of cytokine signaling proteins are keys to recovering observed CD4+ T cell plastic responses. In conclusion, the observed CD4+ T cell-types and transition patterns emerge from the feedback between the intrinsic or intracellular regulatory core

  14. Tr1 and naturally occurring regulatory T cells induce IgG4 in B cells through GITR/GITR-L interaction, IL-10 and TGF-beta.

    PubMed

    Satoguina, Judith S; Adjobimey, Tomabu; Arndts, Kathrin; Hoch, Jochen; Oldenburg, Johannes; Layland, Laura E; Hoerauf, Achim

    2008-11-01

    Regulatory T cells exert their function through the modulation of both T and B cell responses. Our previous studies demonstrated that IL-10-producing Treg (Tr1) can induce B cells to secrete IgG4 in a cell-contact-dependent manner. The benefit of such non-inflammatory B-cell responses is apparent in the hyporesponsive state of patients with helminth infections such as Onchocerciasis. Here, we investigated the mechanisms involved to induce IgG4, within B:Tr-cell co-cultures, using IL-10-producing tetanus-toxoid-specific regulatory T cell lines and clones (Tr-TCC) from human PBMC. During the generation process, we found that increasing Foxp3 levels in regulatory T cell lines correlated with their ability to induce IgG4 in B cells. Using Tr-TCC, we found that blocking glucocorticoid-induced tumour necrosis factor receptor-related protein (GITR) molecules selectively prevented IgG4 production as did neutralizing Ab to glucocorticoid-induced tumour necrosis factor receptor-related protein ligand (GITR-L), IL-10 and TGF-beta. Furthermore, the prevention of IgG4 induction by anti-GITR Ab was reversed by excess rIL-10 but not rTGF-beta. In contrast, anti-ICOS and anti-CTLA-4 Abs had no effect. When compared with Tr-TCC, freshly isolated CD4+CD25+ T cells, but not effector T cell populations, induced low levels of IgG4, which were also blocked by anti-GITR and anti-GITR-L Ab. Thus, the mechanism of IgG4 induction by regulatory cells involves GITR-GITR-L interactions, IL-10 and TGF-beta. PMID:18924213

  15. Insulin counter-regulatory factors, fibrinogen and C-reactive protein during olanzapine administration: effects of the antidiabetic metformin.

    PubMed

    Baptista, Trino; Sandia, Ignacio; Lacruz, Anny; Rangel, Nairy; de Mendoza, Soaira; Beaulieu, Serge; Contreras, Quilianio; Galeazzi, Tatiana; Vargas, Doritza

    2007-03-01

    In this study, the Authors assessed some insulin counter-regulatory factors, fibrinogen and C-reactive protein after olanzapine administration, and the effect of metformin on these variables, 37 patients with chronic schizophrenia were given olanzapine (10 mg/day for 14 weeks). Nineteen patients received metformin (850-2550 mg/day) and 18 received placebo in a randomized, double-blind protocol. The following variables were quantified before and after olanzapine: cortisol, leptin, tumor necrosis factor-alpha, glucagon, growth hormone, fibrinogen and C-reactive protein. Results were correlated with the changes in body weight and the insulin resistance index. We have reported elsewhere that metformin did not prevent olanzapine-induced weight gain, and the insulin resistance index significantly decreased after metformin and placebo; Baptista T, et al. Can J Psychiatry 2006; 51: 192-196. Cortisol, tumor necrosis factor-alpha and fibrinogen levels significantly decreased in both groups. Glucagon significantly increased after metformin (P=0.03). Leptin tended to increase after placebo (P=0.1) and displayed a small nonsignificant reduction after metformin. The C-reactive protein did not change significantly in any group. Contrarily to most published studies, olanzapine was associated with decreased insulin resistance. Decrements in cortisol, fibrinogen and tumor necrosis factor-alpha levels point to an improvement in the metabolic profile. The trend for leptin to increase after placebo, but not after metformin in spite of similar weight gain suggests a beneficial effect of this antidiabetic agent. PMID:17293706

  16. Translation regulatory factor RBM3 is a proto-oncogene that prevents mitotic catastrophe

    PubMed Central

    Sureban, SM; Ramalingam, S; Natarajan, G; May, R; Subramaniam, D; Bishnupuri, KS; Morrison, AR; Dieckgraefe, BK; Brackett, DJ; Postier, RG; Houchen, CW; Anant, S

    2009-01-01

    RNA-binding proteins play a key role in post-transcriptional regulation of mRNA stability and translation. We have identified that RBM3, a translation regulatory protein, is significantly upregulated in human tumors, including a stage-dependent increase in colorectal tumors. Forced RBM3 overexpression in NIH3T3 mouse fibro-blasts and SW480 human colon epithelial cells increases cell proliferation and development of compact multicellular spheroids in soft agar suggesting the ability to induce anchorage-independent growth. In contrast, down-regulating RBM3 in HCT116 colon cancer cells with specific siRNA decreases cell growth in culture, which was partially overcome when treated with prostaglandin E2, a product of cyclooxygenase (COX)-2 enzyme activity. Knockdown also resulted in the growth arrest of tumor xenografts. We have also identified that RBM3 knockdown increases caspase-mediated apoptosis coupled with nuclear cyclin B1, and phosphorylated Cdc25c, Chk1 and Chk2 kinases, implying that under conditions of RBM3 downregulation, cells undergo mitotic catastrophe. RBM3 enhances COX-2, IL-8 and VEGF mRNA stability and translation. Conversely, RBM3 knockdown results in loss in the translation of these transcripts. These data demonstrate that the RNA stabilizing and translation regulatory protein RBM3 is a novel proto-oncogene that induces transformation when overexpressed and is essential for cells to progress through mitosis. PMID:18427544

  17. Antibody Targeting of “Steady-State” Dendritic Cells Induces Tolerance Mediated by Regulatory T Cells

    PubMed Central

    Mahnke, Karsten; Ring, Sabine; Enk, Alexander H.

    2016-01-01

    Dendritic cells (DCs) are often defined as pivotal inducers of immunity, but these proinflammatory properties only develop after stimulation or ex vivo manipulation of DCs. Under non-inflammatory conditions in vivo, DCs are embedded into a tissue environment and encounter a plethora of self-antigens derived from apoptotic material. This material is transported to secondary lymphoid organs. As DCs maintain their non-activated phenotype in a sterile tissue environment, interaction with T cells will induce rather regulatory T cells than effector T cells. Thus, DCs are not only inducers of immunity but are also critical for maintenance of peripheral tolerance. Therapeutically, intervention for the induction of long-lasting tolerance in several autoimmune conditions may therefore be possible by manipulating DC activation and/or targeting of DCs in their “natural” tissue environment. PMID:26941742

  18. Regulatory T Cells Promote β-Catenin–Mediated Epithelium-to-Mesenchyme Transition During Radiation-Induced Pulmonary Fibrosis

    SciTech Connect

    Xiong, Shanshan; Pan, Xiujie; Xu, Long; Yang, Zhihua; Guo, Renfeng; Gu, Yongqing; Li, Ruoxi; Wang, Qianjun; Xiao, Fengjun; Du, Li; Zhou, Pingkun; Zhu, Maoxiang

    2015-10-01

    Purpose: Radiation-induced pulmonary fibrosis results from thoracic radiation therapy and severely limits radiation therapy approaches. CD4{sup +}CD25{sup +}FoxP3{sup +} regulatory T cells (Tregs) as well as epithelium-to-mesenchyme transition (EMT) cells are involved in pulmonary fibrosis induced by multiple factors. However, the mechanisms of Tregs and EMT cells in irradiation-induced pulmonary fibrosis remain unclear. In the present study, we investigated the influence of Tregs on EMT in radiation-induced pulmonary fibrosis. Methods and Materials: Mice thoraxes were irradiated (20 Gy), and Tregs were depleted by intraperitoneal injection of a monoclonal anti-CD25 antibody 2 hours after irradiation and every 7 days thereafter. Mice were treated on days 3, 7, and 14 and 1, 3, and 6 months post irradiation. The effectiveness of Treg depletion was assayed via flow cytometry. EMT and β-catenin in lung tissues were detected by immunohistochemistry. Tregs isolated from murine spleens were cultured with mouse lung epithelial (MLE) 12 cells, and short interfering RNA (siRNA) knockdown of β-catenin in MLE 12 cells was used to explore the effects of Tregs on EMT and β-catenin via flow cytometry and Western blotting. Results: Anti-CD25 antibody treatment depleted Tregs efficiently, attenuated the process of radiation-induced pulmonary fibrosis, hindered EMT, and reduced β-catenin accumulation in lung epithelial cells in vivo. The coculture of Tregs with irradiated MLE 12 cells showed that Tregs could promote EMT in MLE 12 cells and that the effect of Tregs on EMT was partially abrogated by β-catenin knockdown in vitro. Conclusions: Tregs can promote EMT in accelerating radiation-induced pulmonary fibrosis. This process is partially mediated through β-catenin. Our study suggests a new mechanism for EMT, promoted by Tregs, that accelerates radiation-induced pulmonary fibrosis.

  19. Selection of terrestrial transfer factors for radioecological assessment models and regulatory guides

    SciTech Connect

    Ng, Y.C.; Hoffman, F.O.

    1983-01-01

    A parameter value for a radioecological assessment model is not a single value but a distribution of values about a central value. The sources that contribute to the variability of transfer factors to predict foodchain transport of radionuclides are enumerated. Knowledge of these sources, judgement in interpreting the available data, consideration of collateral information, and established criteria that specify the desired level of conservatism in the resulting predictions are essential elements when selecting appropriate parameter values for radioecological assessment models and regulatory guides. 39 references, 4 figures, 5 tables.

  20. Mining regulatory network connections by ranking transcription factor target genes using time series expression data.

    PubMed

    Honkela, Antti; Rattray, Magnus; Lawrence, Neil D

    2013-01-01

    Reverse engineering the gene regulatory network is challenging because the amount of available data is very limited compared to the complexity of the underlying network. We present a technique addressing this problem through focussing on a more limited problem: inferring direct targets of a transcription factor from short expression time series. The method is based on combining Gaussian process priors and ordinary differential equation models allowing inference on limited potentially unevenly sampled data. The method is implemented as an R/Bioconductor package, and it is demonstrated by ranking candidate targets of the p53 tumour suppressor.

  1. Direct lineage reprogramming via pioneer factors; a detour through developmental gene regulatory networks.

    PubMed

    Morris, Samantha A

    2016-08-01

    Although many approaches have been employed to generate defined fate in vitro, the resultant cells often appear developmentally immature or incompletely specified, limiting their utility. Growing evidence suggests that current methods of direct lineage conversion may rely on the transition through a developmental intermediate. Here, I hypothesize that complete conversion between cell fates is more probable and feasible via reversion to a developmentally immature state. I posit that this is due to the role of pioneer transcription factors in engaging silent, unmarked chromatin and activating hierarchical gene regulatory networks responsible for embryonic patterning. Understanding these developmental contexts will be essential for the precise engineering of cell identity. PMID:27486230

  2. How microRNA and transcription factor co-regulatory networks affect osteosarcoma cell proliferation.

    PubMed

    Poos, Kathrin; Smida, Jan; Nathrath, Michaela; Maugg, Doris; Baumhoer, Daniel; Korsching, Eberhard

    2013-01-01

    Osteosarcomas (OS) are complex bone tumors with various genomic alterations. These alterations affect the expression and function of several genes due to drastic changes in the underlying gene regulatory network. However, we know little about critical gene regulators and their functional consequences on the pathogenesis of OS. Therefore, we aimed to determine microRNA and transcription factor (TF) co-regulatory networks in OS cell proliferation. Cell proliferation is an essential part in the pathogenesis of OS and deeper understanding of its regulation might help to identify potential therapeutic targets. Based on expression data of OS cell lines divided according to their proliferative activity, we obtained 12 proliferation-related microRNAs and corresponding target genes. Therewith, microRNA and TF co-regulatory networks were generated and analyzed regarding their structure and functional influence. We identified key co-regulators comprising the microRNAs miR-9-5p, miR-138, and miR-214 and the TFs SP1 and MYC in the derived networks. These regulators are implicated in NFKB- and RB1-signaling and focal adhesion processes based on their common or interacting target genes (e.g., CDK6, CTNNB1, E2F4, HES1, ITGA6, NFKB1, NOTCH1, and SIN3A). Thus, we proposed a model of OS cell proliferation which is primarily co-regulated through the interactions of the mentioned microRNA and TF combinations. This study illustrates the benefit of systems biological approaches in the analysis of complex diseases. We integrated experimental data with publicly available information to unravel the coordinated (post)-transcriptional control of microRNAs and TFs to identify potential therapeutic targets in OS. The resulting microRNA and TF co-regulatory networks are publicly available for further exploration to generate or evaluate own hypotheses of the pathogenesis of OS (http://www.complex-systems.uni-muenster.de/co_networks.html).

  3. Identification of High-Impact cis-Regulatory Mutations Using Transcription Factor Specific Random Forest Models

    PubMed Central

    Svetlichnyy, Dmitry; Imrichova, Hana; Fiers, Mark; Kalender Atak, Zeynep; Aerts, Stein

    2015-01-01

    Cancer genomes contain vast amounts of somatic mutations, many of which are passenger mutations not involved in oncogenesis. Whereas driver mutations in protein-coding genes can be distinguished from passenger mutations based on their recurrence, non-coding mutations are usually not recurrent at the same position. Therefore, it is still unclear how to identify cis-regulatory driver mutations, particularly when chromatin data from the same patient is not available, thus relying only on sequence and expression information. Here we use machine-learning methods to predict functional regulatory regions using sequence information alone, and compare the predicted activity of the mutated region with the reference sequence. This way we define the Predicted Regulatory Impact of a Mutation in an Enhancer (PRIME). We find that the recently identified driver mutation in the TAL1 enhancer has a high PRIME score, representing a “gain-of-target” for MYB, whereas the highly recurrent TERT promoter mutation has a surprisingly low PRIME score. We trained Random Forest models for 45 cancer-related transcription factors, and used these to score variations in the HeLa genome and somatic mutations across more than five hundred cancer genomes. Each model predicts only a small fraction of non-coding mutations with a potential impact on the function of the encompassing regulatory region. Nevertheless, as these few candidate driver mutations are often linked to gains in chromatin activity and gene expression, they may contribute to the oncogenic program by altering the expression levels of specific oncogenes and tumor suppressor genes. PMID:26562774

  4. Increments and duplication events of enzymes and transcription factors influence metabolic and regulatory diversity in prokaryotes.

    PubMed

    Martínez-Núñez, Mario Alberto; Poot-Hernandez, Augusto Cesar; Rodríguez-Vázquez, Katya; Perez-Rueda, Ernesto

    2013-01-01

    In this work, the content of enzymes and DNA-binding transcription factors (TFs) in 794 non-redundant prokaryotic genomes was evaluated. The identification of enzymes was based on annotations deposited in the KEGG database as well as in databases of functional domains (COG and PFAM) and structural domains (Superfamily). For identifications of the TFs, hidden Markov profiles were constructed based on well-known transcriptional regulatory families. From these analyses, we obtained diverse and interesting results, such as the negative rate of incremental changes in the number of detected enzymes with respect to the genome size. On the contrary, for TFs the rate incremented as the complexity of genome increased. This inverse related performance shapes the diversity of metabolic and regulatory networks and impacts the availability of enzymes and TFs. Furthermore, the intersection of the derivatives between enzymes and TFs was identified at 9,659 genes, after this point, the regulatory complexity grows faster than metabolic complexity. In addition, TFs have a low number of duplications, in contrast to the apparent high number of duplications associated with enzymes. Despite the greater number of duplicated enzymes versus TFs, the increment by which duplicates appear is higher in TFs. A lower proportion of enzymes among archaeal genomes (22%) than in the bacterial ones (27%) was also found. This low proportion might be compensated by the interconnection between the metabolic pathways in Archaea. A similar proportion was also found for the archaeal TFs, for which the formation of regulatory complexes has been proposed. Finally, an enrichment of multifunctional enzymes in Bacteria, as a mechanism of ecological adaptation, was detected. PMID:23922780

  5. Increments and Duplication Events of Enzymes and Transcription Factors Influence Metabolic and Regulatory Diversity in Prokaryotes

    PubMed Central

    Martínez-Núñez, Mario Alberto; Poot-Hernandez, Augusto Cesar; Rodríguez-Vázquez, Katya; Perez-Rueda, Ernesto

    2013-01-01

    In this work, the content of enzymes and DNA-binding transcription factors (TFs) in 794 non-redundant prokaryotic genomes was evaluated. The identification of enzymes was based on annotations deposited in the KEGG database as well as in databases of functional domains (COG and PFAM) and structural domains (Superfamily). For identifications of the TFs, hidden Markov profiles were constructed based on well-known transcriptional regulatory families. From these analyses, we obtained diverse and interesting results, such as the negative rate of incremental changes in the number of detected enzymes with respect to the genome size. On the contrary, for TFs the rate incremented as the complexity of genome increased. This inverse related performance shapes the diversity of metabolic and regulatory networks and impacts the availability of enzymes and TFs. Furthermore, the intersection of the derivatives between enzymes and TFs was identified at 9,659 genes, after this point, the regulatory complexity grows faster than metabolic complexity. In addition, TFs have a low number of duplications, in contrast to the apparent high number of duplications associated with enzymes. Despite the greater number of duplicated enzymes versus TFs, the increment by which duplicates appear is higher in TFs. A lower proportion of enzymes among archaeal genomes (22%) than in the bacterial ones (27%) was also found. This low proportion might be compensated by the interconnection between the metabolic pathways in Archaea. A similar proportion was also found for the archaeal TFs, for which the formation of regulatory complexes has been proposed. Finally, an enrichment of multifunctional enzymes in Bacteria, as a mechanism of ecological adaptation, was detected. PMID:23922780

  6. Increments and duplication events of enzymes and transcription factors influence metabolic and regulatory diversity in prokaryotes.

    PubMed

    Martínez-Núñez, Mario Alberto; Poot-Hernandez, Augusto Cesar; Rodríguez-Vázquez, Katya; Perez-Rueda, Ernesto

    2013-01-01

    In this work, the content of enzymes and DNA-binding transcription factors (TFs) in 794 non-redundant prokaryotic genomes was evaluated. The identification of enzymes was based on annotations deposited in the KEGG database as well as in databases of functional domains (COG and PFAM) and structural domains (Superfamily). For identifications of the TFs, hidden Markov profiles were constructed based on well-known transcriptional regulatory families. From these analyses, we obtained diverse and interesting results, such as the negative rate of incremental changes in the number of detected enzymes with respect to the genome size. On the contrary, for TFs the rate incremented as the complexity of genome increased. This inverse related performance shapes the diversity of metabolic and regulatory networks and impacts the availability of enzymes and TFs. Furthermore, the intersection of the derivatives between enzymes and TFs was identified at 9,659 genes, after this point, the regulatory complexity grows faster than metabolic complexity. In addition, TFs have a low number of duplications, in contrast to the apparent high number of duplications associated with enzymes. Despite the greater number of duplicated enzymes versus TFs, the increment by which duplicates appear is higher in TFs. A lower proportion of enzymes among archaeal genomes (22%) than in the bacterial ones (27%) was also found. This low proportion might be compensated by the interconnection between the metabolic pathways in Archaea. A similar proportion was also found for the archaeal TFs, for which the formation of regulatory complexes has been proposed. Finally, an enrichment of multifunctional enzymes in Bacteria, as a mechanism of ecological adaptation, was detected.

  7. Expression of Streptococcus pneumoniae Bacteriocins Is Induced by Antibiotics via Regulatory Interplay with the Competence System

    PubMed Central

    Slager, Jelle; Lake, Frank B.; Gericke, Oliver; Roberts, Ian S.; Rozen, Daniel E.; Veening, Jan-Willem

    2016-01-01

    Pneumococcal bacteriocins (pneumocins) are antibacterial toxins that mediate intra-species competition within the human host. However, the triggers of pneumocin expression are poorly understood. Using RNA-sequencing, we mapped the regulon of the pneumocin cluster (blp) of Streptococcus pneumoniae D39. Furthermore, by analogy with pneumococcal competence, we show that several antibiotics activate the blp-genes. Using real-time gene expression measurements we show that while the promoter driving expression of the two-component regulatory system blpR/H is constitutive, the remaining blp-promoters that control pneumocin expression, immunity and the inducer peptide BlpC, are pH-dependent and induced in the late exponential phase. Intriguingly, competence for genetic transformation, mediated by the paralogous ComD/E two-component quorum system, is induced by the same environmental cues. To test for interplay between these regulatory systems, we quantified the regulatory response to the addition of synthetic BlpC and competence-stimulating peptide (CSP). Supporting the idea of such interplay, we found that immediately upon addition of CSP, the blp-promoters were activated in a comD/E-dependent manner. After a delay, blp-expression was highly induced and was strictly dependent on blpRH and blpC. This raised the question of the mechanism of BlpC export, since bioinformatic analysis showed that the genes encoding the putative exporter for BlpC, blpAB, are not intact in strain D39 and most other strains. By contrast, all sequenced pneumococcal strains contain intact comAB genes, encoding the transport system for CSP. Consistent with the idea that comAB mediate BlpC export, we finally show that high-level expression of the blp-genes requires comAB. Together, our results demonstrate that regulation of pneumocin expression is intertwined with competence, explaining why certain antibiotics induce blp-expression. Antibiotic-induced pneumocin expression might therefore have

  8. Methylphenidate blocks effort-induced depletion of regulatory control in healthy volunteers.

    PubMed

    Sripada, Chandra; Kessler, Daniel; Jonides, John

    2014-06-01

    A recent wave of studies--more than 100 conducted over the last decade--has shown that exerting effort at controlling impulses or behavioral tendencies leaves a person depleted and less able to engage in subsequent rounds of regulation. Regulatory depletion is thought to play an important role in everyday problems (e.g., excessive spending, overeating) as well as psychiatric conditions, but its neurophysiological basis is poorly understood. Using a placebo-controlled, double-blind design, we demonstrated that the psychostimulant methylphenidate (commonly known as Ritalin), a catecholamine reuptake blocker that increases dopamine and norepinephrine at the synaptic cleft, fully blocks effort-induced depletion of regulatory control. Spectral analysis of trial-by-trial reaction times revealed specificity of methylphenidate effects on regulatory depletion in the slow-4 frequency band. This band is associated with the operation of resting-state brain networks that produce mind wandering, which raises potential connections between our results and recent brain-network-based models of control over attention.

  9. Transcription factor MITF and remodeller BRG1 define chromatin organisation at regulatory elements in melanoma cells.

    PubMed

    Laurette, Patrick; Strub, Thomas; Koludrovic, Dana; Keime, Céline; Le Gras, Stéphanie; Seberg, Hannah; Van Otterloo, Eric; Imrichova, Hana; Siddaway, Robert; Aerts, Stein; Cornell, Robert A; Mengus, Gabrielle; Davidson, Irwin

    2015-03-24

    Microphthalmia-associated transcription factor (MITF) is the master regulator of the melanocyte lineage. To understand how MITF regulates transcription, we used tandem affinity purification and mass spectrometry to define a comprehensive MITF interactome identifying novel cofactors involved in transcription, DNA replication and repair, and chromatin organisation. We show that MITF interacts with a PBAF chromatin remodelling complex comprising BRG1 and CHD7. BRG1 is essential for melanoma cell proliferation in vitro and for normal melanocyte development in vivo. MITF and SOX10 actively recruit BRG1 to a set of MITF-associated regulatory elements (MAREs) at active enhancers. Combinations of MITF, SOX10, TFAP2A, and YY1 bind between two BRG1-occupied nucleosomes thus defining both a signature of transcription factors essential for the melanocyte lineage and a specific chromatin organisation of the regulatory elements they occupy. BRG1 also regulates the dynamics of MITF genomic occupancy. MITF-BRG1 interplay thus plays an essential role in transcription regulation in melanoma.

  10. Transcription factor MITF and remodeller BRG1 define chromatin organisation at regulatory elements in melanoma cells

    PubMed Central

    Laurette, Patrick; Strub, Thomas; Koludrovic, Dana; Keime, Céline; Le Gras, Stéphanie; Seberg, Hannah; Van Otterloo, Eric; Imrichova, Hana; Siddaway, Robert; Aerts, Stein; Cornell, Robert A; Mengus, Gabrielle; Davidson, Irwin

    2015-01-01

    Microphthalmia-associated transcription factor (MITF) is the master regulator of the melanocyte lineage. To understand how MITF regulates transcription, we used tandem affinity purification and mass spectrometry to define a comprehensive MITF interactome identifying novel cofactors involved in transcription, DNA replication and repair, and chromatin organisation. We show that MITF interacts with a PBAF chromatin remodelling complex comprising BRG1 and CHD7. BRG1 is essential for melanoma cell proliferation in vitro and for normal melanocyte development in vivo. MITF and SOX10 actively recruit BRG1 to a set of MITF-associated regulatory elements (MAREs) at active enhancers. Combinations of MITF, SOX10, TFAP2A, and YY1 bind between two BRG1-occupied nucleosomes thus defining both a signature of transcription factors essential for the melanocyte lineage and a specific chromatin organisation of the regulatory elements they occupy. BRG1 also regulates the dynamics of MITF genomic occupancy. MITF-BRG1 interplay thus plays an essential role in transcription regulation in melanoma. DOI: http://dx.doi.org/10.7554/eLife.06857.001 PMID:25803486

  11. Expression of myogenic regulatory factors and myo-endothelial remodeling in sporadic inclusion body myositis

    PubMed Central

    Wanschitz, Julia V.; Dubourg, Odile; Lacene, Emmanuelle; Fischer, Michael B.; Höftberger, Romana; Budka, Herbert; Romero, Norma B.; Eymard, Bruno; Herson, Serge; Butler-Browne, Gillian S.; Voit, Thomas; Benveniste, Olivier

    2013-01-01

    Muscle repair relies on coordinated activation and differentiation of satellite cells, a process that is unable to counterbalance progressive degeneration in sporadic inclusion body myositis (s-IBM). To explore features of myo regeneration, the expression of myogenic regulatory factors Pax7, MyoD and Myogenin and markers of regenerating fibers was analyzed by immunohistochemistry in s-IBM muscle compared with polymyositis, dermatomyositis, muscular dystrophy and age-matched controls. In addition, the capillary density and number of interstitial CD34+ hematopoietic progenitor cells was determined by double-immunoflourescence staining. Satellite cells and regenerating fibers were significantly increased in s-IBM similar to other inflammatory myopathies and correlated with the intensity of inflammation (R > 0.428). Expression of MyoD, visualizing activated satellite cells and proliferating myoblasts, was lower in s-IBM compared to polymyosits. In contrast, Myogenin a marker of myogenic cell differentiation was strongly up-regulated in s-IBM muscle. The microvascular architecture in s-IBM was distorted, although the capillary density was normal. Notably, CD34+ hematopoietic cells were significantly increased in the interstitial compartment. Our findings indicate profound myo-endothelial remodeling of s-IBM muscle concomitant to inflammation. An altered expression of myogenic regulatory factors involved in satellite cell activation and differentiation, however, might reflect perturbations of muscle repair in s-IBM. PMID:23058947

  12. Interferon regulatory factors: at the crossroads of immunity, metabolism, and disease.

    PubMed

    Zhao, Guang-Nian; Jiang, Ding-Sheng; Li, Hongliang

    2015-02-01

    The interferon-regulatory factor (IRF) family comprises nine members in mammals. Although this transcription factor family was originally thought to function primarily in the immune system, contributing to both the innate immune response and the development of immune cells, recent advances have revealed that IRFs plays critical roles in other biological processes, such as metabolism. Accordingly, abnormalities in the expression and/or function of IRFs have increasingly been linked to disease. Herein, we provide an update on the recent progress regarding the regulation of immune responses and immune cell development associated with IRFs. Additionally, we discuss the relationships between IRFs and immunity, metabolism, and disease, with a particular focus on the role of IRFs as stress sensors. This article is part of a Special Issue entitled: Autophagy and protein quality control in cardiometabolic diseases.

  13. Commensal microbe-derived butyrate induces the differentiation of colonic regulatory T cells.

    PubMed

    Furusawa, Yukihiro; Obata, Yuuki; Fukuda, Shinji; Endo, Takaho A; Nakato, Gaku; Takahashi, Daisuke; Nakanishi, Yumiko; Uetake, Chikako; Kato, Keiko; Kato, Tamotsu; Takahashi, Masumi; Fukuda, Noriko N; Murakami, Shinnosuke; Miyauchi, Eiji; Hino, Shingo; Atarashi, Koji; Onawa, Satoshi; Fujimura, Yumiko; Lockett, Trevor; Clarke, Julie M; Topping, David L; Tomita, Masaru; Hori, Shohei; Ohara, Osamu; Morita, Tatsuya; Koseki, Haruhiko; Kikuchi, Jun; Honda, Kenya; Hase, Koji; Ohno, Hiroshi

    2013-12-19

    Gut commensal microbes shape the mucosal immune system by regulating the differentiation and expansion of several types of T cell. Clostridia, a dominant class of commensal microbe, can induce colonic regulatory T (Treg) cells, which have a central role in the suppression of inflammatory and allergic responses. However, the molecular mechanisms by which commensal microbes induce colonic Treg cells have been unclear. Here we show that a large bowel microbial fermentation product, butyrate, induces the differentiation of colonic Treg cells in mice. A comparative NMR-based metabolome analysis suggests that the luminal concentrations of short-chain fatty acids positively correlates with the number of Treg cells in the colon. Among short-chain fatty acids, butyrate induced the differentiation of Treg cells in vitro and in vivo, and ameliorated the development of colitis induced by adoptive transfer of CD4(+) CD45RB(hi) T cells in Rag1(-/-) mice. Treatment of naive T cells under the Treg-cell-polarizing conditions with butyrate enhanced histone H3 acetylation in the promoter and conserved non-coding sequence regions of the Foxp3 locus, suggesting a possible mechanism for how microbial-derived butyrate regulates the differentiation of Treg cells. Our findings provide new insight into the mechanisms by which host-microbe interactions establish immunological homeostasis in the gut.

  14. A regulatory role for the memory B cell as suppressor-inducer of feedback control

    SciTech Connect

    Kennedy, M.W.; Thomas, D.B.

    1983-02-01

    A regulatory role is proposed for the antigen-responsive B cell, as suppressor-inducer of feedback control during the secondary response in vivo. In a double adoptive transfer of memory cells primed to a thymus-dependent antigen from one irradiated host to another, antigen-specific suppressors are generated after a critical time in the primary recipient, able to entirely ablate a secondary anti-hapten response. Positive cell selection in the fluorescence-activated cell sorter confirmed that suppression was mediated by an Lyt-2+ T cell; however, positively selected B cells were also inhibitory and able to induce suppressors in a carrier-specific manner: Bhapten induced suppressors in a carrier-primed population, and Bcarrier induced suppressors in a hapten-carrier population. At the peak of the antibody response in the primary host, memory B cells and their progeny were unable to differentiate further to plasma cells due to their intrinsic suppressor-inducer activity, but this autoregulatory circuit could be severed by adoptive transfer to carrier-primed, X-irradiated recipients.

  15. Oxidant-induced apoptosis is mediated by oxidation of the actin-regulatory protein cofilin

    PubMed Central

    Klamt, Fábio; Zdanov, Stéphanie; Levine, Rodney L.; Pariser, Ashley; Zhang, Yaqin; Zhang, Baolin; Yu, Li-Rong; Veenstra, Timothy D.; Shacter, Emily

    2012-01-01

    Physiological oxidants that are generated by activated phagocytes comprise the main source of oxidative stress during inflammation1,2. Oxidants such as taurine chloramine (TnCl) and hydrogen peroxide (H2O2) can damage proteins and induce apoptosis, but the role of specific protein oxidation in this process has not been defined. We found that the actin-binding protein cofilin is a key target of oxidation. When oxidation of this single regulatory protein is prevented, oxidant-induced apoptosis is inhibited. Oxidation of cofilin causes it to lose its affinity for actin and to translocate to the mitochondria, where it induces swelling and cytochrome c release by mediating opening of the permeability transition pore (PTP). This occurs independently of Bax activation and requires both oxidation of cofilin Cys residues and dephosphorylation at Ser 3. Knockdown of endogenous cofilin using targeted siRNA inhibits oxidant-induced apoptosis, which is restored by re-expression of wild-type cofilin but not by cofilin containing Cys to Ala mutations. Exposure of cofilin to TnCl results in intramolecular disulphide bonding and oxidation of Met residues to Met sulphoxide, but only Cys oxidation causes cofilin to induce mitochondrial damage. PMID:19734890

  16. Arabidopsis Ensemble Reverse-Engineered Gene Regulatory Network Discloses Interconnected Transcription Factors in Oxidative Stress[W

    PubMed Central

    Vermeirssen, Vanessa; De Clercq, Inge; Van Parys, Thomas; Van Breusegem, Frank; Van de Peer, Yves

    2014-01-01

    The abiotic stress response in plants is complex and tightly controlled by gene regulation. We present an abiotic stress gene regulatory network of 200,014 interactions for 11,938 target genes by integrating four complementary reverse-engineering solutions through average rank aggregation on an Arabidopsis thaliana microarray expression compendium. This ensemble performed the most robustly in benchmarking and greatly expands upon the availability of interactions currently reported. Besides recovering 1182 known regulatory interactions, cis-regulatory motifs and coherent functionalities of target genes corresponded with the predicted transcription factors. We provide a valuable resource of 572 abiotic stress modules of coregulated genes with functional and regulatory information, from which we deduced functional relationships for 1966 uncharacterized genes and many regulators. Using gain- and loss-of-function mutants of seven transcription factors grown under control and salt stress conditions, we experimentally validated 141 out of 271 predictions (52% precision) for 102 selected genes and mapped 148 additional transcription factor-gene regulatory interactions (49% recall). We identified an intricate core oxidative stress regulatory network where NAC13, NAC053, ERF6, WRKY6, and NAC032 transcription factors interconnect and function in detoxification. Our work shows that ensemble reverse-engineering can generate robust biological hypotheses of gene regulation in a multicellular eukaryote that can be tested by medium-throughput experimental validation. PMID:25549671

  17. Functional analysis of microRNA and transcription factor synergistic regulatory network based on identifying regulatory motifs in non-small cell lung cancer

    PubMed Central

    2013-01-01

    Background Lung cancer, especially non-small cell lung cancer, is a leading cause of malignant tumor death worldwide. Understanding the mechanisms employed by the main regulators, such as microRNAs (miRNAs) and transcription factors (TFs), still remains elusive. The patterns of their cooperation and biological functions in the synergistic regulatory network have rarely been studied. Results Here, we describe the first miRNA-TF synergistic regulation network in human lung cancer. We identified important regulators (MYC, NFKB1, miR-590, and miR-570) and significant miRNA-TF synergistic regulatory motifs by random simulations. The two most significant motifs were the co-regulation of miRNAs and TFs, and TF-mediated cascade regulation. We also developed an algorithm to uncover the biological functions of the human lung cancer miRNA-TF synergistic regulatory network (regulation of apoptosis, cellular protein metabolic process, and cell cycle), and the specific functions of each miRNA-TF synergistic subnetwork. We found that the miR-17 family exerted important effects in the regulation of non-small cell lung cancer, such as in proliferation and cell cycle regulation by targeting the retinoblastoma protein (RB1) and forming a feed forward loop with the E2F1 TF. We proposed a model for the miR-17 family, E2F1, and RB1 to demonstrate their potential roles in the occurrence and development of non-small cell lung cancer. Conclusions This work will provide a framework for constructing miRNA-TF synergistic regulatory networks, function analysis in diseases, and identification of the main regulators and regulatory motifs, which will be useful for understanding the putative regulatory motifs involving miRNAs and TFs, and for predicting new targets for cancer studies. PMID:24200043

  18. UV-B-induced DNA damage mediates expression changes of cell cycle regulatory genes in Arabidopsis root tips.

    PubMed

    Jiang, Lei; Wang, Yan; Björn, Lars Olof; Li, Shaoshan

    2011-04-01

    Even though a number of studies have shown that UV-B radiation inhibits plant growth and regulates the cell cycle progress, little is known about the molecular and cellular mechanisms. Here, we developed a synchronous root-tip cell system to investigate expression changes of cell cycle marker genes and DNA damage under UV-B radiation. Expression analysis of cell cycle marker genes revealed that G1-to-S transition in root-tip cells was accomplished within 6 h. In the in vivo synchronous root-tip cells, high level of UV-B radiation (0.45 W m(-2)) induced expression changes of the cell cycle regulatory genes. Genes involved in G1-to-S transition, Histone H4 and E2Fa, were down-regulated by UV-B radiation during 2-6 h; whereas transcripts for KRP2, a negative regulator of G1-to-S transition, were up-regulated by UV-B at 2 h. The peak time for transcript level of CYCD3;1, a positive factor in G1-to-S transition, was delayed by UV-B radiation. Interestingly, a medium level of UV-B radiation (0.25 W m(-2)) did not change the expression of these genes in root tip cells from wild type. However, cell cycle regulatory genes were greatly affected in uvh1 mutant, which exhibited higher content of cyclobutane pyrimidine dimers (CPDs). Ascorbic acid treatment did not change the expression pattern of cell cycle regulatory genes that were affected by high-level UV-B. Our results implied that UV-B-induced DNA damage results in the delay of G1-to-S transition of plant cell cycle. UV-B-induced G1-to-S arrest may be a protective mechanism that prevents cells with damaged DNA from dividing and may explain the plant growth inhibition under increased solar UV-B radiation.

  19. A Consensus Network of Gene Regulatory Factors in the Human Frontal Lobe.

    PubMed

    Berto, Stefano; Perdomo-Sabogal, Alvaro; Gerighausen, Daniel; Qin, Jing; Nowick, Katja

    2016-01-01

    Cognitive abilities, such as memory, learning, language, problem solving, and planning, involve the frontal lobe and other brain areas. Not much is known yet about the molecular basis of cognitive abilities, but it seems clear that cognitive abilities are determined by the interplay of many genes. One approach for analyzing the genetic networks involved in cognitive functions is to study the coexpression networks of genes with known importance for proper cognitive functions, such as genes that have been associated with cognitive disorders like intellectual disability (ID) or autism spectrum disorders (ASD). Because many of these genes are gene regulatory factors (GRFs) we aimed to provide insights into the gene regulatory networks active in the human frontal lobe. Using genome wide human frontal lobe expression data from 10 independent data sets, we first derived 10 individual coexpression networks for all GRFs including their potential target genes. We observed a high level of variability among these 10 independently derived networks, pointing out that relying on results from a single study can only provide limited biological insights. To instead focus on the most confident information from these 10 networks we developed a method for integrating such independently derived networks into a consensus network. This consensus network revealed robust GRF interactions that are conserved across the frontal lobes of different healthy human individuals. Within this network, we detected a strong central module that is enriched for 166 GRFs known to be involved in brain development and/or cognitive disorders. Interestingly, several hubs of the consensus network encode for GRFs that have not yet been associated with brain functions. Their central role in the network suggests them as excellent new candidates for playing an essential role in the regulatory network of the human frontal lobe, which should be investigated in future studies. PMID:27014338

  20. A Consensus Network of Gene Regulatory Factors in the Human Frontal Lobe

    PubMed Central

    Berto, Stefano; Perdomo-Sabogal, Alvaro; Gerighausen, Daniel; Qin, Jing; Nowick, Katja

    2016-01-01

    Cognitive abilities, such as memory, learning, language, problem solving, and planning, involve the frontal lobe and other brain areas. Not much is known yet about the molecular basis of cognitive abilities, but it seems clear that cognitive abilities are determined by the interplay of many genes. One approach for analyzing the genetic networks involved in cognitive functions is to study the coexpression networks of genes with known importance for proper cognitive functions, such as genes that have been associated with cognitive disorders like intellectual disability (ID) or autism spectrum disorders (ASD). Because many of these genes are gene regulatory factors (GRFs) we aimed to provide insights into the gene regulatory networks active in the human frontal lobe. Using genome wide human frontal lobe expression data from 10 independent data sets, we first derived 10 individual coexpression networks for all GRFs including their potential target genes. We observed a high level of variability among these 10 independently derived networks, pointing out that relying on results from a single study can only provide limited biological insights. To instead focus on the most confident information from these 10 networks we developed a method for integrating such independently derived networks into a consensus network. This consensus network revealed robust GRF interactions that are conserved across the frontal lobes of different healthy human individuals. Within this network, we detected a strong central module that is enriched for 166 GRFs known to be involved in brain development and/or cognitive disorders. Interestingly, several hubs of the consensus network encode for GRFs that have not yet been associated with brain functions. Their central role in the network suggests them as excellent new candidates for playing an essential role in the regulatory network of the human frontal lobe, which should be investigated in future studies. PMID:27014338

  1. Transcription profile of Escherichia coli: genomic SELEX search for regulatory targets of transcription factors

    PubMed Central

    Ishihama, Akira; Shimada, Tomohiro; Yamazaki, Yukiko

    2016-01-01

    Bacterial genomes are transcribed by DNA-dependent RNA polymerase (RNAP), which achieves gene selectivity through interaction with sigma factors that recognize promoters, and transcription factors (TFs) that control the activity and specificity of RNAP holoenzyme. To understand the molecular mechanisms of transcriptional regulation, the identification of regulatory targets is needed for all these factors. We then performed genomic SELEX screenings of targets under the control of each sigma factor and each TF. Here we describe the assembly of 156 SELEX patterns of a total of 116 TFs performed in the presence and absence of effector ligands. The results reveal several novel concepts: (i) each TF regulates more targets than hitherto recognized; (ii) each promoter is regulated by more TFs than hitherto recognized; and (iii) the binding sites of some TFs are located within operons and even inside open reading frames. The binding sites of a set of global regulators, including cAMP receptor protein, LeuO and Lrp, overlap with those of the silencer H-NS, suggesting that certain global regulators play an anti-silencing role. To facilitate sharing of these accumulated SELEX datasets with the research community, we compiled a database, ‘Transcription Profile of Escherichia coli’ (www.shigen.nig.ac.jp/ecoli/tec/). PMID:26843427

  2. Transcription profile of Escherichia coli: genomic SELEX search for regulatory targets of transcription factors.

    PubMed

    Ishihama, Akira; Shimada, Tomohiro; Yamazaki, Yukiko

    2016-03-18

    Bacterial genomes are transcribed by DNA-dependent RNA polymerase (RNAP), which achieves gene selectivity through interaction with sigma factors that recognize promoters, and transcription factors (TFs) that control the activity and specificity of RNAP holoenzyme. To understand the molecular mechanisms of transcriptional regulation, the identification of regulatory targets is needed for all these factors. We then performed genomic SELEX screenings of targets under the control of each sigma factor and each TF. Here we describe the assembly of 156 SELEX patterns of a total of 116 TFs performed in the presence and absence of effector ligands. The results reveal several novel concepts: (i) each TF regulates more targets than hitherto recognized; (ii) each promoter is regulated by more TFs than hitherto recognized; and (iii) the binding sites of some TFs are located within operons and even inside open reading frames. The binding sites of a set of global regulators, including cAMP receptor protein, LeuO and Lrp, overlap with those of the silencer H-NS, suggesting that certain global regulators play an anti-silencing role. To facilitate sharing of these accumulated SELEX datasets with the research community, we compiled a database, 'Transcription Profile of Escherichia coli' (www.shigen.nig.ac.jp/ecoli/tec/). PMID:26843427

  3. Construction and analysis of dynamic transcription factor regulatory networks in the progression of glioma.

    PubMed

    Li, Yongsheng; Shao, Tingting; Jiang, Chunjie; Bai, Jing; Wang, Zishan; Zhang, Jinwen; Zhang, Lili; Zhao, Zheng; Xu, Juan; Li, Xia

    2015-11-03

    The combinatorial cross-regulation of transcription factors (TFs) plays an important role in cellular identity and function; however, the dynamic regulation of TFs during glioma progression remains largely unknown. Here, we used the genome-wide expression of TFs to construct an extensive human TF network comprising interactions among 513 TFs and to analyse the dynamics of the TF-TF network during glioma progression. We found that the TF regulatory networks share a common architecture and that the topological structures are conserved. Strikingly, despite the conservation of the network architecture, TF regulatory networks are highly grade specific, and TF circuitry motifs are dynamically rewired during glioma progression. In addition, the most frequently observed structure in the grade-specific TF networks was the feedforward loop (FFL). We described applications that show how investigating the behaviour of FFLs in glioblastoma can reveal FFLs (such as RARG-NR1I2-CDX2) that are associated with prognosis. We constructed comprehensive TF-TF networks and systematically analysed the circuitry, dynamics, and topological principles of the networks during glioma progression, which will further enhance our understanding of the functions of TFs in glioma.

  4. Medusa structure of the gene regulatory network: dominance of transcription factors in cancer subtype classification.

    PubMed

    Guo, Yuchun; Feng, Ying; Trivedi, Niraj S; Huang, Sui

    2011-05-01

    Gene expression profiles consisting of ten thousands of transcripts are used for clustering of tissue, such as tumors, into subtypes, often without considering the underlying reason that the distinct patterns of expression arise because of constraints in the realization of gene expression profiles imposed by the gene regulatory network. The topology of this network has been suggested to consist of a regulatory core of genes represented most prominently by transcription factors (TFs) and microRNAs, that influence the expression of other genes, and of a periphery of 'enslaved' effector genes that are regulated but not regulating. This 'medusa' architecture implies that the core genes are much stronger determinants of the realized gene expression profiles. To test this hypothesis, we examined the clustering of gene expression profiles into known tumor types to quantitatively demonstrate that TFs, and even more pronounced, microRNAs, are much stronger discriminators of tumor type specific gene expression patterns than a same number of randomly selected or metabolic genes. These findings lend support to the hypothesis of a medusa architecture and of the canalizing nature of regulation by microRNAs. They also reveal the degree of freedom for the expression of peripheral genes that are less stringently associated with a tissue type specific global gene expression profile.

  5. TMREC: A Database of Transcription Factor and MiRNA Regulatory Cascades in Human Diseases

    PubMed Central

    Wang, Shuyuan; Li, Wei; Lian, Baofeng; Liu, Xinyi; Zhang, Yan; Dai, Enyu; Yu, Xuexin; Meng, Fanlin; Jiang, Wei; Li, Xia

    2015-01-01

    Over the past decades, studies have reported that the combinatorial regulation of transcription factors (TFs) and microRNAs (miRNAs) is essential for the appropriate execution of biological events and developmental processes. Dysregulations of these regulators often cause diseases. However, there are no available resources on the regulatory cascades of TFs and miRNAs in the context of human diseases. To fulfill this vacancy, we established the TMREC database in this study. First, we integrated curated transcriptional and post-transcriptional regulations to construct the TF and miRNA regulatory network. Next, we identified all linear paths using the Breadth First Search traversal method. Finally, we used known disease-related genes and miRNAs to measure the strength of association between cascades and diseases. Currently, TMREC consists of 74,248 cascades and 25,194 cascade clusters, involving in 412 TFs, 266 miRNAs and 545 diseases. With the expanding of experimental support regulation data, we will regularly update the database. TMREC aims to help experimental biologists to comprehensively analyse gene expression regulation, to understand the aetiology and to predict novel therapeutic targets.TMREC is freely available at http://bioinfo.hrbmu.edu.cn/TMREC/. PMID:25932650

  6. Molecular characterization of interferon regulatory factor 2 (IRF-2) homolog in pearl oyster Pinctada fucata.

    PubMed

    Huang, Xian-De; Liu, Wen-Guang; Wang, Qi; Zhao, Mi; Wu, Shan-Zeng; Guan, Yun-Yan; Shi, Yu; He, Mao-Xian

    2013-05-01

    Interferon regulatory factors (IRFs) control many facets of the innate and adaptive immune responses, regulate the development of the immune system itself and involve in reproduction and morphogenesis. In the present study, the IRF-2 homology gene, PfIRF-2 from pearl oyster Pinctada fucata was cloned and its genomic structure and promoter were analyzed. PfIRF-2 encodes a putative protein of 350 amino acids, and contains a highly conserved N-terminal DNA-binding domain and a variable C-terminal regulatory domain. Comparison and phylogenetic analysis revealed that PfIRF-2 shared a relatively higher identity with other mollusk but relatively lower identity with vertebrate IRF-2, and was clustered with IRF-1 subfamily composed of IRF-2 and IRF-1. Furthermore, gene expression analysis revealed that PfIRF-2 involved in the immune response to LPS and poly(I:C) stimulation. Immunofluorescence assay showed that the expressed PfIRF-2 was translocated into the nucleus and dual-luciferase reporter assays indicated that PfIRF-2 could involved and activate interferon signaling or NF-κB signal pathway in HEK293 cells. The study of PfIRF-2 may help better understand the innate immune in mollusk.

  7. Regulation of the germinal center gene program by interferon (IFN) regulatory factor 8/IFN consensus sequence-binding protein

    PubMed Central

    Lee, Chang Hoon; Melchers, Mark; Wang, Hongsheng; Torrey, Ted A.; Slota, Rebecca; Qi, Chen-Feng; Kim, Ji Young; Lugar, Patricia; Kong, Hee Jeong; Farrington, Lila; van der Zouwen, Boris; Zhou, Jeff X.; Lougaris, Vassilios; Lipsky, Peter E.; Grammer, Amrie C.; Morse, Herbert C.

    2006-01-01

    Interferon (IFN) consensus sequence-binding protein/IFN regulatory factor 8 (IRF8) is a transcription factor that regulates the differentiation and function of macrophages, granulocytes, and dendritic cells through activation or repression of target genes. Although IRF8 is also expressed in lymphocytes, its roles in B cell and T cell maturation or function are ill defined, and few transcriptional targets are known. Gene expression profiling of human tonsillar B cells and mouse B cell lymphomas showed that IRF8 transcripts were expressed at highest levels in centroblasts, either from secondary lymphoid tissue or transformed cells. In addition, staining for IRF8 was most intense in tonsillar germinal center (GC) dark-zone centroblasts. To discover B cell genes regulated by IRF8, we transfected purified primary tonsillar B cells with enhanced green fluorescent protein–tagged IRF8, generated small interfering RNA knockdowns of IRF8 expression in a mouse B cell lymphoma cell line, and examined the effects of a null mutation of IRF8 on B cells. Each approach identified activation-induced cytidine deaminase (AICDA) and BCL6 as targets of transcriptional activation. Chromatin immunoprecipitation studies demonstrated in vivo occupancy of 5′ sequences of both genes by IRF8 protein. These results suggest previously unappreciated roles for IRF8 in the transcriptional regulation of B cell GC reactions that include direct regulation of AICDA and BCL6. PMID:16380510

  8. Regulation of the germinal center gene program by interferon (IFN) regulatory factor 8/IFN consensus sequence-binding protein.

    PubMed

    Lee, Chang Hoon; Melchers, Mark; Wang, Hongsheng; Torrey, Ted A; Slota, Rebecca; Qi, Chen-Feng; Kim, Ji Young; Lugar, Patricia; Kong, Hee Jeong; Farrington, Lila; van der Zouwen, Boris; Zhou, Jeff X; Lougaris, Vassilios; Lipsky, Peter E; Grammer, Amrie C; Morse, Herbert C

    2006-01-23

    Interferon (IFN) consensus sequence-binding protein/IFN regulatory factor 8 (IRF8) is a transcription factor that regulates the differentiation and function of macrophages, granulocytes, and dendritic cells through activation or repression of target genes. Although IRF8 is also expressed in lymphocytes, its roles in B cell and T cell maturation or function are ill defined, and few transcriptional targets are known. Gene expression profiling of human tonsillar B cells and mouse B cell lymphomas showed that IRF8 transcripts were expressed at highest levels in centroblasts, either from secondary lymphoid tissue or transformed cells. In addition, staining for IRF8 was most intense in tonsillar germinal center (GC) dark-zone centroblasts. To discover B cell genes regulated by IRF8, we transfected purified primary tonsillar B cells with enhanced green fluorescent protein-tagged IRF8, generated small interfering RNA knockdowns of IRF8 expression in a mouse B cell lymphoma cell line, and examined the effects of a null mutation of IRF8 on B cells. Each approach identified activation-induced cytidine deaminase (AICDA) and BCL6 as targets of transcriptional activation. Chromatin immunoprecipitation studies demonstrated in vivo occupancy of 5' sequences of both genes by IRF8 protein. These results suggest previously unappreciated roles for IRF8 in the transcriptional regulation of B cell GC reactions that include direct regulation of AICDA and BCL6.

  9. HIV-1, interferon and the interferon regulatory factor system: an interplay between induction, antiviral responses and viral evasion.

    PubMed

    Marsili, Giulia; Remoli, Anna Lisa; Sgarbanti, Marco; Perrotti, Edvige; Fragale, Alessandra; Battistini, Angela

    2012-01-01

    Thirty years after the first isolation of the etiological agent of AIDS, the virus HIV-1 is still a major threat worldwide with millions of individuals currently infected. Although current combination therapies allow viral replication to be controlled, HIV-1 is not eradicated and persists in drug- and immune system-insensitive reservoirs and a cure is still lacking. Pathogens such as HIV-1 that cause chronic infections are able to adapt to the host in a manner that ensures long term residence and survival, via the evolution of numerous mechanisms that evade various aspects of the innate and adaptive immune response. One such mechanism is targeted to members of the interferon (IFN) regulatory factor (IRF) family of proteins. These transcription factors regulate a variety of biological processes including interferon induction, immune cell activation and downstream pattern recognition receptors (PRRs). HIV-1 renders IRFs harmless and hijacks them to its own advantage in order to facilitate its replication and evasion of immune responses. Type I interferon (IFN), the canonical antiviral innate response, can be induced in both acute and chronic HIV-1 infection in vivo, but in the majority of individuals this initial response is not protective and can contribute to disease progression. Type I IFN expression is largely inhibited in T cells and macrophages in order to successfully establish productive infection, whereas sustained IFN production by plasmacytoid dendritic cells is considered an important source of chronic immune activation, a hallmark to AIDS progression.

  10. Impact of treadmill locomotor training on skeletal muscle IGF1 and myogenic regulatory factors in spinal cord injured rats.

    PubMed

    Liu, Min; Stevens-Lapsley, Jennifer E; Jayaraman, Arun; Ye, Fan; Conover, Christine; Walter, Glenn A; Bose, Prodip; Thompson, Floyd J; Borst, Stephen E; Vandenborne, Krista

    2010-07-01

    The objective of this study was to determine the impact of treadmill locomotor training on the expression of insulin-like growth factor I (IGF1) and changes in myogenic regulatory factors (MRFs) in rat soleus muscle following spinal cord injury (SCI). Moderate, midthoracic (T(8)) contusion SCIs were produced using a NYU (New York University) impactor. Animals were randomly assigned to treadmill training or untrained groups. Rats in the training group were trained starting at 1 week after SCI, for either 3 bouts of 20 min over 1.5 days or 10 bouts over 5 days. Five days of treadmill training completely prevented the decrease in soleus fiber size resulting from SCI. In addition, treadmill training triggered increases in IGF1, MGF and IGFBP4 mRNA expression, and a concurrent reduction of IGFBP5 mRNA in skeletal muscle. Locomotor training also caused an increase in markers of muscle regeneration, including small muscle fibers expressing embryonic myosin and Pax7 positive nuclei and increased expression of the MRFs, myogenin and MyoD. We concluded that treadmill locomotor training ameliorated muscle atrophy in moderate contusion SCI rats. Training-induced muscle regeneration and fiber hypertrophy following SCI was associated with an increase in IGF1, an increase in Pax7 positive nuclei, and upregulation of MRFs.

  11. The Transcription Factor Interferon Regulatory Factor 1 Is Expressed after Cerebral Ischemia and Contributes to Ischemic Brain Injury

    PubMed Central

    Iadecola, Costantino; Salkowski, Cindy A.; Zhang, Fangyi; Aber, Tracy; Nagayama, Masao; Vogel, Stefanie N.; Elizabeth Ross, M.

    1999-01-01

    The transcription factor interferon regulatory factor 1 (IRF-1) is involved in the molecular mechanisms of inflammation and apoptosis, processes that contribute to ischemic brain injury. In this study, the induction of IRF-1 in response to cerebral ischemia and its role in ischemic brain injury were investigated. IRF-1 gene expression was markedly upregulated within 12 h of occlusion of the middle cerebral artery in C57BL/6 mice. The expression reached a peak 4 d after ischemia (6.0 ± 1.8-fold; P < 0.001) and was restricted to the ischemic regions of the brain. The volume of ischemic injury was reduced by 23 ± 3% in IRF-1+/− and by 46 ± 9% in IRF-1−/− mice (P < 0.05). The reduction in infarct volume was paralleled by a substantial attenuation in neurological deficits. Thus, IRF-1 is the first nuclear transacting factor demonstrated to contribute directly to cerebral ischemic damage and may be a novel therapeutic target in ischemic stroke. PMID:9989987

  12. Dendritic Cells in the Periphery Control Antigen-Specific Natural and Induced Regulatory T Cells

    PubMed Central

    Yamazaki, Sayuri; Morita, Akimichi

    2013-01-01

    Dendritic cells (DCs) are specialized antigen-presenting cells that regulate both immunity and tolerance. DCs in the periphery play a key role in expanding naturally occurring Foxp3+ CD25+ CD4+ regulatory T cells (Natural T-regs) and inducing Foxp3 expression (Induced T-regs) in Foxp3− CD4+ T cells. DCs are phenotypically and functionally heterogeneous, and further classified into several subsets depending on distinct marker expression and their location. Recent findings indicate the presence of specialized DC subsets that act to expand Natural T-regs or induce Foxp3+ T-regs from Foxp3− CD4+ T cells. For example, two major subsets of DCs in lymphoid organs act differentially in inducing Foxp3+ T-regs from Foxp3− cells or expanding Natural T-regs with model-antigen delivery by anti-DC subset monoclonal antibodies in vivo. Furthermore, DCs expressing CD103 in the intestine induce Foxp3+ T-regs from Foxp3− CD4+ T cells with endogenous TGF-β and retinoic acid. In addition, antigen-presenting DCs have a capacity to generate Foxp3+ T-regs in the oral cavity where many antigens and commensals exist, similar to intestine and skin. In skin and skin-draining lymph nodes, at least six DC subsets have been identified, suggesting a complex DC-T-reg network. Here, we will review the specific activity of DCs in expanding Natural T-regs and inducing Foxp3+ T-regs from Foxp3− precursors, and further discuss the critical function of DCs in maintaining tolerance at various locations including skin and oral cavity. PMID:23801989

  13. Somite subdomains, muscle cell origins, and the four muscle regulatory factor proteins

    PubMed Central

    1994-01-01

    We show by immunohistology that distinct expression patterns of the four muscle regulatory factor (MRF) proteins identify subdomains of mouse somites. Myf-5 and MyoD are, at specific stages, each expressed in both myotome and dermatome cells. Myf-5 expression is initially restricted to dorsal cells in all somites, as is MyoD expression in neck somites. In trunk somites, however, MyoD is initially expressed in ventral cells. Myogenin and MRF4 are restricted to myotome cells, though the MRF4-expressing cells are initially less widely distributed than the myogenin-expressing cells, which are at all stages found throughout the myotome. All somitic myocytes express one or more MRFs. The transiently distinct expression patterns of the four MRF proteins identify dorsal and ventral subdomains of somites, and suggest that skeletal muscle cells in somites originate at multiple sites and via multiple molecular pathways. PMID:7929574

  14. Machine learning for regulatory analysis and transcription factor target prediction in yeast

    PubMed Central

    Holloway, Dustin T.; Kon, Mark

    2006-01-01

    High throughput technologies, including array-based chromatin immunoprecipitation, have rapidly increased our knowledge of transcriptional maps—the identity and location of regulatory binding sites within genomes. Still, the full identification of sites, even in lower eukaryotes, remains largely incomplete. In this paper we develop a supervised learning approach to site identification using support vector machines (SVMs) to combine 26 different data types. A comparison with the standard approach to site identification using position specific scoring matrices (PSSMs) for a set of 104 Saccharomyces cerevisiae regulators indicates that our SVM-based target classification is more sensitive (73 vs. 20%) when specificity and positive predictive value are the same. We have applied our SVM classifier for each transcriptional regulator to all promoters in the yeast genome to obtain thousands of new targets, which are currently being analyzed and refined to limit the risk of classifier over-fitting. For the purpose of illustration we discuss several results, including biochemical pathway predictions for Gcn4 and Rap1. For both transcription factors SVM predictions match well with the known biology of control mechanisms, and possible new roles for these factors are suggested, such as a function for Rap1 in regulating fermentative growth. We also examine the promoter melting temperature curves for the targets of YJR060W, and show that targets of this TF have potentially unique physical properties which distinguish them from other genes. The SVM output automatically provides the means to rank dataset features to identify important biological elements. We use this property to rank classifying k-mers, thereby reconstructing known binding sites for several TFs, and to rank expression experiments, determining the conditions under which Fhl1, the factor responsible for expression of ribosomal protein genes, is active. We can see that targets of Fhl1 are differentially expressed in

  15. Drug-induced QT interval shortening: potential harbinger of proarrhythmia and regulatory perspectives.

    PubMed

    Shah, Rashmi R

    2010-01-01

    ATP-dependent potassium channel openers such as pinacidil and levcromakalim have long been known to shorten action potential duration and to be profibrillatory in non-clinical models, raising concerns on the clinical safety of drugs that shorten QT interval. Routine non-clinical evaluation of new drugs for their potential to affect cardiac repolarization has revealed that drugs may also shorten QT interval. The description of congenital short QT syndrome in 2000, together with the associated arrhythmias, suggests that drug-induced short QT interval may be proarrhythmic, and an uncanny parallel is evolving between our appreciation of the short and the long QT intervals. Epidemiological studies report an over-representation of short QT interval values in patients with idiopathic ventricular fibrillation. Therefore, as new compounds that shorten QT interval are progressed further into clinical development, questions will inevitably arise on their safety. Arising from the current risk-averse clinical and regulatory environment and concerns on proarrhythmic safety of drugs, together with our lack of a better understanding of the clinical significance of short QT interval, new drugs that substantially shorten QT interval will likely receive an unfavourable regulatory review unless these drugs fulfil an unmet clinical need. This review provides estimates of parameters of QT shortening that may be of potential clinical significance. Rufinamide, a recently approved anticonvulsant, illustrates the current regulatory approach to drugs that shorten QT interval. However, to further substantiate or confirm the safety of these drugs, their approval may well be conditional upon large-scale post-marketing studies with a focus on cardiac safety. PMID:19563537

  16. Decidual vascular endothelial cells promote maternal-fetal immune tolerance by inducing regulatory T cells through canonical Notch1 signaling.

    PubMed

    Yao, Yanyi; Song, Jieping; Wang, Weipeng; Liu, Nian

    2016-05-01

    Adaptation of the maternal immune response to accommodate the semiallogeneic fetus is necessary for pregnancy success. However, the mechanisms by which the fetus avoids rejection despite expression of paternal alloantigens remain incompletely understood. Regulatory T cells (Treg cells) are pivotal for maintaining immune homeostasis, preventing autoimmune disease and fetus rejection. In this study, we found that maternal decidual vascular endothelial cells (DVECs) sustained Foxp3 expression in resting Treg cells in vitro. Moreover, under in vitro Treg cell induction condition with agonistic antibodies and transforming growth factor (TGF)-β, DVECs promoted Treg cell differentiation from non-Treg conventional T cells. Consistent with the promotion of Treg cell maintenance and differentiation, Treg cell-associated gene expression such as TGF-β, Epstein-Barr-induced gene-3, CD39 and glucocorticoid-induced tumor necrosis factor receptor was also increased in the presence of DVECs. Further study revealed that DVECs expressed Notch ligands such as Jagged-1, Delta-like protein 1 (DLL-1) and DLL-4, while Treg cells expressed Notch1 on their surface. The effects of DVECs on Treg cells was inhibited by siRNA-induced knockdown of expression of Jagged-1 and DLL-1 in DVECs. Downregulation of Notch1 in Treg cells using lentiviral shRNA transduction decreased Foxp3 expression in Treg cells. Adoptive transfer of Notch1-deficient Treg cells increased abortion rate in a murine semiallogeneic pregnancy model. Taken together, our study suggests that maternal DVECs are able to maintain decidual Treg cell identity and promote Treg cell differentiation through activation of Notch1 signal pathway in Treg cells and subsequently inhibit the immune response against semiallogeneic fetuses and preventing spontaneous abortion. PMID:26714886

  17. Structural integration in hypoxia-inducible factors

    SciTech Connect

    Wu, Dalei; Potluri, Nalini; Lu, Jingping; Kim, Youngchang; Rastinejad, Fraydoon

    2015-08-20

    The hypoxia-inducible factors (HIFs) coordinate cellular adaptations to low oxygen stress by regulating transcriptional programs in erythropoiesis, angiogenesis and metabolism. These programs promote the growth and progression of many tumours, making HIFs attractive anticancer targets. Transcriptionally active HIFs consist of HIF-alpha and ARNT (also called HIF-1 beta) subunits. Here we describe crystal structures for each of mouse HIF-2 alpha-ARNT and HIF-1 alpha-ARNT heterodimers in states that include bound small molecules and their hypoxia response element. A highly integrated quaternary architecture is shared by HIF-2 alpha-ARNT and HIF-1 alpha-ARNT, wherein ARNT spirals around the outside of each HIF-alpha subunit. Five distinct pockets are observed that permit small-molecule binding, including PAS domain encapsulated sites and an interfacial cavity formed through subunit heterodimerization. The DNA-reading head rotates, extends and cooperates with a distal PAS domain to bind hypoxia response elements. HIF-alpha mutations linked to human cancers map to sensitive sites that establish DNA binding and the stability of PAS domains and pockets.

  18. Intracellular Concentrations of 65 Species of Transcription Factors with Known Regulatory Functions in Escherichia coli

    PubMed Central

    Kori, Ayako; Koshio, Etsuko; Yamada, Kayoko; Maeda, Hiroto; Shimada, Tomohiro; Makinoshima, Hideki; Iwata, Akira; Fujita, Nobuyuki

    2014-01-01

    The expression pattern of the Escherichia coli genome is controlled in part by regulating the utilization of a limited number of RNA polymerases among a total of its approximately 4,600 genes. The distribution pattern of RNA polymerase changes from modulation of two types of protein-protein interactions: the interaction of core RNA polymerase with seven species of the sigma subunit for differential promoter recognition and the interaction of RNA polymerase holoenzyme with about 300 different species of transcription factors (TFs) with regulatory functions. We have been involved in the systematic search for the target promoters recognized by each sigma factor and each TF using the newly developed Genomic SELEX system. In parallel, we developed the promoter-specific (PS)-TF screening system for identification of the whole set of TFs involved in regulation of each promoter. Understanding the regulation of genome transcription also requires knowing the intracellular concentrations of the sigma subunits and TFs under various growth conditions. This report describes the intracellular levels of 65 species of TF with known function in E. coli K-12 W3110 at various phases of cell growth and at various temperatures. The list of intracellular concentrations of the sigma factors and TFs provides a community resource for understanding the transcription regulation of E. coli under various stressful conditions in nature. PMID:24837290

  19. Autoimmune manifestations in human myelodysplasia: a positive correlation with interferon regulatory factor-1 (IRF-1) expression

    PubMed Central

    Giannouli, S; Tzoanopoulos, D; Ritis, K; Kartalis, G; Moutsopoulos, H; Voulgarelis, M

    2004-01-01

    Background: Patients with myelodysplasia may have autoimmune manifestations (AIM). Interferon regulatory factor-1 (IRF-1) is a transcription factor involved in interferon signalling, leukaemogenesis, and the development of the immune system. Objectives: To determine whether IRF-1 is implicated in the pathophysiology of AIM in myelodysplasia. Methods: 14 patients with myelodysplasia were studied, seven with AIM and seven without. Five patients with vasculitis and seven normal subjects served as controls. The expression of IRF-1 was studied in bone marrow mononuclear cells taken from patients and controls, using a relative quantitative reverse transcriptase polymerase chain reaction. Results: A 10-fold reduction in full length IRF-1 mRNA was detected in the myelodysplasia patients without AIM compared with the normal controls. In contrast, the group with AIM had increased IRF-1 transcripts, to a level almost equal to that observed in patients with vasculitis and normal controls. Conclusions: Myelodysplasia patients without IRF-1 expression had a decreased incidence of AIM. Thus the absence of IRF-1 transcription factor appears to protect against the development of autoimmunity in myelodysplasia. PMID:15082491

  20. Inferring regulatory element landscapes and transcription factor networks from cancer methylomes.

    PubMed

    Yao, Lijing; Shen, Hui; Laird, Peter W; Farnham, Peggy J; Berman, Benjamin P

    2015-05-21

    Recent studies indicate that DNA methylation can be used to identify transcriptional enhancers, but no systematic approach has been developed for genome-wide identification and analysis of enhancers based on DNA methylation. We describe ELMER (Enhancer Linking by Methylation/Expression Relationships), an R-based tool that uses DNA methylation to identify enhancers and correlates enhancer state with expression of nearby genes to identify transcriptional targets. Transcription factor motif analysis of enhancers is coupled with expression analysis of transcription factors to infer upstream regulators. Using ELMER, we investigated more than 2,000 tumor samples from The Cancer Genome Atlas. We identified networks regulated by known cancer drivers such as GATA3 and FOXA1 (breast cancer), SOX17 and FOXA2 (endometrial cancer), and NFE2L2, SOX2, and TP63 (squamous cell lung cancer). We also identified novel networks with prognostic associations, including RUNX1 in kidney cancer. We propose ELMER as a powerful new paradigm for understanding the cis-regulatory interface between cancer-associated transcription factors and their functional target genes.

  1. Behavioral and regulatory abnormalities in mice deficient in the NPAS1 and NPAS3 transcription factors.

    PubMed

    Erbel-Sieler, Claudia; Dudley, Carol; Zhou, Yudong; Wu, Xinle; Estill, Sandi Jo; Han, Tina; Diaz-Arrastia, Ramon; Brunskill, Eric W; Potter, S Steven; McKnight, Steven L

    2004-09-14

    Laboratory mice bearing inactivating mutations in the genes encoding the NPAS1 and NPAS3 transcription factors have been shown to exhibit a spectrum of behavioral and neurochemical abnormalities. Behavioral abnormalities included diminished startle response, as measured by prepulse inhibition, and impaired social recognition. NPAS1/NPAS3-deficient mice also exhibited stereotypic darting behavior at weaning and increased locomotor activity. Immunohistochemical staining assays showed that the NPAS1 and NPAS3 proteins are expressed in inhibitory interneurons and that the viability and anatomical distribution of these neurons are unaffected by the absence of either transcription factor. Adult brain tissues from NPAS3- and NPAS1/NPAS3-deficient mice exhibited a distinct reduction in reelin, a large, secreted protein whose expression has been reported to be attenuated in the postmortem brain tissue of patients with schizophrenia. These observations raise the possibility that a regulatory program controlled in inhibitory interneurons by the NPAS1 and NPAS3 transcription factors may be either substantively or tangentially relevant to psychosis.

  2. Behavioral and regulatory abnormalities in mice deficient in the NPAS1 and NPAS3 transcription factors.

    PubMed

    Erbel-Sieler, Claudia; Dudley, Carol; Zhou, Yudong; Wu, Xinle; Estill, Sandi Jo; Han, Tina; Diaz-Arrastia, Ramon; Brunskill, Eric W; Potter, S Steven; McKnight, Steven L

    2004-09-14

    Laboratory mice bearing inactivating mutations in the genes encoding the NPAS1 and NPAS3 transcription factors have been shown to exhibit a spectrum of behavioral and neurochemical abnormalities. Behavioral abnormalities included diminished startle response, as measured by prepulse inhibition, and impaired social recognition. NPAS1/NPAS3-deficient mice also exhibited stereotypic darting behavior at weaning and increased locomotor activity. Immunohistochemical staining assays showed that the NPAS1 and NPAS3 proteins are expressed in inhibitory interneurons and that the viability and anatomical distribution of these neurons are unaffected by the absence of either transcription factor. Adult brain tissues from NPAS3- and NPAS1/NPAS3-deficient mice exhibited a distinct reduction in reelin, a large, secreted protein whose expression has been reported to be attenuated in the postmortem brain tissue of patients with schizophrenia. These observations raise the possibility that a regulatory program controlled in inhibitory interneurons by the NPAS1 and NPAS3 transcription factors may be either substantively or tangentially relevant to psychosis. PMID:15347806

  3. Individual interferon regulatory factor-3 thiol residues are not critical for its activation following virus infection.

    PubMed

    Zucchini, Nicolas; Williams, Virginie; Grandvaux, Nathalie

    2012-09-01

    The interferon regulatory factor (IRF)-3 transcription factor plays a central role in the capacity of the host to mount an efficient innate antiviral immune defense, mainly through the regulation of type I Interferon genes. A tight regulation of IRF-3 is crucial for an adapted intensity and duration of the response. Redox-dependent processes are now well known to regulate signaling cascades. Recent reports have revealed that signaling molecules upstream of IRF-3, including the mitochondrial antiviral-signalling protein (MAVS) and the TNF receptor associated factors (TRAFs) adaptors, are sensitive to redox regulation. In the present study, we assessed whether redox regulation of thiol residues contained in IRF-3, which are priviledged redox sensors, play a role in its regulation following Sendai virus infection, using a combination of mutation of Cysteine (Cys) residues into Alanine and thiols alkylation using N-ethyl maleimide. Alkylation of IRF-3 on Cys289 appears to destabilize IRF-3 dimer in vitro. However, a detailed analysis of IRF-3 phosphorylation, dimerization, nuclear accumulation, and induction of target gene promoter in vivo led us to conclude that IRF-3 specific, individual Cys residues redox status does not play an essential role in its activation in vivo.

  4. Abnormal skin, limb and craniofacial morphogenesis in mice deficient for interferon regulatory factor 6 (Irf6)

    PubMed Central

    Ingraham, Christopher R; Kinoshita, Akira; Kondo, Shinji; Yang, Baoli; Sajan, Samin; Trout, Kurt J; Malik, Margaret I; Dunnwald, Martine; Goudy, Stephen L; Lovett, Michael; Murray, Jeffrey C; Schutte, Brian C

    2007-01-01

    Transcription factor paralogs may share a common role in staged or overlapping expression in specific tissues, as in the Hox family. In other cases, family members have distinct roles in a range of embryologic, differentiation or response pathways (as in the Tbx and Pax families). For the interferon regulatory factor (IRF) family of transcription factors, mice deficient in Irf1, Irf2, Irf3, Irf4, Irf5, Irf7, Irf8 or Irf9 have defects in the immune response but show no embryologic abnormalities1–7. Mice deficient for Irf6 have not been reported, but in humans, mutations in IRF6 cause two mendelian orofacial clefting syndromes8–10, and genetic variation in IRF6 confers risk for isolated cleft lip and palate11–15. Here we report that mice deficient for Irf6 have abnormal skin, limb and craniofacial development. Histological and gene expression analyses indicate that the primary defect is in keratinocyte differentiation and proliferation. This study describes a new role for an IRF family member in epidermal development. PMID:17041601

  5. Regulation of Peroxisome Proliferator-Activated Receptor γ Expression by Adipocyte Differentiation and Determination Factor 1/Sterol Regulatory Element Binding Protein 1: Implications for Adipocyte Differentiation and Metabolism

    PubMed Central

    Fajas, Lluis; Schoonjans, Kristina; Gelman, Laurent; Kim, Jae B.; Najib, Jamila; Martin, Genevieve; Fruchart, Jean-Charles; Briggs, Michael; Spiegelman, Bruce M.; Auwerx, Johan

    1999-01-01

    Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor implicated in adipocyte differentiation and insulin sensitivity. We investigated whether PPARγ expression is dependent on the activity of adipocyte differentiation and determination factor 1/sterol regulatory element binding protein 1 (ADD-1/SREBP-1), another transcription factor associated with both adipocyte differentiation and cholesterol homeostasis. Ectopic expression of ADD-1/SREBP-1 in 3T3-L1 and HepG2 cells induced endogenous PPARγ mRNA levels. The related transcription factor SREBP-2 likewise induced PPARγ expression. In addition, cholesterol depletion, a condition known to result in proteolytic activation of transcription factors of the SREBP family, induced PPARγ expression and improved PPRE-driven transcription. The effect of the SREBPs on PPARγ expression was mediated through the PPARγ1 and -3 promoters. Both promoters contain a consensus E-box motif that mediates the regulation of the PPARγ gene by ADD-1/SREBP-1 and SREBP-2. These results suggest that PPARγ expression can be controlled by the SREBP family of transcription factors and demonstrate new interactions between transcription factors that can regulate different pathways of lipid metabolism. PMID:10409739

  6. Xanthohumol Improves Diet-induced Obesity and Fatty Liver by Suppressing Sterol Regulatory Element-binding Protein (SREBP) Activation*

    PubMed Central

    Miyata, Shingo; Inoue, Jun; Shimizu, Makoto; Sato, Ryuichiro

    2015-01-01

    Sterol regulatory element-binding proteins (SREBPs) are key transcription factors that stimulate the expression of genes involved in fatty acid and cholesterol biosynthesis. Here, we demonstrate that a prenylated flavonoid in hops, xanthohumol (XN), is a novel SREBP inactivator that reduces the de novo synthesis of fatty acid and cholesterol. XN independently suppressed the maturation of SREBPs of insulin-induced genes in a manner different from sterols. Our results suggest that XN impairs the endoplasmic reticulum-to-Golgi translocation of the SREBP cleavage-activating protein (SCAP)-SREBP complex by binding to Sec23/24 and blocking SCAP/SREBP incorporation into common coated protein II vesicles. Furthermore, in diet-induced obese mice, dietary XN suppressed SREBP-1 target gene expression in the liver accompanied by a reduction of the mature form of hepatic SREBP-1, and it inhibited the development of obesity and hepatic steatosis. Altogether, our data suggest that XN attenuates the function of SREBP-1 by repressing its maturation and that it has the potential of becoming a nutraceutical food or pharmacological agent for improving metabolic syndrome. PMID:26140926

  7. Production of Tuber-Inducing Factor

    NASA Technical Reports Server (NTRS)

    Stutte, Gary W.; Yorio, Neil C.

    2006-01-01

    A process for making a substance that regulates the growth of potatoes and some other economically important plants has been developed. The process also yields an economically important by-product: potatoes. The particular growth-regulating substance, denoted tuber-inducing factor (TIF), is made naturally by, and acts naturally on, potato plants. The primary effects of TIF on potato plants are reducing the lengths of the main shoots, reducing the numbers of nodes on the main stems, reducing the total biomass, accelerating the initiation of potatoes, and increasing the edible fraction (potatoes) of the overall biomass. To some extent, these effects of TIF can override environmental effects that typically inhibit the formation of tubers. TIF can be used in the potato industry to reduce growth time and increase harvest efficiency. Other plants that have been observed to be affected by TIF include tomatoes, peppers, radishes, eggplants, marigolds, and morning glories. In the present process, potatoes are grown with their roots and stolons immersed in a nutrient solution in a recirculating hydroponic system. From time to time, a nutrient replenishment solution is added to the recirculating nutrient solution to maintain the required nutrient concentration, water is added to replace water lost from the recirculating solution through transpiration, and an acid or base is added, as needed, to maintain the recirculating solution at a desired pH level. The growing potato plants secrete TIF into the recirculating solution. The concentration of TIF in the solution gradually increases to a range in which the TIF regulates the growth of the plants.

  8. Imbalanced signal transduction in regulatory T cells expressing the transcription factor FoxP3

    PubMed Central

    Yan, Dapeng; Farache, Julia; Mingueneau, Michael; Mathis, Diane; Benoist, Christophe

    2015-01-01

    FoxP3+ T regulatory (Treg) cells have a fundamental role in immunological tolerance, with transcriptional and functional phenotypes that demarcate them from conventional CD4+ T cells (Tconv). Differences between these two lineages in the signaling downstream of T-cell receptor-triggered activation have been reported, and there are different requirements for some signaling factors. Seeking a comprehensive view, we found that Treg cells have a broadly dampened activation of several pathways and signaling nodes upon TCR-mediated activation, with low phosphorylation of CD3ζ, SLP76, Erk1/2, AKT, or S6 and lower calcium flux. In contrast, STAT phosphorylation triggered by interferons, IL2 or IL6, showed variations between Treg and Tconv in magnitude or choice of preferential STAT activation but no general Treg signaling defect. Much, but not all, of the Treg/Tconv difference in TCR-triggered responses could be attributed to lower responsiveness of antigen-experienced cells with CD44hi or CD62Llo phenotypes, which form a greater proportion of the Treg pool. Candidate regulators were tested, but the Treg/Tconv differential could not be explained by overexpression in Treg cells of the signaling modulator CD5, the coinhibitors PD-1 and CTLA4, or the regulatory phosphatase DUSP4. However, transcriptome profiling in Dusp4-deficient mice showed that DUSP4 enhances the expression of a segment of the canonical Treg transcriptional signature, which partially overlaps with the TCR-dependent Treg gene set. Thus, Treg cells, likely because of their intrinsically higher reactivity to self, tune down TCR signals but seem comparatively more attuned to cytokines or other intercellular signals. PMID:26627244

  9. Discovering transcription factor regulatory targets using gene expression and binding data

    PubMed Central

    Maienschein-Cline, Mark; Zhou, Jie; White, Kevin P.; Sciammas, Roger; Dinner, Aaron R.

    2012-01-01

    Motivation: Identifying the target genes regulated by transcription factors (TFs) is the most basic step in understanding gene regulation. Recent advances in high-throughput sequencing technology, together with chromatin immunoprecipitation (ChIP), enable mapping TF binding sites genome wide, but it is not possible to infer function from binding alone. This is especially true in mammalian systems, where regulation often occurs through long-range enhancers in gene-rich neighborhoods, rather than proximal promoters, preventing straightforward assignment of a binding site to a target gene. Results: We present EMBER (Expectation Maximization of Binding and Expression pRofiles), a method that integrates high-throughput binding data (e.g. ChIP-chip or ChIP-seq) with gene expression data (e.g. DNA microarray) via an unsupervised machine learning algorithm for inferring the gene targets of sets of TF binding sites. Genes selected are those that match overrepresented expression patterns, which can be used to provide information about multiple TF regulatory modes. We apply the method to genome-wide human breast cancer data and demonstrate that EMBER confirms a role for the TFs estrogen receptor alpha, retinoic acid receptors alpha and gamma in breast cancer development, whereas the conventional approach of assigning regulatory targets based on proximity does not. Additionally, we compare several predicted target genes from EMBER to interactions inferred previously, examine combinatorial effects of TFs on gene regulation and illustrate the ability of EMBER to discover multiple modes of regulation. Availability: All code used for this work is available at http://dinner-group.uchicago.edu/downloads.html Contact: dinner@uchicago.edu Supplementary Information: Supplementary data are available at Bioinformatics online. PMID:22084256

  10. Restricted maternal nutrition alters myogenic regulatory factor expression in satellite cells of ovine offspring.

    PubMed

    Raja, J S; Hoffman, M L; Govoni, K E; Zinn, S A; Reed, S A

    2016-07-01

    Poor maternal nutrition inhibits muscle development and postnatal muscle growth. Satellite cells are myogenic precursor cells that contribute to postnatal muscle growth, and their activity can be evaluated by the expression of several transcription factors. Paired-box (Pax)7 is expressed in quiescent and active satellite cells. MyoD is expressed in activated and proliferating satellite cells and myogenin is expressed in terminally differentiating cells. Disruption in the expression pattern or timing of expression of myogenic regulatory factors negatively affects muscle development and growth. We hypothesized that poor maternal nutrition during gestation would alter the in vitro temporal expression of MyoD and myogenin in satellite cells from offspring at birth and 3 months of age. Ewes were fed 100% or 60% of NRC requirements from day 31±1.3 of gestation. Lambs from control-fed (CON) or restricted-fed (RES) ewes were euthanized within 24 h of birth (birth; n=5) or were fed a control diet until 3 months of age (n=5). Satellite cells isolated from the semitendinosus muscle were used for gene expression analysis or cultured for 24, 48 or 72 h and immunostained for Pax7, MyoD or myogenin. Fusion index was calculated from a subset of cells allowed to differentiate. Compared with CON, temporal expression of MyoD and myogenin was altered in cultured satellite cells isolated from RES lambs at birth. The percent of cells expressing MyoD was greater in RES than CON (P=0.03) after 24 h in culture. After 48 h of culture, there was a greater percent of cells expressing myogenin in RES compared with CON (P0.05). In satellite cells from RES lambs at 3 months of age, the percent of cells expressing MyoD and myogenin were greater than CON after 72 h in culture (P<0.05). Fusion index was reduced in RES lambs at 3 months of age compared with CON (P<0.001). Restricted nutrition during gestation alters the temporal expression of myogenic regulatory factors in satellite cells of the

  11. Multiple Ets factors and interferon regulatory factor-4 modulate CD68 expression in a cell type-specific manner.

    PubMed

    O'Reilly, Dawn; Quinn, Carmel M; El-Shanawany, Tariq; Gordon, Siamon; Greaves, David R

    2003-06-13

    CD68 is a transmembrane glycoprotein expressed in all cells of the mononuclear phagocyte lineage including monocytes and tissue resident macrophages. Deletion analysis of the 5'-flanking sequences of the gene demonstrated that the proximal -150-bp sequence of the CD68 promoter exhibits high level promoter activity in macrophages. Mutations that abolish Ets factor binding at positions -106 and -89 reduce promoter activity in macrophages to 12 and 30%, respectively. Band shift experiments show that PU.1 associates with the -89 site whereas, Elf-1 preferentially binds the -106 Ets binding site and enhances CD68 activity in vitro. Furthermore, chromatin immunoprecipitation experiments confirm that Elf-1 and PU.1 associate with the CD68 proximal promoter in vivo in THP-1 cells. PU.1 does not bind to the CD68 promoter alone but instead forms heterocomplexes with members of the interferon regulatory factor family (IRF) including IRF-4 and IRF-8. IRF-4 and IRF-8 typically mediate transcriptional activation when associated with PU.1 on composite elements. However, our data show that PU.1/IRF-4 and IRF-8 heterocomplexes down-regulate CD68 promoter activity in macrophages and repression is dependent on the integrity of both the IRF and PU.1 half-sites of this composite element. Chromatin immunoprecipitation data reveal that neither IRF-4 nor IRF-8 associate with the CD68 proximal promoter in macrophages in vivo but IRF-4 is associated with the promoter in B lymphocytes. We propose that expression of CD68 in myeloid cells requires the Ets transcription factors Elf-1 and PU.1 and CD68 expression is down-regulated in lymphoid cells by combinatorial interactions between PU.1 and IRF-4.

  12. Interferon Regulatory Factor 4 controls TH1 cell effector function and metabolism

    PubMed Central

    Mahnke, Justus; Schumacher, Valéa; Ahrens, Stefanie; Käding, Nadja; Feldhoff, Lea Marie; Huber, Magdalena; Rupp, Jan; Raczkowski, Friederike; Mittrücker, Hans-Willi

    2016-01-01

    The transcription factor Interferon Regulatory Factor 4 (IRF4) is essential for TH2 and TH17 cell formation and controls peripheral CD8+ T cell differentiation. We used Listeria monocytogenes infection to characterize the function of IRF4 in TH1 responses. IRF4−/− mice generated only marginal numbers of listeria-specific TH1 cells. After transfer into infected mice, IRF4−/− CD4+ T cells failed to differentiate into TH1 cells as indicated by reduced T-bet and IFN-γ expression, and showed limited proliferation. Activated IRF4−/− CD4+ T cells exhibited diminished uptake of the glucose analog 2-NBDG, limited oxidative phosphorylation and strongly reduced aerobic glycolysis. Insufficient metabolic adaptation contributed to the limited proliferation and TH1 differentiation of IRF4−/− CD4+ T cells. Our study identifies IRF4 as central regulator of TH1 responses and cellular metabolism. We propose that this function of IRF4 is fundamental for the initiation and maintenance of all TH cell responses. PMID:27762344

  13. Specific detection of interferon regulatory factor 5 (IRF5): A case of antibody inequality

    PubMed Central

    Li, Dan; De, Saurav; Li, Dan; Song, Su; Matta, Bharati; Barnes, Betsy J.

    2016-01-01

    Interferon regulatory factor 5 (IRF5) is a member of the IRF family of transcription factors. IRF5 was first identified and characterized as a transcriptional regulator of type I interferon expression after virus infection. In addition to its critical role(s) in the regulation and development of host immunity, subsequent studies revealed important roles for IRF5 in autoimmunity, cancer, obesity, pain, cardiovascular disease, and metabolism. Based on these important disease-related findings, a large number of commercial antibodies have become available to study the expression and function of IRF5. Here we validate a number of these antibodies for the detection of IRF5 by immunoblot, flow cytometry, and immunofluorescence or immunohistochemistry using well-established positive and negative controls. Somewhat surprising, the majority of commercial antibodies tested were unable to specifically recognize human or mouse IRF5. We present data on antibodies that do specifically recognize human or mouse IRF5 in a particular application. These findings reiterate the importance of proper controls and molecular weight standards for the analysis of protein expression. Given that dysregulated IRF5 expression has been implicated in the pathogenesis of numerous diseases, including autoimmune and cancer, results indicate that caution should be used in the evaluation and interpretation of IRF5 expression analysis. PMID:27481535

  14. Aberrant expression of interferon regulatory factor 3 in human lung cancer

    SciTech Connect

    Tokunaga, Takayuki; Naruke, Yuki; Shigematsu, Sayuri; Kohno, Tomoko; Yasui, Kiyoshi; Ma, Yuhua; Chua, Koon Jiew; Katayama, Ikuo; Nakamura, Takashi; Hishikawa, Yoshitaka; Koji, Takehiko; Yatabe, Yasushi; Nagayasu, Takeshi; Fujita, Takashi; Matsuyama, Toshifumi; and others

    2010-06-25

    We analyzed the subcellular distributions and gene structures of interferon regulatory factor 3 (IRF3) transcription factor in 50 cases of human primary lung cancer. The immunohistochemical analyses revealed substantially aberrant IRF3 expression specific to the cancer lesions (2 and 6 tumors with nuclear staining, and 4 and 5 tumors with negative staining, in adenocarcinoma and squamous cell carcinoma, respectively), while the morphologically normal region around the tumors exhibited only cytoplasmic staining. In addition, we determined the sequence of the entire IRF3 coding region, and found two novel variants with the amino acid changes (S{sup 175}(AGC) {yields} R{sup 175}(CGC) and A{sup 208}(GCC) {yields} D{sup 208}(GAC)). The R{sup 175} variant was also detected in a morphologically normal region around the nuclear staining squamous cell carcinoma, and exhibited almost the same functions as the wild type IRF3. On the other hand, the D{sup 208} variant, found in the negative staining squamous cell carcinoma cases, reduced the nuclear translocation in response to I{kappa}B kinase {epsilon} stimulation, as compared to the wild type IRF3, but the same variant was detected in the surrounding morphologically normal region. The aberrant expression of IRF3 and the novel D{sup 208} variant may provide clues to elucidate the etiology of primary lung cancer.

  15. Origin of a novel regulatory module by duplication and degeneration of an ancient plant transcription factor.

    PubMed

    Floyd, Sandra K; Ryan, Joseph G; Conway, Stephanie J; Brenner, Eric; Burris, Kellie P; Burris, Jason N; Chen, Tao; Edger, Patrick P; Graham, Sean W; Leebens-Mack, James H; Pires, J Chris; Rothfels, Carl J; Sigel, Erin M; Stevenson, Dennis W; Neal Stewart, C; Wong, Gane Ka-Shu; Bowman, John L

    2014-12-01

    It is commonly believed that gene duplications provide the raw material for morphological evolution. Both the number of genes and size of gene families have increased during the diversification of land plants. Several small proteins that regulate transcription factors have recently been identified in plants, including the LITTLE ZIPPER (ZPR) proteins. ZPRs are post-translational negative regulators, via heterodimerization, of class III Homeodomain Leucine Zipper (C3HDZ) proteins that play a key role in directing plant form and growth. We show that ZPR genes originated as a duplication of a C3HDZ transcription factor paralog in the common ancestor of euphyllophytes (ferns and seed plants). The ZPRs evolved by degenerative mutations resulting in loss all of the C3HDZ functional domains, except the leucine zipper that modulates dimerization. ZPRs represent a novel regulatory module of the C3HDZ network unique to the euphyllophyte lineage, and their origin correlates to a period of rapid morphological changes and increased complexity in land plants. The origin of the ZPRs illustrates the significance of gene duplications in creating developmental complexity during land plant evolution that likely led to morphological evolution. PMID:25263420

  16. The Transcription Factor Interferon Regulatory Factor-1 (IRF1) Plays a Key Role in the Terminal Effector Pathways of Human Preterm Labor.

    PubMed

    Lim, Ratana; Tran, Ha Thi; Liong, Stella; Barker, Gillian; Lappas, Martha

    2016-02-01

    Preterm birth is the largest single cause of neonatal death and morbidity. By activating cytokine- and Toll-like receptor (TLR)-signaling pathways, infection and/or inflammation are strongly associated with preterm delivery. Interferon regulatory factor-1 (IRF1) is an important regulator of the inflammatory response. The aims of this study were to establish the effect of 1) labor on IRF1 expression in human fetal membranes and myometrium, 2) prolabor mediators on IRF1 expression and activity, and 3) IRF1 small interfering RNA on the expression of prolabor mediators. IRF1 expression was higher in fetal membranes and myometrium after spontaneous term labor and in preterm fetal membranes with infection. The proinflammatory cytokine IL1B, the bacterial product fsl-1, and viral analog polyinosinic:polycytidylic acid (poly [I:C]) significantly increased IRF1 mRNA expression and transcriptional activity in human primary myometrial cells. In addition, IL1B increased IRF1 activity in primary amnion cells. IRF1 silencing in myometrial cells decreased IL1B-, fsl-1-, and poly (I:C)-induced cytokine (IL6, TNF, IL1B) and chemokine (CXCL8, CCL2) mRNA expression and IL6, CXCL8, and CCL2 release. IL1B-, fsl-1-, and poly (I:C)-induced PTGS2 mRNA expression and IL1B-induced prostaglandin release was also decreased by IRF1 silencing. In conclusion, IRF1 upregulation in fetal membranes and myometrium after term labor indicates a proinflammatory role for IRF1 in human parturition. IRF1 is involved in TLR- and cytokine-mediated signaling in human myometrium. These data provide new insights into the mechanisms associated with inflammation- and infection-associated preterm birth. IRF1 inhibitors as therapeutics for the management of spontaneous preterm birth warrants further investigation. PMID:26674566

  17. Tissue-specific targeting of cell fate regulatory genes by E2f factors.

    PubMed

    Julian, L M; Liu, Y; Pakenham, C A; Dugal-Tessier, D; Ruzhynsky, V; Bae, S; Tsai, S-Y; Leone, G; Slack, R S; Blais, A

    2016-04-01

    Cell cycle proteins are important regulators of diverse cell fate decisions, and in this capacity have pivotal roles in neurogenesis and brain development. The mechanisms by which cell cycle regulation is integrated with cell fate control in the brain and other tissues are poorly understood, and an outstanding question is whether the cell cycle machinery regulates fate decisions directly or instead as a secondary consequence of proliferative control. Identification of the genes targeted by E2 promoter binding factor (E2f) transcription factors, effectors of the pRb/E2f cell cycle pathway, will provide essential insights into these mechanisms. We identified the promoter regions bound by three neurogenic E2f factors in neural precursor cells in a genome-wide manner. Through bioinformatic analyses and integration of published genomic data sets we uncovered hundreds of transcriptionally active E2f-bound promoters corresponding to genes that control cell fate processes, including key transcriptional regulators and members of the Notch, fibroblast growth factor, Wnt and Tgf-β signaling pathways. We also demonstrate a striking enrichment of the CCCTC binding factor transcription factor (Ctcf) at E2f3-bound nervous system-related genes, suggesting a potential regulatory co-factor for E2f3 in controlling differentiation. Finally, we provide the first demonstration of extensive tissue specificity among E2f target genes in mammalian cells, whereby E2f3 promoter binding is well conserved between neural and muscle precursors at genes associated with cell cycle processes, but is tissue-specific at differentiation-associated genes. Our findings implicate the cell cycle pathway as a widespread regulator of cell fate genes, and suggest that E2f3 proteins control cell type-specific differentiation programs by regulating unique sets of target genes. This work significantly enhances our understanding of how the cell cycle machinery impacts cell fate and differentiation, and will

  18. Regulatory role of the cannabinoid CB2 receptor in stress-induced neuroinflammation in mice

    PubMed Central

    Zoppi, S; Madrigal, J L; Caso, J R; García-Gutiérrez, M S; Manzanares, J; Leza, J C; García-Bueno, B

    2014-01-01

    Background and Purpose Stress exposure produces excitotoxicity and neuroinflammation, contributing to the cellular damage observed in stress-related neuropathologies. The endocannabinoids provide a homeostatic system, present in stress-responsive neural circuits. Here, we have assessed the possible regulatory role of cannabinoid CB2 receptors in stress-induced excitotoxicity and neuroinflammation. Experimental Approach We used wild type (WT), transgenic overexpressing CB2 receptors (CB2xP) and CB2 receptor knockout (CB2-KO) mice exposed to immobilization and acoustic stress (2 h·day−1 for 4 days). The CB2 receptor agonist JWH-133 was administered daily (2 mg·kg−1, i.p.) to WT and CB2-KO animals. Glutamate uptake was measured in synaptosomes from frontal cortex; Western blots and RT-PCR were used to measure proinflammatory cytokines, enzymes and mediators in homogenates of frontal cortex. Key Results Increased plasma corticosterone induced by stress was not modified by manipulating CB2 receptors. JWH-133 treatment or overexpression of CB2 receptors increased control levels of glutamate uptake, which were reduced by stress back to control levels. JWH-133 prevented the stress-induced increase in proinflammatory cytokines (TNF-α and CCL2), in NF-κB, and in NOS-2 and COX-2 and in the consequent cellular oxidative and nitrosative damage (lipid peroxidation). CB2xP mice exhibited anti-inflammatory or neuroprotective actions similar to those in JWH-133 pretreated animals. Conversely, lack of CB2 receptors (CB2-KO mice) exacerbated stress-induced neuroinflammatory responses and confirmed that effects of JWH-133 were mediated through CB2 receptors. Conclusions and Implications Pharmacological manipulation of CB2 receptors is a potential therapeutic strategy for the treatment of stress-related pathologies with a neuroinflammatory component, such as depression. PMID:24467609

  19. A functional SNP in the regulatory region of the decay-accelerating factor gene associates with extraocular muscle pareses in myasthenia gravis

    PubMed Central

    Heckmann, J M; Uwimpuhwe, H; Ballo, R; Kaur, M; Bajic, V B; Prince, S

    2009-01-01

    Complement activation in myasthenia gravis (MG) may damage muscle endplate and complement regulatory proteins such as decay-accelerating factor (DAF) or CD55 may be protective. We hypothesize that the increased prevalence of severe extraocular muscle (EOM) dysfunction among African MG subjects reported earlier may result from altered DAF expression. To test this hypothesis, we screened the DAF gene sequences relevant to the classical complement pathway and found an association between myasthenics with EOM paresis and the DAF regulatory region c.-198C>G SNP (odds ratio=8.6; P=0.0003). This single nucleotide polymorphism (SNP) results in a twofold activation of a DAF 5′-flanking region luciferase reporter transfected into three different cell lines. Direct matching of the surrounding SNP sequence within the DAF regulatory region with the known transcription factor-binding sites suggests a loss of an Sp1-binding site. This was supported by the observation that the c.-198C>G SNP did not show the normal lipopolysaccharide-induced DAF transcriptional upregulation in lymphoblasts from four patients. Our findings suggest that at critical periods during autoimmune MG, this SNP may result in inadequate DAF upregulation with consequent complement-mediated EOM damage. Susceptible individuals may benefit from anti-complement therapy in addition to immunosuppression. PMID:19675582

  20. Three novel acetylation sites in the Foxp3 transcription factor regulate the suppressive activity of regulatory T cells

    PubMed Central

    Kwon, Hye-Sook; Lim, Hyung W.; Wu, Jessica; Schnoelzer, Martina; Verdin, Eric; Ott, Melanie

    2012-01-01

    The Foxp3 transcription factor is the master regulator of regulatory T cell (Treg) differentiation and function. Its activity is regulated by reversible acetylation. Using mass spectrometry of immunoprecipitated proteins, we identify three novel acetylation sites in murine Foxp3 (K31, K262, and K267) and the corresponding sites in human FoxP3 proteins. Newly raised modification-specific antibodies against acetylated K31 and K267 confirm acetylation of these residues in murine Tregs. Mutant Foxp3 proteins carrying arginine substitutions at the three acetylation sites (3KR) accumulate in T cells to higher levels than wildtype Foxp3 and exert better suppressive activity in co-culture experiments. Acetylation and stability of wildtype, but not mutant, Foxp3 is enhanced when cells are treated with Ex-527, an inhibitor of the NAD+-dependent deacetylase SIRT1. Treatment with Ex-527 promotes Foxp3 expression during induced Treg differentiation, enhances Foxp3 levels in natural Tregs, and prevents loss of Foxp3 expression in adoptively transferred Tregs in mice. Our data identify SIRT1 as a negative regulator of Treg function via deacetylation of three novel target sites in Foxp3. SIRT1 inhibitors strengthen the suppressive activity of Tregs and may be useful in enhancing Treg-based therapeutic approaches to autoimmune diseases or graft rejections. PMID:22312127

  1. Dynamic control of gene regulatory logic by seemingly redundant transcription factors

    PubMed Central

    AkhavanAghdam, Zohreh; Sinha, Joydeb; Tabbaa, Omar P; Hao, Nan

    2016-01-01

    Many transcription factors co-express with their homologs to regulate identical target genes, however the advantages of such redundancies remain elusive. Using single-cell imaging and microfluidics, we study the yeast general stress response transcription factor Msn2 and its seemingly redundant homolog Msn4. We find that gene regulation by these two factors is analogous to logic gate systems. Target genes with fast activation kinetics can be fully induced by either factor, behaving as an 'OR' gate. In contrast, target genes with slow activation kinetics behave as an 'AND' gate, requiring distinct contributions from both factors, upon transient stimulation. Furthermore, such genes become an 'OR' gate when the input duration is prolonged, suggesting that the logic gate scheme is not static but rather dependent on the input dynamics. Therefore, Msn2 and Msn4 enable a time-based mode of combinatorial gene regulation that might be applicable to homologous transcription factors in other organisms. DOI: http://dx.doi.org/10.7554/eLife.18458.001 PMID:27690227

  2. Cleavage of interferon regulatory factor 7 by enterovirus 71 3C suppresses cellular responses.

    PubMed

    Lei, Xiaobo; Xiao, Xia; Xue, Qinghua; Jin, Qi; He, Bin; Wang, Jianwei

    2013-02-01

    Enterovirus 71 (EV71) is a positive-stranded RNA virus which is capable of inhibiting innate immunity. Among virus-encoded proteins, the 3C protein compromises the type I interferon (IFN-I) response mediated by retinoid acid-inducible gene-I (RIG-I) or Toll-like receptor 3 that activates interferon regulatory 3 (IRF3) and IRF7. In the present study, we report that enterovirus 71 downregulates IRF7 through the 3C protein, which inhibits the function of IRF7. When expressed in mammalian cells, the 3C protein mediates cleavage of IRF7 rather than that of IRF3. This process is insensitive to inhibitors of caspase, proteasome, lysosome, and autophagy. H40D substitution in the 3C active site abolishes its activity, whereas R84Q or V154S substitution in the RNA binding motif has no effect. Furthermore, 3C-mediated cleavage occurs at the Q189-S190 junction within the constitutive activation domain of IRF7, resulting in two cleaved IRF7 fragments that are incapable of activating IFN expression. Ectopic expression of wild-type IRF7 limits EV71 replication. On the other hand, expression of the amino-terminal domain of IRF7 enhances EV71 infection, which correlates with its ability to interact with and inhibit IRF3. These results suggest that control of IRF7 by the 3C protein may represent a viral mechanism to escape cellular responses. PMID:23175366

  3. Regulatory T-cell Trafficking: From Thymic Development to Tumor-Induced Immune Suppression

    PubMed Central

    Mailloux, Adam W.; Young, M. Rita I.

    2011-01-01

    Regulatory T cells (Tregs) have become a priority for many investigators in immunology due to their potent immunosuppressive and tolerogenic effects. While Treg activity is required for normal immune homeostasis, dysregulation of their numbers can induce autoimmunity or aid in the pathogenesis of disease. Therefore, great effort has been made to understand the mechanisms by which Tregs accumulate in different areas of the body. Like other lymphocytes, Tregs migrate in response to a network of chemotactic stimuli involving chemokines, chemokine receptors, integrins, and their corresponding ligands. However, many of these stimuli are exclusive to Tregs, inducing their migration while leaving conventional populations unaffected. It is these selective stimuli that result in increased ratios of Tregs among conventional effector populations, leading to changes in immune suppression and homeostasis. This review explores selective Treg trafficking during thymic Treg development, migration to secondary lymphoid tissues and emigration into the periphery during homeostatic conditions, inflammation, and the tumor microenvironment, placing emphasis on stimuli that selectively recruits Tregs to target locations. PMID:21083525

  4. Extra-thymically induced T regulatory cell subsets: the optimal target for antigen-specific immunotherapy

    PubMed Central

    Verhagen, Johan; Wegner, Anja; Wraith, David C

    2015-01-01

    Antigen-specific immunotherapy aims to selectively restore tolerance to innocuous antigens in cases of autoimmune or allergic disease, without the need for general immune suppression. Although the principle of antigen-specific immunotherapy was discovered more than a century ago, its clinical application to date is limited, particularly in the control of autoimmunity. This has resulted mainly from a lack of in-depth understanding of the underlying mechanism. More recently, the differentiation of extra-thymically induced T regulatory (Treg) cell subsets has been shown to be instrumental in peripheral tolerance induction. Two main types of inducible Treg cells, interleukin-10-secreting or Foxp3+, have now been described, each with distinct characteristics and methods of therapeutic induction. It is crucial, therefore, to identify the suitability of either subset in the control of specific immune disorders. This review explores their natural function, the known mechanisms of therapeutic differentiation of either subset as well as their in vivo functionality and discusses new developments that may aid their use in antigen-specific immunotherapy, with a focus on autoimmune disease. PMID:25716063

  5. Low-Dose IL-2 Induces Regulatory T Cell-Mediated Control of Experimental Food Allergy.

    PubMed

    Bonnet, Benjamin; Vigneron, James; Levacher, Béatrice; Vazquez, Thomas; Pitoiset, Fabien; Brimaud, Faustine; Churlaud, Guillaume; Klatzmann, David; Bellier, Bertrand

    2016-07-01

    Regulatory T cells (Tregs) are pivotal for maintenance of immune self-tolerance and also regulate immune responses to exogenous Ags, including allergens. Both decreased Treg number and function have been reported in allergic patients, offering new therapeutic perspectives. We previously demonstrated that Tregs can be selectively expanded and activated by low doses of IL-2 (ld-IL-2) inducing immunoregulation without immunosuppression and established its protective effect in autoimmune diseases. In this study, we evaluated the ability of ld-IL-2 to control allergy in an experimental model of food allergy. Ld-IL-2 induced Treg expansion and activation that elicited protection against clinical manifestations of food allergy in two mouse models with OVA and peanut. This clinical effect was lost in Treg-depleted mice, demonstrating the major contribution of Tregs in ld-IL-2 efficacy. Mechanistic studies further indicated that protection from allergy could be explained by a Treg-dependent local modification of the Th1/Th2 balance and an inhibition of mast cell recruitment and activation. Preventive and therapeutic effects of ld-IL-2 were observed over a 7-mo-period, highlighting its long-term efficacy. This study demonstrated that ld-IL-2 is efficient to prevent and to treat allergic immune responses, and thus represents a promising therapeutic strategy for managing allergic diseases. PMID:27259854

  6. BTat, a trans-acting regulatory protein, contributes to bovine immunodeficiency virus-induced apoptosis.

    PubMed

    Xuan, Chenghao; Qiao, Wentao; Li, Jian; Peng, Guoyuan; Liu, Min; Chen, Qimin; Zhou, Jun; Geng, Yunqi

    2008-01-01

    Bovine immunodeficiency virus (BIV) is a member of the lentivirus subfamily of retroviruses highly related to human immunodeficiency virus in morphologic, antigenic and genomic features. BIV is known to induce chronic pathological changes in infected hosts, which are often associated with the development of immune-mediated lesions. However, the molecular events underlying the cytopathic effect of BIV remain poorly understood. In this study, BIV was found to induce apoptotic cell death, and a small trans-acting regulatory protein encoded by BIV, BTat, was found to participate in the pro-apoptotic action of BIV. Introduction of exogenous BTat to cells triggered apoptosis dramatically, as revealed by assays such as terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling, nuclear morphology analysis, flow cytometry, and cleavages of caspases and poly(ADP-ribose)polymerase. Interestingly, the pro-apoptotic effect of BTat was found to be mediated through its interaction with cellular microtubules and its interference with microtubule dynamics. These results provide the first evidence that induction of apoptosis may contribute to the cytopathic effect of BIV. In addition, these results uncover a novel role for BTat in regulating microtubule dynamics in addition to its conventional role in regulating gene transcription.

  7. Identifying Gene Regulatory Networks in Arabidopsis by In Silico Prediction, Yeast-1-Hybrid, and Inducible Gene Profiling Assays.

    PubMed

    Sparks, Erin E; Benfey, Philip N

    2016-01-01

    A system-wide understanding of gene regulation will provide deep insights into plant development and physiology. In this chapter we describe a threefold approach to identify the gene regulatory networks in Arabidopsis thaliana that function in a specific tissue or biological process. Since no single method is sufficient to establish comprehensive and high-confidence gene regulatory networks, we focus on the integration of three approaches. First, we describe an in silico prediction method of transcription factor-DNA binding, then an in vivo assay of transcription factor-DNA binding by yeast-1-hybrid and lastly the identification of co-expression clusters by transcription factor induction in planta. Each of these methods provides a unique tool to advance our understanding of gene regulation, and together provide a robust model for the generation of gene regulatory networks.

  8. Poly I:C enhances production of nitric oxide in response to interferon-γ via upregulation of interferon regulatory factor 7 in vascular endothelial cells.

    PubMed

    Mori, Daiki; Koide, Naoki; Tsolmongyn, Bilegtsaikhan; Nagata, Hiroshi; Sano, Tsuyoshi; Nonami, Toshiaki; Yokochi, Takashi

    2015-03-01

    The effect of poly I:C on interferon (IFN)-γ-induced nitric oxide (NO) production in vascular endothelial cells was examined using murine aortic endothelial END-D cells. Poly I:C augmented IFN-γ-induced NO production although it alone did not induce the NO production. Poly I:C augmented the NO production via enhanced expression of an inducible NO synthase protein. Poly I:C did not affect the activation of Janus kinase (JAK) 1/2, and signal transducer and activator of transcription (STAT) 1 in IFN-γ signaling. Moreover, there was no significant difference in the IFN-γ-induced interferon regulatory factor (IRF) 1 expression between the presence and absence of poly I:C. Poly I:C led to the activation of IRF7 in END-D cells. Inhibition of poly I:C signaling by amlexanox, an inhibitor of TANK-binding kinase (TBK) 1 and IκB kinase (IKK) ε, abolished the augmentation of IFN-γ-induced NO production. Therefore, poly I:C was suggested to augment IFN-γ-induced NO production at the transcriptional level via enhanced IRF7 activation.

  9. Expression regulation of zebrafish interferon regulatory factor 9 by promoter analysis.

    PubMed

    Shi, Jun; Zhang, Yi-Bing; Zhang, Jian-She; Gui, Jian-Fang

    2013-12-01

    We previously showed that a fish interferon (IFN) regulatory factor 9 (IRF9) homologue, crucian carp Carassius auratus IRF9, displays constitutively nuclear localization and involvement in fish IFN-dependent JAK-STAT signaling; however, little is known about the expression regulation of fish IRF9. Here, we characterized the expression of zebrafish IRF9 by promoter analysis. Zebrafish IRF9 gene promoter contained several putative transcription factor binding sites, including one ISRE (IFN-stimulated response element), one GAS (IFN gamma activation sequence) and three GATEs (IFNγ activated transcriptional element, GATE1/2/3). Further sequence analyses revealed that GAS and GATE motifs existed in all promoters of IRF9 from mammals and fishes. Luciferase assays confirmed that zebrafish IRF9 promoter could be activated by zebrafish IFNφs and zebrafish IFNγ2, as well as transcription factors IRF3, IRF7, and combination of IRF9 and STAT2. Treatment of recombinant crucian carp IFN protein or overexpression of zebrafish IFNγ2 both led to significant increase in crucian carp IRF9 mRNA and protein in cultured fish cells. Comparison of IFN-stimulated promoter activity revealed much more significant induction of zebrafish IRF9 by zebrafish IFNγ2 than by zebrafish IFNφs. Mutation analyses showed that the putative GAS and GATE3 contributed to zebrafish IFNγ2-triggered IRF9 expression, whereas the putative ISRE and the other two GATEs were not functional for induction of zebrafish IRF9. These results together indicated that the expression property of IRF9 might be conserved from fish to mammals and that some not yet identified mechanisms could exist in IRF9 gene transcription regulation in response to IFNs.

  10. T regulatory-1 cells induce IgG4 production by B cells: role of IL-10.

    PubMed

    Satoguina, Judith S; Weyand, Esther; Larbi, John; Hoerauf, Achim

    2005-04-15

    The study was aimed to find out whether T cells with a regulatory profile could regulate the secretion of IgG4. Using tetanus Ag we found that PBMC of healthy human donors responded to exogenous IL-10 by down-regulating IgG1 and increasing IgG4 secretion. IgE was not affected. To investigate the direct effect of IL-10-producing T cells on B cells, we generated T cell clones (TCC) with two different cytokine profiles: first, IL-10high, IL-2low, IL-4low TCC, and second, IL-10low, IL-2high, IL-4high. The T cell-dependent Ab secretion was measured by coculturing purified CD19+ B cells and the TCC. Interestingly, we found that IgG4 production in the coculture correlated with the TCC production of IL-10 (r2 = 0.352, p = 0.0001), but not with IL-2, IL-4, nor IFN-gamma. IgE showed only a trend with regard to IL-4. Further, there was decreased Ab secretion in the absence of T-B cell contact. IL-10 also induced IgG4 when added to a Th1 TCC-B cell coculture system. The present study thus shows that in T-B cell coculture, IL-10, if induced by the TCC or added to the system, down-regulates the immune response by inducing IgG4 secretion. This establishes a direct implication of IL-10 in humoral hyporesponsiveness, particularly in compartments where the T-B cell interplay determines the subsequent immune response. The correlation between IgG4 and IL-10 (r2 = 0.352) indicates that IL-10 is an important but not the only factor for IgG4 induction. PMID:15814696

  11. The regulatory T cell effector soluble fibrinogen-like protein 2 induces tubular epithelial cell apoptosis in renal transplantation.

    PubMed

    Zhao, Zitong; Yang, Cheng; Wang, Lingyan; Li, Long; Zhao, Tian; Hu, Linkun; Rong, Ruiming; Xu, Ming; Zhu, Tongyu

    2014-02-01

    Acute rejection (AR) hinders renal allograft survival. Tubular epithelial cell (TEC) apoptosis contributes to premature graft loss in AR, while the mechanism remains unclear. Soluble fibrinogen-like protein 2 (sFGL2), a novel effector of regulatory T cells (Treg), induces apoptosis to mediate tissue injury. We previously found that serum sFGL2 significantly increased in renal allograft rejection patients. In this study, the role of sFGL2 in AR was further investigated both in vivo and in vitro. The serum level of sFGL2 and the percentage of CD4(+)CD25(+)Foxp3(+) Treg in the peripheral blood were measured in renal allograft recipients with AR or stable renal function (n = 30 per group). The human TEC was stimulated with sFGL2, tumor necrosis factor (TNF)-α, or phosphate buffered saline and investigated for apoptosis in vitro. Apoptosis-associated genes expression in TEC was further assessed. Approval for this study was obtained from the Ethics Committee of Fudan University. Our results showed that the serum level of sFGL2, correlated with Treg in the peripheral blood, was significantly increased in the AR patients. In vitro, sFGL2 remarkably induced TEC apoptosis, with a significant up-regulation of proapoptotic genes, including CASP-3, CASP-8, CASP-9, CASP-10, TRADD, TNFSF10, FADD, FAS, FASLG, BAK1, BAD, BAX, and NF-KB1. However, no significant changes were observed in the expression of antiapoptotic genes, including CARD-18, NAIP, BCL2, IKBKB, and TBK1. Therefore, sFGL2, an effector of Treg, induces TEC apoptosis. Our study suggests that sFGL2 is a potential mediator in the pathogenesis of allograft rejection and provides novel insights into the role of Treg in AR. PMID:24414480

  12. Mechanisms of immune suppression by interleukin-10 and transforming growth factor-β: the role of T regulatory cells

    PubMed Central

    Taylor, Alison; Verhagen, Johan; Blaser, Kurt; Akdis, Mübeccel; Akdis, Cezmi A

    2006-01-01

    Specific immune suppression and induction of tolerance are essential processes in the regulation and circumvention of immune defence. The balance between allergen-specific type 1 regulatory (Tr1) cells and T helper (Th) 2 cells appears to be decisive in the development of allergy. Tr1 cells consistently represent the dominant subset specific for common environmental allergens in healthy individuals. In contrast, there is a high frequency of allergen-specific interleukin-4 (IL-4)-secreting T cells in allergic individuals. Allergen-specific immunotherapy can induce specific Tr1 cells that abolish allergen-induced proliferation of Th1 and Th2 cells, as well as their cytokine production. Tr1 cells utilize multiple suppressor mechanisms, such as IL-10 and transforming growth factor-β (TGF-β) as secreted cytokines and various surface molecules, such as cytotoxic T-lymphocyte antigen 4 and programmed death-1. IL-10 only inhibits T cells stimulated by low numbers of triggered T-cell receptors, which depend on CD28 costimulation. IL-10 inhibits CD28 tyrosine phosphorylation, preventing the binding of phosphatidylinositol 3-kinase p85 and consequently inhibiting the CD28 signalling pathway. In addition, IL-10 and TGF-β secreted by Tr1 cells skew the antibody production from immunoglobulin E (IgE) towards the non-inflammatory isotypes IgG4 and IgA, respectively. Induction of antigen-specific Tr1 cells can thus re-direct an inappropriate immune response against allergens or auto-antigens using a broad range of suppressor mechanisms. PMID:16556256

  13. Using graphical adaptive lasso approach to construct transcription factor and microRNA's combinatorial regulatory network in breast cancer.

    PubMed

    Su, Naifang; Dai, Ding; Deng, Chao; Qian, Minping; Deng, Minghua

    2014-06-01

    Discovering the regulation of cancer-related gene is of great importance in cancer biology. Transcription factors and microRNAs are two kinds of crucial regulators in gene expression, and they compose a combinatorial regulatory network with their target genes. Revealing the structure of this network could improve the authors' understanding of gene regulation, and further explore the molecular pathway in cancer. In this article, the authors propose a novel approach graphical adaptive lasso (GALASSO) to construct the regulatory network in breast cancer. GALASSO use a Gaussian graphical model with adaptive lasso penalties to integrate the sequence information as well as gene expression profiles. The simulation study and the experimental profiles verify the accuracy of the authors' approach. The authors further reveal the structure of the regulatory network, and explore the role of feedforward loops in gene regulation. In addition, the authors discuss the combinatorial regulatory effect between transcription factors and microRNAs, and select miR-155 for detailed analysis of microRNA's role in cancer. The proposed GALASSO approach is an efficient method to construct the combinatorial regulatory network. It also provides a new way to integrate different data sources and could find more applications in meta-analysis problem.

  14. GATA3-Driven Th2 Responses Inhibit TGF-β1–Induced FOXP3 Expression and the Formation of Regulatory T Cells

    PubMed Central

    Mantel, Pierre-Yves; Kuipers, Harmjan; Boyman, Onur; Rhyner, Claudio; Ouaked, Nadia; Rückert, Beate; Karagiannidis, Christian; Lambrecht, Bart N; Hendriks, Rudolf W; Crameri, Reto; Akdis, Cezmi A; Blaser, Kurt; Schmidt-Weber, Carsten B

    2007-01-01

    Transcription factors act in concert to induce lineage commitment towards Th1, Th2, or T regulatory (Treg) cells, and their counter-regulatory mechanisms were shown to be critical for polarization between Th1 and Th2 phenotypes. FOXP3 is an essential transcription factor for natural, thymus-derived (nTreg) and inducible Treg (iTreg) commitment; however, the mechanisms regulating its expression are as yet unknown. We describe a mechanism controlling iTreg polarization, which is overruled by the Th2 differentiation pathway. We demonstrated that interleukin 4 (IL-4) present at the time of T cell priming inhibits FOXP3. This inhibitory mechanism was also confirmed in Th2 cells and in T cells of transgenic mice overexpressing GATA-3 in T cells, which are shown to be deficient in transforming growth factor (TGF)-β–mediated FOXP3 induction. This inhibition is mediated by direct binding of GATA3 to the FOXP3 promoter, which represses its transactivation process. Therefore, this study provides a new understanding of tolerance development, controlled by a type 2 immune response. IL-4 treatment in mice reduces iTreg cell frequency, highlighting that therapeutic approaches that target IL-4 or GATA3 might provide new preventive strategies facilitating tolerance induction particularly in Th2-mediated diseases, such as allergy. PMID:18162042

  15. Vpu Mediates Depletion of Interferon Regulatory Factor 3 during HIV Infection by a Lysosome-Dependent Mechanism

    PubMed Central

    Doehle, Brian P.; Chang, Kristina; Rustagi, Arjun; McNevin, John; McElrath, M. Juliana

    2012-01-01

    HIV has evolved sophisticated mechanisms to avoid restriction by intracellular innate immune defenses that otherwise serve to control acute viral infection and virus dissemination. Innate defenses are triggered when pattern recognition receptor (PRR) proteins of the host cell engage pathogen-associated molecule patterns (PAMPs) present in viral products. Interferon regulatory factor 3 (IRF3) plays a central role in PRR signaling of innate immunity to drive the expression of type I interferon (IFN) and interferon-stimulated genes (ISGs), including a variety of HIV restriction factors, that serve to limit viral replication directly and/or program adaptive immunity. Productive infection of T cells by HIV is dependent upon the targeted proteolysis of IRF3 that occurs through a virus-directed mechanism that results in suppression of innate immune defenses. However, the mechanisms by which HIV controls innate immune signaling and IRF3 function are not defined. Here, we examined the innate immune response induced by HIV strains identified through their differential control of PRR signaling. We identified viruses that, unlike typical circulating HIV strains, lack the ability to degrade IRF3. Our studies show that IRF3 regulation maps specifically to the HIV accessory protein Vpu. We define a molecular interaction between Vpu and IRF3 that redirects IRF3 to the endolysosome for proteolytic degradation, thus allowing HIV to avoid the innate antiviral immune response. Our studies reveal that Vpu is an important IRF3 regulator that supports acute HIV infection through innate immune suppression. These observations define the Vpu-IRF3 interface as a novel target for therapeutic strategies aimed at enhancing the immune response to HIV. PMID:22593165

  16. Vpu mediates depletion of interferon regulatory factor 3 during HIV infection by a lysosome-dependent mechanism.

    PubMed

    Doehle, Brian P; Chang, Kristina; Rustagi, Arjun; McNevin, John; McElrath, M Juliana; Gale, Michael

    2012-08-01

    HIV has evolved sophisticated mechanisms to avoid restriction by intracellular innate immune defenses that otherwise serve to control acute viral infection and virus dissemination. Innate defenses are triggered when pattern recognition receptor (PRR) proteins of the host cell engage pathogen-associated molecule patterns (PAMPs) present in viral products. Interferon regulatory factor 3 (IRF3) plays a central role in PRR signaling of innate immunity to drive the expression of type I interferon (IFN) and interferon-stimulated genes (ISGs), including a variety of HIV restriction factors, that serve to limit viral replication directly and/or program adaptive immunity. Productive infection of T cells by HIV is dependent upon the targeted proteolysis of IRF3 that occurs through a virus-directed mechanism that results in suppression of innate immune defenses. However, the mechanisms by which HIV controls innate immune signaling and IRF3 function are not defined. Here, we examined the innate immune response induced by HIV strains identified through their differential control of PRR signaling. We identified viruses that, unlike typical circulating HIV strains, lack the ability to degrade IRF3. Our studies show that IRF3 regulation maps specifically to the HIV accessory protein Vpu. We define a molecular interaction between Vpu and IRF3 that redirects IRF3 to the endolysosome for proteolytic degradation, thus allowing HIV to avoid the innate antiviral immune response. Our studies reveal that Vpu is an important IRF3 regulator that supports acute HIV infection through innate immune suppression. These observations define the Vpu-IRF3 interface as a novel target for therapeutic strategies aimed at enhancing the immune response to HIV.

  17. Dynamic changes in binding of immunoglobulin heavy chain 3' regulatory region to protein factors during class switching.

    PubMed

    Chatterjee, Sanjukta; Ju, Zhongliang; Hassan, Rabih; Volpi, Sabrina A; Emelyanov, Alexander V; Birshtein, Barbara K

    2011-08-19

    The 3' regulatory region (3' RR) of the Igh locus works at long distances on variable region (V(H)) and switch region (I) region promoters to initiate germ line (non-coding) transcription (GT) and promote class switch recombination (CSR). The 3' RR contains multiple elements, including enhancers (hs3a, hs1.2, hs3b, and hs4) and a proposed insulator region containing CTCF (CCCTC-binding factor) binding sites, i.e. hs5/6/7 and the downstream region ("38"). Notably, deletion of each individual enhancer (hs3a-hs4) has no significant phenotypic consequence, suggesting that the 3' RR has considerable structural flexibility in its function. To better understand how the 3' RR functions, we identified transcription factor binding sites and used chromatin immunoprecipitation (ChIP) assays to monitor their occupancy in splenic B cells that initiate GT and undergo CSR (LPS±IL4), are deficient in GT and CSR (p50(-/-)), or do not undergo CSR despite efficient GT (anti-IgM+IL4). Like 3' RR enhancers, hs5-7 and the 38 region were observed to contain multiple Pax5 binding sites (in addition to multiple CTCF sites). We found that the Pax5 binding profile to the 3' RR dynamically changed during CSR independent of the specific isotype to which switching was induced, and binding focused on hs1.2, hs4, and hs7. CTCF-associated and CTCF-independent cohesin interactions were also identified. Our observations are consistent with a scaffold model in which a platform of active protein complexes capable of facilitating GT and CSR can be formed by varying constellations of 3' RR elements.

  18. Interferon Regulatory Factor 8-Deficiency Determines Massive Neutrophil Recruitment but T Cell Defect in Fast Growing Granulomas during Tuberculosis

    PubMed Central

    Sali, Michela; Anfossi, Antonio Giovanni; Abalsamo, Laura; Palucci, Ivana; Mattei, Fabrizio; Sanchez, Massimo; Giagu, Anna; Antuofermo, Elisabetta; Fadda, Giovanni; Belardelli, Filippo; Delogu, Giovanni; Gabriele, Lucia

    2013-01-01

    Following Mycobacterium tuberculosis (Mtb) infection, immune cell recruitment in lungs is pivotal in establishing protective immunity through granuloma formation and neogenesis of lymphoid structures (LS). Interferon regulatory factor-8 (IRF-8) plays an important role in host defense against Mtb, although the mechanisms driving anti-mycobacterial immunity remain unclear. In this study, IRF-8 deficient mice (IRF-8−/−) were aerogenously infected with a low-dose Mtb Erdman virulent strain and the course of infection was compared with that induced in wild-type (WT-B6) counterparts. Tuberculosis (TB) progression was examined in both groups using pathological, microbiological and immunological parameters. Following Mtb exposure, the bacterial load in lungs and spleens progressed comparably in the two groups for two weeks, after which IRF-8−/− mice developed a fatal acute TB whereas in WT-B6 the disease reached a chronic stage. In lungs of IRF-8−/−, uncontrolled growth of pulmonary granulomas and impaired development of LS were observed, associated with unbalanced homeostatic chemokines, progressive loss of infiltrating T lymphocytes and massive prevalence of neutrophils at late infection stages. Our data define IRF-8 as an essential factor for the maintenance of proper immune cell recruitment in granulomas and LS required to restrain Mtb infection. Moreover, IRF-8−/− mice, relying on a common human and mouse genetic mutation linked to susceptibility/severity of mycobacterial diseases, represent a valuable model of acute TB for comparative studies with chronically-infected congenic WT-B6 for dissecting protective and pathological immune reactions. PMID:23717393

  19. Interferon regulatory factor 8-deficiency determines massive neutrophil recruitment but T cell defect in fast growing granulomas during tuberculosis.

    PubMed

    Rocca, Stefano; Schiavoni, Giovanna; Sali, Michela; Anfossi, Antonio Giovanni; Abalsamo, Laura; Palucci, Ivana; Mattei, Fabrizio; Sanchez, Massimo; Giagu, Anna; Antuofermo, Elisabetta; Fadda, Giovanni; Belardelli, Filippo; Delogu, Giovanni; Gabriele, Lucia

    2013-01-01

    Following Mycobacterium tuberculosis (Mtb) infection, immune cell recruitment in lungs is pivotal in establishing protective immunity through granuloma formation and neogenesis of lymphoid structures (LS). Interferon regulatory factor-8 (IRF-8) plays an important role in host defense against Mtb, although the mechanisms driving anti-mycobacterial immunity remain unclear. In this study, IRF-8 deficient mice (IRF-8⁻/⁻) were aerogenously infected with a low-dose Mtb Erdman virulent strain and the course of infection was compared with that induced in wild-type (WT-B6) counterparts. Tuberculosis (TB) progression was examined in both groups using pathological, microbiological and immunological parameters. Following Mtb exposure, the bacterial load in lungs and spleens progressed comparably in the two groups for two weeks, after which IRF-8⁻/⁻ mice developed a fatal acute TB whereas in WT-B6 the disease reached a chronic stage. In lungs of IRF-8⁻/⁻, uncontrolled growth of pulmonary granulomas and impaired development of LS were observed, associated with unbalanced homeostatic chemokines, progressive loss of infiltrating T lymphocytes and massive prevalence of neutrophils at late infection stages. Our data define IRF-8 as an essential factor for the maintenance of proper immune cell recruitment in granulomas and LS required to restrain Mtb infection. Moreover, IRF-8⁻/⁻ mice, relying on a common human and mouse genetic mutation linked to susceptibility/severity of mycobacterial diseases, represent a valuable model of acute TB for comparative studies with chronically-infected congenic WT-B6 for dissecting protective and pathological immune reactions. PMID:23717393

  20. Signals and myogenic regulatory factors restrict pax3 and pax7 expression to dermomyotome-like tissue in zebrafish

    PubMed Central

    Hammond, Christina L.; Hinits, Yaniv; Osborn, Daniel P.S.; Minchin, James E.N.; Tettamanti, Gianluca; Hughes, Simon M.

    2014-01-01

    Pax3/7 paired homeodomain transcription factors are important markers of muscle stem cells. Pax3 is required upstream of myod for lateral dermomyotomal cells in the amniote somite to form particular muscle cells. Later Pax3/7-dependent cells generate satellite cells and most body muscle. Here we analyse early myogenesis from, and regulation of, a population of Pax3-expressing dermomyotome-like cells in the zebrafish. Zebrafish pax3 is widely expressed in the lateral somite and, along with pax7, becomes restricted anteriorly and then to the external cells on the lateral somite surface. Midline-derived Hedgehog signals appear to act directly on lateral somite cells to repress Pax3/7. Both Hedgehog and Fgf8, signals that induce muscle formation within the somite, suppress Pax3/7 and promote expression of myogenic regulatory factors (MRFs) myf5 and myod in specific muscle precursor cell populations. Loss of MRF function leads to loss of myogenesis by specific populations of muscle fibres, with parallel up-regulation of Pax3/7. Myod is required for lateral fast muscle differentiation from pax3-expressing cells. In contrast, either Myf5 or Myod is sufficient to promote slow muscle formation from adaxial cells. Thus, myogenic signals act to drive somite cells to a myogenic fate through up-regulation of distinct combinations of MRFs. Our data show that the relationship between Pax3/7 genes and myogenesis is evolutionarily ancient, but that changes in the MRF targets for particular signals contribute to myogenic differences between species. PMID:17094960

  1. Affinity Density: a novel genomic approach to the identification of transcription factor regulatory targets

    PubMed Central

    Hazelett, Dennis J.; Lakeland, Daniel L.; Weiss, Joseph B.

    2009-01-01

    Methods: A new method was developed for identifying novel transcription factor regulatory targets based on calculating Local Affinity Density. Techniques from the signal-processing field were used, in particular the Hann digital filter, to calculate the relative binding affinity of different regions based on previously published in vitro binding data. To illustrate this approach, the complete genomes of Drosophila melanogaster and D.pseudoobscura were analyzed for binding sites of the homeodomain proteinc Tinman, an essential heart development gene in both Drosophila and Mouse. The significant binding regions were identified relative to genomic background and assigned to putative target genes. Valid candidates common to both species of Drosophila were selected as a test of conservation. Results: The new method was more sensitive than cluster searches for conserved binding motifs with respect to positive identification of known Tinman targets. Our Local Affinity Density method also identified a significantly greater proportion of Tinman-coexpressed genes than equivalent, optimized cluster searching. In addition, this new method predicted a significantly greater than expected number of genes with previously published RNAi phenotypes in the heart. Availability: Algorithms were implemented in Python, LISP, R and maxima, using MySQL to access locally mirrored sequence data from Ensembl (D.melanogaster release 4.3) and flybase (D.pseudoobscura). All code is licensed under GPL and freely available at http://www.ohsu.edu/cellbio/dev_biol_prog/affinitydensity/. Contact: hazelett@ohsu.edu PMID:19401399

  2. Are good ideas enough? The impact of socio-economic and regulatory factors on GMO commercialisation.

    PubMed

    Vàzquez-Salat, Núria

    2013-01-01

    In recent years scientific literature has seen an increase in publications describing new transgenic applications. Although technically-sound, these promising developments might not necessarily translate into products available to the consumer. This article highlights the impact of external factors on the commercial viability of Genetically Modified (GM) animals in the pharmaceutical and food sectors. Through the division of the production chain into three Policy Domains -Science, Market and Public- I present an overview of the broad range of regulatory and socio-economic components that impacts on the path towards commercialisation of GM animals. To further illustrate the unique combination of forces that influence each application, I provide an in-depth analysis of two real cases: GM rabbits producing human polyclonal antibodies (pharmaceutical case study) and GM cows producing recombinant human lactoferrin (food case study). The inability to generalise over the commercial success of a given transgenic application should encourage researchers to perform these type of exercises early in the R & D process. Furthermore, through the analysis of these case studies we can observe a change in the biopolitics of Genetically Modified Organisms (GMOs). Contrary to the GM plant biopolitical landscape, developing states such as China and Argentina are placing themselves as global leaders in GM animals. The pro-GM attitude of these states is likely to cause a shift in the political evolution of global GMO governance. PMID:24510133

  3. Na+/H+ Exchanger Regulatory Factor 1 (NHERF1) Directly Regulates Osteogenesis*

    PubMed Central

    Liu, Li; Alonso, Veronica; Guo, Lida; Tourkova, Irina; Henderson, Sarah E.; Almarza, Alejandro J.; Friedman, Peter A.; Blair, Harry C.

    2012-01-01

    Bone formation requires synthesis, secretion, and mineralization of matrix. Deficiencies in these processes produce bone defects. The absence of the PDZ domain protein Na+/H+ exchange regulatory factor 1 (NHERF1) in mice, or its mutation in humans, causes osteomalacia believed to reflect renal phosphate wasting. We show that NHERF1 is expressed by mineralizing osteoblasts and organizes Na+/H+ exchangers (NHEs) and the PTH receptor. NHERF1-null mice display reduced bone formation and wide mineralizing fronts despite elimination of phosphate wasting by dietary supplementation. Bone mass was normal, reflecting coordinated reduction of bone resorption and formation. NHERF1-null bone had decreased strength, consistent with compromised matrix quality. Mesenchymal stem cells from NHERF1-null mice showed limited osteoblast differentiation but enhanced adipocyte differentiation. PTH signaling and Na+/H+ exchange were dysregulated in these cells. Osteoclast differentiation from monocytes was unaffected. Thus, NHERF1 is required for normal osteoblast differentiation and matrix synthesis. In its absence, compensatory mechanisms maintain bone mass, but bone strength is reduced. PMID:23109343

  4. FootprintDB: Analysis of Plant Cis-Regulatory Elements, Transcription Factors, and Binding Interfaces.

    PubMed

    Contreras-Moreira, Bruno; Sebastian, Alvaro

    2016-01-01

    FootprintDB is a database and search engine that compiles regulatory sequences from open access libraries of curated DNA cis-elements and motifs, and their associated transcription factors (TFs). It systematically annotates the binding interfaces of the TFs by exploiting protein-DNA complexes deposited in the Protein Data Bank. Each entry in footprintDB is thus a DNA motif linked to the protein sequence of the TF(s) known to recognize it, and in most cases, the set of predicted interface residues involved in specific recognition. This chapter explains step-by-step how to search for DNA motifs and protein sequences in footprintDB and how to focus the search to a particular organism. Two real-world examples are shown where this software was used to analyze transcriptional regulation in plants. Results are described with the aim of guiding users on their interpretation, and special attention is given to the choices users might face when performing similar analyses. PMID:27557773

  5. Are good ideas enough? The impact of socio-economic and regulatory factors on GMO commercialisation.

    PubMed

    Vàzquez-Salat, Núria

    2013-01-01

    In recent years scientific literature has seen an increase in publications describing new transgenic applications. Although technically-sound, these promising developments might not necessarily translate into products available to the consumer. This article highlights the impact of external factors on the commercial viability of Genetically Modified (GM) animals in the pharmaceutical and food sectors. Through the division of the production chain into three Policy Domains -Science, Market and Public- I present an overview of the broad range of regulatory and socio-economic components that impacts on the path towards commercialisation of GM animals. To further illustrate the unique combination of forces that influence each application, I provide an in-depth analysis of two real cases: GM rabbits producing human polyclonal antibodies (pharmaceutical case study) and GM cows producing recombinant human lactoferrin (food case study). The inability to generalise over the commercial success of a given transgenic application should encourage researchers to perform these type of exercises early in the R & D process. Furthermore, through the analysis of these case studies we can observe a change in the biopolitics of Genetically Modified Organisms (GMOs). Contrary to the GM plant biopolitical landscape, developing states such as China and Argentina are placing themselves as global leaders in GM animals. The pro-GM attitude of these states is likely to cause a shift in the political evolution of global GMO governance.

  6. Activation of interferon regulatory factor 3 is inhibited by the influenza A virus NS1 protein.

    PubMed

    Talon, J; Horvath, C M; Polley, R; Basler, C F; Muster, T; Palese, P; García-Sastre, A

    2000-09-01

    We present a novel mechanism by which viruses may inhibit the alpha/beta interferon (IFN-alpha/beta) cascade. The double-stranded RNA (dsRNA) binding protein NS1 of influenza virus is shown to prevent the potent antiviral interferon response by inhibiting the activation of interferon regulatory factor 3 (IRF-3), a key regulator of IFN-alpha/beta gene expression. IRF-3 activation and, as a consequence, IFN-beta mRNA induction are inhibited in wild-type (PR8) influenza virus-infected cells but not in cells infected with an isogenic virus lacking the NS1 gene (delNS1 virus). Furthermore, NS1 is shown to be a general inhibitor of the interferon signaling pathway. Inhibition of IRF-3 activation can be achieved by the expression of wild-type NS1 in trans, not only in delNS1 virus-infected cells but also in cells infected with a heterologous RNA virus (Newcastle disease virus). We propose that inhibition of IRF-3 activation by a dsRNA binding protein significantly contributes to the virulence of influenza A viruses and possibly to that of other viruses.

  7. Interferon regulatory factor-1 polymorphisms are associated with the control of Plasmodium falciparum infection

    PubMed Central

    Mangano, Valentina D; Luoni, Gaia; Rockett, Kirk A; Sirima, Bienvenu S; Konaté, Amadou; Forton, Julian; Clark, Taane; Bancone, Germana; Akha, Elham Sadighi; Kwiatkowski, Dominic P; Modiano, David

    2010-01-01

    We describe the haplotypic structure of the Interferon Regulatory Factor-1 (IRF-1) locus in two West African ethnic groups, Fulani and Mossi, that differ in their susceptibility and immune response to Plasmodium falciparum malaria. Both populations showed significant associations between IRF-1 polymorphisms and carriage of P. falciparum infection, with different patterns of association that may reflect their different haplotypic architecture. Genetic variation at this locus does not therefore account for the Fulani-specific resistance to malaria while it could contribute to parasite clearance's ability in populations living in endemic areas. We then conducted a case-control study of three haplotype-tagging Single Nucleotide Polymorphisms (htSNPs) in 370 hospitalized malaria patients (160 severe and 210 uncomplicated) and 410 healthy population controls, all from the Mossi ethnic group. All 3 htSNPs showed correlation with blood infection levels in malaria patients, and the rs10065633 polymorphism was associated with severe disease (p=0.02). These findings provide the first evidence of the involvement in malaria susceptibility of a specific locus within the 5q31 region, previously shown to be linked with P. falciparum infection levels. PMID:18200030

  8. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells

    PubMed Central

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C.; Côté, Maxime C.; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-01-01

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors. PMID:27739523

  9. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells

    NASA Astrophysics Data System (ADS)

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C.; Côté, Maxime C.; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-10-01

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors.

  10. Characterization of Amphioxus IFN Regulatory Factor Family Reveals an Archaic Signaling Framework for Innate Immune Response.

    PubMed

    Yuan, Shaochun; Zheng, Tingting; Li, Peiyi; Yang, Rirong; Ruan, Jie; Huang, Shengfeng; Wu, Zhenxin; Xu, Anlong

    2015-12-15

    The IFN regulatory factor (IRF) family encodes transcription factors that play important roles in immune defense, stress response, reproduction, development, and carcinogenesis. Although the origin of the IRF family has been dated back to multicellular organisms, invertebrate IRFs differ from vertebrate IRFs in genomic structure and gene synteny, and little is known about their functions. Through comparison of multiple amphioxus genomes, in this study we suggested that amphioxus contains nine IRF members, whose orthologs are supposed to be shared among three amphioxus species. As the orthologs to the vertebrate IRF1 and IRF4 subgroups, Branchiostoma belcheri tsingtauense (bbt)IRF1 and bbtIRF8 bind the IFN-stimulated response element (ISRE) and were upregulated when amphioxus intestinal cells were stimulated with poly(I:C). As amphioxus-specific IRFs, both bbtIRF3 and bbtIRF7 bind ISRE. When activated, they can be phosphorylated by bbtTBK1 and then translocate into nucleus for target gene transcription. As transcriptional repressors, bbtIRF2 and bbtIRF4 can inhibit the transcriptional activities of bbtIRF1, 3, 7, and 8 by competing for the binding of ISRE. Interestingly, amphioxus IRF2, IRF8, and Rel were identified as target genes of bbtIRF1, bbtIRF7, and bbtIRF3, respectively, suggesting a dynamic feedback regulation among amphioxus IRF and NF-κB. Collectively, to our knowledge we present for the first time an archaic IRF signaling framework in a basal chordate, shedding new insights into the origin and evolution of vertebrate IFN-based antiviral networks.

  11. Contribution of Ser386 and Ser396 to activation of interferon regulatory factor 3.

    PubMed

    Chen, Weijun; Srinath, Hema; Lam, Suvana S; Schiffer, Celia A; Royer, William E; Lin, Kai

    2008-05-30

    IRF-3, a member of the interferon regulatory factor (IRF) family of transcription factors, functions in innate immune defense against viral infection. Upon infection, host cell IRF-3 is activated by phosphorylation at its seven C-terminal Ser/Thr residues: (385)SSLENTVDLHISNSHPLSLTS(405). This phosphoactivation triggers IRF-3 to react with the coactivators, CREB-binding protein (CBP)/p300, to form a complex that activates target genes in the nucleus. However, the role of each phosphorylation site for IRF-3 phosphoactivation remains unresolved. To address this issue, all seven Ser/Thr potential phosphorylation sites were screened by mutational studies, size-exclusion chromatography, and isothermal titration calorimetry. Using purified proteins, we show that CBP (amino acid residues 2067-2112) interacts directly with IRF-3 (173-427) and six of its single-site mutants to form heterodimers, but when CBP interacts with IRF-3 S396D, oligomerization is evident. CBP also interacts in vitro with IRF-3 double-site mutants to form different levels of oligomerization. Among all the single-site mutants, IRF-3 S396D showed the strongest binding to CBP. Although IRF-3 S386D alone did not interact as strongly with CBP as did other mutants, it strengthened the interaction and oligomerization of IRF-3 S396D with CBP. In contrast, IRF-3 S385D weakened the interaction and oligomerization of IRF-3 S396D and S386/396D with CBP. Thus, it appears that Ser385 and Ser386 serve antagonistic functions in regulating IRF-3 phosphoactivation. These results indicate that Ser386 and Ser396 are critical for IRF-3 activation, and support a phosphorylation-oligomerization model for IRF-3 activation.

  12. Identification of the Staphylococcus aureus vfrAB Operon, a Novel Virulence Factor Regulatory Locus

    PubMed Central

    Daly, Seth M.; Hall, Pamela R.; Bayles, Kenneth W.

    2014-01-01

    During a screen of the Nebraska Transposon Mutant Library, we identified 71 mutations in the Staphylococcus aureus genome that altered hemolysis on blood agar medium. Although many of these mutations disrupted genes known to affect the production of alpha-hemolysin, two of them were associated with an apparent operon, designated vfrAB, that had not been characterized previously. Interestingly, a ΔvfrB mutant exhibited only minor effects on the transcription of the hla gene, encoding alpha-hemolysin, when grown in broth, as well as on RNAIII, a posttranscriptional regulatory RNA important for alpha-hemolysin translation, suggesting that VfrB may function at the posttranscriptional level. Indeed, a ΔvfrB mutant had increased aur and sspAB protease expression under these conditions. However, disruption of the known secreted proteases in the ΔvfrB mutant did not restore hemolytic activity in the ΔvfrB mutant on blood agar. Further analysis revealed that, in contrast to the minor effects of VfrB on hla transcription when strains were cultured in liquid media, the level of hla transcription was decreased 50-fold in the absence of VfrB on solid media. These results demonstrate that while VfrB represses protease expression when strains are grown in broth, hla regulation is highly responsive to factors associated with growth on solid media. Intriguingly, the ΔvfrB mutant displayed increased pathogenesis in a model of S. aureus dermonecrosis, further highlighting the complexity of VfrB-dependent virulence regulation. The results of this study describe a phenotype associated with a class of highly conserved yet uncharacterized proteins found in Gram-positive bacteria, and they shed new light on the regulation of virulence factors necessary for S. aureus pathogenesis. PMID:24549328

  13. Identification of the Staphylococcus aureus vfrAB operon, a novel virulence factor regulatory locus.

    PubMed

    Bose, Jeffrey L; Daly, Seth M; Hall, Pamela R; Bayles, Kenneth W

    2014-05-01

    During a screen of the Nebraska Transposon Mutant Library, we identified 71 mutations in the Staphylococcus aureus genome that altered hemolysis on blood agar medium. Although many of these mutations disrupted genes known to affect the production of alpha-hemolysin, two of them were associated with an apparent operon, designated vfrAB, that had not been characterized previously. Interestingly, a ΔvfrB mutant exhibited only minor effects on the transcription of the hla gene, encoding alpha-hemolysin, when grown in broth, as well as on RNAIII, a posttranscriptional regulatory RNA important for alpha-hemolysin translation, suggesting that VfrB may function at the posttranscriptional level. Indeed, a ΔvfrB mutant had increased aur and sspAB protease expression under these conditions. However, disruption of the known secreted proteases in the ΔvfrB mutant did not restore hemolytic activity in the ΔvfrB mutant on blood agar. Further analysis revealed that, in contrast to the minor effects of VfrB on hla transcription when strains were cultured in liquid media, the level of hla transcription was decreased 50-fold in the absence of VfrB on solid media. These results demonstrate that while VfrB represses protease expression when strains are grown in broth, hla regulation is highly responsive to factors associated with growth on solid media. Intriguingly, the ΔvfrB mutant displayed increased pathogenesis in a model of S. aureus dermonecrosis, further highlighting the complexity of VfrB-dependent virulence regulation. The results of this study describe a phenotype associated with a class of highly conserved yet uncharacterized proteins found in Gram-positive bacteria, and they shed new light on the regulation of virulence factors necessary for S. aureus pathogenesis. PMID:24549328

  14. Regulatory SNPs Alter the Gene Expression of Diabetic Retinopathy Associated Secretary Factors

    PubMed Central

    Chen, Chian-Feng; Liou, Shiow-Wen; Wu, Hsin-Han; Lin, Chin-Hui; Huang, Li-Shan; Woung, Lin-Chung; Tsai, Ching-Yao

    2016-01-01

    Objectives: Diabetic retinopathy (DR) is a common microvascular complication in both type I and type II diabetes. Several previous reports indicated the serum centration of some secretary factors were highly associated with DR. Therefore, we hypothesis regulatory SNPs (rSNPs) genotype in secretary factors may alter these gene expression and lead to DR. Methods: At first, pyrosequencing were applying to screen the SNPs which present allele frequency different in DR and DNR. Then individual genotyping was processed by Taqman assays in Taiwanese DR and DNR patients. To evaluate the effect of SNP allele on transcriptional activity, we measured promoter activity using luciferase reporter constructs. Results: We found the frequencies of the CC, CG, and GG genotype of the rs2010963 polymorphism were 15.09%, 47.14%, and 37.74% in DR and 12.90%, 19.35%, and 67.74% in DNR, respectively (p = 0.0205). The prevalence of DR was higher (p = 0.00793) in patients with the CC or CG genotype (62.26% and 32.26% for DR and DNR, respectively) compared with the patients with the GG genotype. To evaluate the effect of rs2010963-C allele on transcriptional activity, we measured promoter activity using luciferase reporter constructs. The rs2010963-C reporter showed 1.6 to 2-fold higher luciferase activity than rs2010963-G in 3 cell lines. Conclusion: Our data proposed rs2010963-C altered the expression level of VEGFA in different tissues. We suggested small increase but long term exposure to VEGFA may lead to DR finally.

  15. Characterization of Amphioxus IFN Regulatory Factor Family Reveals an Archaic Signaling Framework for Innate Immune Response.

    PubMed

    Yuan, Shaochun; Zheng, Tingting; Li, Peiyi; Yang, Rirong; Ruan, Jie; Huang, Shengfeng; Wu, Zhenxin; Xu, Anlong

    2015-12-15

    The IFN regulatory factor (IRF) family encodes transcription factors that play important roles in immune defense, stress response, reproduction, development, and carcinogenesis. Although the origin of the IRF family has been dated back to multicellular organisms, invertebrate IRFs differ from vertebrate IRFs in genomic structure and gene synteny, and little is known about their functions. Through comparison of multiple amphioxus genomes, in this study we suggested that amphioxus contains nine IRF members, whose orthologs are supposed to be shared among three amphioxus species. As the orthologs to the vertebrate IRF1 and IRF4 subgroups, Branchiostoma belcheri tsingtauense (bbt)IRF1 and bbtIRF8 bind the IFN-stimulated response element (ISRE) and were upregulated when amphioxus intestinal cells were stimulated with poly(I:C). As amphioxus-specific IRFs, both bbtIRF3 and bbtIRF7 bind ISRE. When activated, they can be phosphorylated by bbtTBK1 and then translocate into nucleus for target gene transcription. As transcriptional repressors, bbtIRF2 and bbtIRF4 can inhibit the transcriptional activities of bbtIRF1, 3, 7, and 8 by competing for the binding of ISRE. Interestingly, amphioxus IRF2, IRF8, and Rel were identified as target genes of bbtIRF1, bbtIRF7, and bbtIRF3, respectively, suggesting a dynamic feedback regulation among amphioxus IRF and NF-κB. Collectively, to our knowledge we present for the first time an archaic IRF signaling framework in a basal chordate, shedding new insights into the origin and evolution of vertebrate IFN-based antiviral networks. PMID:26573836

  16. Regulatory SNPs Alter the Gene Expression of Diabetic Retinopathy Associated Secretary Factors

    PubMed Central

    Chen, Chian-Feng; Liou, Shiow-Wen; Wu, Hsin-Han; Lin, Chin-Hui; Huang, Li-Shan; Woung, Lin-Chung; Tsai, Ching-Yao

    2016-01-01

    Objectives: Diabetic retinopathy (DR) is a common microvascular complication in both type I and type II diabetes. Several previous reports indicated the serum centration of some secretary factors were highly associated with DR. Therefore, we hypothesis regulatory SNPs (rSNPs) genotype in secretary factors may alter these gene expression and lead to DR. Methods: At first, pyrosequencing were applying to screen the SNPs which present allele frequency different in DR and DNR. Then individual genotyping was processed by Taqman assays in Taiwanese DR and DNR patients. To evaluate the effect of SNP allele on transcriptional activity, we measured promoter activity using luciferase reporter constructs. Results: We found the frequencies of the CC, CG, and GG genotype of the rs2010963 polymorphism were 15.09%, 47.14%, and 37.74% in DR and 12.90%, 19.35%, and 67.74% in DNR, respectively (p = 0.0205). The prevalence of DR was higher (p = 0.00793) in patients with the CC or CG genotype (62.26% and 32.26% for DR and DNR, respectively) compared with the patients with the GG genotype. To evaluate the effect of rs2010963-C allele on transcriptional activity, we measured promoter activity using luciferase reporter constructs. The rs2010963-C reporter showed 1.6 to 2-fold higher luciferase activity than rs2010963-G in 3 cell lines. Conclusion: Our data proposed rs2010963-C altered the expression level of VEGFA in different tissues. We suggested small increase but long term exposure to VEGFA may lead to DR finally. PMID:27648002

  17. Myogenic regulatory factor (MRF) expression is affected by exercise in postnatal chicken skeletal muscles.

    PubMed

    Yin, Huadong; Li, Diyan; Wang, Yan; Zhao, Xiaoling; Liu, Yiping; Yang, Zhiqin; Zhu, Qing

    2015-05-01

    The MyoD1, MyoG, Myf5, and Mrf4 proteins belong to the family of muscle regulatory factors (MRFs) and play important roles in skeletal muscle hyperplasia and hypertrophy. We hypothesized that exercise would affect MRF mRNA and protein abundance in postnatal chicken skeletal muscle driving molecular changes that could ultimately lead to increased muscle fiber diameter. At day (d) 43, twelve hundred chickens with similar body weight were randomly assigned to cage, pen, and free-range groups. The MRF mRNA abundance was measured in the pectoralis major and thigh muscle at d56, d70, and d84, and the protein levels of MRFs were determined from the thigh muscle at d84. The results showed no significant difference in mRNA of the MRFs among the three groups at d56 (P>0.05). At d84, chicken in the pen and free-range group showed higher MyoD1, MyoG, Myf5, and Mrf4 mRNA abundance compared to the caged chickens (P<0.05). Free-range chickens had higher Mrf4 and MyoG expression than those in penned ones (P<0.05). Protein abundances of all four factors were lowest in the caged group, and Mrf4 and MyoG protein quantities were greatest in free-range chickens (P<0.05), but Myf5 and MyoD1 protein abundance did not differ between penned and caged groups. The results suggested that exercise up-regulated MRF expression in the postnatal skeletal muscles, which led to an increase in muscle fiber diameter, and eventually affected the meat quality of the skeletal muscles in adult chickens.

  18. Hypoxia inducible factor 1 alpha down-regulates type i collagen through Sp3 transcription factor in human chondrocytes.

    PubMed

    Duval, Elise; Bouyoucef, Mouloud; Leclercq, Sylvain; Baugé, Catherine; Boumédiene, Karim

    2016-09-01

    Cartilage engineering is one challenging issue in regenerative medicine. Low oxygen tension or hypoxia inducible factor-1 (HIF-1α) gene therapy are promising strategies in the field of cartilage repair. Previously, we showed that hypoxia and its mediator HIF-1 regulate matrix genes expression (collagens and aggrecan). Here, we investigated the molecular mechanism involved in the regulation of type I collagen (COL1A1) by HIF-1 in human articular chondrocytes. We show that HIF-1α reduces COL1A1 transcription, through a distal promoter (-2300 to -1816 bp upstream transcription initiation site), containing two GC boxes that bind Sp transcription factors (Sp1/Sp3). Sp1 acts as a positive regulator but is not induced by HIF-1. COL1A1 inhibition caused by HIF-1 implies only Sp3, which accumulates and competes Sp1 binding on COL1A1 promoter. Additionally, Sp3 ectopic expression inhibits COL1A1, while Sp3 knockdown counteracts the downregulation of COL1A1 induced by HIF-1. In conclusion, we established a new regulatory model of COL1A1 regulation by HIF-1, and bring out its relationship with Sp3 transcription factor. In a fundamental level, these findings give insights in the mechanisms controlling COL1A1 gene expression. This may be helpful to improve strategies to impair type I collagen expression during chondrocyte differentiation for cartilage engineering. © 2016 IUBMB Life, 68(9):756-763, 2016. PMID:27521280

  19. Regulatory and functional connection of microphthalmia-associated transcription factor and anti-metastatic pigment epithelium derived factor in melanoma.

    PubMed

    Fernández-Barral, Asunción; Orgaz, Jose Luis; Baquero, Pablo; Ali, Zaheer; Moreno, Alberto; Tiana, María; Gómez, Valentí; Riveiro-Falkenbach, Erica; Cañadas, Carmen; Zazo, Sandra; Bertolotto, Corine; Davidson, Irwin; Rodríguez-Peralto, Jose Luis; Palmero, Ignacio; Rojo, Federico; Jensen, Lasse Dahl; del Peso, Luis; Jiménez, Benilde

    2014-06-01

    Pigment epithelium-derived factor (PEDF), a member of the serine protease inhibitor superfamily, has potent anti-metastatic effects in cutaneous melanoma through its direct actions on endothelial and melanoma cells. Here we show that PEDF expression positively correlates with microphthalmia-associated transcription factor (MITF) in melanoma cell lines and human samples. High PEDF and MITF expression is characteristic of low aggressive melanomas classified according to molecular and pathological criteria, whereas both factors are decreased in senescent melanocytes and naevi. Importantly, MITF silencing down-regulates PEDF expression in melanoma cell lines and primary melanocytes, suggesting that the correlation in the expression reflects a causal relationship. In agreement, analysis of Chromatin immunoprecipitation coupled to high throughput sequencing (ChIP-seq) data sets revealed three MITF binding regions within the first intron of SERPINF1, and reporter assays demonstrated that the binding of MITF to these regions is sufficient to drive transcription. Finally, we demonstrate that exogenous PEDF expression efficiently halts in vitro migration and invasion, as well as in vivo dissemination of melanoma cells induced by MITF silencing. In summary, these results identify PEDF as a novel transcriptional target of MITF and support a relevant functional role for the MITF-PEDF axis in the biology of melanoma.

  20. Scatter factor induces blood vessel formation in vivo.

    PubMed Central

    Grant, D S; Kleinman, H K; Goldberg, I D; Bhargava, M M; Nickoloff, B J; Kinsella, J L; Polverini, P; Rosen, E M

    1993-01-01

    Scatter factor (also known as hepatocyte growth factor) is a glycoprotein secreted by stromal cells that stimulates cell motility and proliferation. In vitro, scatter factor stimulates vascular endothelial cell migration, proliferation, and organization into capillary-like tubes. Using two different in vivo assays, we showed that physiologic quantities of purified native mouse scatter factor and recombinant human hepatocyte growth factor induce angiogenesis (the formation of new blood vessels). The angiogenic activity was blocked by specific anti-scatter factor antibodies. Scatter factor induced cultured microvascular endothelial cells to accumulate and secrete significantly increased quantities of urokinase, an enzyme associated with development of an invasive endothelial phenotype during angiogenesis. We further showed that immunoreactive scatter factor is present surrounding sites of blood vessel formation in psoriatic skin. These findings suggest that scatter factor may act as a paracrine mediator in pathologic angiogenesis associated with human inflammatory disease. Images Fig. 1 Fig. 2 Fig. 3 Fig. 5 PMID:7680481

  1. Therapeutic Effect of Ergotope Peptides on Collagen-Induced Arthritis by Downregulation of Inflammatory and Th1/Th17 Responses and Induction of Regulatory T Cells

    PubMed Central

    Niu, Xiaoyin; Deng, Shaohua; Li, Shan; Xi, Yebin; Li, Chengzhen; Wang, Li; He, Dongyi; Wang, Zhaojun; Chen, Guangjie

    2016-01-01

    Rheumatoid arthritis (RA) is a systemic autoimmune disease that results in a chronic and inflammatory disorder. Dynamic balance of helper T cells (Th) 1 and 17 and regulatory T cells (Treg) is broken in RA. Since there is no cure for RA at present, it is necessary to find a truly effective and convenient treatment. Several studies have intended to induce ergotopic regulation to treat autoimmune diseases. This study was undertaken to find potential ergotope peptides and investigate their effects in treating the animal model of RA and their underlying regulatory mechanisms. First, we selected functional ergotope peptides from 25 overlapping peptides derived from the interleukin 2 receptor (IL-2R) α chain, and then used these peptides to treat collagen-induced arthritis (CIA). We showed ergotope peptides as immunomodulatory factors with great benefits at the clinical and pathologic levels. This effect was associated with inhibition of type II collagen (CII)-specific proliferation and autoantibody production as well as induction of antiergotypic immune response, downregulation of both Th1 and Th17 cells and their related components, and emergence of Treg cells that had suppressive action on autoreactive T cells. We also proved that cytotoxic T lymphocyte–associated antigen-4 (CTLA-4) and IL-10 are two important mediators that are critical to Treg suppressive function. Inhibition of Th1 and Th17 in established CIA could be attributed to ergotope-induced Treg cells. Our findings reveal that ergotope peptides induce regulatory immune responses and restore immune tolerance, suggesting that treatment with ergotope peptides may be a novel approach to therapy for RA patients and has good application prospects, with cheap, effective, convenient, wide-spectrum features. PMID:27579476

  2. CD25+ regulatory T cell inhibition enhances vaccine-induced immunity to neuroblastoma.

    PubMed

    Johnson, Bryon D; Jing, Weiqing; Orentas, Rimas J

    2007-01-01

    Evidence that CD4CD25 regulatory T (Treg) cells play a role in the progression of cancer continues to mount. There is a great deal of interest as to whether transient elimination or functional inhibition of these cells can improve the efficacy of immunotherapy for cancer. Our goals in this study were to test whether treatment of mice with anti-CD25 monoclonal antibody (mAb) (PC61) could induce rejection of a murine neuroblastoma, whether anti-CD25 treatment could increase tumor immunity when administered just before cell-based vaccination, and to learn how anti-CD25 treatment influences the vaccine-induced antitumor response. Treatment of mice with anti-CD25 mAb induced rejection of the mouse neuroblastoma, Neuro-2a, as 90% of anti-CD25-treated mice survived challenge with a lethal dose of tumor cells. In vivo anti-CD25 mAb treatment before the first of 2 weekly vaccines significantly improved the survival of tumor-vaccinated/challenged mice (75% vs. 33% survival), whereas antibody treatment before each of the 2 vaccines did not, suggesting that excessive treatment with anti-CD25 mAb interferes with activated antitumor effector cells. A detailed phenotypic analysis of tissues from anti-CD25-treated mice indicated that the antibody partially depletes CD4Foxp3 Treg cells (25% to 40%) in A/J mice, and that the antibody may inhibit the remaining cells by inducing loss of CD25 expression and blocking CD25 molecules, partially confirming recent data from other investigators. Importantly, we found that in vivo anti-CD25 mAb treatment significantly decreased the contribution of asialo GM1 cells in the antitumor response. As we did not see a direct effect of anti-CD25 mAb on in vitro assays of immune cell function in spleen cells from treated animals, this indicates that inhibition of Treg cells amplifies the immune response in vivo in a manner that bypasses the requirement for innate immune activation, potentially mediated by natural killer cells, and allows for protective

  3. PKA regulatory subunit 1A inactivating mutation induces serotonin signaling in primary pigmented nodular adrenal disease

    PubMed Central

    Bram, Zakariae; Louiset, Estelle; Renouf, Sylvie; Duparc, Céline; Boutelet, Isabelle; Rizk-Rabin, Marthe; Libé, Rossella; Young, Jacques; Carson, Dennis; Vantyghem, Marie-Christine; Szarek, Eva; Martinez, Antoine; Stratakis, Constantine A.; Bertherat, Jérôme

    2016-01-01

    Primary pigmented nodular adrenocortical disease (PPNAD) is a rare cause of ACTH-independent hypercortisolism. The disease is primarily caused by germline mutations of the protein kinase A (PKA) regulatory subunit 1A (PRKAR1A) gene, which induces constitutive activation of PKA in adrenocortical cells. Hypercortisolism is thought to result from PKA hyperactivity, but PPNAD tissues exhibit features of neuroendocrine differentiation, which may lead to stimulation of steroidogenesis by abnormally expressed neurotransmitters. We hypothesized that serotonin (5-HT) may participate in the pathophysiology of PPNAD-associated hypercortisolism. We show that PPNAD tissues overexpress the 5-HT synthesizing enzyme tryptophan hydroxylase type 2 (Tph2) and the serotonin receptors types 4, 6, and 7, leading to formation of an illicit stimulatory serotonergic loop whose pharmacological inhibition in vitro decreases cortisol production. In the human PPNAD cell line CAR47, the PKA inhibitor H-89 decreases 5-HT4 and 5-HT7 receptor expression. Moreover, in the human adrenocortical cell line H295R, inhibition of PRKAR1A expression increases the expression of Tph2 and 5-HT4/6/7 receptors, an effect that is blocked by H-89. These findings show that the serotonergic process observed in PPNAD tissues results from PKA activation by PRKAR1A mutations. They also suggest that Tph inhibitors may represent efficient treatments of hypercortisolism in patients with PPNAD.

  4. PKA regulatory subunit 1A inactivating mutation induces serotonin signaling in primary pigmented nodular adrenal disease

    PubMed Central

    Bram, Zakariae; Louiset, Estelle; Renouf, Sylvie; Duparc, Céline; Boutelet, Isabelle; Rizk-Rabin, Marthe; Libé, Rossella; Young, Jacques; Carson, Dennis; Vantyghem, Marie-Christine; Szarek, Eva; Martinez, Antoine; Stratakis, Constantine A.; Bertherat, Jérôme

    2016-01-01

    Primary pigmented nodular adrenocortical disease (PPNAD) is a rare cause of ACTH-independent hypercortisolism. The disease is primarily caused by germline mutations of the protein kinase A (PKA) regulatory subunit 1A (PRKAR1A) gene, which induces constitutive activation of PKA in adrenocortical cells. Hypercortisolism is thought to result from PKA hyperactivity, but PPNAD tissues exhibit features of neuroendocrine differentiation, which may lead to stimulation of steroidogenesis by abnormally expressed neurotransmitters. We hypothesized that serotonin (5-HT) may participate in the pathophysiology of PPNAD-associated hypercortisolism. We show that PPNAD tissues overexpress the 5-HT synthesizing enzyme tryptophan hydroxylase type 2 (Tph2) and the serotonin receptors types 4, 6, and 7, leading to formation of an illicit stimulatory serotonergic loop whose pharmacological inhibition in vitro decreases cortisol production. In the human PPNAD cell line CAR47, the PKA inhibitor H-89 decreases 5-HT4 and 5-HT7 receptor expression. Moreover, in the human adrenocortical cell line H295R, inhibition of PRKAR1A expression increases the expression of Tph2 and 5-HT4/6/7 receptors, an effect that is blocked by H-89. These findings show that the serotonergic process observed in PPNAD tissues results from PKA activation by PRKAR1A mutations. They also suggest that Tph inhibitors may represent efficient treatments of hypercortisolism in patients with PPNAD. PMID:27699247

  5. Haploinsufficiency of interferon regulatory factor 6 alters brain morphology in the mouse.

    PubMed

    Aerts, Andrea; DeVolder, Ian; Weinberg, Seth M; Thedens, Dan; Dunnwald, Martine; Schutte, Brian C; Nopoulos, Peg

    2014-03-01

    Orofacial clefts are among the commonest birth defects. Among many genetic contributors to orofacial clefting, Interferon Regulatory Factor 6 (IRF6) is unique since mutations in this gene cause Van der Woude (VWS), the most common clefting syndrome. Furthermore, variants in IRF6 contribute to increased risk for non-syndromic cleft lip and/or palate (NSCL/P). Our previous work shows that individuals with either VWS or NSCL/P may have cerebral anomalies (larger anterior, smaller posterior regions), and a smaller cerebellum. The objective of this study was to test the hypothesis that disrupting Irf6 in the mouse will result in quantitative brain changes similar to those reported for humans with VWS and NSCL/P. Male mice heterozygous for Irf6 (Irf6(gt1/+); n = 9) and wild-type (Irf6(+/+) ; n = 6) mice at comparable age underwent a 4.7-T MRI scan to obtain quantitative measures of cortical and subcortical brain structures. There was no difference in total brain volume between groups. However, the frontal cortex was enlarged in the Irf6(gt1/+) mice compared to that of wild types (P = 0.028) while the posterior cortex did not differ. In addition, the volume of the cerebellum of Irf6(gt1/+) mice was decreased (P = 0.004). Mice that were heterozygous for Irf6 showed a similar pattern of brain anomalies previously reported in humans with VWS and NSCL/P. These structural differences were present in the absence of overt oral clefts. These results support a role for IRF6 in brain morphometry and provide evidence for a potential genetic link to abnormal brain development in orofacial clefting. PMID:24357509

  6. Activation of Sterol Regulatory Element Binding Factors by Fenofibrate and Gemfibrozil Stimulates Myelination in Zebrafish.

    PubMed

    Ashikawa, Yoshifumi; Nishimura, Yuhei; Okabe, Shiko; Sasagawa, Shota; Murakami, Soichiro; Yuge, Mizuki; Kawaguchi, Koki; Kawase, Reiko; Tanaka, Toshio

    2016-01-01

    Oligodendrocytes are major myelin-producing cells and play essential roles in the function of a healthy nervous system. However, they are also one of the most vulnerable neural cell types in the central nervous system (CNS), and myelin abnormalities in the CNS are found in a wide variety of neurological disorders, including multiple sclerosis, adrenoleukodystrophy, and schizophrenia. There is an urgent need to identify small molecular weight compounds that can stimulate myelination. In this study, we performed comparative transcriptome analysis to identify pharmacodynamic effects common to miconazole and clobetasol, which have been shown to stimulate myelination by mouse oligodendrocyte progenitor cells (OPCs). Of the genes differentially expressed in both miconazole- and clobetasol-treated mouse OPCs compared with untreated cells, we identified differentially expressed genes (DEGs) common to both drug treatments. Gene ontology analysis revealed that these DEGs are significantly associated with the sterol biosynthetic pathway, and further bioinformatics analysis suggested that sterol regulatory element binding factors (SREBFs) might be key upstream regulators of the DEGs. In silico screening of a public database for chemicals associated with SREBF activation identified fenofibrate, a peroxisome proliferator-activated receptor α (PPARα) agonist, as a drug that increases the expression of known SREBF targets, raising the possibility that fenofibrate may also stimulate myelination. To test this, we performed in vivo imaging of zebrafish expressing a fluorescent reporter protein under the control of the myelin basic protein (mbp) promoter. Treatment of zebrafish with fenofibrate significantly increased expression of the fluorescent reporter compared with untreated zebrafish. This increase was attenuated by co-treatment with fatostatin, a specific inhibitor of SREBFs, confirming that the fenofibrate effect was mediated via SREBFs. Furthermore, incubation of zebrafish

  7. Regulatory mechanisms underlying sepsis progression in patients with tumor necrosis factor-α genetic variations

    PubMed Central

    LIU, YANGZHOU; HAN, NING; LI, QINCHUAN; LI, ZENGCHUN

    2016-01-01

    The present study aimed to investigate the regulatory mechanisms underlying sepsis progression in patients with tumor necrosis factor (TNF)-α genetic variations. The GSE5760 expression profile data, which was downloaded from the Gene Expression Omnibus database, contained 30 wild-type (WT) and 28 mutation (MUT) samples. Differentially expressed genes (DEGs) between the two types of samples were identified using the Student's t-test, and the corresponding microRNAs (miRNAs) were screened using WebGestalt software. An integrated miRNA-DEG network was constructed using the Cytoscape software, based on the interactions between the DEGs, as identified using the Search Tool for the Retrieval of Interacting Genes/Proteins database, and the correlation between miRNAs and their target genes. Furthermore, Gene Ontology and pathway enrichment analyses were conducted for the DEGs using the Database for Annotation, Visualization and Integrated Discovery and the KEGG Orthology Based Annotation System, respectively. A total of 390 DEGS between the WT and MUT samples, along with 11 -associated miRNAs, were identified. The integrated miRNA-DEG network consisted of 38 DEGs and 11 miRNAs. Within this network, COPS2 was found to be associated with transcriptional functions, while FUS was found to be involved in mRNA metabolic processes. Other DEGs, including FBXW7 and CUL3, were enriched in the ubiquitin-mediated proteolysis pathway. In addition, miR-15 was predicted to target COPS2 and CUL3. The results of the present study suggested that COPS2, FUS, FBXW7 and CUL3 may be associated with sepsis in patients with TNF-α genetic variations. In the progression of sepsis, FBXW7 and CUL3 may participate in the ubiquitin-mediated proteolysis pathway, whereas COPS2 may regulate the phosphorylation and ubiquitination of the FUS protein. Furthermore, COPS2 and CUL3 may be novel targets of miR-15. PMID:27347057

  8. Activation of Sterol Regulatory Element Binding Factors by Fenofibrate and Gemfibrozil Stimulates Myelination in Zebrafish

    PubMed Central

    Ashikawa, Yoshifumi; Nishimura, Yuhei; Okabe, Shiko; Sasagawa, Shota; Murakami, Soichiro; Yuge, Mizuki; Kawaguchi, Koki; Kawase, Reiko; Tanaka, Toshio

    2016-01-01

    Oligodendrocytes are major myelin-producing cells and play essential roles in the function of a healthy nervous system. However, they are also one of the most vulnerable neural cell types in the central nervous system (CNS), and myelin abnormalities in the CNS are found in a wide variety of neurological disorders, including multiple sclerosis, adrenoleukodystrophy, and schizophrenia. There is an urgent need to identify small molecular weight compounds that can stimulate myelination. In this study, we performed comparative transcriptome analysis to identify pharmacodynamic effects common to miconazole and clobetasol, which have been shown to stimulate myelination by mouse oligodendrocyte progenitor cells (OPCs). Of the genes differentially expressed in both miconazole- and clobetasol-treated mouse OPCs compared with untreated cells, we identified differentially expressed genes (DEGs) common to both drug treatments. Gene ontology analysis revealed that these DEGs are significantly associated with the sterol biosynthetic pathway, and further bioinformatics analysis suggested that sterol regulatory element binding factors (SREBFs) might be key upstream regulators of the DEGs. In silico screening of a public database for chemicals associated with SREBF activation identified fenofibrate, a peroxisome proliferator-activated receptor α (PPARα) agonist, as a drug that increases the expression of known SREBF targets, raising the possibility that fenofibrate may also stimulate myelination. To test this, we performed in vivo imaging of zebrafish expressing a fluorescent reporter protein under the control of the myelin basic protein (mbp) promoter. Treatment of zebrafish with fenofibrate significantly increased expression of the fluorescent reporter compared with untreated zebrafish. This increase was attenuated by co-treatment with fatostatin, a specific inhibitor of SREBFs, confirming that the fenofibrate effect was mediated via SREBFs. Furthermore, incubation of zebrafish

  9. Activation of Sterol Regulatory Element Binding Factors by Fenofibrate and Gemfibrozil Stimulates Myelination in Zebrafish.

    PubMed

    Ashikawa, Yoshifumi; Nishimura, Yuhei; Okabe, Shiko; Sasagawa, Shota; Murakami, Soichiro; Yuge, Mizuki; Kawaguchi, Koki; Kawase, Reiko; Tanaka, Toshio

    2016-01-01

    Oligodendrocytes are major myelin-producing cells and play essential roles in the function of a healthy nervous system. However, they are also one of the most vulnerable neural cell types in the central nervous system (CNS), and myelin abnormalities in the CNS are found in a wide variety of neurological disorders, including multiple sclerosis, adrenoleukodystrophy, and schizophrenia. There is an urgent need to identify small molecular weight compounds that can stimulate myelination. In this study, we performed comparative transcriptome analysis to identify pharmacodynamic effects common to miconazole and clobetasol, which have been shown to stimulate myelination by mouse oligodendrocyte progenitor cells (OPCs). Of the genes differentially expressed in both miconazole- and clobetasol-treated mouse OPCs compared with untreated cells, we identified differentially expressed genes (DEGs) common to both drug treatments. Gene ontology analysis revealed that these DEGs are significantly associated with the sterol biosynthetic pathway, and further bioinformatics analysis suggested that sterol regulatory element binding factors (SREBFs) might be key upstream regulators of the DEGs. In silico screening of a public database for chemicals associated with SREBF activation identified fenofibrate, a peroxisome proliferator-activated receptor α (PPARα) agonist, as a drug that increases the expression of known SREBF targets, raising the possibility that fenofibrate may also stimulate myelination. To test this, we performed in vivo imaging of zebrafish expressing a fluorescent reporter protein under the control of the myelin basic protein (mbp) promoter. Treatment of zebrafish with fenofibrate significantly increased expression of the fluorescent reporter compared with untreated zebrafish. This increase was attenuated by co-treatment with fatostatin, a specific inhibitor of SREBFs, confirming that the fenofibrate effect was mediated via SREBFs. Furthermore, incubation of zebrafish

  10. Cloning and expression analysis of interferon regulatory factor 7 in the Pacific cod, Gadus macrocephalus.

    PubMed

    Sun, Hang; Jiang, Zhiqiang; Mao, Mingguang; Huo, Yuan; Han, Yuzhe; Zhang, Saisai

    2016-02-01

    Interferon regulatory factor 7 (IRF7) plays an important role in regulating the response of type I interferon (IFN) to viral infection. To understand the mechanisms underlying immune reactions in the Pacific cod, Gadus macrocephalus, the gene encoding G. macrocephalus IRF7 was cloned and characterized. The cDNA of G. macrocephalus IRF7 was also cloned and sequenced. A cDNA sequence of 2032 bp was assembled using polymerase chain reaction (PCR) products. It contains an open reading frame of 1323 bp in length, which encoded a 440-amino acid polypeptide that comprised a DNA-binding domain (DBD), an IRF association domain (IAD), and a serine-rich domain (SRD). In the DBD, the tryptophan cluster consisted of only four tryptophans, which is a unique characteristic in fish IRF7. The mRNA of IRF7 was detected in various tissues, including in the spleen, thymus, kidney, intestine, and gills, using relative quantification PCR (R-qPCR). Dynamic expression of IRF7 was observed in larvae throughout post-hatching (ph) development, with the highest level detected at day of ph (dph) 25. Response to immune stimulation was examined by challenging larvae with polyriboinosinic polyribocytidylic acid (pIC) to mimic viral infection and elicit an immune reaction. R-qPCR revealed that the expression of IRF7 significantly increased in pIC-treated groups relative to that in the control groups, in a time-dependent manner, with peak responses at 48 and 72 h after pIC-treatment. These results show that IRF7 is expressed in various tissues of adult fish and larvae and is sensitive to viral infection, suggesting that it plays a role in antiviral immune defense in G. macrocephalus. PMID:26702560

  11. Muscle regulatory factors regulate T1R3 taste receptor expression.

    PubMed

    Kokabu, Shoichiro; Lowery, Jonathan W; Toyono, Takashi; Seta, Yuji; Hitomi, Suzuro; Sato, Tsuyoshi; Enoki, Yuichiro; Okubo, Masahiko; Fukushima, Yosuke; Yoda, Tetsuya

    2015-12-25

    T1R3 is a T1R class of G protein-coupled receptors, composing subunit of the umami taste receptor when complexed with T1R1. T1R3 was originally discovered in gustatory tissue but is now known to be expressed in a wide variety of tissues and cell types such the intestine, pancreatic β-cells, skeletal muscle, and heart. In addition to taste recognition, the T1R1/T1R3 complex functions as an amino acid sensor and has been proposed to be a control mechanism for the secretion of hormones, such as cholecystokinin, insulin, and duodenal HCO3(-) and activates the mammalian rapamycin complex 1 (MTORC1) to inhibit autophagy. T1R3 knockout mice have increased rate of autophagy in the heart, skeletal muscle and liver. Thus, T1R3 has multiple physiological functions and is widely expressed in vivo. However, the exact mechanisms regulating T1R3 expression are largely unknown. Here, we used comparative genomics and functional analyses to characterize the genomic region upstream of the annotated transcriptional start of human T1R3. This revealed that the T1R3 promoter in human and mouse resides in an evolutionary conserved region (ECR). We also identified a repressive element located upstream of the human T1R3 promoter that has relatively high degree of conservation with rhesus macaque. Additionally, the muscle regulatory factors MyoD and Myogenin regulate T1R3 expression and T1R3 expression increases with skeletal muscle differentiation of murine myoblast C2C12 cells. Taken together, our study raises the possibility that MyoD and Myogenin might control skeletal muscle metabolism and homeostasis through the regulation of T1R3 promoter activity. PMID:26545778

  12. Molecular cloning and characterization of interferon regulatory factor 9 (IRF9) in Japanese flounder, Paralichthys olivaceus.

    PubMed

    Hu, Guo-Bin; Zhao, Ming-Yu; Lin, Jing-Yun; Liu, Qiu-Ming; Zhang, Shi-Cui

    2014-08-01

    Interferon regulatory factor 9 (IRF9) in mammals is known to be involved in antiviral response. In this study, we studied the structure, mRNA tissue distribution and regulation of IRF9 from Japanese flounder, Paralichthys olivaceus. The cDNA sequence of IRF9 is 3305 bp long, containing an open reading frame (ORF) of 1308 bp that encodes a peptide of 435 amino acids. The predicted protein sequence shares 33.7-72.0% identity to other fish IRF9s. Japanese flounder IRF9 possesses a DNA-binding domain (DBD), an IRF association domain (IAD), two nuclear localization signals (NLSs) and a proline-rich domain (PRD). The IRF9 transcripts were detectable in all examined tissues of healthy Japanese flounders, with higher levels in the head kidney, kidney, liver and spleen. The IRF9 mRNA levels were up-regulated in the gills, head kidney, spleen and muscle when challenged with polyinosinic:polycytidylic acid (poly I:C) or lymphocystis disease virus (LCDV). The up-regulations were stronger and arose earlier in the case of poly I:C treatment in most tested organs in a 7-day time course, with maximum increases ranging from 1.37- to 8.59-fold and peak time points from 3 h to 3 d post injection depending on different organs, relative to those in the case of LCDV treatment which ranged from 1.32- to 3.21-fold and from 18 h to 3 d post injection, respectively. The highest and earliest inductions were detected in the spleen in both challenge cases, while the inductions by LCDV in the muscle were quite faint. These results demonstrate a role of Japanese flounder IRF9 in the host's antiviral responses. PMID:24837327

  13. Protective Effects of Total Glucosides of Paeony on N-nitrosodiethylamine-induced Hepatocellular Carcinoma in Rats via Down-regulation of Regulatory B Cells.

    PubMed

    Song, S S; Yuan, P F; Li, P P; Wu, H X; Ni, W J; Lu, J T; Wei, W

    2015-01-01

    Total glucoside of paeony (TGP), extracted from the root of Paeonia Lactiflora, has been known to show anti-inflammatory, anti-oxidative, hepato-protective and immuno-regulatory activities. The aim of this present study was to determine the anti-tumor effect of TGP against N-nitrosodiethylamine (DEN)-induced hepatocellular carcinoma (HCC) in rats, and to find the related mechanisms. Rat HCC model was established by intragastrically administrating with DEN (8 mg/kg). We found the number of tumor nodules and the index of liver and spleen were increased in the model group compared with the normal group, and was significantly decreased by TGP. Additionally, TGP obviously improved the hepatic pathological lesions induced by DEN, and decreased the elevated levels of serum alanine aminotransferase (ALT), glutamic oxalacetic transaminase (AST), alkaline phosphatase (ALP) and alpha fetoprotein (AFP) by DEN. Moreover, TGP decreased the level of B cell-activating factor (BAFF) and the proportion of IL-10-producing regulatory B cells (Bregs), and the decrease of BAFF by TGP is positively correlated to the decrease of IL-10-producing Bregs by TGP. These results suggest that TGP had a good therapeutic action on DEN-induced HCC rats, which might be due to its down-regulation of Bregs through reducing the level of BAFF. PMID:26207789

  14. Construction of pancreatic cancer double-factor regulatory network based on chip data on the transcriptional level.

    PubMed

    Zhao, Li-Li; Zhang, Tong; Liu, Bing-Rong; Liu, Tie-Fu; Tao, Na; Zhuang, Li-Wei

    2014-05-01

    Transcription factor (TF) and microRNA (miRNA) have been discovered playing crucial roles in cancer development. However, the effect of TFs and miRNAs in pancreatic cancer pathogenesis remains vague. We attempted to reveal the possible mechanism of pancreatic cancer based on transcription level. Using GSE16515 datasets downloaded from gene expression omnibus database, we first identified the differentially expressed genes (DEGs) in pancreatic cancer by the limma package in R. Then the DEGs were mapped into DAVID to conduct the kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis. TFs and miRNAs that DEGs significantly enriched were identified by Fisher's test, and then the pancreatic cancer double-factor regulatory network was constructed. In our study, total 1117 DEGs were identified and they significantly enriched in 4 KEGG pathways. A double-factor regulatory network was established, including 29 DEGs, 24 TFs, 25 miRNAs. In the network, LAMC2, BRIP1 and miR155 were identified which may be involved in pancreatic cancer development. In conclusion, the double-factor regulatory network was found to play an important role in pancreatic cancer progression and our results shed new light on the molecular mechanism of pancreatic cancer.

  15. LGP2 Downregulates Interferon Production during Infection with Seasonal Human Influenza A Viruses That Activate Interferon Regulatory Factor 3

    PubMed Central

    Malur, Meghana; Gale, Michael

    2012-01-01

    LGP2, a member of the RIG-I-like receptor family, lacks the amino-terminal caspase activation recruitment domains (CARDs) required for initiating the activation of interferon regulatory factor 3 (IRF3) and interferon (IFN) transcription. The role of LGP2 in virus infection is controversial, and the only LGP2 experiments previously carried out with mammalian influenza A viruses employed an attenuated, mouse-adapted H1N1 A/PR/8/34 (PR8) virus that does not encode the NS1 protein. Here we determine whether LGP2 has a role during infection with wild-type, nonattenuated influenza A viruses that have circulated in the human population, specifically two types of seasonal influenza A viruses: (i) H3N2 and H1N1 viruses that activate IRF3 and IFN transcription and (ii) recent H1N1 viruses that block these two activations. In human cells infected with an H3N2 virus that activates IRF3, overexpression of LGP2 or its repressor domain decreased STAT1 activation and IFN-β transcription approximately 10-fold. Overexpression of LGP2 also caused a 10-fold decrease of STAT1 activation during infection with other seasonal influenza A viruses that activate IRF3. Using LGP2+/+ and LGP2−/− mouse cells, we show that endogenous LGP2 decreased IFN production during H3N2 virus infection 3- to 4-fold. In contrast, in both mouse and human cells infected with H1N1 viruses that do not activate IRF3, LGP2 had no detectable role. These results demonstrate that LGP2 downregulates IFN production during infection by seasonal influenza A viruses that activate IRF3 and IFN transcription. It is intriguing that LGP2, a host protein induced during influenza A virus infection, downregulates the host antiviral IFN response. PMID:22837208

  16. Inhibition of mRNA export and dimerization of interferon regulatory factor 3 by Theiler's virus leader protein.

    PubMed

    Ricour, Céline; Delhaye, Sophie; Hato, Stanleyson V; Olenyik, Tamara D; Michel, Bénédicte; van Kuppeveld, Frank J M; Gustin, Kurt E; Michiels, Thomas

    2009-01-01

    Theiler's murine encephalomyelitis virus (TMEV or Theiler's virus) is a neurotropic picornavirus that can persist lifelong in the central nervous system of infected mice, causing a chronic inflammatory demyelinating disease. The leader (L) protein of the virus is an important determinant of viral persistence and has been shown to inhibit transcription of type I interferon (IFN) genes and to cause nucleocytoplasmic redistribution of host proteins. In this study, it was shown that expression of the L protein shuts off synthesis of the reporter proteins green fluorescent protein and firefly luciferase, suggesting that it induces a global shut-off of host protein expression. The L protein did not inhibit transcription or translation of the reporter genes, but blocked cellular mRNA export from the nucleus. This activity correlated with the phosphorylation of nucleoporin 98 (Nup98), an essential component of the nuclear pore complex. In contrast, the data confirmed that the L protein inhibited IFN expression at the transcriptional level, and showed that transcription of other chemokine or cytokine genes was affected by the L protein. This transcriptional inhibition correlated with inhibition of interferon regulatory factor 3 (IRF-3) dimerization. Whether inhibition of IRF-3 dimerization and dysfunction of the nuclear pore complex are related phenomena remains an open question. In vivo, IFN antagonism appears to be an important role of the L protein early in infection, as a virus bearing a mutation in the zinc finger of the L protein replicated as efficiently as the wild-type virus in type I IFN receptor-deficient mice, but had impaired fitness in IFN-competent mice.

  17. LGP2 downregulates interferon production during infection with seasonal human influenza A viruses that activate interferon regulatory factor 3.

    PubMed

    Malur, Meghana; Gale, Michael; Krug, Robert M

    2012-10-01

    LGP2, a member of the RIG-I-like receptor family, lacks the amino-terminal caspase activation recruitment domains (CARDs) required for initiating the activation of interferon regulatory factor 3 (IRF3) and interferon (IFN) transcription. The role of LGP2 in virus infection is controversial, and the only LGP2 experiments previously carried out with mammalian influenza A viruses employed an attenuated, mouse-adapted H1N1 A/PR/8/34 (PR8) virus that does not encode the NS1 protein. Here we determine whether LGP2 has a role during infection with wild-type, nonattenuated influenza A viruses that have circulated in the human population, specifically two types of seasonal influenza A viruses: (i) H3N2 and H1N1 viruses that activate IRF3 and IFN transcription and (ii) recent H1N1 viruses that block these two activations. In human cells infected with an H3N2 virus that activates IRF3, overexpression of LGP2 or its repressor domain decreased STAT1 activation and IFN-β transcription approximately 10-fold. Overexpression of LGP2 also caused a 10-fold decrease of STAT1 activation during infection with other seasonal influenza A viruses that activate IRF3. Using LGP2(+/+) and LGP2(-/-) mouse cells, we show that endogenous LGP2 decreased IFN production during H3N2 virus infection 3- to 4-fold. In contrast, in both mouse and human cells infected with H1N1 viruses that do not activate IRF3, LGP2 had no detectable role. These results demonstrate that LGP2 downregulates IFN production during infection by seasonal influenza A viruses that activate IRF3 and IFN transcription. It is intriguing that LGP2, a host protein induced during influenza A virus infection, downregulates the host antiviral IFN response.

  18. Dissection of the regulatory mechanism of a heat-shock responsive promoter in Haloarchaea: a new paradigm for general transcription factor directed archaeal gene regulation

    PubMed Central

    Lu, Qiuhe; Han, Jing; Zhou, Ligang; Coker, James A.; DasSarma, Priya; DasSarma, Shiladitya; Xiang, Hua

    2008-01-01

    Multiple general transcription factors (GTFs), TBP and TFB, are present in many haloarchaea, and are deemed to accomplish global gene regulation. However, details and the role of GTF-directed transcriptional regulation in stress response are still not clear. Here, we report a comprehensive investigation of the regulatory mechanism of a heat-induced gene (hsp5) from Halobacterium salinarum. We demonstrated by mutation analysis that the sequences 5′ and 3′ to the core elements (TATA box and BRE) of the hsp5 promoter (Phsp5) did not significantly affect the basal and heat-induced gene expression, as long as the transcription initiation site was not altered. Moreover, the BRE and TATA box of Phsp5 were sufficient to render a nonheat-responsive promoter heat-inducible, in both Haloferax volcanii and Halobacterium sp. NRC-1. DNA–protein interactions revealed that two heat-inducible GTFs, TFB2 from H. volcanii and TFBb from Halobacterium sp. NRC-1, could specifically bind to Phsp5 likely in a temperature-dependent manner. Taken together, the heat-responsiveness of Phsp5 was mainly ascribed to the core promoter elements that were efficiently recognized by specific heat-induced GTFs at elevated temperature, thus providing a new paradigm for GTF-directed gene regulation in the domain of Archaea. PMID:18390887

  19. Dependence on nuclear factor of activated T-cells (NFAT) levels discriminates conventional T cells from Foxp3+ regulatory T cells.

    PubMed

    Vaeth, Martin; Schliesser, Ulrike; Müller, Gerd; Reissig, Sonja; Satoh, Kazuki; Tuettenberg, Andrea; Jonuleit, Helmut; Waisman, Ari; Müller, Martin R; Serfling, Edgar; Sawitzki, Birgit S; Berberich-Siebelt, Friederike

    2012-10-01

    Several lines of evidence suggest nuclear factor of activated T-cells (NFAT) to control regulatory T cells: thymus-derived naturally occurring regulatory T cells (nTreg) depend on calcium signals, the Foxp3 gene harbors several NFAT binding sites, and the Foxp3 (Fork head box P3) protein interacts with NFAT. Therefore, we investigated the impact of NFAT on Foxp3 expression. Indeed, the generation of peripherally induced Treg (iTreg) by TGF-β was highly dependent on NFAT expression because the ability of CD4(+) T cells to differentiate into iTreg diminished markedly with the number of NFAT family members missing. It can be concluded that the expression of Foxp3 in TGF-β-induced iTreg depends on the threshold value of NFAT rather than on an individual member present. This is specific for iTreg development, because frequency of nTreg remained unaltered in mice lacking NFAT1, NFAT2, or NFAT4 alone or in combination. Different from expectation, however, the function of both nTreg and iTreg was independent on robust NFAT levels, reflected by less nuclear NFAT in nTreg and iTreg. Accordingly, absence of one or two NFAT members did not alter suppressor activity in vitro or during colitis and transplantation in vivo. This scenario emphasizes an inhibition of high NFAT activity as treatment for autoimmune diseases and in transplantation, selectively targeting the proinflammatory conventional T cells, while keeping Treg functional.

  20. Expression of NAC1 up-stream regulatory region and its relationship to the lateral root initiation induced by gibberellins and auxins.

    PubMed

    Wang, Youhua; Duan, Liusheng; Lu, Mengzhu; Li, Zhaohu; Wang, Minjie; Zhai, Zhixi

    2006-10-01

    A 1050 bp up-stream regulatory fragment of the transcription factor gene NAC1 in Arabidopsis thaliana was isolated using polymerase chain reaction (PCR) based techniques. The fragment was used to substitute the 35S promoter of the pBI121 plasmid to construct a beta-glucuronidase gene (GUS) expression system. The construct was introduced into tobacco (Nicotiana tabaccum) plants by the Agrobacterium-mediated transferring method. GUS expression pattern was studied by using the transgenic lines. The results showed that the GUS driven by the NAC1 up-stream regulatory region was specifically expressed in the root meristem region, basal areas of the lateral root primordium and the lateral roots. The GUS expression was induced by 3-indolebutyric acid (IBA) and gibberellins (GA3 and GA4+7). The results indicated that the up-stream regulatory fragment of NAC1 responded to plant hormones. The fragment might be involved in both auxins and gibberellins signaling in promoting the development of lateral roots.

  1. A Regulatory Circuit Composed of a Transcription Factor, IscR, and a Regulatory RNA, RyhB, Controls Fe-S Cluster Delivery

    PubMed Central

    Chareyre, Sylvia; Barras, Frédéric

    2016-01-01

    ABSTRACT Fe-S clusters are cofactors conserved through all domains of life. Once assembled by dedicated ISC and/or SUF scaffolds, Fe-S clusters are conveyed to their apo-targets via A-type carrier proteins (ATCs). Escherichia coli possesses four such ATCs. ErpA is the only ATC essential under aerobiosis. Recent studies reported a possible regulation of the erpA mRNA by the small RNA (sRNA) RyhB, which controls the expression of many genes under iron starvation. Surprisingly, erpA has not been identified in recent transcriptomic analysis of the iron starvation response, thus bringing into question the actual physiological significance of the putative regulation of erpA by RyhB. Using an sRNA library, we show that among 26 sRNAs, only RyhB represses the expression of an erpA-lacZ translational fusion. We further demonstrate that this repression occurs during iron starvation. Using mutational analysis, we show that RyhB base pairs to the erpA mRNA, inducing its disappearance. In addition, IscR, the master regulator of Fe-S homeostasis, represses expression of erpA at the transcriptional level when iron is abundant, but depleting iron from the medium alleviates this repression. The conjunction of transcriptional derepression by IscR and posttranscriptional repression by RyhB under Fe-limiting conditions is best described as an incoherent regulatory circuit. This double regulation allows full expression of erpA at iron concentrations for which Fe-S biogenesis switches from the ISC to the SUF system. We further provide evidence that this regulatory circuit coordinates ATC usage to iron availability. PMID:27651365

  2. Functional Improvement of Regulatory T Cells from Rheumatoid Arthritis Subjects Induced by Capsular Polysaccharide Glucuronoxylomannogalactan

    PubMed Central

    Alunno, Alessia; Bartoloni Bocci, Elena; Perito, Stefano; Chow, Siu-Kei; Cenci, Elio; Casadevall, Arturo; Gerli, Roberto; Vecchiarelli, Anna

    2014-01-01

    Objective Regulatory T cells (Treg) play a critical role in the prevention of autoimmunity, and the suppressive activity of these cells is impaired in rheumatoid arthritis (RA). The aim of the present study was to investigate function and properties of Treg of RA patients in response to purified polysaccharide glucuronoxylomannogalactan (GXMGal). Methods Flow cytometry and western blot analysis were used to investigate the frequency, function and properties of Treg cells. Results GXMGal was able to: i) induce strong increase of FOXP3 on CD4+ T cells without affecting the number of CD4+CD25+FOXP3+ Treg cells with parallel increase in the percentage of non-conventional CD4+CD25−FOXP3+ Treg cells; ii) increase intracellular levels of TGF-β1 in CD4+CD25−FOXP3+ Treg cells and of IL-10 in both CD4+CD25+FOXP3+ and CD4+CD25−FOXP3+ Treg cells; iii) enhance the suppressive activity of CD4+CD25+FOXP3+ and CD4+CD25−FOXP3+ Treg cells in terms of inhibition of effector T cell activity and increased secretion of IL-10; iv) decrease Th1 response as demonstrated by inhibition of T-bet activation and down-regulation of IFN-γ and IL-12p70 production; v) decrease Th17 differentiation by down-regulating pSTAT3 activation and IL-17A, IL-23, IL-21, IL-22 and IL-6 production. Conclusion These data show that GXMGal improves Treg functions and increases the number and function of CD4+CD25−FOXP3+ Treg cells of RA patients. It is suggested that GXMGal may be potentially useful for restoring impaired Treg functions in autoimmune disorders and for developing Treg cell-based strategies for the treatment of these diseases. PMID:25338013

  3. TGF-β-Induced Regulatory T Cells Directly Suppress B Cell Responses through a Noncytotoxic Mechanism.

    PubMed

    Xu, Anping; Liu, Ya; Chen, Weiqian; Wang, Julie; Xue, Youqiu; Huang, Feng; Rong, Liming; Lin, Jin; Liu, Dahai; Yan, Mei; Li, Quan-Zhen; Li, Bin; Song, Jianxun; Olsen, Nancy; Zheng, Song Guo

    2016-05-01

    Foxp3(+) regulatory T cells (Treg) playing a crucial role in the maintenance of immune tolerance and prevention of autoimmune diseases consist of thymus-derived naturally occurring CD4(+)Foxp3(+) Treg cells (nTreg) and those that can be induced ex vivo with TGF-β (iTreg). Although both Treg subsets share similar phenotypes and functional characteristics, they also have potential biologic differences on their biology. The role of iTreg in regulating B cells remains unclear so far. The suppression assays of Treg subsets on activation, proliferation, and Abs production of B cells were measured using a Treg and B cell coculture system in vitro. Transwell and Ab blockade experiments were performed to assess the roles of cell contact and soluble cytokines. Treg were adoptively transferred to lupus mice to assess in vivo effects on B cells. Like nTreg, iTreg subset also directly suppressed activation and proliferation of B cells. nTreg subset suppressed B cell responses through cytotoxic manner related to expression of granzyme A, granzyme B, and perforin, whereas the role of iTreg subset on B cells did not involve in cytotoxic action but depending on TGF-β signaling. Furthermore, iTreg subset can significantly suppress Ab produced by lupus B cells in vitro. Comparison experiments using autoantibodies microarrays demonstrated that adoptive transfer of iTreg had a superior effect than nTreg subset on suppressing lupus B cell responses in vivo. Our data implicate a role and advantage of iTreg subset in treating B cell-mediated autoimmune diseases, boosting the translational potential of these findings. PMID:27001954

  4. Physiological and receptor-selective retinoids modulate interferon gamma signaling by increasing the expression, nuclear localization, and functional activity of interferon regulatory factor-1.

    PubMed

    Luo, Xin M; Ross, A Catharine

    2005-10-28

    Synergistic actions between all-trans-retinoic acid (atRA) and interferon gamma (IFNgamma) on modulation of cellular functions have been reported both in vitro and in vivo. However, the mechanism of atRA-mediated regulation of IFNgamma signaling is poorly understood. In this study, we have used the human lung epithelial cell line A549 to examine the effect of atRA on IFNgamma-induced expression of IFN regulatory factor-1 (IRF-1), an important transcription factor involved in cell growth and apoptosis, differentiation, and antiviral and antibacterial immune responses. At least 4 h of pretreatment with atRA followed by suboptimal concentrations of IFNgamma induced a faster, higher, and more stable expression of IRF-1 than IFNgamma alone. Actinomycin D completely blocked the induction of IRF-1 by the combination, suggesting regulation at the transcriptional level. Further, we found that activation of signal transducer and activator of transcription-1 was induced more dramatically by atRA and IFNgamma than by IFNgamma alone. Expression of IFNgamma receptor-1 on the cell surface was also increased upon atRA pretreatment. Experiments using receptor-selective retinoids revealed that ligands for retinoic acid receptor-alpha (RARalpha), including atRA, 9-cis-retinoic acid, and Am580, sequentially increased the levels of IFNgamma receptor-1, activated signal transducer and activator of transcription-1, and IRF-1 and that an RARalpha antagonist was able to inhibit the effects of atRA and Am580. In addition, atRA pretreatment affected the transcriptional functions of IFNgamma-induced IRF-1, increasing its nuclear localization and DNA binding activity as well as the transcript levels of IRF-1 target genes. These results suggest that atRA, an RARalpha ligand, regulates IFNgamma-induced IRF-1 by affecting multiple components of the IFNgamma signaling pathway, from the plasma membrane to the nuclear transcription factors.

  5. IL-10 plays a central regulatory role in the cytokines induced by hepatitis C virus core protein and polyinosinic acid:polycytodylic acid.

    PubMed

    Pang, Xiaoli; Wang, Zhaoxia; Zhai, Naicui; Zhang, Qianqian; Song, Hongxiao; Zhang, Yujiao; Li, Tianyang; Li, Haijun; Su, Lishan; Niu, Junqi; Tu, Zhengkun

    2016-09-01

    Hepatitis C virus (HCV) can cause persistent infection and chronic liver disease, and viral factors are involved in HCV persistence. HCV core protein, a highly conserved viral protein, not only elicits an immunoresponse, but it also regulates it. In addition, HCV core protein interacts with toll-like receptors (TLRs) on monocytes, inducing them to produce cytokines. Polyinosinic acid:polycytodylic acid (polyI:C) is a synthetic analogue of double-stranded RNA that binds to TLR3 and can induce secretion of type I IFN from monocytes. Cytokine response against HCV is likely to affect the natural course of infection as well as HCV persistence. However, possible effects of cytokines induced by HCV core protein and polyI:C remain to be investigated. In this study, we isolated CD14(+) monocytes from healthy donors, cultured them in the presence of HCV core protein and/or polyI:C, and characterized the induced cytokines, phenotypes and mechanisms. We demonstrated that HCV core protein- and polyI:C-stimulated CD14(+) monocytes secreted tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-10, and type I interferon (IFN). Importantly, TNF-α and IL-1β regulated the secretion of IL-10, which then influenced the expression of signal transducer and activator of transcription 1 (STAT1) and interferon regulatory factor 1 (IRF1) and subsequently the production of type I IFN. Interestingly, type I IFN also regulated the production of IL-10, which in turn inhibited the nuclear factor (NF)-κB subunit, reducing TNF-α and IL-1β levels. Therefore, IL-10 appears to play a central role in regulating the production of cytokines induced by HCV core protein and polyI:C. PMID:27337528

  6. SNPs in putative regulatory regions identified by human mouse comparative sequencing and transcription factor binding site data

    SciTech Connect

    Banerjee, Poulabi; Bahlo, Melanie; Schwartz, Jody R.; Loots, Gabriela G.; Houston, Kathryn A.; Dubchak, Inna; Speed, Terence P.; Rubin, Edward M.

    2002-01-01

    Genome wide disease association analysis using SNPs is being explored as a method for dissecting complex genetic traits and a vast number of SNPs have been generated for this purpose. As there are cost and throughput limitations of genotyping large numbers of SNPs and statistical issues regarding the large number of dependent tests on the same data set, to make association analysis practical it has been proposed that SNPs should be prioritized based on likely functional importance. The most easily identifiable functional SNPs are coding SNPs (cSNPs) and accordingly cSNPs have been screened in a number of studies. SNPs in gene regulatory sequences embedded in noncoding DNA are another class of SNPs suggested for prioritization due to their predicted quantitative impact on gene expression. The main challenge in evaluating these SNPs, in contrast to cSNPs is a lack of robust algorithms and databases for recognizing regulatory sequences in noncoding DNA. Approaches that have been previously used to delineate noncoding sequences with gene regulatory activity include cross-species sequence comparisons and the search for sequences recognized by transcription factors. We combined these two methods to sift through mouse human genomic sequences to identify putative gene regulatory elements and subsequently localized SNPs within these sequences in a 1 Megabase (Mb) region of human chromosome 5q31, orthologous to mouse chromosome 11 containing the Interleukin cluster.

  7. Genome-wide survey of tissue-specific microRNA and transcription factor regulatory networks in 12 tissues.

    PubMed

    Guo, Zhiyun; Maki, Miranda; Ding, Ruofan; Yang, Yalan; Zhang, Bao; Xiong, Lili

    2014-06-03

    Tissue-specific miRNAs (TS miRNA) specifically expressed in particular tissues play an important role in tissue identity, differentiation and function. However, transcription factor (TF) and TS miRNA regulatory networks across multiple tissues have not been systematically studied. Here, we manually extracted 116 TS miRNAs and systematically investigated the regulatory network of TF-TS miRNA in 12 human tissues. We identified 2,347 TF-TS miRNA regulatory relations and revealed that most TF binding sites tend to enrich close to the transcription start site of TS miRNAs. Furthermore, we found TS miRNAs were regulated widely by non-tissue specific TFs and the tissue-specific expression level of TF have a close relationship with TF-genes regulation. Finally, we describe TSmiR (http://bioeng.swjtu.edu.cn/TSmiR), a novel and web-searchable database that houses interaction maps of TF-TS miRNA in 12 tissues. Taken together, these observations provide a new suggestion to better understand the regulatory network and mechanisms of TF-TS miRNAs underlying different tissues.

  8. Genetic Variation of Goat Interferon Regulatory Factor 3 Gene and Its Implication in Goat Evolution

    PubMed Central

    Shu, Liping; Zhang, Yesheng; Wang, Yangzi; Sanni, Timothy M.; Imumorin, Ikhide G.; Peters, Sunday O.; Zhang, Jiajin; Dong, Yang; Wang, Wen

    2016-01-01

    The immune systems are fundamentally vital for evolution and survival of species; as such, selection patterns in innate immune loci are of special interest in molecular evolutionary research. The interferon regulatory factor (IRF) gene family control many different aspects of the innate and adaptive immune responses in vertebrates. Among these, IRF3 is known to take active part in very many biological processes. We assembled and evaluated 1356 base pairs of the IRF3 gene coding region in domesticated goats from Africa (Nigeria, Ethiopia and South Africa) and Asia (Iran and China) and the wild goat (Capra aegagrus). Five segregating sites with θ value of 0.0009 for this gene demonstrated a low diversity across the goats’ populations. Fu and Li tests were significantly positive but Tajima’s D test was significantly negative, suggesting its deviation from neutrality. Neighbor joining tree of IRF3 gene in domesticated goats, wild goat and sheep showed that all domesticated goats have a closer relationship than with the wild goat and sheep. Maximum likelihood tree of the gene showed that different domesticated goats share a common ancestor and suggest single origin. Four unique haplotypes were observed across all the sequences, of which, one was particularly common to African goats (MOCH-K14-0425, Poitou and WAD). In assessing the evolution mode of the gene, we found that the codon model dN/dS ratio for all goats was greater than one. Phylogenetic Analysis by Maximum Likelihood (PAML) gave a ω0 (dN/dS) value of 0.067 with LnL value of -6900.3 for the first Model (M1) while ω2 = 1.667 in model M2 with LnL value of -6900.3 with positive selection inferred in 3 codon sites. Mechanistic empirical combination (MEC) model for evaluating adaptive selection pressure on particular codons also confirmed adaptive selection pressure in three codons (207, 358 and 408) in IRF3 gene. Positive diversifying selection inferred with recent evolutionary changes in domesticated goat

  9. Interferon regulatory factor 1 and histone H4 acetylation in systemic lupus erythematosus

    PubMed Central

    Leung, Yiu Tak; Shi, Lihua; Maurer, Kelly; Song, Li; Zhang, Zhe; Petri, Michelle; Sullivan, Kathleen E

    2015-01-01

    Histone acetylation modulates gene expression and has been described as increased in systemic lupus erythematosus (SLE). We investigated interferon regulatory factor 1 (IRF1) interactions that influence H4 acetylation (H4ac) in SLE. Intracellular flow cytometry for H4 acetylated lysine (K) 5, K8, K12, and K16 was performed. Histone acetylation was defined in monocytes and T cells from controls and SLE patients. RNA-Seq studies were performed on monocytes to look for an imbalance in histone acetyltransferases and histone deacetylase enzyme expression. Expression levels were validated using real-time quantitative RT-PCR. IRF1 induction of H4ac was evaluated using D54MG cells overexpressing IRF1. IRF1 protein interactions were studied using co-immunoprecipitation assays. IRF1-dependent recruitment of histone acetyltransferases to target genes was examined by ChIP assays using p300 antibody. Flow cytometry data showed significantly increased H4K5, H4K8, H4K12, and H4K16 acetylation in SLE monocytes. HDAC3 and HDAC11 gene expression were decreased in SLE monocytes. PCAF showed significantly higher gene expression in SLE than controls. IRF1-overexpressing D54MG cells were associated with significantly increased H4K5, H4K8, and H4K12 acetylation compared to vector-control D54MG cells both globally and at specific target genes. Co-immunoprecipitation studies using D54MG cells revealed IRF1 protein-protein interactions with PCAF, P300, CBP, GCN5, ATF2, and HDAC3. ChIP experiments demonstrated increased p300 recruitment to known IRF1 targets in D54MG cells overexpressing IRF1. In contrast, p300 binding to IRF1 targets decreased in D54MG cells with IRF1 knockdown. SLE appears to be associated with an imbalance in histone acetyltransferases and histone deacetylase enzymes favoring pathologic H4 acetylation. Furthermore, IRF1 directly interacts with chromatin modifying enzymes, supporting a model where recruitment to specific target genes is mediated in part by IRF1. PMID

  10. Genetic Variation of Goat Interferon Regulatory Factor 3 Gene and Its Implication in Goat Evolution.

    PubMed

    Okpeku, Moses; Esmailizadeh, Ali; Adeola, Adeniyi C; Shu, Liping; Zhang, Yesheng; Wang, Yangzi; Sanni, Timothy M; Imumorin, Ikhide G; Peters, Sunday O; Zhang, Jiajin; Dong, Yang; Wang, Wen

    2016-01-01

    The immune systems are fundamentally vital for evolution and survival of species; as such, selection patterns in innate immune loci are of special interest in molecular evolutionary research. The interferon regulatory factor (IRF) gene family control many different aspects of the innate and adaptive immune responses in vertebrates. Among these, IRF3 is known to take active part in very many biological processes. We assembled and evaluated 1356 base pairs of the IRF3 gene coding region in domesticated goats from Africa (Nigeria, Ethiopia and South Africa) and Asia (Iran and China) and the wild goat (Capra aegagrus). Five segregating sites with θ value of 0.0009 for this gene demonstrated a low diversity across the goats' populations. Fu and Li tests were significantly positive but Tajima's D test was significantly negative, suggesting its deviation from neutrality. Neighbor joining tree of IRF3 gene in domesticated goats, wild goat and sheep showed that all domesticated goats have a closer relationship than with the wild goat and sheep. Maximum likelihood tree of the gene showed that different domesticated goats share a common ancestor and suggest single origin. Four unique haplotypes were observed across all the sequences, of which, one was particularly common to African goats (MOCH-K14-0425, Poitou and WAD). In assessing the evolution mode of the gene, we found that the codon model dN/dS ratio for all goats was greater than one. Phylogenetic Analysis by Maximum Likelihood (PAML) gave a ω0 (dN/dS) value of 0.067 with LnL value of -6900.3 for the first Model (M1) while ω2 = 1.667 in model M2 with LnL value of -6900.3 with positive selection inferred in 3 codon sites. Mechanistic empirical combination (MEC) model for evaluating adaptive selection pressure on particular codons also confirmed adaptive selection pressure in three codons (207, 358 and 408) in IRF3 gene. Positive diversifying selection inferred with recent evolutionary changes in domesticated goat IRF3

  11. [Risk factors and subjective symptoms of drug-induced leucopenia].

    PubMed

    Hayashi, Kyoko; Ohtsu, Fumiko; Yano, Reiko; Sakakibara, Jinsaku; Goto, Nobuyuki

    2011-01-01

    The present study investigated risk factors and subjective symptoms associated with drug-induced leucopenia. We selected 248 patients with drug-induced leucopenia from the Case Reports of Adverse Drug Reactions and Poisoning Information System (CARPIS) database of over 47000 case reports of adverse drug reactions and assigned them to a case group. We also randomly selected 743 cases of adverse drug reactions not associated with leucopenia as a control group. A comparison of patient characteristic data between the two groups using logistic-regression analysis revealed that female sex, autoimmune disease and renal damage were background risk factors for drug-induced leucopenia. In addition, thiamazole, ritodrine, propylthiouracil, ticlopidine, allopurinol, minocycline and captopril administration significantly increased the risk of drug-induced leucopenia. A significant association was also found for fever, chills and pharyngeal abnormalities. Based on these findings, we developed two estimated regression equations to help prevent drug-induced leucopenia in the community pharmacy setting. PMID:21212623

  12. Transient cefuroxime/metronidazole treatment induced factor V antibodies.

    PubMed

    Van den Berg, Sjoerd Adrianus Antonius; Verwer, Patricia E; Idema, René N; Van Guldener, Coen

    2014-01-01

    A 29-year-old patient presented with an appendicular infiltrate, initially treated with intravenous antibiotics, but later requiring percutaneous drainage. Both prothrombin time (PT) and activated partial thromboplastin time (aPTT) were prolonged on 3 days of antibiotic treatment and unresponsive to vitamin K or prothrombin complex concentrate. Laboratory investigation ultimately showed reduced factor V activity and factor V antibodies. In contrast to previously described cases of factor V antibodies, PT and aPTT were only mildly prolonged and residual factor V activity was still >20%. Draining of the abscess did not induce significant bleeding. Afterwards, no haemostatic medication was required. The patient was discharged from the hospital without complications. One week after cessation of the antibiotic treatment, PT and aPTT were within normal range again, with a factor V activity level of 36%. In conclusion, we present a patient with transient factor V antibodies, induced by antibiotics, without clinical bleeding tendency.

  13. Genetic factors and manganese-induced neurotoxicity

    PubMed Central

    Chen, Pan; Parmalee, Nancy; Aschner, Michael

    2014-01-01

    Manganese (Mn), is a trace metal required for normal physiological processes in humans. Mn levels are tightly regulated, as high levels of Mn result in accumulation in the brain and cause a neurological disease known as manganism. Manganism shares many similarities with Parkinson’s disease (PD), both at the physiological level and the cellular level. Exposure to high Mn-containing environments increases the risk of developing manganism. Mn is absorbed primarily through the intestine and then released in the blood. Excessive Mn is secreted in the bile and excreted in feces. Mn enters and exits cells through a number of non-specific importers localized on the cell membrane. Mutations in one of the Mn exporters, SLC30A10 (solute carrier family 30, member 10), result in Mn induced toxicity with liver impairments and neurological dysfunction. Four PD genes have been identified in connection to regulation of Mn toxicity, shedding new light on potential links between manganism and PD. PMID:25136353

  14. Abdominal {gamma}-Radiation Induces an Accumulation of Function-Impaired Regulatory T Cells in the Small Intestine

    SciTech Connect

    Billiard, Fabienne; Buard, Valerie; Benderitter, Marc; Linard, Christine

    2011-07-01

    Purpose: To assess the frequency and the functional characteristics of one major component of immune tolerance, the CD4{sup +}FoxP3{sup +} regulatory T cells (Tregs) in a mouse model of abdominal irradiation. Methods and Materials: Mice were exposed to a single abdominal dose of {gamma}-radiation (10 Gy). We evaluated small intestine Treg infiltration by Foxp3 immunostaining and the functional suppressive activity of Tregs isolated from mesenteric lymph nodes. Results: Foxp3 immunostaining showed that radiation induced a long-term infiltration of the intestine by Tregs (levels 5.5 times greater than in controls). Co-culture of Tregs from mesenteric lymph nodes with CD4{sup +} effector cells showed that the Tregs had lost their suppressive function. This loss was associated with a significant decrease in the levels of Foxp3, TGF-{beta}, and CTLA-4 mRNA, all required for optimal Treg function. At Day 90 after irradiation, Tregs regained their suppressive activity as forkhead box P3 (Foxp3), transforming growth factor beta (TGF-{beta}), and cytotoxic T-lymphocyte antigen 4 (CTLA-4) expression returned to normal. Analysis of the secretory function of mesenteric lymph node Tregs, activated in vitro with anti-CD3/anti-CD28 Abs, showed that this dysfunction was independent of a defect in interleukin-10 secretion. Conclusion: Radiation caused a long-term accumulation of function-impaired Foxp3{sup +}CD4{sup +} Tregs in the intestine. Our study provides new insights into how radiation affects the immune tolerance in peripheral tissues.

  15. Immunosuppressive Effects of Streptozotocin-Induced Diabetes Result in Absolute Lymphopenia and a Relative Increase of T Regulatory Cells

    PubMed Central

    Muller, Yannick D.; Golshayan, Déla; Ehirchiou, Driss; Wyss, Jean Christophe; Giovannoni, Laurianne; Meier, Raphael; Serre-Beinier, Véronique; Puga Yung, Gisella; Morel, Philippe; Bühler, Leo H.; Seebach, Jörg D.

    2011-01-01

    OBJECTIVE Streptozotocin (STZ) is the most widely used diabetogenic agent in animal models of islet transplantation. However, the immunomodifying effects of STZ and the ensuing hyperglycemia on lymphocyte subsets, particularly on T regulatory cells (Tregs), remain poorly understood. RESEARCH DESIGN AND METHODS This study evaluated how STZ-induced diabetes affects adaptive immunity and the consequences thereof on allograft rejection in murine models of islet and skin transplantation. The respective toxicity of STZ and hyperglycemia on lymphocyte subsets was tested in vitro. The effect of hyperglycemia was assessed independently of STZ in vivo by the removal of transplanted syngeneic islets, using an insulin pump, and with rat insulin promoter diphtheria toxin receptor transgenic mice. RESULTS Early lymphopenia in both blood and spleen was demonstrated after STZ administration. Direct toxicity of STZ on lymphocytes, particularly on CD8+ cells and B cells, was shown in vitro. Hyperglycemia also correlated with blood and spleen lymphopenia in vivo but was not lymphotoxic in vitro. Independently of hyperglycemia, STZ led to a relative increase of Tregs in vivo, with the latter retaining their suppressive capacity in vitro. The higher frequency of Tregs was associated with Treg proliferation in the blood, but not in the spleen, and higher blood levels of transforming growth factor-β. Finally, STZ administration delayed islet and skin allograft rejection compared with naive mice. CONCLUSIONS These data highlight the direct and indirect immunosuppressive effects of STZ and acute hyperglycemia, respectively. Thus, these results have important implications for the future development of tolerance-based protocols and their translation from the laboratory to the clinic. PMID:21752956

  16. Phosphorylation of CRTC3 by the salt-inducible kinases controls the interconversion of classically activated and regulatory macrophages.

    PubMed

    Clark, Kristopher; MacKenzie, Kirsty F; Petkevicius, Kasparas; Kristariyanto, Yosua; Zhang, Jiazhen; Choi, Hwan Geun; Peggie, Mark; Plater, Lorna; Pedrioli, Patrick G A; McIver, Ed; Gray, Nathanael S; Arthur, J Simon C; Cohen, Philip

    2012-10-16

    Macrophages acquire strikingly different properties that enable them to play key roles during the initiation, propagation, and resolution of inflammation. Classically activated (M1) macrophages produce proinflammatory mediators to combat invading pathogens and respond to tissue damage in the host, whereas regulatory macrophages (M2b) produce high levels of anti-inflammatory molecules, such as IL-10, and low levels of proinflammatory cytokines, like IL-12, and are important for the resolution of inflammatory responses. A central problem in this area is to understand how the formation of regulatory macrophages can be promoted at sites of inflammation to prevent and/or alleviate chronic inflammatory and autoimmune diseases. Here, we demonstrate that the salt-inducible kinases (SIKs) restrict the formation of regulatory macrophages and that their inhibition induces striking increases in many of the characteristic markers of regulatory macrophages, greatly stimulating the production of IL-10 and other anti-inflammatory molecules. We show that SIK inhibitors elevate IL-10 production by inducing the dephosphorylation of cAMP response element-binding protein (CREB)-regulated transcriptional coactivator (CRTC) 3, its dissociation from 14-3-3 proteins and its translocation to the nucleus where it enhances a gene transcription program controlled by CREB. Importantly, the effects of SIK inhibitors on IL-10 production are lost in macrophages that express a drug-resistant mutant of SIK2. These findings identify SIKs as a key molecular switch whose inhibition reprograms macrophages to an anti-inflammatory phenotype. The remarkable effects of SIK inhibitors on macrophage function suggest that drugs that target these protein kinases may have therapeutic potential for the treatment of inflammatory and autoimmune diseases.

  17. T regulatory cells and B cells cooperate to form a regulatory loop that maintains gut homeostasis and suppresses dextran sulfate sodium-induced colitis.

    PubMed

    Wang, L; Ray, A; Jiang, X; Wang, J-y; Basu, S; Liu, X; Qian, T; He, R; Dittel, B N; Chu, Y

    2015-11-01

    Regulatory T cells (Tregs) and B cells present in gut-associated lymphoid tissues (GALT) are both implicated in the resolution of colitis. However, how the functions of these cells are coordinated remains elusive. We used the dextran sulfate sodium (DSS)-induced colitis model combined with gene-modified mice to monitor the progression of colitis, and simultaneously examine the number of Tregs and B cells, and the production of IgA antibodies. We found that DSS-treated mice exhibited more severe colitis in the absence of B cells, and that the adoptive transfer of B cells attenuated the disease. Moreover, the transfer of IL-10(-/-) B cells also attenuated colitis, suggesting that B cells inhibited colitis through an interleukin-10 (IL-10)-independent pathway. Furthermore, antibody depletion of Tregs resulted in exacerbated colitis. Intriguingly, the number of GALT Tregs in B cell-deficient mice was significantly decreased during colitis and the adoptive transfer of B cells into these mice restored the Treg numbers, indicating that B cells contribute to Treg homeostasis. We also found that B cells induced the proliferation of Tregs that in turn promoted B-cell differentiation into IgA-producing plasma cells. These results demonstrate that B cells and Tregs interact and cooperate to prevent excessive immune responses that can lead to colitis.

  18. T regulatory cells and B cells cooperate to form a regulatory loop that maintains gut homeostasis and suppresses dextran sulfate sodium-induced colitis

    PubMed Central

    Wang, L; Ray, A; Jiang, X; Wang, J-y; Basu, S; Liu, X; Qian, T; He, R; Dittel, B N; Chu, Y

    2015-01-01

    Regulatory T cells (Tregs) and B cells present in gut-associated lymphoid tissues (GALT) are both implicated in the resolution of colitis. However, how the functions of these cells are coordinated remains elusive. We used the dextran sulfate sodium (DSS)-induced colitis model combined with gene-modified mice to monitor the progression of colitis, and simultaneously examine the number of Tregs and B cells, and the production of IgA antibodies. We found that DSS-treated mice exhibited more severe colitis in the absence of B cells, and that the adoptive transfer of B cells attenuated the disease. Moreover, the transfer of IL-10−/− B cells also attenuated colitis, suggesting that B cells inhibited colitis through an interleukin-10 (IL-10)-independent pathway. Furthermore, antibody depletion of Tregs resulted in exacerbated colitis. Intriguingly, the number of GALT Tregs in B cell-deficient mice was significantly decreased during colitis and the adoptive transfer of B cells into these mice restored the Treg numbers, indicating that B cells contribute to Treg homeostasis. We also found that B cells induced the proliferation of Tregs that in turn promoted B-cell differentiation into IgA-producing plasma cells. These results demonstrate that B cells and Tregs interact and cooperate to prevent excessive immune responses that can lead to colitis. PMID:25807185

  19. Hepatic carcinoma-associated fibroblasts induce IDO-producing regulatory dendritic cells through IL-6-mediated STAT3 activation.

    PubMed

    Cheng, J-T; Deng, Y-N; Yi, H-M; Wang, G-Y; Fu, B-S; Chen, W-J; Liu, W; Tai, Y; Peng, Y-W; Zhang, Q

    2016-02-22

    Although carcinoma-associated fibroblasts (CAFs) in tumor microenvironments have a critical role in immune cell modulation, their effects on the generation of regulatory dendritic cells (DCs) are still unclear. In this study, we initially show that CAFs derived from hepatocellular carcinoma (HCC) tumors facilitate the generation of regulatory DCs, which are characterized by low expression of costimulatory molecules, high suppressive cytokines production and enhanced regulation of immune responses, including T-cell proliferation impairment and promotion of regulatory T-cell (Treg) expansion via indoleamine 2,3-dioxygenase (IDO) upregulation. Our findings also indicate that STAT3 activation in DCs, as mediated by CAF-derived interleukin (IL)-6, is essential to IDO production. Moreover, IDO inhibitor, STAT3 and IL-6 blocking antibodies can reverse this hepatic CAF-DC regulatory function. Therefore, our results provide new insights into the mechanisms by which CAFs induce tumor immune escape as well as a novel cancer immunotherapeutic approach (for example, targeting CAFs, IDO or IL-6).

  20. The Regulatory Role of Nuclear Factor Kappa B in the Heart of Hereditary Hypertriglyceridemic Rat

    PubMed Central

    Vranková, Stanislava; Barta, Andrej; Klimentová, Jana; Dovinová, Ima; Líšková, Silvia; Dobešová, Zdenka; Pecháňová, Oľga; Kuneš, Jaroslav; Zicha, Josef

    2016-01-01

    Activation of nuclear factor-κB (NF-κB) by increased production of reactive oxygen species (ROS) might induce transcription and expression of different antioxidant enzymes and also of nitric oxide synthase (NOS) isoforms. Thus, we aimed at studying the effect of NF-κB inhibition, caused by JSH-23 (4-methyl-N1-(3-phenyl-propyl)-benzene-1,2-diamine) injection, on ROS and NO generation in hereditary hypertriglyceridemic (HTG) rats. 12-week-old, male Wistar and HTG rats were treated with JSH-23 (bolus, 10 μmol, i.v.). After one week, blood pressure (BP), superoxide dismutase (SOD) activity, SOD1, endothelial NOS (eNOS), and NF-κB (p65) protein expressions were higher in the heart of HTG rats compared to control rats. On the other hand, NOS activity was decreased. In HTG rats, JSH-23 treatment increased BP and heart conjugated dienes (CD) concentration (measured as the marker of tissue oxidative damage). Concomitantly, SOD activity together with SOD1 expression was decreased, while NOS activity and eNOS protein expression were increased significantly. In conclusion, NF-κB inhibition in HTG rats led to decreased ROS degradation by SOD followed by increased oxidative damage in the heart and BP elevation. In these conditions, increased NO generation may represent rather a counterregulatory mechanism activated by ROS. Nevertheless, this mechanism was not sufficient enough to compensate BP increase in HTG rats. PMID:27148433

  1. The Regulatory T Cell Lineage Factor Foxp3 Regulates Gene Expression through Several Distinct Mechanisms Mostly Independent of Direct DNA Binding

    PubMed Central

    Andersen, Kristian G.; Hebenstreit, Daniel; Teichmann, Sarah A.; Betz, Alexander G.

    2015-01-01

    The lineage factor Foxp3 is essential for the development and maintenance of regulatory T cells, but little is known about the mechanisms involved. Here, we demonstrate that an N-terminal proline-rich interaction region is crucial for Foxp3’s function. Subdomains within this key region link Foxp3 to several independent mechanisms of transcriptional regulation. Our study suggests that Foxp3, even in the absence of its DNA-binding forkhead domain, acts as a bridge between DNA-binding interaction partners and proteins with effector function permitting it to regulate a large number of genes. We show that, in one such mechanism, Foxp3 recruits class I histone deacetylases to the promoters of target genes, counteracting activation-induced histone acetylation and thereby suppressing their expression. PMID:26107960

  2. Risk factors for ganciclovir-induced thrombocytopenia and leukopenia.

    PubMed

    Matsumoto, Kazuaki; Shigemi, Akari; Ikawa, Kazuro; Kanazawa, Naoko; Fujisaki, Yuko; Morikawa, Norifumi; Takeda, Yasuo

    2015-01-01

    Ganciclovir is a nucleoside guanosine analogue that exhibits therapeutic activity against human cytomegalovirus infection, and is primarily excreted via glomerular filtration and active tubular secretion. The adverse effects induced by ganciclovir therapy are generally of a hematological nature and include thrombocytopenia and leukopenia. Low marrow cellularity and elevated serum creatinine have been identified as risk factors for ganciclovir-induced neutropenia. However, the risk factors for thrombocytopenia have yet to be determined. Therefore, this study investigated patients administered ganciclovir to determine the risk factors for thrombocytopenia and leukopenia. Thrombocytopenia occurred in 41 of these patients (30.6%). Multivariate logistic regression analysis identified three independent risk factors for thrombocytopenia: cancer chemotherapy (odds ratio (OR)=3.1), creatinine clearance (<20 mL/min) (OR=12.8), and the ganciclovir dose (≥12 mg/kg/d) (OR=15.1). Leukopenia occurred in 36 patients (28.6%), and white blood cell count (<6000 cells/mm(3)) (OR=3.7) and the ganciclovir dose (≥12 mg/kg/d) (OR=7.8) were identified as risk factors. These results demonstrated that several factors influenced the occurrence of ganciclovir-induced thrombocytopenia and leukopenia, and suggest that special attention should be paid to patients receiving cancer chemotherapy with a low creatinine clearance (<20 mL/min) and high dose (≥12 mg/kg/d) in order to avoid ganciclovir-induced thrombocytopenia. PMID:25747982

  3. Regulatory T-cell suppressor program co-opts transcription factor IRF4 to control TH2 responses

    PubMed Central

    Zheng, Ye; Chaudhry, Ashutosh; Kas, Arnold; deRoos, Paul; Kim, Jeong M.; Chu, Tin-Tin; Corcoran, Lynn; Treuting, Piper; Klein, Ulf; Rudensky, Alexander Y.

    2010-01-01

    In the course of infection or autoimmunity, particular transcription factors orchestrate the differentiation of TH1, TH2 or TH17 effector cells, the responses of which are limited by a distinct lineage of suppressive regulatory T cells (Treg). Treg cell differentiation and function are guided by the transcription factor Foxp3, and their deficiency due to mutations in Foxp3 results in aggressive fatal autoimmune disease associated with sharply augmented TH1 and TH2 cytokine production1–3. Recent studies suggested that Foxp3 regulates the bulk of the Foxp3-dependent transcriptional program indirectly through a set of transcriptional regulators serving as direct Foxp3 targets4,5. Here we show that in mouse Treg cells, high amounts of interferon regulatory factor-4 (IRF4), a transcription factor essential for TH2 effector cell differentiation, is dependent on Foxp3 expression. We proposed that IRF4 expression endows Treg cells with the ability to suppress TH2 responses. Indeed, ablation of a conditional Irf4 allele in Treg cells resulted in selective dysregulation of TH2 responses, IL4-dependent immunoglobulin isotype production, and tissue lesions with pronounced plasma cell infiltration, in contrast to the mononuclear-cell-dominated pathology typical of mice lacking Treg cells. Our results indicate that Treg cells use components of the transcriptional machinery, promoting a particular type of effector CD4+ T cell differentiation, to efficiently restrain the corresponding type of the immune response. PMID:19182775

  4. Regulation of endogenous human gene expression by ligand-inducible TALE transcription factors.

    PubMed

    Mercer, Andrew C; Gaj, Thomas; Sirk, Shannon J; Lamb, Brian M; Barbas, Carlos F

    2014-10-17

    The construction of increasingly sophisticated synthetic biological circuits is dependent on the development of extensible tools capable of providing specific control of gene expression in eukaryotic cells. Here, we describe a new class of synthetic transcription factors that activate gene expression in response to extracellular chemical stimuli. These inducible activators consist of customizable transcription activator-like effector (TALE) proteins combined with steroid hormone receptor ligand-binding domains. We demonstrate that these ligand-responsive TALE transcription factors allow for tunable and conditional control of gene activation and can be used to regulate the expression of endogenous genes in human cells. Since TALEs can be designed to recognize any contiguous DNA sequence, the conditional gene regulatory system described herein will enable the design of advanced synthetic gene networks.

  5. Programmed Death-1 Antibody Blocks Therapeutic Effects of T-Regulatory Cells in Cockroach Antigen-Induced Allergic Asthma

    PubMed Central

    McGee, Halvor S.; Yagita, Hideo; Shao, Zhifei; Agrawal, Devendra K.

    2010-01-01

    We recently reported that the adoptive transfer of T-regulatory cells (Tregs) isolated from lung and spleen tissue of green fluorescent protein–transgenic mice reversed airway hyperresponsiveness and airway inflammation. Because Programmed Death-1 (PD-1) is a pivotal receptor regulating effector T-cell activation by Tregs, we evaluated whether PD-1 is involved in the therapeutic effect of naturally occurring Tregs (NTregs) and inducible Tregs (iTregs) in cockroach (CRA)-sensitized and challenged mice. The CD4+CD25+ NTregs and CD4+CD25− iTregs isolated from the lungs and spleens of BALB/c mice were adoptively transferred into CRA-sensitized and CRA-challenged mice with and without anti–PD-1 antibody (100 μg/mice). The CD4+CD25+ T cells in the lung were phenotyped after adoptive transfer. Concentrations of IL-4, IL-5, IL-10, IFN-γ, and IL-13 in bronchoalveolar lavage fluid (BALF) were measured using ELISA. The NTregs and iTregs from either lung or spleen tissue reversed airway hyperresponsiveness for at least 4 wk. However, the therapeutic effect was blocked by administering the anti–PD-1 antibody. The administration of Tregs-recipient mice with anti–PD-1 antibody significantly decreased cytotoxic T-lymphocyte antigen-4 expression, with low concentrations of Forkhead-winged transcriptional factor box 3 (Foxp3) mRNA transcripts in lung CD4+CD25+ T cells. These mice had substantially higher concentrations of BALF IL-4, IL-5, and IL-13, but significantly decreased levels of BALF IL-10. Adoptive therapy recipients without the anti–PD-1 antibody exhibited high levels of CTLA-4 expression and Foxp3 transcripts in lung CD4+CD25+ T cells, with a significant decrease in BALF IL-4, IL-5, and IL-13 concentrations and a substantial increase in BALF IL-10 concentrations. These data suggest that the reversal of airway hyperresponsiveness and airway inflammation by Tregs is mediated in part by PD-1, because other costimulatory molecules (e.g., inducible costimulatory

  6. Characterization of the microRNA pool and the factors affecting its regulatory potential.

    PubMed

    Cui, Kai; Lyu, Qing; Xu, Naihan; Liu, Qing; Zhang, Jiarong; Xing, Wei; Bai, Linfu; Liao, Meijian; He, Jie; Yuan, Bo; Chen, Deheng; Xie, Weidong; Zhang, Yaou

    2014-12-01

    The regulation of gene expression by microRNAs (miRNAs) is complex due to a number of variables involved. The potential for one miRNA to target many genes, the presence of multiple miRNA response elements (MREs) in one mRNA molecule and the interplay between RNAs that share common MREs each add a layer of complexity to the process; making it difficult to determine how regulation of gene expression by miRNAs works within the context of the system as a whole. In this study, we used luciferase report vectors inserted with different 3'UTR fragments as probes to detect the repressive effect of the miRNA pool on gene expression and uncovered some essential characteristics of gene regulation mediated by the miRNA pool, such as the nonlinear correlative relationship between the regulatory potential of a miRNA pool and the number of potential MREs, the buffering effect and the saturating effect of the miRNA pool, and the restrictive effect caused by the density of MREs. Through expressing gradient concentration of 3'UTR fragments, we indirectly detected the regulatory potential of the competing endogenous RNA (ceRNA) pool and analysed its effect on the regulatory potential of the miRNA pool. Our results provide some new insights into miRNA pool mediated gene regulation.

  7. Sites of Predicted Stress-Induced DNA Duplex Destabilization Occur Preferentially at Regulatory Loci

    NASA Astrophysics Data System (ADS)

    Benham, Craig J.

    1993-04-01

    This paper describes a computational method to predict the sites on a DNA molecule where imposed superhelical stresses destabilize the duplex. Several DNA sequences are analyzed in this way, including the pBR322 and ColE1 plasmids, bacteriophage f1, and the polyoma and bovine papilloma virus genomes. Superhelical destabilization in these molecules is predicted to occur at small numbers of discrete sites, most of which are within regulatory regions. The most destabilized sites include the terminator and promoter regions of specific plasmid operons, the LexA binding sites of genes under SOS control, the intergenic control region of bacteriophage f1, and the polyadenylylation sites in eukaryotic viruses. These results demonstrate the existence of close correspondences between sites of predicted superhelical duplex destabilization and specific types of regulatory regions. The use of these correspondences to supplement string-matching techniques in the search for regulatory loci is discussed.

  8. Brugia malayi Microfilariae Induce a Regulatory Monocyte/Macrophage Phenotype That Suppresses Innate and Adaptive Immune Responses

    PubMed Central

    Venugopal, Gopinath; Rao, Gopala B.; Lucius, Richard; Srikantam, Aparna; Hartmann, Susanne

    2014-01-01

    Background Monocytes and macrophages contribute to the dysfunction of immune responses in human filariasis. During patent infection monocytes encounter microfilariae in the blood, an event that occurs in asymptomatically infected filariasis patients that are immunologically hyporeactive. Aim To determine whether blood microfilariae directly act on blood monocytes and in vitro generated macrophages to induce a regulatory phenotype that interferes with innate and adaptive responses. Methodology and principal findings Monocytes and in vitro generated macrophages from filaria non-endemic normal donors were stimulated in vitro with Brugia malayi microfilarial (Mf) lysate. We could show that monocytes stimulated with Mf lysate develop a defined regulatory phenotype, characterised by expression of the immunoregulatory markers IL-10 and PD-L1. Significantly, this regulatory phenotype was recapitulated in monocytes from Wuchereria bancrofti asymptomatically infected patients but not patients with pathology or endemic normals. Monocytes from non-endemic donors stimulated with Mf lysate directly inhibited CD4+ T cell proliferation and cytokine production (IFN-γ, IL-13 and IL-10). IFN-γ responses were restored by neutralising IL-10 or PD-1. Furthermore, macrophages stimulated with Mf lysate expressed high levels of IL-10 and had suppressed phagocytic abilities. Finally Mf lysate applied during the differentiation of macrophages in vitro interfered with macrophage abilities to respond to subsequent LPS stimulation in a selective manner. Conclusions and significance Conclusively, our study demonstrates that Mf lysate stimulation of monocytes from healthy donors in vitro induces a regulatory phenotype, characterized by expression of PD-L1 and IL-10. This phenotype is directly reflected in monocytes from filarial patients with asymptomatic infection but not patients with pathology or endemic normals. We suggest that suppression of T cell functions typically seen in lymphatic

  9. Hypoxia inducible factor-1 alpha and multiple myeloma

    PubMed Central

    Tiwary, Bhupendra Nath

    2016-01-01

    Rapid tumor growth creates a state of hypoxia in the tumor microenvironment and results in release of hypoxia inducible factor-1 alpha (HiF-1α) in the local milieu. Hypoxia inducible factor activity is deregulated in many human cancers, especially those that are highly hypoxic. In multiple myeloma (MM) in initial stages of disease establishment, the hypoxic bone marrow microenvironment supports the initial survival and growth of the myeloma cells. Hypoxic tumour cells are usually resistant to radiotherapy and most conventional chemotherapeutic agents, rendering them highly aggressive and metastatic. Therefore, HIF is an attractive, although challenging, therapeutic target in MM directly or indirectly in recent years. PMID:26900575

  10. MicroRNA-23a downregulates the expression of interferon regulatory factor-1 in hepatocellular carcinoma cells.

    PubMed

    Yan, Yihe; Liang, Zhihai; Du, Qiang; Yang, Muqing; Geller, David A

    2016-08-01

    Interferon regulatory factor-1 (IRF-1) is a tumor-suppressor gene induced by interferon-γ (IFNγ) and plays an important role in the cell death of hepatocellular carcinoma (HCC). HCC tumors evade death in part by downregulating IRF-1 expression, yet the molecular mechanisms accounting for IRF-1 suppression in HCC have not yet been characterized. Previous studies have shown that microRNA-23a (miR-23a) can suppress apoptosis by targeting IRF-1. Therefore, we hypothesized that miR-23a promotes HCC growth by downregulating IRF-1. For the in vivo studies, 7 cases of resected HCC and adjacent liver samples were analyzed. For the in vitro studies, IRF-1 mRNA and protein were examined in HepG2 and Huh-7 HCC cells after IFNγ stimulation by real-time PCR and western blotting, respectively. To determine the role of miR-23a in regulating IRF-1, HepG2 cells were transfected with an miR-23a mimic or inhibitor, and IRF-1 expression was examined. Binding of miR-23a was assessed by cloning the 528-bp human IRF-1 3'-untranslated region (3'UTR) into luciferase reporter plasmid pMIR-IRF-1-3'UTR. The results showed that IRF-1 mRNA expression was downregulated in the human HCC tumor tissues compared to that in the adjacent background liver tissues. IFNγ-induced IRF-1 protein was less in the HepG2 tumor cells compared to that in the primary human hepatocytes. miR-23a expression was inversely correlated with IRF-1, and addition of the miR-23a inhibitor increased basal IRF-1 mRNA and protein. Likewise, the miR-23a mimic downregulated IFNγ-induced IRF-1 protein expression, while the miR-23a inhibitor increased IRF-1. Furthermore, the miR-23a mimic repressed IRF-1-3'UTR reporter activity, while the miR-23a inhibitor increased the reporter activity. These results demonstrated that IRF-1 expression is downregulated in human HCC tumors compared to that noted in the background liver. miR-23a downregulates the expression of IRF-1 in HCC cells, and the IRF-1 3'UTR has an miR‑23a binding

  11. MicroRNA-23a downregulates the expression of interferon regulatory factor-1 in hepatocellular carcinoma cells

    PubMed Central

    Yan, Yihe; Liang, Zhihai; Du, Qiang; Yang, Muqing; Geller, David A.

    2016-01-01

    Interferon regulatory factor-1 (IRF-1) is a tumor-suppressor gene induced by interferon-γ (IFNγ) and plays an important role in the cell death of hepatocellular carcinoma (HCC). HCC tumors evade death in part by downregulating IRF-1 expression, yet the molecular mechanisms accounting for IRF-1 suppression in HCC have not yet been characterized. Previous studies have shown that microRNA-23a (miR-23a) can suppress apoptosis by targeting IRF-1. Therefore, we hypothesized that miR-23a promotes HCC growth by down-regulating IRF-1. For the in vivo studies, 7 cases of resected HCC and adjacent liver samples were analyzed. For the in vitro studies, IRF-1 mRNA and protein were examined in HepG2 and Huh-7 HCC cells after IFNγ stimulation by real-time PCR and western blotting, respectively. To determine the role of miR-23a in regulating IRF-1, HepG2 cells were transfected with an miR-23a mimic or inhibitor, and IRF-1 expression was examined. Binding of miR-23a was assessed by cloning the 528-bp human IRF-1 3′-untranslated region (3′UTR) into luciferase reporter plasmid pMIR-IRF-1-3′UTR. The results showed that IRF-1 mRNA expression was down-regulated in the human HCC tumor tissues compared to that in the adjacent background liver tissues. IFNγ-induced IRF-1 protein was less in the HepG2 tumor cells compared to that in the primary human hepatocytes. miR-23a expression was inversely correlated with IRF-1, and addition of the miR-23a inhibitor increased basal IRF-1 mRNA and protein. Likewise, the miR-23a mimic downregulated IFNγ-induced IRF-1 protein expression, while the miR-23a inhibitor increased IRF-1. Furthermore, the miR-23a mimic repressed IRF-1-3′UTR reporter activity, while the miR-23a inhibitor increased the reporter activity. These results demonstrated that IRF-1 expression is downregulated in human HCC tumors compared to that noted in the background liver. miR-23a downregulates the expression of IRF-1 in HCC cells, and the IRF-1 3′UTR has an miR-23a

  12. Andrographolide prevents high-fat diet-induced obesity in C57BL/6 mice by suppressing the sterol regulatory element-binding protein pathway.

    PubMed

    Ding, Lili; Li, Jinmei; Song, Baoliang; Xiao, Xu; Huang, Wendong; Zhang, Binfeng; Tang, Xiaowen; Qi, Meng; Yang, Qiming; Yang, Qiaoling; Yang, Li; Wang, Zhengtao

    2014-11-01

    Sterol regulatory element-binding proteins (SREBPs) are major transcription factors regulating the expression of genes involved in biosynthesis of cholesterol, fatty acids, and triglycerides. We investigated the effect of the specific SREBP suppressor andrographolide, a natural compound isolated from Andrographis paniculata, on the regulation of SREBP signaling by use of Western blot, reporter gene assay, and quantitative real-time polymerase chain reaction analysis. In addition, the antiobesity effects of andrographolide were evaluated in C57BL/6 mice with high-fat diet (HFD)-induced obesity. Our results showed that andrographolide downregulated the expressions of SREBPs target genes and decreased cellular lipid accumulation in vitro. Further, andrographolide (100 mg/kg per day) attenuated HFD-induced body weight gain and fat accumulation in liver or adipose tissues, and improved serum lipid levels and insulin or glucose sensitivity in HFD-induced obese mice. Andrographolide effectively suppressed the respiratory quotient, energy expenditure, and oxygen consumption, which may have contributed to the decreased body-weight gain of the obese mice fed with a HFD. Consistently, andrographolide regulated SREBP target genes and metabolism-associated genes in liver or brown adipose tissue, which may have directly contributed to the lower lipid levels and enhanced insulin sensitivity. Taken together, our results indicated that andrographolide ameliorated lipid metabolism and improved glucose use in mice with HFD-induced obesity. Andrographolide has potential as a leading compound in the prevention or treatment of obesity and insulin resistance.

  13. TOP1 and 2, polysaccharides from Taraxacum officinale, attenuate CCl(4)-induced hepatic damage through the modulation of NF-kappaB and its regulatory mediators.

    PubMed

    Park, Chung Mu; Youn, Hyun Joo; Chang, Hee Kyung; Song, Young Sun

    2010-05-01

    In this work, we estimate the inhibitory effect of two polysaccharides from Taraxacum officinale (TOP) on CCl(4)-induced oxidative stress and inflammation in Sprague-Dawley rats. TOP1 and 2 (304, 92 mg/kg bw) were administered for 7 days via a stomach sonde, and hepatitis was induced by a single dose of CCl(4) (50% CCl(4)/olive oil; 0.5 mL/kg bw) administration. CCl(4) significantly elevated serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities. Histopathological observation further revealed that CCl(4)-induced moderate levels of inflammatory cell infiltration, centrilobular fatty change, apoptosis, and necrosis. However, TOPs pretreatment markedly decreased AST and ALT activities as well as hepatic lesions. TOPs also increased free radical scavenging activity, as exhibited by a lowered TBARS concentration. TOPs pretreatment also reversed other hepatitis-associated symptoms, including GSH depletion, inhibited anti-oxidative enzyme activities, up-regulation of NF-kappaB and increased expression of its regulatory inflammatory mediators, such as inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-1beta. These results suggest that TOPs have a hepatoprotective effect by modulating inflammatory responses and ameliorating oxidative stress.

  14. Population genetic and phylogenetic evidence for positive selection on regulatory mutations at the factor VII locus in humans.

    PubMed Central

    Hahn, Matthew W; Rockman, Matthew V; Soranzo, Nicole; Goldstein, David B; Wray, Gregory A

    2004-01-01

    The abundance of cis-regulatory polymorphisms in humans suggests that many may have been important in human evolution, but evidence for their role is relatively rare. Four common polymorphisms in the 5' promoter region of factor VII (F7), a coagulation factor, have been shown to affect its transcription and protein abundance both in vitro and in vivo. Three of these polymorphisms have low-frequency alleles that decrease expression of F7 and may provide protection against myocardial infarction (heart attacks). The fourth polymorphism has a minor allele that increases the level of transcription. To look for evidence of natural selection on the cis-regulatory variants flanking F7, we genotyped three of the polymorphisms in six Old World populations for which we also have data from a group of putatively neutral SNPs. Our population genetic analysis shows evidence for selection within humans; surprisingly, the strongest evidence is due to a large increase in frequency of the high-expression variant in Singaporean Chinese. Further characterization of a Japanese population shows that at least part of the increase in frequency of the high-expression allele is found in other East Asian populations. In addition, to examine interspecific patterns of selection we sequenced the homologous 5' noncoding region in chimpanzees, bonobos, a gorilla, an orangutan, and a baboon. Analysis of these data reveals an excess of fixed differences within transcription factor binding sites along the human lineage. Our results thus further support the hypothesis that regulatory mutations have been important in human evolution. PMID:15238535

  15. The Third Intron of the Interferon Regulatory Factor-8 Is an Initiator of Repressed Chromatin Restricting Its Expression in Non-Immune Cells

    PubMed Central

    Barnea-Yizhar, Ofer; Ram, Sigal; Kovalev, Ekaterina; Azriel, Aviva; Rand, Ulfert; Nakayama, Manabu; Hauser, Hansjörg; Gepstein, Lior; Levi, Ben-Zion

    2016-01-01

    Interferon Regulatory Factor-8 (IRF-8) serves as a key factor in the hierarchical differentiation towards monocyte/dendritic cell lineages. While much insight has been accumulated into the mechanisms essential for its hematopoietic specific expression, the mode of restricting IRF-8 expression in non-hematopoietic cells is still unknown. Here we show that the repression of IRF-8 expression in restrictive cells is mediated by its 3rd intron. Removal of this intron alleviates the repression of Bacterial Artificial Chromosome (BAC) IRF-8 reporter gene in these cells. Fine deletion analysis points to conserved regions within this intron mediating its restricted expression. Further, the intron alone selectively initiates gene silencing only in expression-restrictive cells. Characterization of this intron’s properties points to its role as an initiator of sustainable gene silencing inducing chromatin condensation with suppressive histone modifications. This intronic element cannot silence episomal transgene expression underlining a strict chromatin-dependent silencing mechanism. We validated this chromatin-state specificity of IRF-8 intron upon in-vitro differentiation of induced pluripotent stem cells (iPSCs) into cardiomyocytes. Taken together, the IRF-8 3rd intron is sufficient and necessary to initiate gene silencing in non-hematopoietic cells, highlighting its role as a nucleation core for repressed chromatin during differentiation. PMID:27257682

  16. Antibodies targeting human OX40 expand effector T cells and block inducible and natural regulatory T cell function

    PubMed Central

    Voo, Kui S.; Bover, Laura; Harline, Megan L.; Vien, Long T.; Facchinetti, Valeria; Arima, Kazuhiko; Kwak, Larry W.; Liu, Yong J.

    2013-01-01

    Current cancer vaccines induce tumor-specific T cell responses without sustained tumor regression because immunosuppressive elements within the tumor induce exhaustion of effector T cells and infiltration of immune-suppressive regulatory T cells (Tregs). Therefore, much effort has been made to generate agonistic Abs targeting members of the TNFR superfamily, such as OX40, 4- 1BB, and GITR, expressed on effector T cells and Tregs, to reinvigorate T cell effector function and block Treg-suppressive function. In this article, we describe the development of a panel of anti-human OX40 agonistic mouse mAbs that could promote effector CD4+ and CD8+ T cell proliferation, inhibit the induction of CD4+ IL-10 -producing type 1 regulatory T cells, inhibit the expansion of ICOS+IL-10+ Tregs, inhibit TGF-b–induced FOXP3 expression on naive CD4+ T cells, and block natural Treg–suppressive function. We humanized two anti–human OX40 mAb clones, and they retained the potency of their parental clones. These Abs should provide broad opportunities for potential combination therapy to treat a wide realm of cancers and preventative vaccines against infectious diseases. PMID:24014877

  17. Antibodies targeting human OX40 expand effector T cells and block inducible and natural regulatory T cell function.

    PubMed

    Voo, Kui S; Bover, Laura; Harline, Megan L; Vien, Long T; Facchinetti, Valeria; Arima, Kazuhiko; Kwak, Larry W; Liu, Yong J

    2013-10-01

    Current cancer vaccines induce tumor-specific T cell responses without sustained tumor regression because immunosuppressive elements within the tumor induce exhaustion of effector T cells and infiltration of immune-suppressive regulatory T cells (Tregs). Therefore, much effort has been made to generate agonistic Abs targeting members of the TNFR superfamily, such as OX40, 4-1BB, and GITR, expressed on effector T cells and Tregs, to reinvigorate T cell effector function and block Treg-suppressive function. In this article, we describe the development of a panel of anti-human OX40 agonistic mouse mAbs that could promote effector CD4(+) and CD8(+) T cell proliferation, inhibit the induction of CD4(+) IL-10 -producing type 1 regulatory T cells, inhibit the expansion of ICOS(+)IL-10(+) Tregs, inhibit TGF-β-induced FOXP3 expression on naive CD4(+) T cells, and block natural Treg-suppressive function. We humanized two anti-human OX40 mAb clones, and they retained the potency of their parental clones. These Abs should provide broad opportunities for potential combination therapy to treat a wide realm of cancers and preventative vaccines against infectious diseases. PMID:24014877

  18. An integrated approach to characterize transcription factor and microRNA regulatory networks involved in Schwann cell response to peripheral nerve injury

    PubMed Central

    2013-01-01

    Background The regenerative response of Schwann cells after peripheral nerve injury is a critical process directly related to the pathophysiology of a number of neurodegenerative diseases. This SC injury response is dependent on an intricate gene regulatory program coordinated by a number of transcription factors and microRNAs, but the interactions among them remain largely unknown. Uncovering the transcriptional and post-transcriptional regulatory networks governing the Schwann cell injury response is a key step towards a better understanding of Schwann cell biology and may help develop novel therapies for related diseases. Performing such comprehensive network analysis requires systematic bioinformatics methods to integrate multiple genomic datasets. Results In this study we present a computational pipeline to infer transcription factor and microRNA regulatory networks. Our approach combined mRNA and microRNA expression profiling data, ChIP-Seq data of transcription factors, and computational transcription factor and microRNA target prediction. Using mRNA and microRNA expression data collected in a Schwann cell injury model, we constructed a regulatory network and studied regulatory pathways involved in Schwann cell response to injury. Furthermore, we analyzed network motifs and obtained insights on cooperative regulation of transcription factors and microRNAs in Schwann cell injury recovery. Conclusions This work demonstrates a systematic method for gene regulatory network inference that may be used to gain new information on gene regulation by transcription factors and microRNAs. PMID:23387820

  19. Growth factor involvement in tension-induced skeletal muscle growth

    NASA Technical Reports Server (NTRS)

    Vandenburgh, Herman H.

    1993-01-01

    Long-term manned space travel will require a better understanding of skeletal muscle atrophy which results from microgravity. Astronaut strength and dexterity must be maintained for normal mission operations and for emergency situations. Although exercise in space slows the rate of muscle loss, it does not prevent it. A biochemical understanding of how gravity/tension/exercise help to maintain muscle size by altering protein synthesis and/or degradation rate should ultimately allow pharmacological intervention to prevent muscle atrophy in microgravity. The overall objective is to examine some of the basic biochemical processes involved in tension-induced muscle growth. With an experimental in vitro system, the role of exogenous and endogenous muscle growth factors in mechanically stimulated muscle growth are examined. Differentiated avian skeletal myofibers can be 'exercised' in tissue culture using a newly developed dynamic mechanical cell stimulator device which simulates different muscle activity patterns. Patterns of mechanical activity which significantly affect muscle growth and metabolic characteristics were found. Both exogenous and endogenous growth factors are essential for tension-induced muscle growth. Exogenous growth factors found in serum, such as insulin, insulin-like growth factors, and steroids, are important regulators of muscle protein turnover rates and mechanically-induced muscle growth. Endogenous growth factors are synthesized and released into the culture medium when muscle cells are mechanically stimulated. At least one family of mechanically induced endogenous factors, the prostaglandins, help to regulate the rates of protein turnover in muscle cells. Endogenously synthesized IGF-1 is another. The interaction of muscle mechanical activity and these growth factors in the regulation of muscle protein turnover rates with our in vitro model system is studied.

  20. Growth factor induced proliferation, migration, and lumen formation of rat endometrial epithelial cells in vitro

    PubMed Central

    ISLAM, Md. Rashedul; YAMAGAMI, Kazuki; YOSHII, Yuka; YAMAUCHI, Nobuhiko

    2016-01-01

    Endometrial modulation is essential for the preservation of normal uterine physiology, and this modulation is driven by a number of growth factors. The present study investigated the mitogenic, motogenic, and morphogenic effects of epidermal growth factor (EGF) and hepatocyte growth factor (HGF) on rat endometrial epithelial (REE) cells. The REE cells were isolated and cultured and then characterized based on their morphology and their expression of epithelial cell markers. The MTT assay revealed that EGF and HGF induce proliferation of REE cells. Consistent with increased proliferation, we found that the cell cycle regulatory factor Cyclin D1 was also upregulated upon EGF and HGF addition. REE cell migration was prompted by EGF, as observed with the Oris Cell Migration Assay. The morphogenic impact of growth factors on REE cells was studied in a three-dimensional BD Matrigel cell culture system, wherein these growth factors also increased the frequency of lumen formation. In summary, we show that EGF and HGF have a stimulatory effect on REE cells, promoting proliferation, cell migration, and lumen formation. Our findings provide important insights that further the understanding of endometrial regeneration and its regulation. PMID:26946922

  1. CLINICAL FACTORS FOR DEVELOPING SHOCK IN RADIOCONTRAST MEDIA INDUCED ANAPHYLAXIS.

    PubMed

    Kim, Sang Min; Ko, Byuk Sung; Kim, Ji Yeon; Ha, Sang Ook; Ahn, Shin; Sohn, Chang Hwan; Seo, Dong Woo; Kim, Tae-Bum; Kim, Won Young

    2016-03-01

    The aim of this study was to investigate the time interval between radiocontrast media (RCM) administration and the development of anaphylactic shock, and risk factors associated with RCM-induced anaphylactic shock. We reviewed the medical records of 154 patients with RCM-induced anaphylaxis presenting to the emergency department of a tertiary care hospital between January 2005 and December 2014. Clinical features of RCM-induced anaphylaxis were analyzed, and patients were categorized into shock and non-shock groups to identify associated factors in affected patients. Of the 154 cases of RCM-induced anaphylaxis, 101 (65.9%) patients experienced shock. The median time between RCM exposure and the onset of shock was 11 min (interquartile range, 7.0-18.8). In patients with RCM-induced anaphylaxis accompanying shock, the median time from RCM to the first symptom onset was 6 min (interquartile range, 5.0-10.0). In the multivariate analysis, age, neurological manifestations, and allergy history except RCM were associated with the development of shock. RCM-induced anaphylaxis was commonly accompanied with shock, and the time interval between RCM exposure and the onset of shock was short. Physicians should pay attention to the development of potential cardiovascular collapse in anaphylaxis patients of old age and with neurologic manifestations. PMID:26506069

  2. Hypoxia Inducible Factors and Hypertension: Lessons from Sleep Apnea Syndrome

    PubMed Central

    Nanduri, Jayasri; Peng, Ying-Jie; Yuan, Guoxiang; Kumar, Ganesh K.; Prabhakar, Nanduri R.

    2015-01-01

    Systemic hypertension is one of the most prevalent cardiovascular diseases. Sleep disordered breathing (SDB) with recurrent apnea is a major risk factor for developing essential hypertension. Chronic intermittent hypoxia (CIH) is a hallmark manifestation of recurrent apnea. Rodent models patterned after the O2 profiles seen with SDB patients showed that CIH is the major stimulus for causing systemic hypertension. This article reviews the physiological and molecular basis of CIH-induced hypertension. Physiological studies have identified that augmented carotid body chemosensory reflex and the resulting increase in sympathetic nerve activity is a major contributor to CIH-induced hypertension. Analysis of molecular mechanisms revealed that CIH activates hypoxia-inducible factor (HIF)-1 and suppresses HIF-2- mediated transcription. Dysregulation of HIF-1- and HIF-2- mediated transcription leads to imbalance of pro-oxidant and anti-oxidant enzyme gene expression resulting in increased reactive species (ROS) generation in the chemosensory reflex which is central for developing hypertension. PMID:25772710

  3. Interruption of dendritic cell-mediated TIM-4 signaling induces regulatory T cells and promotes skin allograft survival.

    PubMed

    Yeung, Melissa Y; McGrath, Martina M; Nakayama, Masafumi; Shimizu, Tetsunosuke; Boenisch, Olaf; Magee, Ciara N; Abdoli, Rozita; Akiba, Hisaya; Ueno, Takuya; Turka, Laurence A; Najafian, Nader

    2013-10-15

    Dendritic cells (DCs) are the central architects of the immune response, inducing inflammatory or tolerogenic immunity, dependent on their activation status. As such, DCs are highly attractive therapeutic targets and may hold the potential to control detrimental immune responses. TIM-4, expressed on APCs, has complex functions in vivo, acting both as a costimulatory molecule and a phosphatidylserine receptor. The effect of TIM-4 costimulation on T cell activation remains unclear. In this study, we demonstrate that Ab blockade of DC-expressed TIM-4 leads to increased induction of induced regulatory T cells (iTregs) from naive CD4(+) T cells, both in vitro and in vivo. iTreg induction occurs through suppression of IL-4/STAT6/Gata3-induced Th2 differentiation. In addition, blockade of TIM-4 on previously activated DCs still leads to increased iTreg induction. iTregs induced under TIM-4 blockade have equivalent potency to control and, upon adoptive transfer, significantly prolong skin allograft survival in vivo. In RAG(-/-) recipients of skin allografts adoptively transferred with CD4(+) T cells, we show that TIM-4 blockade in vivo is associated with a 3-fold prolongation in allograft survival. Furthermore, in this mouse model of skin transplantation, increased induction of allospecific iTregs and a reduction in T effector responses were observed, with decreased Th1 and Th2 responses. This enhanced allograft survival and protolerogenic skewing of the alloresponse is critically dependent on conversion of naive CD4(+) to Tregs in vivo. Collectively, these studies identify blockade of DC-expressed TIM-4 as a novel strategy that holds the capacity to induce regulatory immunity in vivo.

  4. Immunotherapy of Murine Retrovirus-Induced Acquired Immunodeficiency by CD4 T Regulatory Cell Depletion and PD-1 Blockade ▿

    PubMed Central

    Li, Wen; Green, William R.

    2011-01-01

    LP-BM5 retrovirus induces a complex disease featuring an acquired immunodeficiency syndrome termed murine AIDS (MAIDS) in susceptible strains of mice, such as C57BL/6 (B6). CD4 T helper effector cells are required for MAIDS induction and progression of viral pathogenesis. CD8 T cells are not needed for viral pathogenesis, but rather, are essential for protection from disease in resistant strains, such as BALB/c. We have discovered an immunodominant cytolytic T lymphocyte (CTL) epitope encoded in a previously unrecognized LP-BM5 retroviral alternative (+1 nucleotide [nt]) gag translational open reading frame. CTLs specific for this cryptic gag epitope are the basis of protection from LP-BM5-induced immunodeficiency in BALB/c mice, and the inability of B6 mice to mount an anti-gag CTL response appears critical to the initiation and progression of LP-BM5-induced MAIDS. However, uninfected B6 mice primed by LP-BM5-induced tumors can generate CTL responses to an LP-BM5 retrovirus infection-associated epitope(s) that is especially prevalent on such MAIDS tumor cells, indicating the potential to mount a protective CD8 T-cell response. Here, we utilized this LP-BM5 retrovirus-induced disease system to test whether modulation of normal immune down-regulatory mechanisms can alter retroviral pathogenesis. Thus, following in vivo depletion of CD4 T regulatory (Treg) cells and/or selective interruption of PD-1 negative signaling in the CD8 T-cell compartment, retroviral pathogenesis was significantly decreased, with the combined treatment of CD4 Treg cell depletion and PD-1 blockade working in a synergistic fashion to substantially reduce the induction of MAIDS. PMID:21917983

  5. [Butanol as a regulatory factor of ompC gene expression in E. coli cells].

    PubMed

    Seregina, T A; Shakulov, R S; Mironov, A S

    2012-11-01

    The influence of butanol on the expression of ompC gene encoding synthesis of OmpC porin in the MG 1655 strain of E. coli and butanol-tolerant mutant ButR was studied. It was shown that in the case of wild bacteria, the addition of butanol to the growth medium results in an increased level of ompC transcription. However, OmpC porin is not detected in the membrane fraction of cells. ButR mutant exhibits a higher level of ompC gene expression. A direct correlation is observed between the level of OmpC porin expression and its content in the membrane fraction of ButR mutant cells. In the regulatory region of the ompC gene of the ButR mutant, three nucleotide substitutions located in the binding sites of OmpR and CpxR activator proteins were identified. It was shown that mutations in the regulatory region of the ompC gene in the ButR mutant are responsible for the decreased level of OmpC porin expression under normal growth conditions. However, these mutations lead to an increased level of OmpC porin synthesis in the presence of butanol. These data suggest an additional mechanism of ompC gene regulation with the participation of butanol as a positive transcription effector.

  6. PcFKH1, a novel regulatory factor from the forkhead family, controls the biosynthesis of penicillin in Penicillium chrysogenum.

    PubMed

    Domínguez-Santos, Rebeca; García-Estrada, Carlos; Kosalková, Katarina; Prieto, Carlos; Santamarta, Irene; Martín, Juan-Francisco

    2015-08-01

    Penicillin biosynthesis in Penicillium chrysogenum (re-identified as Penicillium rubens) is a good example of a biological process subjected to complex global regulatory networks and serves as a model to study fungal secondary metabolism. The winged-helix family of transcription factors recently described, which includes the forkhead type of proteins, is a key type of regulatory proteins involved in this process. In yeasts and humans, forkhead transcription factors are involved in different processes (cell cycle regulation, cell death control, pre-mRNA processing and morphogenesis); one member of this family of proteins has been identified in the P. chrysogenum genome (Pc18g00430). In this work, we have characterized this novel transcription factor (named PcFKH1) by generating knock-down mutants and overexpression strains. Results clearly indicate that PcFKH1 positively controls antibiotic biosynthesis through the specific interaction with the promoter region of the penDE gene, thus regulating penDE mRNA levels. PcFKH1 also binds to the pcbC promoter, but with low affinity. In addition, it also controls other ancillary genes of the penicillin biosynthetic process, such as phlA (encoding phenylacetyl CoA ligase) and ppt (encoding phosphopantetheinyl transferase). PcFKH1 also plays a role in conidiation and spore pigmentation, but it does not seem to be involved in hyphal morphology or cell division in the improved laboratory reference strain Wisconsin 54-1255. A genome-wide analysis of processes putatively coregulated by PcFKH1 and PcRFX1 (another winged-helix transcription factor) in P. chrysogenum provided evidence of the global effect of these transcription factors in P. chrysogenum metabolism.

  7. Overexpression of gankyrin in mouse hepatocytes induces hemangioma by suppressing factor inhibiting hypoxia-inducible factor-1 (FIH-1) and activating hypoxia-inducible factor-1.

    PubMed

    Liu, Yu; Higashitsuji, Hiroaki; Higashitsuji, Hisako; Itoh, Katsuhiko; Sakurai, Toshiharu; Koike, Kazuhiko; Hirota, Kiichi; Fukumoto, Manabu; Fujita, Jun

    2013-03-01

    Gankyrin (also called p28 or PSMD10) is an oncoprotein commonly overexpressed in hepatocellular carcinomas. It consists of 7 ankyrin repeats and interacts with multiple proteins including Rb, Cdk4, MDM2 and NF-κB. To assess the oncogenic activity in vivo, we produced transgenic mice that overexpress gankyrin specifically in the hepatocytes. Unexpectedly, 5 of 7 F2 transgenic mice overexpressing hepatitis B virus X protein (HBX) promoter-driven gankyrin, and one of 3 founder mice overexpressing serum amyloid P component (SAP) promoter-driven gankyrin developed hepatic vascular neoplasms (hemangioma/hemangiosarcomas) whereas none of the wild-type mice did. Endothelial overgrowth was more frequent in the livers of diethylnitrosamine-treated transgenic mice than wild-type mice. Mouse hepatoma Hepa1-6 cells overexpressing gankyrin formed tumors with more vascularity than parental Hepa1-6 cells in the transplanted mouse skin. We found that gankyrin binds to and sequester factor inhibiting hypoxia-inducible factor-1 (FIH-1), which results in decreased interaction between FIH-1 and hypoxia-inducible factor-1α (HIF-1α) and increased activity of HIF-1 to promote VEGF production. The effects of gankyrin were more prominent under 3% O2 than 1% or 20% O2 conditions. Thus, the present study clarified, at least partly, mechanisms of vascular tumorigenesis, and suggests that gankyrin might play a physiological role in hypoxic responses besides its roles as an oncoprotein. PMID:23376718

  8. Mutational analysis of the [4Fe-4S]-cluster converting iron regulatory factor from its RNA-binding form to cytoplasmic aconitase.

    PubMed Central

    Hirling, H; Henderson, B R; Kühn, L C

    1994-01-01

    The control of cellular iron homeostasis involves the coordinate post-transcriptional regulation of ferritin mRNA translation and transferring receptor mRNA stability. These regulatory events are mediated by a soluble cytoplasmic protein, iron regulatory factor (IRF), which binds specifically to mRNA hairpin structures, termed iron-responsive elements (IREs), in the respective mRNAs. IRF is modulated by variations of cellular iron levels and exists as either an apo-protein or a [4Fe-4S]-cluster protein. The two conformations show distinct, mutually exclusive functions. High-affinity IRE binding is observed with the apo-form induced by iron deprivation, but is lost under high iron conditions when IRF is converted to the [4Fe-4S]-cluster form which shows cytoplasmic aconitase activity. Moreover, IRE binding is inactivated by the sulfhydryl-oxidizing agent diamide and fully activated in vitro by 2% 2-mercapto-ethanol, whereas alkylation of IRF inhibits IRE binding. In the present study, we analyzed each of the above features using site-directed mutants of recombinant human IRF. The results support the bifunctional nature of IRF. We conclude that cysteines 437, 503 and 506 anchor the [4Fe-4S]-cluster, and are essential to the aconitase activity. Mutagenesis changing any of the cysteines to serine leads to constitutive RNA binding in 0.02% 2-mercaptoethanol. Cysteine 437 is particularly critical to the RNA-protein interaction. The spontaneous or diamide-induced formation of disulfide bonds between cysteines 437 and 503 or 437 and 506, in apo-IRF, as well as its alkylation by N-ethylmaleimide, inhibit binding to the IRE.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:7508861

  9. Factors that modify risks of radiation-induced cancer

    SciTech Connect

    Fabrikant, J.I.

    1988-11-01

    The collective influence of biologic and physical factors that modify risks of radiation-induced cancer introduces uncertainties sufficient to deny precision of estimates of human cancer risk that can be calculated for low-dose radiation in exposed populations. The important biologic characteristics include the tissue sites and cell types, baseline cancer incidence, minimum latent period, time-to-tumor recognition, and the influence of individual host (age and sex) and competing etiologic influences. Physical factors include radiation dose, dose rate, and radiation quality. Statistical factors include time-response projection models, risk coefficients, and dose-response relationships. Other modifying factors include other carcinogens, and other biological sources (hormonal status, immune status, hereditary factors).

  10. Mycobacteria-responsive sonic hedgehog signaling mediates programmed death-ligand 1- and prostaglandin E2-induced regulatory T cell expansion

    PubMed Central

    Holla, Sahana; Stephen-Victor, Emmanuel; Prakhar, Praveen; Sharma, Meenu; Saha, Chaitrali; Udupa, Vibha; Kaveri, Srinivas V.; Bayry, Jagadeesh; Balaji, Kithiganahalli Narayanaswamy

    2016-01-01

    CD4+CD25+FoxP3+ regulatory T cells (Tregs) are exploited by mycobacteria to subvert the protective host immune responses. The Treg expansion in the periphery requires signaling by professional antigen presenting cells and in particularly dendritic cells (DC). However, precise molecular mechanisms by which mycobacteria instruct Treg expansion via DCs are not established. Here we demonstrate that mycobacteria-responsive sonic hedgehog (SHH) signaling in human DCs leads to programmed death ligand-1 (PD-L1) expression and cyclooxygenase (COX)-2-catalyzed prostaglandin E2 (PGE2) that orchestrate mycobacterial infection-induced expansion of Tregs. While SHH-responsive transcription factor GLI1 directly arbitrated COX-2 transcription, specific microRNAs, miR-324-5p and miR-338-5p, which target PD-L1 were downregulated by SHH signaling. Further, counter-regulatory roles of SHH and NOTCH1 signaling during mycobacterial-infection of human DCs was also evident. Together, our results establish that Mycobacterium directs a fine-balance of host signaling pathways and molecular regulators in human DCs to expand Tregs that favour immune evasion of the pathogen. PMID:27080341

  11. Enhancer sequence variants and transcription-factor deregulation synergize to construct pathogenic regulatory circuits in B-cell lymphoma.

    PubMed

    Koues, Olivia I; Kowalewski, Rodney A; Chang, Li-Wei; Pyfrom, Sarah C; Schmidt, Jennifer A; Luo, Hong; Sandoval, Luis E; Hughes, Tyler B; Bednarski, Jeffrey J; Cashen, Amanda F; Payton, Jacqueline E; Oltz, Eugene M

    2015-01-20

    Most B-cell lymphomas arise in the germinal center (GC), where humoral immune responses evolve from potentially oncogenic cycles of mutation, proliferation, and clonal selection. Although lymphoma gene expression diverges significantly from GC B cells, underlying mechanisms that alter the activities of corresponding regulatory elements (REs) remain elusive. Here we define the complete pathogenic circuitry of human follicular lymphoma (FL), which activates or decommissions REs from normal GC B cells and commandeers enhancers from other lineages. Moreover, independent sets of transcription factors, whose expression was deregulated in FL, targeted commandeered versus decommissioned REs. Our approach revealed two distinct subtypes of low-grade FL, whose pathogenic circuitries resembled GC B or activated B cells. FL-altered enhancers also were enriched for sequence variants, including somatic mutations, which disrupt transcription-factor binding and expression of circuit-linked genes. Thus, the pathogenic regulatory circuitry of FL reveals distinct genetic and epigenetic etiologies for GC B-cell transformation.

  12. Superior Cervical Ganglia Neurons Induce Foxp3+ Regulatory T Cells via Calcitonin Gene-Related Peptide

    PubMed Central

    Szklany, Kirsten; Ruiter, Evelyn; Mian, Firoz; Kunze, Wolfgang; Bienenstock, John; Forsythe, Paul; Karimi, Khalil

    2016-01-01

    The nervous and immune systems communicate bidirectionally, utilizing diverse molecular signals including cytokines and neurotransmitters to provide an integrated response to changes in the body’s internal and external environment. Although, neuro-immune interactions are becoming better understood under inflammatory circumstances and it has been evidenced that interaction between neurons and T cells results in the conversion of encephalitogenic T cells to T regulatory cells, relatively little is known about the communication between neurons and naïve T cells. Here, we demonstrate that following co-culture of naïve CD4+ T cells with superior cervical ganglion neurons, the percentage of Foxp3 expressing CD4+CD25+ cells significantly increased. This was mediated in part by immune-regulatory cytokines TGF-β and IL-10, as well as the neuropeptide calcitonin gene-related peptide while vasoactive intestinal peptide was shown to play no role in generation of T regulatory cells. Additionally, T cells co-cultured with neurons showed a decrease in the levels of pro-inflammatory cytokine IFN-γ released upon in vitro stimulation. These findings suggest that the generation of Tregs may be promoted by naïve CD4+ T cell: neuron interaction through the release of neuropeptide CGRP. PMID:27022966

  13. Two new monoclonal antibodies for biochemical and flow cytometric analyses of human interferon regulatory factor-3 activation, turnover, and depletion.

    PubMed

    Rustagi, Arjun; Doehle, Brian P; McElrath, M Juliana; Gale, Michael

    2013-02-01

    Interferon regulatory factor-3 (IRF-3) is a master transcription factor that drives the host intracellular innate immune response to virus infection. The importance of IRF-3 in innate immune responses is highlighted by the fact that pathogenic viruses have developed strategies for antagonism of IRF-3. Several tools exist for evaluation of viral regulation of IRF-3 activation and function, but high-quality monoclonal antibodies that mark the differential activation states of human IRF-3 are lacking. To study IRF-3 activation, turnover, and depletion in a high-throughput manner in the context of virus infection, we have developed two new monoclonal antibodies to human IRF-3. These antibodies detect IRF-3 in virus-infected cells in a wide variety of assays and provide a new tool to study virus-host interactions and innate immune signaling.

  14. Two new monoclonal antibodies for biochemical and flow cytometric analyses of human interferon regulatory factor-3 activation, turnover, and depletion

    PubMed Central

    Rustagi, Arjun; Doehle, Brian P.; McElrath, M. Juliana; Gale, Michael

    2012-01-01

    Interferon regulatory factor-3 (IRF-3) is a master transcription factor that drives the host intracellular innate immune response to virus infection. The importance of IRF-3 in innate immune responses is highlighted by the fact that pathogenic viruses have developed strategies for antagonism of IRF-3. Several tools exist for evaluation of viral regulation of IRF-3 activation and function, but high-quality monoclonal antibodies that mark the differential activation states of human IRF-3 are lacking. To study IRF-3 activation, turnover, and depletion in a high-throughput manner in the context of virus infection, we have developed two new monoclonal antibodies to human IRF-3. These antibodies detect IRF-3 in virus-infected cells in a wide variety of assays and provide a new tool to study virus-host interactions and innate immune signaling. PMID:22705311

  15. Inhibitor of apoptosis proteins (IAPs) as regulatory factors of hepatic apoptosis.

    PubMed

    Wang, Kewei; Lin, Bingliang

    2013-10-01

    IAPs are a group of regulatory proteins that are structurally related. Their conserved homologues have been identified in various organisms. In human, eight IAP members have been recognized based on baculoviral IAP repeat (BIR) domains. IAPs are key regulators of apoptosis, cytokinesis and signal transduction. The antiapoptotic property of IAPs depends on their professional role for caspases. IAPs are functionally non-equivalent and regulate effector caspases through distinct mechanisms. IAPs impede apoptotic process via membrane receptor-dependent (extrinsic) cascade and mitochondrial dependent (intrinsic) pathway. IAP-mediated apoptosis affects the progression of liver diseases. Therapeutic options of liver diseases may depend on the understanding toward mechanisms of the IAP-mediated apoptosis.

  16. Suppression of the pancreatic duodenal homeodomain transcription factor-1 (Pdx-1) promoter by sterol regulatory element-binding protein-1c (SREBP-1c).

    PubMed

    Amemiya-Kudo, Michiyo; Oka, Junko; Takeuchi, Yoshinori; Okazaki, Hiroaki; Yamamoto, Takashi; Yahagi, Naoya; Matsuzaka, Kaori; Okazaki, Sachiko; Osuga, Jun-ichi; Yamada, Nobuhiro; Murase, Toshio; Shimano, Hitoshi

    2011-08-12

    Overexpression of sterol regulatory element-binding protein-1c (SREBP-1c) in β cells causes impaired insulin secretion and β cell dysfunction associated with diminished pancreatic duodenal homeodomain transcription factor-1 (PDX-1) expression in vitro and in vivo. To identify the molecular mechanism responsible for this effect, the mouse Pdx-1 gene promoter (2.7 kb) was analyzed in β cell and non-β cell lines. Despite no apparent sterol regulatory element-binding protein-binding sites, the Pdx-1 promoter was suppressed by SREBP-1c in β cells in a dose-dependent manner. PDX-1 activated its own promoter. The E-box (-104/-99 bp) in the proximal region, occupied by ubiquitously expressed upstream stimulatory factors (USFs), was crucial for the PDX-1-positive autoregulatory loop through direct PDX-1·USF binding. This positive feedback activation was a prerequisite for SREBP-1c suppression of the promoter in non-β cells. SREBP-1c and PDX-1 directly interact through basic helix-loop-helix and homeobox domains, respectively. This robust SREBP-1c·PDX-1 complex interferes with PDX-1·USF formation and inhibits the recruitment of PDX-1 coactivators. SREBP-1c also inhibits PDX-1 binding to the previously described PDX-1-binding site (-2721/-2646 bp) in the distal enhancer region of the Pdx-1 promoter. Endogenous up-regulation of SREBP-1c in INS-1 cells through the activation of liver X receptor and retinoid X receptor by 9-cis-retinoic acid and 22-hydroxycholesterol inhibited PDX-1 mRNA and protein expression. Conversely, SREBP-1c RNAi restored Pdx-1 mRNA and protein levels. Through these multiple mechanisms, SREBP-1c, when induced in a lipotoxic state, repressed PDX-1 expression contributing to the inhibition of insulin expression and β cell dysfunction.

  17. Placental Growth Factor Administration Abolishes Placental Ischemia-Induced Hypertension.

    PubMed

    Spradley, Frank T; Tan, Adelene Y; Joo, Woo S; Daniels, Garrett; Kussie, Paul; Karumanchi, S Ananth; Granger, Joey P

    2016-04-01

    Preeclampsia is a pregnancy-specific disorder of new-onset hypertension. Unfortunately, the most effective treatment is early delivery of the fetus and placenta. Placental ischemia appears central to the pathogenesis of preeclampsia because placental ischemia/hypoxia induced in animals by reduced uterine perfusion pressure (RUPP) or in humans stimulates release of hypertensive placental factors into the maternal circulation. The anti-angiogenic factor soluble fms-like tyrosine kinase-1 (sFlt-1), which antagonizes and reduces bioavailable vascular endothelial growth factor and placental growth factor (PlGF), is elevated in RUPP rats and preeclampsia. Although PlGF and vascular endothelial growth factor are both natural ligands for sFlt-1, vascular endothelial growth factor also has high affinity to VEGFR2 (Flk-1) causing side effects like edema. PlGF is specific for sFlt-1. We tested the hypothesis that PlGF treatment reduces placental ischemia-induced hypertension by antagonizing sFlt-1 without adverse consequences to the mother or fetus. On gestational day 14, rats were randomized to 4 groups: normal pregnant or RUPP±infusion of recombinant human PlGF (180 μg/kg per day; AG31, a purified, recombinant human form of PlGF) for 5 days via intraperitoneal osmotic minipumps. On day 19, mean arterial blood pressure and plasma sFlt-1 were higher and glomerular filtration rate lower in RUPP than normal pregnant rats. Infusion of recombinant human PlGF abolished these changes seen with RUPP along with reducing oxidative stress. These data indicate that the increased sFlt-1 and reduced PlGF resulting from placental ischemia contribute to maternal hypertension. Our novel finding that recombinant human PlGF abolishes placental ischemia-induced hypertension, without major adverse consequences, suggests a strong therapeutic potential for this growth factor in preeclampsia. PMID:26831193

  18. Gene structures and promoter characteristics of interferon regulatory factor 1 (IRF-1), IRF-2 and IRF-7 from snakehead Channa argus.

    PubMed

    Jia, Weizhang; Guo, Qionglin

    2008-04-01

    Three interferon regulatory factor (IRF) genes, CaIRF-1, CaIRF-2 and CaIRF-7, and their promoters of snakehead (Channa argus) were cloned and characterized. The CaIRF-1 gene consists of ten exons, spans 4.3 kb and encodes a putative peptide of 299 aa. The CaIRF-2 gene consists of nine exons, spans 8 kb and encodes a putative peptide of 328 aa. The gene organizations of CaIRF-1 and CaIRF-2 are very similar to that of human IRF-1 and IRF-2 except more compact. Comparison of exon-intron organization of the two genes indicated a common evolutionary structure, notably within the exons encoding the DNA binding domain (DBD) of the two factors. The CaIRF-7 gene spans 4.1 kb and encodes a putative peptide of 437 aa. However, the gene organization of CaIRF-7 consisting of ten exons is different to human IRF-7a gene which has an intron in 5' UTR. Three CaIRFs share homology in N-terminal encompassing the DBD that contains a characteristic repeat of tryptophan residues. The promoters of CaIRF-1 and CaIRF-2 genes contain the conserved sites for NF-kappaB and Sp1. The gamma-IFN activation sites (GAS) were found in the promoters of CaIRF-1 and CaIRF-7. The promoter of CaIRF-7 contains conserved interferon stimulating response element (ISRE) which is characteristic of IFN-induced gene promoter, and suggests that there also exist intracellular amplifier circuit in fish IFN signal pathway. Moreover, the element GAAANN oriented in both directions is repeated in CaIRF promoter regions, which confers to further inducibility by IFN. The constitutive expression of CaIRF genes were found to increase obviously in response to induction by the known IFN-inducer poly I:C.

  19. Construction of the influenza A virus infection-induced cell-specific inflammatory regulatory network based on mutual information and optimization

    PubMed Central

    2013-01-01

    Background Influenza A virus (IAV) infection-induced inflammatory regulatory networks (IRNs) are extremely complex and dynamic. Specific biological experiments for investigating the interactions between individual inflammatory factors cannot provide a detailed and insightful multidimensional view of IRNs. Recently, data from high-throughput technologies have permitted system-level analyses. The construction of large and cell-specific IRNs from high-throughput data is essential to understanding the pathogenesis of IAV infection. Results In this study, we proposed a computational method, which combines nonlinear ordinary differential equation (ODE)-based optimization with mutual information, to construct a cell-specific optimized IRN during IAV infection by integrating gene expression data with a prior knowledge of network topology. Moreover, we used the average relative error and sensitivity analysis to evaluate the effectiveness of our proposed approach. Furthermore, from the optimized IRN, we confirmed 45 interactions between proteins in biological experiments and identified 37 new regulatory interactions and 8 false positive interactions, including the following interactions: IL1β regulates TLR3, TLR3 regulates IFN-β and TNF regulates IL6. Most of these regulatory interactions are statistically significant by Z-statistic. The functional annotations of the optimized IRN demonstrated clearly that the defense response, immune response, response to wounding and regulation of cytokine production are the pivotal processes of IAV-induced inflammatory response. The pathway analysis results from the Kyoto Encyclopaedia of Genes and Genomes (KEGG) showed that 8 pathways are enriched significantly. The 5 pathways were validated by experiments, and 3 other pathways, including the intestinal immune network for IgA production, the cytosolic DNA-sensing pathway and the allograft rejection pathway, are the predicted novel pathways involved in the inflammatory response

  20. Methylation-dependent regulation of hypoxia inducible factor-1 alpha gene expression by the transcription factor Kaiso.

    PubMed

    Pierre, Christina C; Longo, Joseph; Bassey-Archibong, Blessing I; Hallett, Robin M; Milosavljevic, Snezana; Beatty, Laura; Hassell, John A; Daniel, Juliet M

    2015-12-01

    Low oxygen tension (hypoxia) is a common characteristic of solid tumors and strongly correlates with poor prognosis and resistance to treatment. In response to hypoxia, cells initiate a cascade of transcriptional events regulated by the hypoxia inducible factor-1 (HIF-1) heterodimer. Since the oxygen-sensitive HIF-1α subunit is stabilized during hypoxia, it functions as the regulatory subunit of the protein. To date, while the mechanisms governing HIF-1α protein stabilization and function have been well studied, those governing HIF1A gene expression are not fully understood. However, recent studies have suggested that methylation of a HIF-1 binding site in the HIF1A promoter prevents its autoregulation. Here we report that the POZ-ZF transcription factor Kaiso modulates HIF1A gene expression by binding to the methylated HIF1A promoter in a region proximal to the autoregulatory HIF-1 binding site. Interestingly, Kaiso's regulation of HIF1A occurs primarily during hypoxia, which is consistent with the finding that Kaiso protein levels peak after 4 h of hypoxic incubation and return to normoxic levels after 24 h. Our data thus support a role for Kaiso in fine-tuning HIF1A gene expression after extended periods of hypoxia.

  1. Hypoxia inducible factors and the response to hypoxic stress

    PubMed Central

    Majmundar, Amar J.; Wong, Waihay J.; Simon, M. Celeste

    2011-01-01

    Oxygen (O2) is an essential nutrient that serves as a key substrate in cellular metabolism and bioenergetics. In a variety of physiological and pathological states, organisms encounter insufficient O2 availability, or hypoxia. In order to cope with this stress, evolutionarily conserved responses are engaged. In mammals, the primary transcriptional response to hypoxic stress is mediated by the Hypoxia-inducible factors (HIFs). While canonically regulated by prolyl hydroxylase domain-containing enzymes (PHDs), the HIFα subunits are intricately responsive to numerous other factors including Factor Inhibiting HIF-1α (FIH1), sirtuins, and metabolites. These transcription factors function in normal tissue homeostasis and impinge on critical aspects of disease progression and recovery. Insights from basic HIF biology are being translated into pharmaceuticals targeting the HIF pathway. PMID:20965423

  2. Genomic analysis of host - Peste des petits ruminants vaccine viral transcriptome uncovers transcription factors modulating immune regulatory pathways.

    PubMed

    Manjunath, Siddappa; Kumar, Gandham Ravi; Mishra, Bishnu Prasad; Mishra, Bina; Sahoo, Aditya Prasad; Joshi, Chaitanya G; Tiwari, Ashok K; Rajak, Kaushal Kishore; Janga, Sarath Chandra

    2015-01-01

    Peste des petits ruminants (PPR), is an acute transboundary viral disease of economic importance, affecting goats and sheep. Mass vaccination programs around the world resulted in the decline of PPR outbreaks. Sungri 96 is a live attenuated vaccine, widely used in Northern India against PPR. This vaccine virus, isolated from goat works efficiently both in sheep and goat. Global gene expression changes under PPR vaccine virus infection are not yet well defined. Therefore, in this study we investigated the host-vaccine virus interactions by infecting the peripheral blood mononuclear cells isolated from goat with PPRV (Sungri 96 vaccine virus), to quantify the global changes in the transcriptomic signature by RNA-sequencing. Viral genome of Sungri 96 vaccine virus was assembled from the PPRV infected transcriptome confirming the infection and demonstrating the feasibility of building a complete non-host genome from the blood transcriptome. Comparison of infected transcriptome with control transcriptome revealed 985 differentially expressed genes. Functional analysis showed enrichment of immune regulatory pathways under PPRV infection. Key genes involved in immune system regulation, spliceosomal and apoptotic pathways were identified to be dysregulated. Network analysis revealed that the protein - protein interaction network among differentially expressed genes is significantly disrupted in infected state. Several genes encoding TFs that govern immune regulatory pathways were identified to co-regulate the differentially expressed genes. These data provide insights into the host - PPRV vaccine virus interactome for the first time. Our findings suggested dysregulation of immune regulatory pathways and genes encoding Transcription Factors (TFs) that govern these pathways in response to viral infection. PMID:25827022

  3. Hypoxia inducible factor pathway inhibitors as anticancer therapeutics

    PubMed Central

    Burroughs, Sarah K; Kaluz, Stefan; Wang, Danzhu; Wang, Ke

    2013-01-01

    Hypoxia is a significant feature of solid tumor cancers. Hypoxia leads to a more malignant phenotype that is resistant to chemotherapy and radiation, is more invasive and has greater metastatic potential. Hypoxia activates the hypoxia inducible factor (HIF) pathway, which mediates the biological effects of hypoxia in tissues. The HIF complex acts as a transcription factor for many genes that increase tumor survival and proliferation. To date, many HIF pathway inhibitors indirectly affect HIF but there have been no clinically approved direct HIF inhibitors. This can be attributed to the complexity of the HIF pathway, as well as to the challenges of inhibiting protein–protein interactions. PMID:23573973

  4. Hypoxia and Hypoxia Inducible Factors: Diverse Roles in Liver Diseases

    PubMed Central

    Nath, Bharath; Szabo, Gyongyi

    2011-01-01

    Hypoxia has been shown to have a role in the pathogenesis of several forms of liver disease. The Hypoxia Inducible Factors (HIFs) are a family of evolutionarily conserved transcriptional regulators that affect a homeostatic response to low oxygen tension and have been identified as key mediators of angiogenesis, inflammation, and metabolism. In this review, we summarize the evidence for a role of HIFs across a range of hepatic pathophysiology. We describe regulation of the hypoxia inducible factors and review investigations that demonstrate a role for HIFs in the development of liver fibrosis, activation of innate immune pathways, hepatocellular carcinoma, as well as other liver diseases in both human disease as well as murine models. PMID:22120903

  5. Comparative analysis of transcription factor gene families from Papaver somniferum: identification of regulatory factors involved in benzylisoquinoline alkaloid biosynthesis.

    PubMed

    Agarwal, Parul; Pathak, Sumya; Lakhwani, Deepika; Gupta, Parul; Asif, Mehar Hasan; Trivedi, Prabodh Kumar

    2016-05-01

    Opium poppy (Papaver somniferum L.), known for biosynthesis of several therapeutically important benzylisoquinoline alkaloids (BIAs), has emerged as the premier organism to study plant alkaloid metabolism. The most prominent molecules produced in opium poppy include narcotic analgesic morphine, the cough suppressant codeine, the muscle relaxant papaverine and the anti-microbial agent sanguinarine and berberine. Despite several health benefits, biosynthesis of some of these molecules is very low due to tight temporal and spatial regulation of the genes committed to their biosynthesis. Transcription factors, one of the prime regulators of secondary plant product biosynthesis, might be involved in controlled biosynthesis of BIAs in P. somniferum. In this study, identification of members of different transcription factor gene families using transcriptome datasets of 10 cultivars of P. somniferum with distinct chemoprofile has been carried out. Analysis suggests that most represented transcription factor gene family in all the poppy cultivars is WRKY. Comparative transcriptome analysis revealed differential expression pattern of the members of a set of transcription factor gene families among 10 cultivars. Through analysis, two members of WRKY and one member of C3H gene family were identified as potential candidates which might regulate thebaine and papaverine biosynthesis, respectively, in poppy.

  6. Comparative analysis of transcription factor gene families from Papaver somniferum: identification of regulatory factors involved in benzylisoquinoline alkaloid biosynthesis.

    PubMed

    Agarwal, Parul; Pathak, Sumya; Lakhwani, Deepika; Gupta, Parul; Asif, Mehar Hasan; Trivedi, Prabodh Kumar

    2016-05-01

    Opium poppy (Papaver somniferum L.), known for biosynthesis of several therapeutically important benzylisoquinoline alkaloids (BIAs), has emerged as the premier organism to study plant alkaloid metabolism. The most prominent molecules produced in opium poppy include narcotic analgesic morphine, the cough suppressant codeine, the muscle relaxant papaverine and the anti-microbial agent sanguinarine and berberine. Despite several health benefits, biosynthesis of some of these molecules is very low due to tight temporal and spatial regulation of the genes committed to their biosynthesis. Transcription factors, one of the prime regulators of secondary plant product biosynthesis, might be involved in controlled biosynthesis of BIAs in P. somniferum. In this study, identification of members of different transcription factor gene families using transcriptome datasets of 10 cultivars of P. somniferum with distinct chemoprofile has been carried out. Analysis suggests that most represented transcription factor gene family in all the poppy cultivars is WRKY. Comparative transcriptome analysis revealed differential expression pattern of the members of a set of transcription factor gene families among 10 cultivars. Through analysis, two members of WRKY and one member of C3H gene family were identified as potential candidates which might regulate thebaine and papaverine biosynthesis, respectively, in poppy. PMID:26108744

  7. Social factors modulate restraint stress induced hyperthermia in mice.

    PubMed

    Watanabe, Shigeru

    2015-10-22

    Stress-induced hyperthermia (SIH) was examined in three different social conditions in mice by thermographic measurement of the body surface temperature. Placing animals in cylindrical holders induced restraint stress. I examined the effect of the social factors in SIH using the thermograph (body surface temperature). Mice restrained in the holders alone showed SIH. Mice restrained in the holders at the same time as other similarly restrained cage mates (social equality condition) showed less hyperthermia. Interestingly, restrained mice with free moving cage mates (social inequality condition) showed the highest hyperthermia. These results are consistent with a previous experiment measuring the memory-enhancing effects of stress and the stress-induced elevation of corticosterone, and suggest that social inequality enhances stress.

  8. Social factors modulate restraint stress induced hyperthermia in mice.

    PubMed

    Watanabe, Shigeru

    2015-10-22

    Stress-induced hyperthermia (SIH) was examined in three different social conditions in mice by thermographic measurement of the body surface temperature. Placing animals in cylindrical holders induced restraint stress. I examined the effect of the social factors in SIH using the thermograph (body surface temperature). Mice restrained in the holders alone showed SIH. Mice restrained in the holders at the same time as other similarly restrained cage mates (social equality condition) showed less hyperthermia. Interestingly, restrained mice with free moving cage mates (social inequality condition) showed the highest hyperthermia. These results are consistent with a previous experiment measuring the memory-enhancing effects of stress and the stress-induced elevation of corticosterone, and suggest that social inequality enhances stress. PMID:26232073

  9. Genetic and environmental factors affecting cryptic variations in gene regulatory networks

    PubMed Central

    2013-01-01

    Background Cryptic genetic variation (CGV) is considered to facilitate phenotypic evolution by producing visible variations in response to changes in the internal and/or external environment. Several mechanisms enabling the accumulation and release of CGVs have been proposed. In this study, we focused on gene regulatory networks (GRNs) as an important mechanism for producing CGVs, and examined how interactions between GRNs and the environment influence the number of CGVs by using individual-based simulations. Results Populations of GRNs were allowed to evolve under various stabilizing selections, and we then measured the number of genetic and phenotypic variations that had arisen. Our results showed that CGVs were not depleted irrespective of the strength of the stabilizing selection for each phenotype, whereas the visible fraction of genetic variation in a population decreased with increasing strength of selection. On the other hand, increasing the number of different environments that individuals encountered within their lifetime (i.e., entailing plastic responses to multiple environments) suppressed the accumulation of CGVs, whereas the GRNs with more genes and interactions were favored in such heterogeneous environments. Conclusions Given the findings that the number of CGVs in a population was largely determined by the size (order) of GRNs, we propose that expansion of GRNs and adaptation to novel environments are mutually facilitating and sustainable sources of evolvability and hence the origins of biological diversity and complexity. PMID:23622056

  10. Inducer effect on the complex formation between rat liver nuclear proteins and cytochrome P450 2B gene regulatory elements.

    PubMed

    Duzhak, T G; Schwartz, E I; Gulyaeva, L F; Lyakhovich, V V

    2002-09-01

    DNA gel retardation assay has been applied to the investigation of complexes between rat liver nuclear proteins and Barbie box positive regulatory element of cytochrome P450 2B (CYP2B) genes. The intensities of B1 and B2 bands detected in the absence of an inducer increased after 30 min protein incubation with phenobarbital (PB) or triphenyldioxane (TPD), but not with 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOPOB). In addition, a new complex (B3 band) was for the first time detected under induction by PB, TPD, and TCPOPOB. Increase in the incubation time up to 2 h facilitated the formation of other new complexes (B4 and B5 bands), which were detected only in the presence of TPD. The use of [3H]TPD in hybridization experiments revealed that this inducer, capable of binding to Barbie box DNA, is also present in B4 and B5 complexes. It is probable that the investigated compounds activate the same proteins at the initial induction steps, which correlates with the formation of B1, B2, and B3 complexes. The further induction step might be inducer-specific, as indicated by the formation of B4 and B5 complexes in the presence of TPD only. Thus, the present data suggest the possibility of specific gene activation signaling pathways that are dependent on a particular inducer. PMID:12387719

  11. Ciliary dyslexia candidate genes DYX1C1 and DCDC2 are regulated by Regulatory Factor X (RFX) transcription factors through X-box promoter motifs

    PubMed Central

    Tammimies, Kristiina; Bieder, Andrea; Lauter, Gilbert; Sugiaman-Trapman, Debora; Torchet, Rachel; Hokkanen, Marie-Estelle; Burghoorn, Jan; Castrén, Eero; Kere, Juha; Tapia-Páez, Isabel; Swoboda, Peter

    2016-01-01

    DYX1C1, DCDC2, and KIAA0319 are three of the most replicated dyslexia candidate genes (DCGs). Recently, these DCGs were implicated in functions at the cilium. Here, we investigate the regulation of these DCGs by Regulatory Factor X transcription factors (RFX TFs), a gene family known for transcriptionally regulating ciliary genes. We identify conserved X-box motifs in the promoter regions of DYX1C1, DCDC2, and KIAA0319 and demonstrate their functionality, as well as the ability to recruit RFX TFs using reporter gene and electrophoretic mobility shift assays. Furthermore, we uncover a complex regulation pattern between RFX1, RFX2, and RFX3 and their significant effect on modifying the endogenous expression of DYX1C1 and DCDC2 in a human retinal pigmented epithelial cell line immortalized with hTERT (hTERT-RPE1). In addition, induction of ciliogenesis increases the expression of RFX TFs and DCGs. At the protein level, we show that endogenous DYX1C1 localizes to the base of the cilium, whereas DCDC2 localizes along the entire axoneme of the cilium, thereby validating earlier localization studies using overexpression models. Our results corroborate the emerging role of DCGs in ciliary function and characterize functional noncoding elements, X-box promoter motifs, in DCG promoter regions, which thus can be targeted for mutation screening in dyslexia and ciliopathies associated with these genes.—Tammimies, K., Bieder, A., Lauter, G., Sugiaman-Trapman, D., Torchet, R., Hokkanen, M.-E., Burghoorn, J., Castrén, E., Kere, J., Tapia-Páez, I., Swoboda, P. Ciliary dyslexia candidate genes DYX1C1 and DCDC2 are regulated by Regulatory Factor (RF) X transcription factors through X-box promoter motifs. PMID:27451412

  12. Dexamethasone impairs hypoxia-inducible factor-1 function

    SciTech Connect

    Wagner, A.E.; Huck, G.; Stiehl, D.P.; Jelkmann, W.; Hellwig-Buergel, T.

    2008-07-25

    Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric transcription-factor composed of {alpha}- and {beta}-subunits. HIF-1 is not only necessary for the cellular adaptation to hypoxia, but it is also involved in inflammatory processes and wound healing. Glucocorticoids (GC) are therapeutically used to suppress inflammatory responses. Herein, we investigated whether GC modulate HIF-1 function using GC receptor (GR) possessing (HepG2) and GR deficient (Hep3B) human hepatoma cell cultures as model systems. Dexamethasone (DEX) treatment increased HIF-1{alpha} levels in the cytosol of HepG2 cells, while nuclear HIF-1{alpha} levels and HIF-1 DNA-binding was reduced. In addition, DEX dose-dependently lowered the hypoxia-induced luciferase activity in a reporter gene system. DEX suppressed the hypoxic stimulation of the expression of the HIF-1 target gene VEGF (vascular endothelial growth factor) in HepG2 cultures. DEX did not reduce hypoxically induced luciferase activity in HRB5 cells, a Hep3B derivative lacking GR. Transient expression of the GR in HRB5 cells restored the susceptibility to DEX. Our study discloses the inhibitory action of GC on HIF-1 dependent gene expression, which may be important with respect to the impaired wound healing in DEX-treated patients.

  13. A trans-acting Variant within the Transcription Factor RIM101 Interacts with Genetic Background to Determine its Regulatory Capacity.

    PubMed

    Read, Timothy; Richmond, Phillip A; Dowell, Robin D

    2016-01-01

    Most genetic variants associated with disease occur within regulatory regions of the genome, underscoring the importance of defining the mechanisms underlying differences in regulation of gene expression between individuals. We discovered a pair of co-regulated, divergently oriented transcripts, AQY2 and ncFRE6, that are expressed in one strain of Saccharomyces cerevisiae, ∑1278b, but not in another, S288c. By combining classical genetics techniques with high-throughput sequencing, we identified a trans-acting single nucleotide polymorphism within the transcription factor RIM101 that causes the background-dependent expression of both transcripts. Subsequent RNA-seq experiments revealed that RIM101 regulates many more targets in S288c than in ∑1278b and that deletion of RIM101 in both backgrounds abrogates the majority of differential expression between the strains. Strikingly, only three transcripts undergo a significant change in expression after swapping RIM101 alleles between backgrounds, implying that the differences in the RIM101 allele lead to a remarkably focused transcriptional response. However, hundreds of RIM101-dependent targets undergo a subtle but consistent shift in expression in the S288c RIM101-swapped strain, but not its ∑1278b counterpart. We conclude that ∑1278b may harbor a variant(s) that buffers against widespread transcriptional dysregulation upon introduction of a non-native RIM101 allele, emphasizing the importance of accounting for genetic background when assessing the impact of a regulatory variant.

  14. Environment-induced epigenetic reprogramming in genomic regulatory elements in smoking mothers and their children.

    PubMed

    Bauer, Tobias; Trump, Saskia; Ishaque, Naveed; Thürmann, Loreen; Gu, Lei; Bauer, Mario; Bieg, Matthias; Gu, Zuguang; Weichenhan, Dieter; Mallm, Jan-Philipp; Röder, Stefan; Herberth, Gunda; Takada, Eiko; Mücke, Oliver; Winter, Marcus; Junge, Kristin M; Grützmann, Konrad; Rolle-Kampczyk, Ulrike; Wang, Qi; Lawerenz, Christian; Borte, Michael; Polte, Tobias; Schlesner, Matthias; Schanne, Michaela; Wiemann, Stefan; Geörg, Christina; Stunnenberg, Hendrik G; Plass, Christoph; Rippe, Karsten; Mizuguchi, Junichiro; Herrmann, Carl; Eils, Roland; Lehmann, Irina

    2016-03-01

    Epigenetic mechanisms have emerged as links between prenatal environmental exposure and disease risk later in life. Here, we studied epigenetic changes associated with maternal smoking at base pair resolution by mapping DNA methylation, histone modifications, and transcription in expectant mothers and their newborn children. We found extensive global differential methylation and carefully evaluated these changes to separate environment associated from genotype-related DNA methylation changes. Differential methylation is enriched in enhancer elements and targets in particular "commuting" enhancers having multiple, regulatory interactions with distal genes. Longitudinal whole-genome bisulfite sequencing revealed that DNA methylation changes associated with maternal smoking persist over years of life. Particularly in children prenatal environmental exposure leads to chromatin transitions into a hyperactive state. Combined DNA methylation, histone modification, and gene expression analyses indicate that differential methylation in enhancer regions is more often functionally translated than methylation changes in promoters or non-regulatory elements. Finally, we show that epigenetic deregulation of a commuting enhancer targeting c-Jun N-terminal kinase 2 (JNK2) is linked to impaired lung function in early childhood.

  15. Expression of hypoxia-inducible factor 1 alpha and oligodendrocyte lineage gene-1 in cultured brain slices after oxygen-glucose deprivation☆

    PubMed Central

    Cui, Hong; Han, Weijuan; Yang, Lijun; Chang, Yanzhong

    2013-01-01

    Oligodendrocyte lineage gene-1 expressed in oligodendrocytes may trigger the repair of neuronal myelin impairment, and play a crucial role in myelin repair. Hypoxia-inducible factor 1α, a transcription factor, is of great significance in premature infants with hypoxic-ischemic brain damage. There is little evidence of direct regulatory effects of hypoxia-inducible factor 1α on oligodendrocyte lineage gene-1. In this study, brain slices of Sprague-Dawley rats were cultured and subjected to oxygen-glucose deprivation. Then, slices were transfected with hypoxia-inducible factor 1α or oligodendrocyte lineage gene-1. The expression levels of hypoxia-inducible factor 1α and oligodendrocyte lineage gene-1 were significantly up-regulated in rat brains prior to transfection, as detected by immunohistochemical staining. Eight hours after transfection of slices with hypoxia-inducible factor 1α, oligodendrocyte lineage gene-1 expression was upregulated, and reached a peak 24 hours after transfection. Oligodendrocyte lineage gene-1 transfection induced no significant differences in hypoxia-inducible factor 1α levels in rat brain tissues with oxygen-glucose deprivation. These experimental findings indicate that hypoxia-inducible factor 1α can regulate oligodendrocyte lineage gene-1 expression in hypoxic brain tissue, thus repairing the neural impairment. PMID:25206673

  16. Regulatory T cells ameliorate acetaminophen-induced immune-mediated liver injury.

    PubMed

    Wang, Xuefu; Sun, Rui; Chen, Yongyan; Lian, Zhe-Xiong; Wei, Haiming; Tian, Zhigang

    2015-04-01

    The contribution of innate immune cells to acetaminophen (APAP)-induced liver injury has been extensively investigated. However, the roles of T cell populations among adaptive immune cells in APAP-induced liver injury remain to be elucidated. Herein, we found that distinct CD4(+) T cell subsets but not CD8(+) T cells modulated APAP-induced liver injury in mice. After APAP challenge, more CD62L(low)CD44(hi)CD4(+) T cells appeared in the liver, accompanied by increased IFN-γ. The removal of CD4(+) T cells by either antibody depletion or genetic deficiency markedly compromised pro-inflammatory cytokine levels and ameliorated liver injury. Meanwhile, we also found that the frequency and absolute number of Treg cells also increased. Treg cell depletion increased hepatic CD62L(low)CD44(hi)CD4(+) T cells, augmented pro-inflammatory cytokines, and exacerbated liver injury, while adoptive transfer of Treg cells ameliorated APAP-induced liver injury. Furthermore, the recruitment of Treg cells into the liver through specific expression of CXCL10 in the liver could ameliorate APAP-induced liver injury. Our investigation suggests that Th1 and Treg subsets are involved in regulating APAP-induced liver injury. Thus, modulating the Th1/Treg balance may be an effective strategy to prevent and/or treat APAP-induced liver injury.

  17. Drug-induced lupus: Including anti-tumour necrosis factor and interferon induced.

    PubMed

    Araújo-Fernández, S; Ahijón-Lana, M; Isenberg, D A

    2014-05-01

    Drug-induced lupus erythematosus is defined as a syndrome with clinical and serological features similar to systemic lupus erythematosus that is temporally related to continuous drug exposure and which resolves after discontinuation of this drug. More than 90 drugs, including biological modulators such as tumour necrosis factor-α inhibitors and interferons, have been identified as likely 'culprits'. While there are no standard diagnostic criteria for drug-induced lupus erythematosus, guidelines that can help to distinguish drug-induced lupus erythematosus from systemic lupus erythematosus have been proposed and several different patterns of drug-induced lupus erythematosus are emerging. Distinguishing drug-induced lupus erythematosus from systemic lupus erythematosus is important because the prognosis of drug-induced lupus erythematosus is usually good when the drug is withdrawn. This review discusses the differences between drug-induced lupus erythematosus and systemic lupus erythematosus, the mechanisms of action of drug-induced lupus erythematosus and drugs that are usually associated with drug-induced lupus erythematosus, with particular focus on the biological treatments.

  18. Nuclear Factor of Activated T Cells Transcription Factor Nfatp Controls Superantigen-Induced Lethal Shock

    PubMed Central

    Tsytsykova, Alla V.; Goldfeld, Anne E.

    2000-01-01

    Tumor necrosis factor α (TNF-α) is the key mediator of superantigen-induced T cell lethal shock. Here, we show that nuclear factor of activated T cells transcription factor, NFATp, controls susceptibility to superantigen-induced lethal shock in mice through its activation of TNF-α gene transcription. In NFATp-deficient mice, T cell stimulation leads to delayed induction and attenuation of TNF-α mRNA levels, decreased TNF-α serum levels, and resistance to superantigen-induced lethal shock. By contrast, after lipopolysaccharide (LPS) challenge, serum levels of TNF-α and susceptibility to shock are unaffected. These results demonstrate that NFATp is an essential activator of immediate early TNF-α gene expression in T cells and they present in vivo evidence of the inducer- and cell type–specific regulation of TNF-α gene expression. Furthermore, they suggest NFATp as a potential selective target in the treatment of superantigen-induced lethal shock. PMID:10952728

  19. Asynchronous combinatorial action of four regulatory factors activates Bcl11b for T cell commitment.

    PubMed

    Kueh, Hao Yuan; Yui, Mary A; Ng, Kenneth K H; Pease, Shirley S; Zhang, Jingli A; Damle, Sagar S; Freedman, George; Siu, Sharmayne; Bernstein, Irwin D; Elowitz, Michael B; Rothenberg, Ellen V

    2016-08-01

    During T cell development, multipotent progenitors relinquish competence for other fates and commit to the T cell lineage by turning on Bcl11b, which encodes a transcription factor. To clarify lineage commitment mechanisms, we followed developing T cells at the single-cell level using Bcl11b knock-in fluorescent reporter mice. Notch signaling and Notch-activated transcription factors collaborate to activate Bcl11b expression irrespectively of Notch-dependent proliferation. These inputs work via three distinct, asynchronous mechanisms: an early locus 'poising' function dependent on TCF-1 and GATA-3, a stochastic-permissivity function dependent on Notch signaling, and a separate amplitude-control function dependent on Runx1, a factor already present in multipotent progenitors. Despite their necessity for Bcl11b expression, these inputs act in a stage-specific manner, providing a multitiered mechanism for developmental gene regulation. PMID:27376470

  20. Bacterial β-(1,3)-glucan prevents DSS-induced IBD by restoring the reduced population of regulatory T cells.

    PubMed

    Lee, Kwang-Ho; Park, Min; Ji, Kon-Young; Lee, Hwa-Youn; Jang, Ji-Hun; Yoon, Il-Joo; Oh, Seung-Su; Kim, Su-Man; Jeong, Yun-Hwa; Yun, Chul-Ho; Kim, Mi-Kyoung; Lee, In-Young; Choi, Ha-Rim; Ko, Ki-sung; Kang, Hyung-Sik

    2014-10-01

    Bacterial β-(1,3)-glucan has more advantages in terms of cost, yield and efficiency than that derived from mushrooms, plants, yeasts and fungi. We have previously developed a novel and high-yield β-(1,3)-glucan produced by Agrobacterium sp. R259. This study aimed to elucidate the functional mechanism and therapeutic efficacy of bacterial β-(1,3)-glucan in dextran sulfate sodium (DSS)-induced inflammatory bowel disease (IBD).Mice were orally pretreated with bacterial β-(1,3)-glucan at daily doses of 2.5 or 5mg/kg for 2 weeks. After 6 days of DSS treatment, clinical assessment of IBD severity and expression of pro-inflammatory cytokines were evaluated. In vivo cell proliferation was examined by immunohistochemistry using Ki-67 and ER-TR7 antibodies. The frequency of regulatory T cells (Tregs) was analyzed by flow cytometry. Natural killer (NK) activity and IgA level were evaluated using NK cytotoxicity assay and ELISA.The deterioration of body weight gain, colonic architecture, disease score and histological score was recovered in DSS-induced IBD mice when pretreated with bacterial β-(1,3)-glucan. The recruitment of macrophages and the gene expression of proinflammatory cytokines, such as IL-1β, IL-6 and IL-17A/F, were markedly decreased in the colon of β-(1,3)-glucan-pretreated mice. β-(1,3)-Glucan induced the recovery of Tregs in terms of their frequency in DSS-induced IBD mice. Intriguingly, β-(1,3)-glucan reversed the functional defects of NK cells and excessive IgA production in DSS-induced IBD mice.We conclude that bacterial β-(1,3)-glucan prevented the progression of DSS-induced IBD by recovering the reduction of Tregs, functional defect of NK cells and excessive IgA production.

  1. Transforming growth factor-alpha-induced transcriptional activation of the vascular permeability factor (VPF/VEGF) gene requires AP-2-dependent DNA binding and transactivation.

    PubMed Central

    Gille, J; Swerlick, R A; Caughman, S W

    1997-01-01

    The endothelial cell-specific mitogen vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) represents a central regulator of cutaneous angiogenesis. Increased VPF/VEGF expression has recently been reported in psoriatic skin and healing wounds, both conditions in which transforming growth factor-alpha (TGF alpha) and its ligand, the epidermal growth factor receptor, are markedly up-regulated. Since TGF alpha strongly induces VPF/VEGF synthesis in keratinocytes, TGF alpha-mediated VPF/VEGF expression is likely to play a significant role in the initiation and maintenance of increased vascular hyperpermeability and hyperproliferation in skin biology. The objectives of the present studies were to determine the molecular mechanisms responsible for TGF alpha-induced transcriptional activation of the VPF/VEGF gene. We have identified a GC-rich TGF alpha-responsive region between -88 bp and -65 bp of the VPF/VEGF promoter that is necessary for constitutive and TGF alpha-inducible transcriptional activation. In electrophoretic mobility shift assays, this region binds Sp1-dependent protein complexes constitutively and an additional TGF alpha-inducible protein complex that is distinct from Sp1 protein. Both AP-2 and Egr-1 transcription factors were detected as components of the TGF alpha-inducible protein complex in supershift EMSA studies. In co-transfection studies, an AP-2 but not an Egr-1 expression vector activated VPF/VEGF transcription, thus indicating that AP-2 protein is functionally important in TGF alpha-induced VPF/VEGF gene expression. By clarifying regulatory mechanisms that are critical for angiogenic processes in the skin, these studies may form the basis for new therapeutic strategies to modulate VPF/VEGF expression in cutaneous inflammation and wound healing. PMID:9049304

  2. Gallium induces the production of virulence factors in Pseudomonas aeruginosa.

    PubMed

    García-Contreras, Rodolfo; Pérez-Eretza, Berenice; Lira-Silva, Elizabeth; Jasso-Chávez, Ricardo; Coria-Jiménez, Rafael; Rangel-Vega, Adrián; Maeda, Toshinari; Wood, Thomas K

    2014-02-01

    The novel antimicrobial gallium is a nonredox iron III analogue with bacteriostatic and bactericidal properties, effective for the treatment of Pseudomonas aeruginosa in vitro and in vivo in mouse and rabbit infection models. It interferes with iron metabolism, transport, and presumably its homeostasis. As gallium exerts its antimicrobial effects by competing with iron, we hypothesized that it ultimately will lead cells to an iron deficiency status. As iron deficiency promotes the expression of virulence factors in vitro and promotes the pathogenicity of P. aeruginosa in animal models, it is anticipated that treatment with gallium will also promote the production of virulence factors. To test this hypothesis, the reference strain PA14 and two clinical isolates from patients with cystic fibrosis were exposed to gallium, and their production of pyocyanin, rhamnolipids, elastase, alkaline protease, alginate, pyoverdine, and biofilm was determined. Gallium treatment induced the production of all the virulence factors tested in the three strains except for pyoverdine. In addition, as the Ga-induced virulence factors are quorum sensing controlled, co-administration of Ga and the quorum quencher brominated furanone C-30 was assayed, and it was found that C-30 alleviated growth inhibition from gallium. Hence, adding both C-30 and gallium may be more effective in the treatment of P. aeruginosa infections.

  3. Hematopoietic growth factors in drug-induced agranulocytosis.

    PubMed

    Pavithran, K; Thomas, M

    2002-05-01

    Drug-induced agranulocytosis (DIA) is a potentially fatal disorder. Hematopoietic growth factors have been used in the treatment of DIA. We report nine cases of DIA treated with granulocyte macrophage - colony stimulating factor (GM-CSF) in a dose of 300 microg/day. All the patients had evidence of systemic infection. Mean time to reach an absolute neutrophil count of 0.5 x 10(9)/L was three days. One patient succumbed to the disease. The cause of death was multiorgan failure. No adverse events were observed with GM-CSF. We conclude that hematopoietic growth factors are useful in shortening the period of neutropenia and reducing morbidity and mortality in these patients.

  4. Transcription factor NFAT1 controls allergic contact hypersensitivity through regulation of activation induced cell death program

    PubMed Central

    Kwon, Ho-Keun; Kim, Gi-Cheon; Hwang, Ji Sun; Kim, Young; Chae, Chang-Suk; Nam, Jong Hee; Jun, Chang-Duk; Rudra, Dipayan; Surh, Charles D.; Im, Sin-Hyeog

    2016-01-01

    Allergic contact hypersensitivity (CHS) is an inflammatory skin disease mediated by allergen specific T cells. In this study, we investigated the role of transcription factor NFAT1 in the pathogenesis of contact hypersensitivity. NFAT1 knock out (KO) mice spontaneously developed CHS-like skin inflammation in old age. Healthy young NFAT1 KO mice displayed enhanced susceptibility to hapten-induced CHS. Both CD4+ and CD8+ T cells from NFAT1 KO mice displayed hyper-activated properties and produced significantly enhanced levels of inflammatory T helper 1(Th1)/Th17 type cytokines. NFAT1 KO T cells were more resistant to activation induced cell death (AICD), and regulatory T cells derived from these mice showed a partial defect in their suppressor activity. NFAT1 KO T cells displayed a reduced expression of apoptosis associated BCL-2/BH3 family members. Ectopic expression of NFAT1 restored the AICD defect in NFAT1 KO T cells and increased AICD in normal T cells. Recipient Rag2−/− mice transferred with NFAT1 KO T cells showed more severe CHS sensitivity due to a defect in activation induced hapten-reactive T cell apoptosis. Collectively, our results suggest the NFAT1 plays a pivotal role as a genetic switch in CD4+/CD8+ T cell tolerance by regulating AICD process in the T cell mediated skin inflammation. PMID:26777750

  5. Intranasal immunization with heat shock protein 60 induces CD4(+) CD25(+) GARP(+) and type 1 regulatory T cells and inhibits early atherosclerosis.

    PubMed

    Zhong, Y; Tang, H; Wang, X; Zeng, Q; Liu, Y; Zhao, X I; Yu, K; Shi, H; Zhu, R; Mao, X

    2016-03-01

    Atherosclerosis is an autoimmune inflammatory disease involving both innate and adaptive immune mechanisms. Immune tolerance induction may have therapeutic potential for the suppression of atherosclerosis. Current interest is directed towards mucosal tolerance induction, especially nasal tolerance. Previous studies have shown that heat shock protein 60 (HSP60) is recognized as an important autoantigen in atherosclerosis, and nasal or oral HSP60 can induce tolerance and ameliorate atherosclerosis by inducing several subsets of regulatory T cells (Tregs ) such as latency-associated peptide (LAP)(+) and forkhead box transcription factor 3 (FoxP3)(+) Tregs. However, little is known regarding the detailed mechanisms of nasal tolerance. Here, we again investigated the impact of nasal HSP60 on atherosclerosis and the mechanisms underlying the anti-atherosclerosis responses. We found that nasal HSP60 caused a significant 33·6% reduction in plaque size at the aortic root in the early stages of atherosclerosis (P < 0·001). Notably, a significant increase in activated CD4(+) CD25(+) glycoprotein A repetitions predominant (GARP)(+) Tregs, type 1 Tregs (Tr1 cells), and CD4(+) CD25(+) FoxP3(+) Tregs, as well as a marked decrease in the numbers of type 1 and 17 T helper cells was detected in the spleens and cervical lymph nodes of HSP60-treated mice. Moreover, nasal HSP60 increases the production of transforming growth factor (TGF)-β and interleukin (IL)-10 and decreases the secretion of IFN-γ and IL-17. Interestingly, the atheroprotective role of nasal HSP60 treatment was abrogated partly by the neutralization of IL-10. Our findings show that nasal administration of HSP60 can attenuate atherosclerotic formation by inducing GARP(+) Tregs, Tr1 cells and FoxP3(+) Tregs, and that these Tregs maintain immune homeostasis by secreting IL-10 and TGF-β.

  6. Myeloid Dendritic Cells (DCs) of Mice Susceptible to Paracoccidioidomycosis Suppress T Cell Responses whereas Myeloid and Plasmacytoid DCs from Resistant Mice Induce Effector and Regulatory T Cells

    PubMed Central

    Pina, Adriana; Frank de Araujo, Eliseu; Felonato, Maíra; Loures, Flávio V.; Feriotti, Claudia; Bernardino, Simone; Barbuto, José Alexandre M.

    2013-01-01

    The protective adaptive immune response in paracoccidioidomycosis, a mycosis endemic among humans, is mediated by T cell immunity, whereas impaired T cell responses are associated with severe, progressive disease. The early host response to Paracoccidioides brasiliensis infection is not known since the disease is diagnosed at later phases of infection. Our laboratory established a murine model of infection where susceptible mice reproduce the severe disease, while resistant mice develop a mild infection. This work aimed to characterize the influence of dendritic cells in the innate and adaptive immunity of susceptible and resistant mice. We verified that P. brasiliensis infection induced in bone marrow-derived dendritic cells (DCs) of susceptible mice a prevalent proinflammatory myeloid phenotype that secreted high levels of interleukin-12 (IL-12), tumor necrosis factor alpha, and IL-β, whereas in resistant mice, a mixed population of myeloid and plasmacytoid DCs secreting proinflammatory cytokines and expressing elevated levels of secreted and membrane-bound transforming growth factor β was observed. In proliferation assays, the proinflammatory DCs from B10.A mice induced anergy of naïve T cells, whereas the mixed DC subsets from resistant mice induced the concomitant proliferation of effector and regulatory T cells (Tregs). Equivalent results were observed during pulmonary infection. The susceptible mice displayed preferential expansion of proinflammatory myeloid DCs, resulting in impaired proliferation of effector T cells. Conversely, the resistant mice developed myeloid and plasmacytoid DCs that efficiently expanded gamma interferon-, IL-4-, and IL-17-positive effector T cells associated with increased development of Tregs. Our work highlights the deleterious effect of excessive innate proinflammatory reactions and provides new evidence for the importance of immunomodulation during pulmonary paracoccidioidomycosis. PMID:23340311

  7. Estrogen induced concentration dependent differential gene expression in human breast cancer (MCF7) cells: Role of transcription factors

    SciTech Connect

    Chandrasekharan, Sabarinath; Kandasamy, Krishna Kumar; Dayalan, Pavithra; Ramamurthy, Viraragavan

    2013-08-02

    Highlights: •Estradiol (E2) at low dose induced cell proliferation in breast cancer cells. •E2 at high concentration induced cell stress in breast cancer cells. •Estrogen receptor physically interacts only with a few transcription factors. •Differential expression of genes with Oct-1 binding sites increased under stress. •Transcription factor binding sites showed distinct spatial distribution on genes. -- Abstract: Background: Breast cancer cells respond to estrogen in a concentration dependent fashion, resulting in proliferation or apoptosis. The mechanism of this concentration dependent differential outcome is not well understood yet. Methodology: Meta-analysis of the expression data of MCF7 cells treated with low (1 nM) or high (100 nM) dose of estradiol (E2) was performed. We identified genes differentially expressed at the low or the high dose, and examined the nature of regulatory elements in the vicinity of these genes. Specifically, we looked for the difference in the presence, abundance and spatial distribution of binding sites for estrogen receptor (ER) and selected transcription factors (TFs) in the genomic region up to 25 kb upstream and downstream from the transcription start site (TSS) of these genes. Results: It was observed that at high dose E2 induced the expression of stress responsive genes, while at low dose, genes involved in cell cycle were induced. We found that the occurrence of transcription factor binding regions (TFBRs) for certain factors such as Sp1 and SREBP1 were higher on regulatory regions of genes expressed at low dose. At high concentration of E2, genes with a higher frequency of Oct-1 binding regions were predominantly involved. In addition, there were differences in the spatial distribution pattern of the TFBRs in the genomic regions among the two sets of genes. Discussion: E2 induced predominantly proliferative/metabolic response at low concentrations; but at high concentration, stress–rescue responses were induced

  8. Deacetylase inhibition increases regulatory T cell function and decreases incidence and severity of collagen-induced arthritis.

    PubMed

    Saouaf, Sandra J; Li, Bin; Zhang, Geng; Shen, Yuan; Furuuchi, Narumi; Hancock, Wayne W; Greene, Mark I

    2009-10-01

    Collagen-induced arthritis (CIA) is an established mouse model of disease with hallmarks of clinical rheumatoid arthritis. Histone/protein deacetylase inhibitors (HDACi) are known to inhibit the pathogenesis of CIA and other models of autoimmune disease, although the mechanisms responsible are unclear. Regulatory T cell (Treg) function is defective in rheumatoid arthritis. FOXP3 proteins in Tregs are present in a dynamic protein complex containing histone acetyltransferase and HDAC enzymes, and FOXP3 itself is acetylated on lysine residues. We therefore investigated the effects of HDACi therapy on regulatory T cell function in the CIA model. Administration of an HDACi, valproic acid (VPA), significantly decreased disease incidence (p<0.005) and severity (p<0.03) in CIA. In addition, VPA treatment increased both the suppressive function of CD4(+)CD25(+) Tregs (p<0.04) and the numbers of CD25(+)FOXP3(+) Tregs in vivo. Hence, clinically approved HDACi such as VPA may limit autoimmune disease in vivo through effects on the production and function of FOXP3(+) Treg cells. PMID:19577564

  9. Deacetylase Inhibition Increases Regulatory T Cell Function and Decreases Incidence and Severity of Collagen-induced Arthritis

    PubMed Central

    Saouaf, Sandra J.; Li, Bin; Zhang, Geng; Shen, Yuan; Furuuchi, Narumi; Hancock, Wayne W.; Greene, Mark I.

    2009-01-01

    Collagen-induced arthritis (CIA) is an established mouse model of disease with hallmarks of clinical rheumatoid arthritis. Histone/protein deacetylase inhibitors (HDACi) are known to inhibit the pathogenesis of CIA and other models of autoimmune disease, although the mechanisms responsible are unclear. Regulatory T cell (Treg) function is defective in rheumatoid arthritis. FOXP3 proteins in Tregs are present in a dynamic protein complex containing histone acetyltransferase and HDAC enzymes, and FOXP3 itself is acetylated on lysine residues. We therefore investigated the effects of HDACi therapy on regulatory T cell function in the CIA model. Administration of an HDACi, valproic acid (VPA), significantly decreased disease incidence (p<0.005) and severity (p<0.03) in CIA. In addition, VPA treatment increased both the suppressive function of CD4+CD25+ Tregs (p<0.04) and the numbers of CD25+FOXP3+ Tregs in vivo. Hence, clinically approved HDACi such as VPA may limit autoimmune disease in vivo through effects on the production and function of FOXP3+ Treg cells. PMID:19577564

  10. Hsp65-producing Lactococcus lactis prevents experimental autoimmune encephalomyelitis in mice by inducing CD4+LAP+ regulatory T cells.

    PubMed

    Rezende, Rafael M; Oliveira, Rafael P; Medeiros, Samara R; Gomes-Santos, Ana C; Alves, Andrea C; Loli, Flávia G; Guimarães, Mauro A F; Amaral, Sylvia S; da Cunha, André P; Weiner, Howard L; Azevedo, Vasco; Miyoshi, Anderson; Faria, Ana M C

    2013-02-01

    Heat shock proteins (Hsps) participate in the cellular response to stress and they are hiperexpressed in inflammatory conditions. They are also known to play a major role in immune modulation, controlling, for instance, autoimmune responses. In this study, we showed that oral administration of a recombinant Lactococcus lactis strain that produces and releases LPS-free Hsp65 prevented the development of experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice. This was confirmed by the reduced inflammatory cell infiltrate and absence of injury signs in the spinal cord. The effect was associated with reduced IL-17 and increased IL-10 production in mesenteric lymph node and spleen cell cultures. Hsp65-producing-L. lactis-fed mice had a remarkable increase in the number of natural and inducible CD4+Foxp3+ regulatory T (Treg) cells and CD4+LAP+ (Latency-associated peptide) Tregs - which express the membrane-bound TGF-β - in spleen, inguinal and mesenteric lymph nodes as well as in spinal cord. Moreover, many Tregs co-expressed Foxp3 and LAP. In vivo depletion of LAP+ cells abrogated the effect of Hsp65-producing L. lactis in EAE prevention and worsened disease in medium-fed mice. Thus, Hsp65-L.lactis seems to boost this critical regulatory circuit involved in controlling EAE development in mice. PMID:22939403

  11. Hsp65-producing Lactococcus lactis prevents experimental autoimmune encephalomyelitis in mice by inducing CD4+LAP+ regulatory T cells

    PubMed Central

    Rezende, Rafael M.; Oliveira, Rafael P.; Medeiros, Samara R.; Gomes-Santos, Ana C.; Alves, Andrea C.; Loli, Flávia G.; Guimarães, Mauro A.F.; Amaral, Sylvia S.; da Cunha, André P.; Weiner, Howard L.; Azevedo, Vasco; Miyoshi, Anderson; Faria, Ana M.C.

    2013-01-01

    Heat shock proteins (Hsps) participate in the cellular response to stress and they are hiperexpressed in inflammatory conditions. They are also known to play a major role in immune modulation, controlling, for instance, autoimmune responses. In this study, we showed that oral administration of a recombinant Lactococcus lactis strain that produces and releases LPS-free Hsp65 prevented the development of experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice. This was confirmed by the reduced inflammatory cell infiltrate and absence of injury signs in the spinal cord. The effect was associated with reduced IL-17 and increased IL-10 production in mesenteric lymph node and spleen cell cultures. Hsp65-producing-L. lactis-fed mice had a remarkable increase in the number of natural and inducible CD4+Foxp3+ regulatory T (Treg) cells and CD4+LAP+ (Latency-associated peptide) Tregs - which express the membrane-bound TGF-β - in spleen, inguinal and mesenteric lymph nodes as well as in spinal cord. Moreover, many Tregs co-expressed Foxp3 and LAP. In vivo depletion of LAP+ cells abrogated the effect of Hsp65-producing L. lactis in EAE prevention and worsened disease in medium-fed mice. Thus, Hsp65-L.lactis seems to boost this critical regulatory circuit involved in controlling EAE development in mice. PMID:22939403

  12. Hsp65-producing Lactococcus lactis prevents experimental autoimmune encephalomyelitis in mice by inducing CD4+LAP+ regulatory T cells.

    PubMed

    Rezende, Rafael M; Oliveira, Rafael P; Medeiros, Samara R; Gomes-Santos, Ana C; Alves, Andrea C; Loli, Flávia G; Guimarães, Mauro A F; Amaral, Sylvia S; da Cunha, André P; Weiner, Howard L; Azevedo, Vasco; Miyoshi, Anderson; Faria, Ana M C

    2013-02-01

    Heat shock proteins (Hsps) participate in the cellular response to stress and they are hiperexpressed in inflammatory conditions. They are also known to play a major role in immune modulation, controlling, for instance, autoimmune responses. In this study, we showed that oral administration of a recombinant Lactococcus lactis strain that produces and releases LPS-free Hsp65 prevented the development of experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice. This was confirmed by the reduced inflammatory cell infiltrate and absence of injury signs in the spinal cord. The effect was associated with reduced IL-17 and increased IL-10 production in mesenteric lymph node and spleen cell cultures. Hsp65-producing-L. lactis-fed mice had a remarkable increase in the number of natural and inducible CD4+Foxp3+ regulatory T (Treg) cells and CD4+LAP+ (Latency-associated peptide) Tregs - which express the membrane-bound TGF-β - in spleen, inguinal and mesenteric lymph nodes as well as in spinal cord. Moreover, many Tregs co-expressed Foxp3 and LAP. In vivo depletion of LAP+ cells abrogated the effect of Hsp65-producing L. lactis in EAE prevention and worsened disease in medium-fed mice. Thus, Hsp65-L.lactis seems to boost this critical regulatory circuit involved in controlling EAE development in mice.

  13. Regulatory role of the 90-kDa-heat-shock protein (Hsp90) and associated factors on gene expression.

    PubMed

    Erlejman, Alejandra G; Lagadari, Mariana; Toneatto, Judith; Piwien-Pilipuk, Graciela; Galigniana, Mario D

    2014-02-01

    The term molecular chaperone was first used to describe the ability of nucleoplasmin to prevent the aggregation of histones with DNA during the assembly of nucleosomes. Subsequently, the name was extended to proteins that mediate the post-translational assembly of oligomeric complexes protecting them from denaturation and/or aggregation. Hsp90 is a 90-kDa molecular chaperone that represents the major soluble protein of the cell. In contrast to most conventional chaperones, Hsp90 functions as a refined sensor of protein function and its principal role in the cell is to facilitate biological activity to properly folded client proteins that already have a preserved tertiary structure. Consequently, Hsp90 is related to basic cell functions such as cytoplasmic transport of soluble proteins, translocation of client proteins to organelles, and regulation of the biological activity of key signaling factors such as protein kinases, ubiquitin ligases, steroid receptors, cell cycle regulators, and transcription factors. A growing amount of evidence links the protective action of this molecular chaperone to mechanisms related to posttranslational modifications of soluble nuclear factors as well as histones. In this article, we discuss some aspects of the regulatory action of Hsp90 on transcriptional regulation and how this effect could have impacted genetic assimilation mechanism in some organisms.

  14. Regulatory role of the 90-kDa-heat-shock protein (Hsp90) and associated factors on gene expression.

    PubMed

    Erlejman, Alejandra G; Lagadari, Mariana; Toneatto, Judith; Piwien-Pilipuk, Graciela; Galigniana, Mario D

    2014-02-01

    The term molecular chaperone was first used to describe the ability of nucleoplasmin to prevent the aggregation of histones with DNA during the assembly of nucleosomes. Subsequently, the name was extended to proteins that mediate the post-translational assembly of oligomeric complexes protecting them from denaturation and/or aggregation. Hsp90 is a 90-kDa molecular chaperone that represents the major soluble protein of the cell. In contrast to most conventional chaperones, Hsp90 functions as a refined sensor of protein function and its principal role in the cell is to facilitate biological activity to properly folded client proteins that already have a preserved tertiary structure. Consequently, Hsp90 is related to basic cell functions such as cytoplasmic transport of soluble proteins, translocation of client proteins to organelles, and regulation of the biological activity of key signaling factors such as protein kinases, ubiquitin ligases, steroid receptors, cell cycle regulators, and transcription factors. A growing amount of evidence links the protective action of this molecular chaperone to mechanisms related to posttranslational modifications of soluble nuclear factors as well as histones. In this article, we discuss some aspects of the regulatory action of Hsp90 on transcriptional regulation and how this effect could have impacted genetic assimilation mechanism in some organisms. PMID:24389346

  15. An Intergenic Regulatory Region Mediates Drosophila Myc -Induced Apoptosis and Blocks Tissue Hyperplasia

    PubMed Central

    Zhang, Can; Tintó, Sergio Casas; Li, Guangyao; Lin, Nianwei; Chung, Michelle; Moreno, Eduardo; Moberg, Kenneth H.; Zhou, Lei

    2014-01-01

    Induction of cell autonomous apoptosis following oncogene-induced overproliferation is a major tumor-suppressive mechanism in vertebrates. However the detailed mechanism mediating this process remains enigmatic. In this study we demonstrate that dMyc-induced cell-autonomous apoptosis in the fruit fly Drosophila melanogaster relies on an intergenic sequence termed the IRER (Irradiation Responsive Enhancer Region). The IRER mediates expression of surrounding pro-apoptotic genes, and we use an in vivo reporter of the IRER chromatin state to gather evidence that epigenetic control of DNA accessibility within the IRER is an important determinant of the strength of this response to excess dMyc. In prior work we showed that the IRER also mediates P53-dependent induction of pro-apoptotic genes following DNA damage, and the chromatin conformation within IRER is regulated by Polycomb group-mediated histone modifications. dMyc-induced apoptosis and the P53-mediated DNA damage response thus overlap in a requirement for the IRER. The epigenetic mechanisms controlling IRER accessibility appear to set thresholds for the P53 and dMyc-induced expression of apoptotic genes in vivo and may have a profound impact on cellular sensitivity to oncogene-induced stress. PMID:24931167

  16. Activated CD8+ T cells induce expansion of Vβ5+ regulatory T cells via TNFR2 signaling

    PubMed Central

    Joedicke, Jara J; Myers, Lara; Carmody, Aaron B; Messer, Ronald J; Wajant, Harald; Lang, Karl S; Lang, Philipp A; Mak, Tak W; Hasenkrug, Kim J; Dittmer, Ulf

    2014-01-01

    Vβ5+ regulatory T cells (Tregs), which are specific for a mouse endogenous retroviral superantigen, become activated and proliferate in response to Friend retrovirus (FV) infection. We previously reported that FV-induced expansion of this Treg subset was dependent on CD8+ T cells and TNFα, but independent of IL-2. We now show that the inflammatory milieu associated with FV infection is not necessary for induction of Vβ5+ Treg expansion. Rather, it is the presence of activated CD8+ T cells that is critical for their expansion. The data indicate that the mechanism involves signaling between the membrane-bound form of TNFα (memTNFα) on activated CD8+ T cells and TNF receptor 2 (TNFR2) on Tregs. CD8+ T cells expressing memTNFα but no soluble TNFα (solTNFα) remained competent to induce strong Vβ5+ Treg expansion in vivo. In addition, Vβ5+ Tregs expressing only TNFR2 but no TNFR1 were still responsive to expansion. Finally, treatment of naïve mice with solTNFα did not induce Vβ5+ Treg expansion, but treatment with a TNFR2-specific agonist did. These results reveal a new mechanism of intercellular communication between activated CD8+ T cell effectors and Tregs that results in the activation and expansion of a Treg subset that subsequently suppresses CD8+ T cell functions. PMID:25098294

  17. Global analysis of induced transcription factors and cofactors identifies Tfdp2 as an essential coregulator during terminal erythropoiesis.

    PubMed

    Chen, Cynthia; Lodish, Harvey F

    2014-06-01

    Key transcriptional regulators of terminal erythropoiesis, such as GATA-binding factor 1 (GATA1) and T-cell acute lymphocytic leukemia protein 1 (TAL1), have been well characterized, but transcription factors and cofactors and their expression modulations have not yet been explored on a global scale. Here, we use global gene expression analysis to identify 28 transcription factors and 19 transcriptional cofactors induced during terminal erythroid differentiation whose promoters are enriched for binding by GATA1 and TAL1. Utilizing protein-protein interaction databases to identify cofactors for each transcription factor, we pinpoint several co-induced pairs, of which E2f2 and its cofactor transcription factor Dp-2 (Tfdp2) were the most highly induced. TFDP2 is a critical cofactor required for proper cell cycle control and gene expression. GATA1 and TAL1 are bound to the regulatory regions of Tfdp2 and upregulate its expression and knockdown of Tfdp2 results in significantly reduced rates of proliferation as well as reduced upregulation of many erythroid-important genes. Loss of Tfdp2 also globally inhibits the normal downregulation of many E2F2 target genes, including those that regulate the cell cycle, causing cells to accumulate in S phase and resulting in increased erythrocyte size. Our findings highlight the importance of TFDP2 in coupling the erythroid cell cycle with terminal differentiation and validate this study as a resource for future work on elucidating the role of diverse transcription factors and coregulators in erythropoiesis.

  18. Identification of hub genes and regulatory factors of glioblastoma multiforme subgroups by RNA-seq data analysis

    PubMed Central

    Li, Yanan; Min, Weijie; Li, Mengmeng; Han, Guosheng; Dai, Dongwei; Zhang, Lei; Chen, Xin; Wang, Xinglai; Zhang, Yuhui; Yue, Zhijian; Liu, Jianmin

    2016-01-01

    Glioblastoma multiforme (GBM) is the most common malignant brain tumor. This study aimed to identify the hub genes and regulatory factors of GBM subgroups by RNA sequencing (RNA-seq) data analysis, in order to explore the possible mechanisms responsbile for the progression of GBM. The dataset RNASeqV2 was downloaded by TCGA-Assembler, containing 169 GBM and 5 normal samples. Gene expression was calculated by the reads per kilobase per million reads measurement, and nor malized with tag count comparison. Following subgroup classification by the non-negative matrix factorization, the differentially expressed genes (DEGs) were screened in 4 GBM subgroups using the method of significance analysis of microarrays. Functional enrichment analysis was performed by DAVID, and the protein-protein interaction (PPI) network was constructed based on the HPRD database. The subgroup-related microRNAs (miRNAs or miRs), transcription factors (TFs) and small molecule drugs were predicted with predefined criteria. A cohort of 19,515 DEGs between the GBM and control samples was screened, which were predominantly enriched in cell cycle- and immunoreaction-related pathways. In the PPI network, lymphocyte cytosolic protein 2 (LCP2), breast cancer 1 (BRCA1), specificity protein 1 (Sp1) and chromodomain-helicase-DNA-binding protein 3 (CHD3) were the hub nodes in subgroups 1–4, respectively. Paired box 5 (PAX5), adipocyte protein 2 (aP2), E2F transcription factor 1 (E2F1) and cAMP-response element-binding protein-1 (CREB1) were the specific TFs in subgroups 1–4, respectively. miR-147b, miR-770-5p, miR-220a and miR-1247 were the particular miRNAs in subgroups 1–4, respectively. Natalizumab was the predicted small molecule drug in subgroup 2. In conclusion, the molecular regulatory mechanisms of GBM pathogenesis were distinct in the different subgroups. Several crucial genes, TFs, miRNAs and small molecules in the different GBM subgroups were identified, which may be used as potential

  19. Identification of hub genes and regulatory factors of glioblastoma multiforme subgroups by RNA-seq data analysis.

    PubMed

    Li, Yanan; Min, Weijie; Li, Mengmeng; Han, Guosheng; Dai, Dongwei; Zhang, Lei; Chen, Xin; Wang, Xinglai; Zhang, Yuhui; Yue, Zhijian; Liu, Jianmin

    2016-10-01

    Glioblastoma multiforme (GBM) is the most common malignant brain tumor. This study aimed to identify the hub genes and regulatory factors of GBM subgroups by RNA sequencing (RNA-seq) data analysis, in order to explore the possible mechanisms responsbile for the progression of GBM. The dataset RNASeqV2 was downloaded by TCGA-Assembler, containing 169 GBM and 5 normal samples. Gene expression was calculated by the reads per kilobase per million reads measurement, and nor malized with tag count comparison. Following subgroup classification by the non-negative matrix factorization, the differentially expressed genes (DEGs) were screened in 4 GBM subgroups using the method of significance analysis of microarrays. Functional enrichment analysis was performed by DAVID, and the protein-protein interaction (PPI) network was constructed based on the HPRD database. The subgroup-related microRNAs (miRNAs or miRs), transcription factors (TFs) and small molecule drugs were predicted with pre-defined criteria. A cohort of 19,515 DEGs between the GBM and control samples was screened, which were predominantly enriched in cell cycle- and immunoreaction-related pathways. In the PPI network, lymphocyte cytosolic protein 2 (LCP2), breast cancer 1 (BRCA1), specificity protein 1 (Sp1) and chromodomain-helicase-DNA-binding protein 3 (CHD3) were the hub nodes in subgroups 1-4, respectively. Paired box 5 (PAX5), adipocyte protein 2 (aP2), E2F transcription factor 1 (E2F1) and cAMP-response element-binding protein-1 (CREB1) were the specific TFs in subgroups 1-4, respectively. miR‑147b, miR‑770-5p, miR‑220a and miR‑1247 were the particular miRNAs in subgroups 1-4, respectively. Natalizumab was the predicted small molecule drug in subgroup 2. In conclusion, the molecular regulatory mechanisms of GBM pathogenesis were distinct in the different subgroups. Several crucial genes, TFs, miRNAs and small molecules in the different GBM subgroups were identified

  20. Factors influencing fluoxetine-induced sexual dysfunction in female rats.

    PubMed

    Adams, Sarah; Heckard, Danyeal; Hassell, James; Uphouse, Lynda

    2012-11-01

    Treatment with selective serotonin reuptake inhibitors, such as fluoxetine, produces sexual side effects with low sexual desire being the most prevalent effect in females. In few studies have preclinical models for such antidepressant-induced sexual dysfunction been fruitful. In the current manuscript, the effects of fluoxetine on multiple measures of female sexual motivation and sexual receptivity were examined. Ovariectomized, Fischer rats were primed with 10 μg estradiol benzoate and 500 μg progesterone. Partner preference, active investigation of the male, and measures of sexual behavior were examined after injection with 15 mg/kg fluoxetine. Factors (pretesting for sexual behavior, size of the test arena, non-contact time with a male) that differ among experiments designed to study antidepressant-induced female rat sexual dysfunction were studied. The male preference ratio was not affected by fluoxetine treatment but active investigation of the male was reduced; lordosis behavior was inhibited and pretesting for sexual receptivity amplified fluoxetine's inhibition; size of the testing arena or non-contact experience with the male had no effect. Regardless of test condition, when given the opportunity to escape from the male, fluoxetine-treated females displayed escape behavior. Measures of male preference and active investigation, but not lordosis behavior, appeared to be affected by fluoxetine's impact on activity. The collective data provided a behavioral profile of fluoxetine-induced sexual dysfunction. These findings reinforce the value of multiple measures when attempting to model antidepressant-induced female sexual dysfunction.

  1. Activated Factor X Induces Endothelial Cell Senescence Through IGFBP-5

    PubMed Central

    Sanada, Fumihiro; Taniyama, Yoshiaki; Muratsu, Jun; Otsu, Rei; Iwabayashi, Masaaki; Carracedo, Miguel; Rakugi, Hiromi; Morishita, Ryuichi

    2016-01-01

    Uncontrolled coagulation contributes to the pathophysiology of several chronic inflammatory diseases. In these conditions, senescent cells are often observed and is involved in the generation of inflammation. The coincidence of hyper-coagulation, cell senescence, and inflammation suggests the existence of a common underlying mechanism. Recent evidence indicates that activated coagulation factor X (FXa) plays a role in the processes beyond blood coagulation. This non-hematologic function entails the mediation of inflammation and tissue remodeling. We therefore tested the hypothesis that FXa induces cell senescence resulting in tissue inflammation and impaired tissue regeneration. Human umbilical vein endothelial cells were stimulated with FXa for 14 days. The proliferation of cells treated with FXa was significantly smaller, and the fraction of senescence-associated β-galactosidase-positive cells was increased as compared to the control group. RT-qPCR array revealed that FXa increased the expression of IGFBP-5, EGR-1, p53, and p16INK4a. Inhibition of FXa by a direct FXa inhibitor, rivaroxaban, or IGFBP-5 by siRNA decreased FXa-induced cell senescence, restoring cell proliferation. Moreover, in an ischemic hind limb mouse model, FXa inhibited neovascularization by endothelial progenitor cell. However, rivaroxaban significantly restored FXa-induced impaired angiogenesis. In summary, FXa induced endothelial cell senescence through IGFBP-5, resulting in impaired angiogenesis. PMID:27752126

  2. The neurohormone orexin stimulates hypoxia-inducible factor-1 activity.

    PubMed

    Sikder, Devanjan; Kodadek, Thomas

    2007-11-15

    Orexin A and Orexin B (also known as hypocretins) are neuropeptides that bind two related G-coupled protein receptors (OXR1 and OXR2) and thus induce wakefulness, food consumption, and locomotion. Conversely, deletion of the orexin gene in mice produces a condition similar to canine and human narcolepsy. Despite the central importance of the orexin system in regulating wakefulness and feeding behavior, little is known about the downstream signaling mechanisms that achieve these effects. In this study, genomics techniques are used to probe this question and reveal that orexin activates the hypoxia-inducible factor 1 (HIF-1), a heterodimeric transcription factor whose pathogenic role in stimulating angiogenesis in hypoxic tumors has been the focus of intense investigation. Orexin-stimulated HIF-1 activity is due to both increased HIF-1alpha gene transcription and a down-regulation of von Hippel-Lindau (VHL), the E3 ubiquitin ligase that mediates the turnover of HIF-1 via the ubiquitin-proteasome pathway. Orexin-mediated activation of HIF-1 results in increased glucose uptake and higher glycolytic activity, as expected from studies of hypoxic cells. However, orexin receptor-expressing cells somehow override the HIF-1-mediated preference for funneling pyruvate into anaerobic glycolysis and instead favor ATP production through the tricarboxylic acid cycle and oxidative phosphorylation. These findings implicate HIF-1 as an important transcription factor in the hormone-mediated regulation of hunger and wakefulness.

  3. Interferon Regulatory Factor 6 (IRF6) and Fibroblast Growth Factor Receptor 1 (FGFR1) Contribute to Human Tooth Agenesis

    PubMed Central

    Vieira, Alexandre R.; Modesto, Adriana; Meira, Raquel; Barbosa, Anna Renata Schneider; Lidral, Andrew C.; Murray, Jeffrey C.

    2008-01-01

    Phenotypic characteristics expressed in syndromes give clues to the factors involved in the cause of isolated forms of the same defects. We investigated two genes responsible for craniofacial syndromes, FGFR1 and IRF6, in a collection of families with isolated tooth agenesis. Cheek swab samples were obtained for DNA analysis from 116 case/parent trios. Probands had at least one developmentally missing tooth, excluding third molars. In addition, we studied 89 cases and 50 controls from Ohio to replicate any positive findings. Genotyping was performed by kinetic polymerase chain-reaction or TaqMan assays. Linkage disequilibrium analysis and transmission distortion of the marker alleles were performed. The same variants in the IRF6 gene that are associated with isolated orofacial clefts are also associated with human tooth agenesis (rs861019, P = 0.058; rs17015215—V274I, P = 0.0006; rs7802, P = 0.004). Mutations in IRF6 cause Van der Woude and popliteal pterygium syndromes. The craniofacial phenotypic characteristics of these syndromes include oral clefts and preferential tooth agenesis of incisors and premolars, besides pits on the lower lips. Also it appears that preferential premolar agenesis is associated with FGFR1 (P = 0.014) and IRF6 (P = 0.002) markers. There were statistically significant data suggesting that IRF6 interacts not only with MSX1 (P = 0.001), but also with TGFA (P = 0.03). PMID:17318851

  4. Tumor necrosis factor-alpha antagonist-induced sarcoidosis.

    PubMed

    Clementine, Rochelle Robicheaux; Lyman, Justin; Zakem, Jerald; Mallepalli, Jyothi; Lindsey, Stephen; Quinet, Robert

    2010-09-01

    Sarcoidosis is a multisystem granulomatous disease of unknown etiology. Tumor necrosis factor (TNF)-alpha is an important player in granuloma formation, and recent clinical trials have investigated the efficacy of TNF-alpha inhibitors in sarcoidosis. Paradoxically, there are several case reports in the medical literature describing the development of sarcoidosis in patients treated with TNF-alpha inhibitors. We describe 3 cases of TNF-alpha antagonist-induced sarcoidosis: 1 case of pulmonary, ocular and cutaneous sarcoidosis developing in a patient receiving infliximab for erosive rheumatoid arthritis, 1 case of etanercept-induced sarcoidosis in a patient with seronegative rheumatoid arthritis, and 1 case of sarcoidosis developing in a patient receiving etanercept for erosive rheumatoid arthritis. We also provide a brief discussion on the role of TNF alpha in granuloma formation and implications in the use of TNF-alpha antagonists in autoimmune disease.

  5. Phosphate starvation induces the sporulation killing factor of Bacillus subtilis.

    PubMed

    Allenby, Nicholas E E; Watts, Carys A; Homuth, Georg; Prágai, Zoltán; Wipat, Anil; Ward, Alan C; Harwood, Colin R

    2006-07-01

    Bacillus subtilis produces and exports a peptide sporulation killing factor (SkfA) that induces lysis of sibling cells. skfA is part of the skf operon (skfA-H), which is responsible for immunity to SkfA, as well as for production and export of SkfA. Here we report that transcription of skfA is markedly induced when cells of B. subtilis are subjected to phosphate starvation. The role of PhoP in regulation of the skf operon was confirmed by in vitro gel shift assays, which showed that this operon is a new member of the PhoP regulon. A putative stem-loop structure in the skfA-skfB intergenic region is proposed to act as a stabilizer of an skfA-specific transcript. PMID:16816204

  6. Phosphate Starvation Induces the Sporulation Killing Factor of Bacillus subtilis

    PubMed Central

    Allenby, Nicholas E. E.; Watts, Carys A.; Homuth, Georg; Prágai, Zoltán; Wipat, Anil; Ward, Alan C.; Harwood, Colin R.

    2006-01-01

    Bacillus subtilis produces and exports a peptide sporulation killing factor (SkfA) that induces lysis of sibling cells. skfA is part of the skf operon (skfA-H), which is responsible for immunity to SkfA, as well as for production and export of SkfA. Here we report that transcription of skfA is markedly induced when cells of B. subtilis are subjected to phosphate starvation. The role of PhoP in regulation of the skf operon was confirmed by in vitro gel shift assays, which showed that this operon is a new member of the PhoP regulon. A putative stem-loop structure in the skfA-skfB intergenic region is proposed to act as a stabilizer of an skfA-specific transcript. PMID:16816204

  7. Phosphate starvation induces the sporulation killing factor of Bacillus subtilis.

    PubMed

    Allenby, Nicholas E E; Watts, Carys A; Homuth, Georg; Prágai, Zoltán; Wipat, Anil; Ward, Alan C; Harwood, Colin R

    2006-07-01

    Bacillus subtilis produces and exports a peptide sporulation killing factor (SkfA) that induces lysis of sibling cells. skfA is part of the skf operon (skfA-H), which is responsible for immunity to SkfA, as well as for production and export of SkfA. Here we report that transcription of skfA is markedly induced when cells of B. subtilis are subjected to phosphate starvation. The role of PhoP in regulation of the skf operon was confirmed by in vitro gel shift assays, which showed that this operon is a new member of the PhoP regulon. A putative stem-loop structure in the skfA-skfB intergenic region is proposed to act as a stabilizer of an skfA-specific transcript.

  8. Regulatory role of neuron-restrictive silencing factor in expression of TRPC1

    SciTech Connect

    Ohba, Takayoshi; Watanabe, Hiroyuki; Takahashi, Yoichiro; Suzuki, Takashi; Miyoshi, Ichiro; Nakayama, Shinnsuke; Satoh, Eisaku; Iino, Kenji; Sasano, Hironobu; Mori, Yasuo; Kuromitsu, Sadao; Imagawa, Keiichi; Saito, Yoshihiko; Iijima, Toshihiko; Ito, Hiroshi; Murakami, Manabu . E-mail: mmura0123@hotmail.co.jp

    2006-12-22

    Neuron-restrictive silencer factor (NRSF) binds its consensus element to repress the transcription of various genes. The dominant-negative form (dnNRSF) has a hypertrophic effect on cardiogenesis through an unidentified mechanism. We examined the involvement of transient receptor potential (TRP) channel proteins, using transgenic mice overexpressing dnNRSF (dnNRSF mice). Electrophoretic mobility-shift assays revealed an interaction between NRSF and a neuron-restrictive silencer element-like sequence in intron 4 of TRPC1 genomic DNA. According to RT-PCR and Western analyses, TRPC1 was up-regulated in dnNRSF mouse heart. Transient overexpression of TRPC1 in HEK 293T cells increased the activity of the nuclear factor in activated T cells (NFAT) promoter and stimulated store-operated Ca{sup 2+} channel (SOCC)-mediated Ca{sup 2+} entry. Transfection of TRPC1 into primary cardiomyocytes increased NFAT activity, indicating a major role for TRPC1 in NFAT activation. Our findings strongly suggest that NRSF regulates TRP1 gene expression and causes changes in the levels of calcium entry through SOCCs.

  9. Dedifferentiation of Adult Human Myoblasts Induced by Ciliary Neurotrophic Factor In Vitro

    PubMed Central

    Chen, Xiaoping; Mao, Zebin; Liu, Shuhong; Liu, Hong; Wang, Xuan; Wu, Haitao; Wu, Yan; Zhao, Tong; Fan, Wenhong; Li, Yong; Yew, David T.; Kindler, Pawel M.; Li, Linsong; He, Qihua; Qian, Lingjia; Wang, Xiaomin; Fan, Ming

    2005-01-01

    Ciliary neurotrophic factor (CNTF) is primarily known for its important cellular effects within the nervous system. However, recent studies indicate that its receptor can be highly expressed in denervated skeletal muscle. Here, we investigated the direct effect of CNTF on skeletal myoblasts of adult human. Surprisingly, we found that CNTF induced the myogenic lineage-committed myoblasts at a clonal level to dedifferentiate into multipotent progenitor cells—they not only could proliferate for over 20 passages with the expression absence of myogenic specific factors Myf5 and MyoD, but they were also capable of differentiating into new phenotypes, mainly neurons, glial cells, smooth muscle cells, and adipocytes. These “progenitor cells” retained their myogenic memory and were capable of redifferentiating into myotubes. Furthermore, CNTF could activate the p44/p42 MAPK and down-regulate the expression of myogenic regulatory factors (MRFs). Finally, PD98059, a specific inhibitor of p44/p42 MAPK pathway, was able to abolish the effects of CNTF on both myoblast fate and MRF expression. Our results demonstrate the myogenic lineage-committed human myoblasts can dedifferentiate at a clonal level and CNTF is a novel regulator of skeletal myoblast dedifferentiation via p44/p42 MAPK pathway. PMID:15843428

  10. Hidden Markov induced Dynamic Bayesian Network for recovering time evolving gene regulatory networks

    NASA Astrophysics Data System (ADS)

    Zhu, Shijia; Wang, Yadong

    2015-12-01

    Dynamic Bayesian Networks (DBN) have been widely used to recover gene regulatory relationships from time-series data in computational systems biology. Its standard assumption is ‘stationarity’, and therefore, several research efforts have been recently proposed to relax this restriction. However, those methods suffer from three challenges: long running time, low accuracy and reliance on parameter settings. To address these problems, we propose a novel non-stationary DBN model by extending each hidden node of Hidden Markov Model into a DBN (called HMDBN), which properly handles the underlying time-evolving networks. Correspondingly, an improved structural EM algorithm is proposed to learn the HMDBN. It dramatically reduces searching space, thereby substantially improving computational efficiency. Additionally, we derived a novel generalized Bayesian Information Criterion under the non-stationary assumption (called BWBIC), which can help significantly improve the reconstruction accuracy and largely reduce over-fitting. Moreover, the re-estimation formulas for all parameters of our model are derived, enabling us to avoid reliance on parameter settings. Compared to the state-of-the-art methods, the experimental evaluation of our proposed method on both synthetic and real biological data demonstrates more stably high prediction accuracy and significantly improved computation efficiency, even with no prior knowledge and parameter settings.

  11. Genetic and molecular analysis of Sn, a light-inducible, tissue specific regulatory gene in maize.

    PubMed

    Tonelli, C; Consonni, G; Dolfini, S F; Dellaporta, S L; Viotti, A; Gavazzi, G

    1991-03-01

    The Sn locus of maize is functionally similar to the R and B loci, in that Sn differentially controls the tissue-specific deposition of anthocyanin pigments in certain seedling and plant cells. We show that Sn shows molecular similarity to the R gene and have used R DNA probes to characterize several Sn alleles. Northern analysis demonstrates that all Sn alleles encode a 2.5 kb transcript, which is expressed in a tissue-specific fashion consistent with the distribution of anthocyanins. Expression of the Sn gene is light-regulated. However, the Sn: bol3 allele allows Sn mRNA transcription to occur in the dark, leading to pigmentation in dark-grown seedlings and cob integuments. We report the isolation of genomic and cDNA clones of the light-independent Sn: bol3 allele. Using Sn cDNA as a probe, the spatial and temporal expression of Sn has been examined. The cell-specific localization of Sn mRNA has been confirmed by in situ hybridization using labelled antisense RNA probes. According to its proposed regulatory role, expression of Sn precedes and, in turn, causes a coordinate and tissue-specific accumulation of mRNA of structural genes for pigment synthesis and deposition, such as A1 and C2. The functional and structural relationship between R, B, Lc and Sn is discussed in terms of an evolutionary derivation from a single ancestral gene which gave rise this diverse gene family by successive duplication events.

  12. Starvation Induces Proteasome Autophagy with Different Pathways for Core and Regulatory Particles*

    PubMed Central

    Waite, Kenrick A.; Mota-Peynado, Alina De-La; Vontz, Gabrielle; Roelofs, Jeroen

    2016-01-01

    The proteasome is responsible for the degradation of many cellular proteins. If and how this abundant and normally stable complex is degraded by cells is largely unknown. Here we show that in yeast, upon nitrogen starvation, proteasomes are targeted for vacuolar degradation through autophagy. Using GFP-tagged proteasome subunits, we observed that autophagy of a core particle (CP) subunit depends on the deubiquitinating enzyme Ubp3, although a regulatory particle (RP) subunit does not. Furthermore, upon blocking of autophagy, RP remained largely nuclear, although CP largely localized to the cytosol as well as granular structures within the cytosol. In all, our data reveal a regulated process for the removal of proteasomes upon nitrogen starvation. This process involves CP and RP dissociation, nuclear export, and independent vacuolar targeting of CP and RP. Thus, in addition to the well characterized transcriptional up-regulation of genes encoding proteasome subunits, cells are also capable of down-regulating cellular levels of proteasomes through proteaphagy. PMID:26670610

  13. Growth factor involvement in tension-induced skeletal muscle growth

    NASA Technical Reports Server (NTRS)

    Vandenburgh, H. H.

    1987-01-01

    Muscle tissue culture techniques were developed to grow skeletal myofibers which differentiate into more adult-like myofibers. Mechanical simulation studies of these muscle cells in a newly developed mechanical cell simulator can now be performed to study growth processes in skeletal muscle. Conditions in the mechanical cell simulator were defined where mechanical activity can either prevent muscle wasting or stimulate muscle growth. The role of endogenous and exogenous growth factors in tension-induced muscle growth is being investigated under the defined conditions of tissue culture.

  14. Regulatory role of the OsWOX3A transcription factor in rice root development.

    PubMed

    Cho, Sung-Hwan; Paek, Nam-Chon

    2016-06-01

    WUSCHEL-related homeobox (WOX) transcription factors play important roles in cell fate determination during plant development and function in the signaling pathways of phytohormones such as gibberellic acid (GA). In a recent study, we demonstrated that OsWOX3A directly binds to the promoter of the KAO gene, which encodes ent-kaurenoic acid oxidase, an enzyme involved in GA biosynthesis, and represses KAO expression. Interestingly, we observed that OsWOX3A overexpression causes not only severe dwarfism, but also an increase in the number of lateral roots. Moreover, the expression of PIN-FORMED 1 (PIN1), involved in polar auxin transport, is significantly upregulated in OsWOX3A-OX plants. These findings indicate that OsWOX3A may be involved in modulation of GA-auxin crosstalk in rice root development. PMID:27171233

  15. The regulatory role of serum response factor pathway in neutrophil inflammatory response

    PubMed Central

    Taylor, Ashley; Halene, Stephanie

    2015-01-01

    Purpose of review Neutrophils rapidly migrate to sites of injury and infection. Egress of neutrophils from the circulation into tissues is a highly regulated process involving several distinct steps. Cell–cell interactions mediated by selectins and integrins and reorganization of the actin cytoskeleton are key mechanisms facilitating appropriate neutrophil recruitment. Neutrophil function is impaired in inherited and acquired disorders, such as leukocyte adhesion deficiency and myelodysplasia. Since the discovery that deletion of all or part of chromosome 5 is the most common genetic aberration in myelodysplasia, the roles of several of the deleted genes have been investigated in hematopoiesis. Several genes encoding proteins of the serum response factor (SRF) pathway are located on 5q. This review focuses, in particular, on the role of SRF in myeloid maturation and neutrophil function. Recent findings SRF and its pathway fulfill multiple complex roles in the regulation of the innate and adaptive immune system. Loss of SRF leads to defects in B-cell and T-cell development. SRF-deficient macrophages fail to spread, transmigrate, and phagocytose bacteria, and SRF-deficient neutrophils show defective chemotaxis in vitro and in vivo with failure of inside-out activation and trafficking of the Mac1 integrin complex. Loss of the formin mammalian Diaphanous 1, a regulator of linear actin polymerization and mediator of Ras homolog family member A signaling to SRF, results in aberrant myeloid differentiation and hyperactivity of the immune system. Summary SRF is an essential transcription factor in hematopoiesis and mature myeloid cell function. SRF regulates neutrophil migration, integrin activation, and trafficking. Disruption of the SRF pathway results in myelodysplasia and immune dysfunction. PMID:25402621

  16. INTERRUPTION OF DENDRITIC CELL-MEDIATED TIM-4 SIGNALING INDUCES REGULATORY T CELLS AND PROMOTES SKIN ALLOGRAFT SURVIVAL

    PubMed Central

    Yeung, Melissa Y.; McGrath, Martina M.; Nakayama, Masafumi; Shimizu, Tetsunosuke; Boenisch, Olaf; Magee, Ciara N.; Abdoli, Rozita; Akiba, Hisaya; Ueno, Takuya; Turka, Laurence A.; Najafian, Nader

    2013-01-01

    Dendritic cells (DCs) are the central architects of the immune response, inducing inflammatory or tolerogenic immunity, dependent upon their activation status. As such, DCs are highly attractive therapeutic targets and may hold the potential to control detrimental immune responses. TIM-4, expressed on antigen presenting cells, has complex functions in vivo, acting both as a costimulatory molecule and a phosphatidylserine (PS) receptor. The effect of TIM-4 costimulation on T cell activation remains unclear. Here, we demonstrate that antibody blockade of DC-expressed TIM-4 leads to increased induction of iTregs from naïve CD4+ T cells, both in vitro and in vivo. iTreg induction occurs through suppression of IL-4/STAT6/Gata3 induced Th2 differentiation. In addition, blockade of TIM-4 on previously activated DCs still leads to increased iTreg induction. iTregs induced under TIM-4 blockade have equivalent potency to control and upon adoptive transfer, significantly prolong skin allograft survival in vivo. In RAG−/− recipients of skin allografts adoptively transferred with CD4+ T cells, we show that TIM-4 blockade in vivo is associated with a three-fold prolongation in allograft survival. Furthermore, in this mouse model of skin transplantation, increased induction of allospecific iTregs and a reduction in T effector responses were observed, with decreased Th1 and Th2 responses. This enhanced allograft survival and pro-tolerogenic skewing of the alloresponse is critically dependent upon conversion of naïve CD4+ to Tregs in vivo. Collectively, these studies identify blockade of DC-expressed TIM-4 as a novel strategy which holds the capacity to induce regulatory immunity in vivo. PMID:24038092

  17. Tumor necrosis factor-inducing activities of Cryptococcus neoformans components.

    PubMed Central

    Delfino, D; Cianci, L; Migliardo, M; Mancuso, G; Cusumano, V; Corradini, C; Teti, G

    1996-01-01

    Cryptococcus neoformans-induced tumor necrosis factor alpha (TNF-alpha) production may lead to increased human immunodeficiency virus replication in patients with AIDS. In order to identify cryptococcal components that are predominantly responsible for stimulating TNF production, various concentrations of glucuronoxylomannan (GXM), galactoxylomannan (GalXM), mannoproteins (MP), and alpha(1-3) [corrected] glucan were added to whole-blood cultures. All of the cryptococcal components tested, as well as whole heat-killed cryptococci, were capable of inducing TNF-alpha release in a dose-dependent manner. MP were significantly more potent than any of the other cryptococcal components tested or heat-killed cryptococci in stimulating TNF-alpha production (P < 0.05). GXM, in contrast, was significantly less potent in this activity than either GalXM or MP (P < 0.05). As little as 0.5 microg of MP per ml was sufficient to produce moderate but significant elevations of TNF-alpha release. Maximal MP-induced TNF-alpha levels were similar to those induced by Salmonella enteritidis lipopolysaccharide, our positive control. Further experiments using isolated leukocytes suggested that monocytes were the cell population mainly responsible for TNF-alpha production, although the participation of other cell types could not be excluded. The presence of complement-sufficient plasma was a necessary requirement for TNF-alpha induction by GXM, GalXM, and low doses of MP. High MP concentrations (100 microg/ml) were also capable of stimulating TNF-alpha production in the absence of plasma. These data indicate that soluble products released by C. neoformans are capable of inducing TNF-alpha secretion in human leukocytes. This may be clinically relevant, since high concentrations of such products are frequently found in the body fluids of AIDS patients infected with C. neoformans. PMID:8945566

  18. Three-phase power factor controller with induced EMF sensing

    NASA Technical Reports Server (NTRS)

    Nola, F. J. (Inventor)

    1984-01-01

    A power factor controller for an ac induction motor is provided which is of the type comprising thyristor switches connected in series with the motor, phase detectors for sensing the motor current and voltage and providing an output proportional to the phase difference between the motor voltage and current, and a control circuit, responsive to the output of the phase detector and to a power factor command signal, for controlling switching of the thyristor. The invention involves sensing the induced emf produced by the motor during the time interval when the thyristor is off and for producing a corresponding feedback signal for controlling switching of the thyristor. The sensed emf is also used to enhance soft starting of the motor.

  19. Vectorology and Factor Delivery in Induced Pluripotent Stem Cell Reprogramming

    PubMed Central

    2014-01-01

    Induced pluripotent stem cell (iPSC) reprogramming requires sustained expression of multiple reprogramming factors for a limited period of time (10–30 days). Conventional iPSC reprogramming was achieved using lentiviral or simple retroviral vectors. Retroviral reprogramming has flaws of insertional mutagenesis, uncontrolled silencing, residual expression and re-activation of transgenes, and immunogenicity. To overcome these issues, various technologies were explored, including adenoviral vectors, protein transduction, RNA transfection, minicircle DNA, excisable PiggyBac (PB) transposon, Cre-lox excision system, negative-sense RNA replicon, positive-sense RNA replicon, Epstein-Barr virus-based episomal plasmids, and repeated transfections of plasmids. This review provides summaries of the main vectorologies and factor delivery systems used in current reprogramming protocols. PMID:24625220

  20. Evaluation of transcription factor that regulates T helper 17 and regulatory T cells function in periodontal health and disease

    PubMed Central

    Karthikeyan, B.; Talwar; Arun, K. V.; Kalaivani, S.

    2015-01-01

    Background: The differentiation of naοve T helper (Th) cells towards Th17 and regulatory T cells (Treg) is regulated by the transcription factors retinoic acid related orphan receptor gamma transcription (RORYt) and Forkhead box p3 (Foxp3), respectively. An imbalance in the activity of these transcription factors could result in the dysregulation of Th17/Treg response. Materials and Methods: Total RNA was isolated from gingival tissue obtained from 10 patients, each from periodontally healthy and diseased groups. The gene expression of RORYt and Foxp3 was measured by real-time reverse transcription polymerization chain reaction using total RNA isolates from gingival tissues group when compared to the healthy group, while Foxp3 demonstrated a 6.68 ± 0.03 fold decrease of expression in diseased group when compared to healthy group. Conclusion: Our results indicate a functional imbalance in the Th17/Treg response in periodontal disease group when compared to the periodontally healthy group. PMID:26538941

  1. Different papillomaviruses have different repertoires of transcription factor binding sites: convergence and divergence in the upstream regulatory region

    PubMed Central

    García-Vallvé, Santiago; Iglesias-Rozas, José R; Alonso, Ángel; Bravo, Ignacio G

    2006-01-01

    Background Papillomaviruses (PVs) infect stratified squamous epithelia in warm-blooded vertebrates and have undergone a complex evolutionary process. The control of the expression of the early ORFs in PVs depends on the binding of cellular and viral transcription factors to the upstream regulatory region (URR) of the virus. It is believed that there is a core of transcription factor binding sites (TFBS) common to all PVs, with additional individual differences, although most of the available information focuses only on a handful of viruses. Results We have studied the URR of sixty-one PVs, covering twenty different hosts. We have predicted the TFBS present in the URR and analysed these results by principal component analysis and genetic algorithms. The number and nature of TFBS in the URR might be much broader than thus far described, and different PVs have different repertoires of TFBS. Conclusion There are common fingerprints in the URR in PVs that infect primates, although the ancestors of these viruses diverged a long time ago. Additionally, there are obvious differences between the URR of alpha and beta PVs, despite these PVs infect similar histological cell types in the same host, i.e. human. A thorough analysis of the TFBS in the URR might provide crucial information about the differential biology of cancer-associated PVs. PMID:16526953

  2. Overexpression of the Dominant-Negative Form of Interferon Regulatory Factor 1 in Oligodendrocytes Protects against Experimental Autoimmune Encephalomyelitis

    PubMed Central

    Ren, Zhihua; Wang, Yan; Tao, Duan; Liebenson, David; Liggett, Thomas; Goswami, Rajendra; Clarke, Robert; Stefoski, Dusan

    2011-01-01

    Interferon regulatory factor 1 (IRF-1) is a transcription factor that has been implicated in the pathogenesis of the human autoimmune demyelinating disease multiple sclerosis (MS) and in its animal model, experimental autoimmune encephalomyelitis (EAE). The goal of the present study was to directly examine the role of IRF-1 in oligodendrocyte injury and inflammatory demyelination. For the purpose of this study, we generated a transgenic mouse line (CNP/dnIRF-1) that overexpresses the dominant-negative form of IRF-1 (dnIRF1) specifically in oligodendrocytes. CNP/dnIRF-1 mice exhibited no phenotypic abnormalities but displayed suppressed IRF-1 signaling in oligodendrocytes. The major finding of our study was that the CNP/dnIRF-1 mice, compared with the wild-type mice, were protected against EAE, a phenomenon associated with significant reduction of inflammatory demyelination and with oligodendrocyte and axonal preservation. The observed protection was related to suppressed IRF-1 signaling and impaired expression of immune and proapoptotic genes in oligodendrocytes. No significant differences in the peripheral immune responses between the wild-type and the CNP/dnIRF-1 mice were identified throughout the experiments. This study indicates that IRF-1 plays a critical role in the pathogenesis of EAE by mediating oligodendrocyte response to inflammation and injury. It also suggests that oligodendrocytes are actively involved in the neuroimmune network, and that exploring oligodendrocyte-related pathogenic mechanisms, in addition to the conventional immune-based ones, may have important therapeutic implications in MS. PMID:21653838

  3. Progesterone as a morphological regulatory factor of the male and female gerbil prostate

    PubMed Central

    Fochi, Ricardo A; Santos, Fernanda C A; Goes, Rejane M; Taboga, Sebastião R

    2013-01-01

    Testosterone (T) and oestrogen are the main active steroid hormones in the male and female reproductive system respectively. In female rodents progesterone (P4), together with testosterone and oestrogen, has an essential role in the regulation of the oestrous cycle, which influences the prostate physiology through their oscillations. In this work we investigated how the male and female prostate gland of Mongolian gerbils responds to surgical castration at the start of puberty and what are the effects of T, oestradiol (E2) and P4 replacement, using both quantitative and qualitative methods. We also examined the location of the main steroid receptors present in the prostate. In the castrated animals of both sexes an intense glandular regression, along with disorganization of the stromal compartment, and abundant hyperplasia was observed. The replacement of P4 secured a mild recovery of the glandular morphology, inducing the growth of secretory cells and restoring the androgen receptor (AR) cells. The administration of P4 and E2 eliminated epithelial hyperplasia and intensified gland hypertrophy, favouring the emergence of prostatic intraepithelial neoplasia (PIN). In animals treated with T and P4, even though there are some inflammatory foci and other lesions, the prostate gland revealed morphology closer to that of control animals. In summary, through the administration of P4, we could demonstrate that this hormone has anabolic characteristics, promoting hyperplasia and hypertrophy, mainly in the epithelial compartment. When combined with E2 and T, there is an accentuation of glandular hypertrophy that interrupts the development of hyperplasia and ensures the presence of a less dysplastic glandular morphology. PMID:24205795

  4. A(1) and A(3) adenosine receptors inhibit LPS-induced hypoxia-inducible factor-1 accumulation in murine astrocytes.

    PubMed

    Gessi, Stefania; Merighi, Stefania; Stefanelli, Angela; Fazzi, Debora; Varani, Katia; Borea, Pier Andrea

    2013-10-01

    Adenosine (Ado) exerts neuroprotective and anti-inflammatory functions by acting through four receptor subtypes A1, A2A, A2B and A3. Astrocytes are one of its targets in the central nervous system. Hypoxia-inducible factor-1 (HIF-1), a master regulator of oxygen homeostasis, is induced after hypoxia, ischemia and inflammation and plays an important role in brain injury. HIF-1 is expressed by astrocytes, however the regulatory role played by Ado on HIF-1α modulation induced by inflammatory and hypoxic conditions has not been investigated. Primary murine astrocytes were activated with lipopolysaccharide (LPS) with or without Ado, Ado receptor agonists, antagonists and receptor silencing, before exposure to normoxia or hypoxia. HIF-1α accumulation and downstream genes regulation were determined. Ado inhibited LPS-increased HIF-1α accumulation under both normoxic and hypoxic conditions, through activation of A1 and A3 receptors. In cells incubated with the blockers of p44/42 MAPK and Akt, LPS-induced HIF-1α accumulation was significantly decreased in normoxia and hypoxia, suggesting the involvement of p44/42 MAPK and Akt in this effect and Ado inhibited kinases phosphorylation. A series of angiogenesis and metabolism related genes were modulated by hypoxia in an HIF-1 dependent way, but not further increased by LPS, with the exception of GLUT-1 and hexochinase II that were elevated by LPS only in normoxia and inhibited by Ado receptors. Instead, genes involved in inflammation, like inducible nitric-oxide synthase (iNOS) and A2B receptors, were increased by LPS in normoxia, strongly stimulated by LPS in concert with hypoxia and inhibited by Ado, through A1 and A3 receptor subtypes. In conclusion A1 and A3 receptors reduce the LPS-mediated HIF-1α accumulation in murine astrocytes, resulting in a downregulation of genes involved in inflammation and hypoxic injury, like iNOS and A2B receptors, in both normoxic and hypoxic conditions.

  5. Identification of temozolomide resistance factors in glioblastoma via integrative miRNA/mRNA regulatory network analysis.

    PubMed

    Hiddingh, Lotte; Raktoe, Rajiv S; Jeuken, Judith; Hulleman, Esther; Noske, David P; Kaspers, Gertjan J L; Vandertop, W Peter; Wesseling, Pieter; Wurdinger, Thomas

    2014-01-01

    Drug resistance is a major issue in the treatment of glioblastoma. Almost all glioblastomas are intrinsically resistant to chemotherapeutic temozolomide (TMZ) or develop resistance during treatment. The interaction networks of microRNAs (miRNAs) and mRNAs likely regulate most biological processes and can be employed to better understand complex processes including drug resistance in cancer. In this study, we examined if integrative miRNA/mRNA network analysis using the web-service tool mirConnX could be used to identify drug resistance factors in glioblastoma. We used TMZ-resistant glioblastoma cells and their integrated miRNA/mRNA networks to identify TMZ-sensitizing factors. TMZ resistance was previously induced in glioblastoma cell lines U87, Hs683, and LNZ308. miRNA/mRNA expression profiling of these cells and integration of the profiles using mirConnX resulted in the identification of plant homeodomain (PHD)-like finger 6 (PHF6) as a potential TMZ-sensitizing factor in resistant glioblastoma cells. Analysis of PHF6 expression showed significant upregulation in glioblastoma as compared to normal tissue. Interference with PHF6 expression in three TMZ-resistant subclones significantly enhanced TMZ-induced cell kill in two of these cell lines. Altogether, these results demonstrate that mirConnX is a feasible and useful tool to investigate miRNA/mRNA interactions in TMZ-resistant cells and has potential to identify drug resistance factors in glioblastoma.

  6. Identification of temozolomide resistance factors in glioblastoma via integrative miRNA/mRNA regulatory network analysis

    PubMed Central

    Hiddingh, Lotte; Raktoe, Rajiv S.; Jeuken, Judith; Hulleman, Esther; Noske, David P.; Kaspers, Gertjan J. L.; Vandertop, W. Peter; Wesseling, Pieter; Wurdinger, Thomas

    2014-01-01

    Drug resistance is a major issue in the treatment of glioblastoma. Almost all glioblastomas are intrinsically resistant to chemotherapeutic temozolomide (TMZ) or develop resistance during treatment. The interaction networks of microRNAs (miRNAs) and mRNAs likely regulate most biological processes and can be employed to better understand complex processes including drug resistance in cancer. In this study, we examined if integrative miRNA/mRNA network analysis using the web-service tool mirConnX could be used to identify drug resistance factors in glioblastoma. We used TMZ-resistant glioblastoma cells and their integrated miRNA/mRNA networks to identify TMZ-sensitizing factors. TMZ resistance was previously induced in glioblastoma cell lines U87, Hs683, and LNZ308. miRNA/mRNA expression profiling of these cells and integration of the profiles using mirConnX resulted in the identification of plant homeodomain (PHD)-like finger 6 (PHF6) as a potential TMZ-sensitizing factor in resistant glioblastoma cells. Analysis of PHF6 expression showed significant upregulation in glioblastoma as compared to normal tissue. Interference with PHF6 expression in three TMZ-resistant subclones significantly enhanced TMZ-induced cell kill in two of these cell lines. Altogether, these results demonstrate that mirConnX is a feasible and useful tool to investigate miRNA/mRNA interactions in TMZ-resistant cells and has potential to identify drug resistance factors in glioblastoma. PMID:24919120

  7. In Vivo Expansion of Regulatory T cells With IL-2/IL-2 mAb Complexes Prevents Anti-factor VIII Immune Responses in Hemophilia A Mice Treated With Factor VIII Plasmid-mediated Gene Therapy

    PubMed Central

    Liu, Chao-Lien; Ye, Peiqing; Yen, Benjamin C; Miao, Carol H

    2011-01-01

    Generation of transgene-specific immune responses can constitute a major complication following gene therapy treatment. An in vivo approach to inducing selective expansion of Regulatory T (Treg) cells by injecting interleukin-2 (IL-2) mixed with a specific IL-2 monoclonal antibody (JES6-1) was adopted to modulate anti-factor VIII (anti-FVIII) immune responses. Three consecutive IL-2 complexes treatments combined with FVIII plasmid injection prevented anti-FVIII formation and achieved persistent, therapeutic-level of FVIII expression in hemophilia A (HemA) mice. The IL-2 complexes treatment expanded CD4+CD25+Foxp3+ Treg cells five- to sevenfold on peak day, and they gradually returned to normal levels within 7–14 days without changing other lymphocyte populations. The transiently expanded Treg cells are highly activated and display suppressive function in vitro. Adoptive transfer of the expanded Treg cells protected recipient mice from generation of high-titer antibodies following FVIII plasmid challenge. Repeated plasmid transfer is applicable in tolerized mice without eliciting immune responses. Mice treated with IL-2 complexes mounted immune responses against both T-dependent and T-independent neoantigens, indicating that IL-2 complexes did not hamper the immune system for long. These results demonstrate the important role of Treg cells in suppressing anti-FVIII immune responses and the potential of developing Treg cell expansion therapies that induce long-term tolerance to FVIII. PMID:21468007

  8. Transcription factor evolution in eukaryotes and the assembly of the regulatory toolkit in multicellular lineages.

    PubMed

    de Mendoza, Alex; Sebé-Pedrós, Arnau; Šestak, Martin Sebastijan; Matejcic, Marija; Torruella, Guifré; Domazet-Loso, Tomislav; Ruiz-Trillo, Iñaki

    2013-12-10

    Transcription factors (TFs) are the main players in transcriptional regulation in eukaryotes. However, it remains unclear what role TFs played in the origin of all of the different eukaryotic multicellular lineages. In this paper, we explore how the origin of TF repertoires shaped eukaryotic evolution and, in particular, their role into the emergence of multicellular lineages. We traced the origin and expansion of all known TFs through the eukaryotic tree of life, using the broadest possible taxon sampling and an updated phylogenetic background. Our results show that the most complex multicellular lineages (i.e., those with embryonic development, Metazoa and Embryophyta) have the most complex TF repertoires, and that these repertoires were assembled in a stepwise manner. We also show that a significant part of the metazoan and embryophyte TF toolkits evolved earlier, in their respective unicellular ancestors. To gain insights into the role of TFs in the development of both embryophytes and metazoans, we analyzed TF expression patterns throughout their ontogeny. The expression patterns observed in both groups recapitulate those of the whole transcriptome, but reveal some important differences. Our comparative genomics and expression data reshape our view on how TFs contributed to eukaryotic evolution and reveal the importance of TFs to the origins of multicellularity and embryonic development. PMID:24277850

  9. Sources and biology of regulatory factors active on mouse myeloid leukemic cells

    SciTech Connect

    Metcalf, D.

    1982-01-01

    The action of serum or cells in enforcing differentiation in mouse myelomonocytic leukemic cells was monitored in agar cultures of WEHI-3B leukemic cells. The repeated intravenous injection of 5 ..mu..g endotoxin initially increased serum differentiating activity but after the third injection responses to further injections decreased markedly. Congenitally athymic (nude) mice exhibited normal rises in serum differentiating activity when injected with endotoxin but C3H HeJ mice failed to respond to challenge with purified lipid A. Whole body irradiation up to 1,200 rads did not increase serum differentiating activity but did not suppress responses to challenge injection of endotoxin. Coculture of WEHI-3B cells with peritoneal cells from normal or irradiated BALB/c mice caused marked granulocytic differentiation in WEHI-3B colonies. This effect was not seen if leukemic cells were cultured with thymus, spleen, or bone marrow cells. The serum halflife of the factor in postendotoxin serum enforcing differentiation of WEHI-3B cells was shown to be 1.5-2.3 hr.

  10. Myogenic regulatory factors during regeneration of skeletal muscle in young, adult, and old rats

    NASA Technical Reports Server (NTRS)

    Marsh, D. R.; Criswell, D. S.; Carson, J. A.; Booth, F. W.

    1997-01-01

    Myogenic factor mRNA expression was examined during muscle regeneration after bupivacaine injection in Fischer 344/Brown Norway F1 rats aged 3, 18, and 31 mo of age (young, adult, and old, respectively). Mass of the tibialis anterior muscle in the young rats had recovered to control values by 21 days postbupivacaine injection but in adult and old rats remained 40% less than that of contralateral controls at 21 and 28 days of recovery. During muscle regeneration, myogenin mRNA was significantly increased in muscles of young, adult, and old rats 5 days after bupivacaine injection. Subsequently, myogenin mRNA levels in young rat muscle decreased to postinjection control values by day 21 but did not return to control values in 28-day regenerating muscles of adult and old rats. The expression of MyoD mRNA was also increased in muscles at day 5 of regeneration in young, adult, and old rats, decreased to control levels by day 14 in young and adult rats, and remained elevated in the old rats for 28 days. In summary, either a diminished ability to downregulate myogenin and MyoD mRNAs in regenerating muscle occurs in old rat muscles, or the continuing myogenic effort includes elevated expression of these mRNAs.

  11. Macrophage-secreted factors induce adipocyte inflammation and insulin resistance

    SciTech Connect

    Permana, Paska A. . E-mail: Paska.Permana@med.va.gov; Menge, Christopher; Reaven, Peter D.

    2006-03-10

    Macrophage infiltration into adipose tissue increases with obesity, a condition associated with low-grade inflammation and insulin resistance. We investigated the direct effects of macrophage-secreted factors on adipocyte inflammation and insulin resistance. 3T3-L1 adipocytes incubated with media conditioned by RAW264.7 macrophages (RAW-CM) showed dramatically increased transcription of several inflammation-related genes, greater nuclear factor kappa B (NF-{kappa}B) activity, and enhanced binding of U937 monocytes. All of these effects were prevented by co-incubation with pyrrolidinedithiocarbamate, an NF-{kappa}B inhibitor. Adipocytes incubated with RAW-CM also released more non-esterified fatty acids and this increased lipolysis was not suppressed by insulin. In addition, RAW-CM treatment decreased insulin-stimulated glucose uptake in adipocytes. Taken together, these results indicate that macrophage-secreted factors induce inflammatory responses and reduce insulin responsiveness in adipocytes. These effects of macrophage-secreted factors on adipocytes may contribute significantly to the systemic inflammation and insulin resistance associated with obesity.

  12. Galectin-9 Ameliorates Con A-Induced Hepatitis by Inducing CD4+CD25low/int Effector T-Cell Apoptosis and Increasing Regulatory T Cell Number

    PubMed Central

    Zhang, Mengying; Zhong, Min; Suo, Qifeng

    2012-01-01

    Background T cell-mediated liver damage is a key event in the pathogenesis of many chronic human liver diseases, such as liver transplant rejection, primary biliary cirrhosis, and sclerosing cholangitis. We and other groups have previously reported that galectin-9, one of the β-galactoside binding animal lectins, might be potentially useful in the treatment of T cell-mediated diseases. To evaluate the direct effect of galectin-9 on hepatitis induced by concanavalin A (Con A) administration in mice and to clarify the mechanisms involved, we administered galectin-9 into mice, and evaluated its therapeutic effect on Con A-induced hepatitis. Methodology/Principal Findings Galectin-9 was administrated i.v. to Balb/c mice 30 min before Con A injection. Compared with no treatment, galectin-9 pretreatment significantly reduced serum ALT and AST levels and improved liver histopathology, suggesting an ameliorated hepatitis. This therapeutic effect was not only attributable to a blunted Th1 immune response, but also to an increased number in regulatory T cells, as reflected in a significantly increased apoptosis of CD4+CD25low/int effector T cells and in reduced proinflammatory cytokine levels. Conclusion/Significance Our findings constitute the first preclinical data indicating that interfering with TIM-3/galectin-9 signaling in vivo could ameliorate Con A-induced hepatitis. This strategy may represent a new therapeutic approach in treating human diseases involving T cell activation. PMID:23118999

  13. Regulatory roles of tumor necrosis factor-alpha and interleukin-1 beta in monocyte chemoattractant protein-1-mediated pulmonary granuloma formation in the rat.

    PubMed Central

    Flory, C. M.; Jones, M. L.; Miller, B. F.; Warren, J. S.

    1995-01-01

    Intravenous infusion of particulate yeast cell wall glucan into rats results in the synchronous development of angiocentric pulmonary granulomas that are composed almost entirely of monocytes and macrophages. Previous studies indicate that locally produced monocyte chemoattractant protein-1 (MCP-1) is required for full granuloma development. Because tumor necrosis factor-alpha (TNF-alpha) and interleukin 1 (IL-1) can induce MCP-1 production in a variety of cell types, we sought to determine their potential regulatory roles in this model. A single infusion of anti-TNF-alpha antibody at the time of glucan infusion (time 0) markedly reduced MCP-1 mRNA levels at 1 and 6 hours but not at later time points; there was no effect on granuloma size or number measured at 48 hours. When multiple infusions of anti-TNF-alpha antibody were administered over a 23-hour period (0 to 23 hours), MCP-1 mRNA was reduced through 24 hours, there was a significant reduction in peak bronchoalveolar lavage fluid MCP-1 activity at 48 hours, and there were marked reductions in granuloma size and number at 48 hours. Similar results were observed in animals that received infusions of anti-IL-1 beta. Infusion of anti-IL-1 beta at time 0 resulted in moderate reductions in MCP-1 mRNA at 1 and 6 hours and had no effect on granuloma size or number measured at 48 hours. When multiple infusions of anti-IL-1 beta were administered over a 23-hour period (0 to 23 hours), MCP-1 mRNA was reduced through 24 hours, there was a moderate reduction in peak bronchoalveolar lavage fluid MCP-1 activity at 48 hours, and there were marked reductions in granuloma size and number at 48 hours. A single infusion of anti-TNF-alpha and anti-IL-1 beta together at time 0 resulted in marked reductions in whole lung MCP-1 and mRNA at 1 and 6 hours, but not at 24 hours. Multiple combined infusions of anti-TNF-alpha and anti-IL-1 beta over a 23-hour period resulted in additive reductions in MCP-1 mRNA through 24 hours

  14. Regulatory T Cell Induced by Poria cocos Bark Exert Therapeutic Effects in Murine Models of Atopic Dermatitis and Food Allergy

    PubMed Central

    See, Hye-Jeong; Choi, Gyeyoung; Shon, Dong-Hwa

    2016-01-01

    The prevalence of allergic disorders including atopic dermatitis (AD) and food allergy (FA) has increased dramatically in pediatric populations, but there is no effective drug available for their management. Therefore, trials are required for the development of safe therapeutic agents such as herbal medicines. We determined whether orally administered Poria cocos bark (PCB) extract could exert immunosuppressive effects on allergic and inflammatory symptoms of AD and FA. For both AD, which was induced using house dust mite extract, and FA, which was induced by exposure to ovalbumin, model mice were orally treated with PCB extract for 62 days and 18 days, respectively. We also investigated the inductive effect of PCB extract on the generation and maintenance of Foxp3+CD4+ regulatory T cells (Tregs). The symptoms of AD and FA were ameliorated by the administration of PCB extract. Furthermore, PCB extract inhibited the Th2-related cytokines and increased the population of Foxp3+CD4+ Tregs in both AD and FA models. In ex vivo experiments, PCB extract promoted the functional differentiation of Foxp3+CD4+ Tregs, which is dependent on aryl hydrocarbon receptor activation. Thus, PCB extract has potential as an oral immune suppressor for the treatment of AD and FA through the generation of Tregs. PMID:27445434

  15. Regulatory T Cell Induced by Poria cocos Bark Exert Therapeutic Effects in Murine Models of Atopic Dermatitis and Food Allergy.

    PubMed

    Bae, Min-Jung; See, Hye-Jeong; Choi, Gyeyoung; Kang, Chang-Yuil; Shon, Dong-Hwa; Shin, Hee Soon

    2016-01-01

    The prevalence of allergic disorders including atopic dermatitis (AD) and food allergy (FA) has increased dramatically in pediatric populations, but there is no effective drug available for their management. Therefore, trials are required for the development of safe therapeutic agents such as herbal medicines. We determined whether orally administered Poria cocos bark (PCB) extract could exert immunosuppressive effects on allergic and inflammatory symptoms of AD and FA. For both AD, which was induced using house dust mite extract, and FA, which was induced by exposure to ovalbumin, model mice were orally treated with PCB extract for 62 days and 18 days, respectively. We also investigated the inductive effect of PCB extract on the generation and maintenance of Foxp3(+)CD4(+) regulatory T cells (Tregs). The symptoms of AD and FA were ameliorated by the administration of PCB extract. Furthermore, PCB extract inhibited the Th2-related cytokines and increased the population of Foxp3(+)CD4(+) Tregs in both AD and FA models. In ex vivo experiments, PCB extract promoted the functional differentiation of Foxp3(+)CD4(+) Tregs, which is dependent on aryl hydrocarbon receptor activation. Thus, PCB extract has potential as an oral immune suppressor for the treatment of AD and FA through the generation of Tregs. PMID:27445434

  16. The hypoxia-inducible factor-1α activates ectopic production of fibroblast growth factor 23 in tumor-induced osteomalacia

    PubMed Central

    Zhang, Qian; Doucet, Michele; Tomlinson, Ryan E; Han, Xiaobin; Quarles, L Darryl; Collins, Michael T; Clemens, Thomas L

    2016-01-01

    Tumor-induced osteomalacia (TIO) is a rare paraneoplastic syndrome in which ectopic production of fibroblast growth factor 23 (FGF23) by non-malignant mesenchymal tumors causes phosphate wasting and bone fractures. Recent studies have implicated the hypoxia-inducible factor-1α (HIF-1α) in other phosphate wasting disorders caused by elevated FGF23, including X-linked hypophosphatemic rickets and autosomal dominant hypophosphatemia. Here we provide evidence that HIF-1α mediates aberrant FGF23 in TIO by transcriptionally activating its promoter. Immunohistochemical studies in phosphaturic mesenchymal tumors resected from patients with documented TIO showed that HIF-1α and FGF23 were co-localized in spindle-shaped cells adjacent to blood vessels. Cultured tumor tissue produced high levels of intact FGF23 and demonstrated increased expression of HIF-1α protein. Transfection of MC3T3-E1 and Saos-2 cells with a HIF-1α expression construct induced the activity of a FGF23 reporter construct. Prior treatment of tumor organ cultures with HIF-1α inhibitors decreased HIF-1α and FGF23 protein accumulation and inhibited HIF-1α-induced luciferase reporter activity in transfected cells. Chromatin immunoprecipitation assays confirmed binding to a HIF-1α consensus sequence within the proximal FGF23 promoter, which was eliminated by treatment with a HIF-1α inhibitor. These results show for the first time that HIF-1α is a direct transcriptional activator of FGF23 and suggest that upregulation of HIF-1α activity in TIO contributes to the aberrant FGF23 production in these patients. PMID:27468359

  17. Placental Induced Growth Factor (PIGf) in Coronary Artery Disease

    NASA Technical Reports Server (NTRS)

    Sundaresan, Alamelu; Carabello, Blaise; Mehta, Satish; Schlegel, Todd; Pellis, Neal; Ott, Mark; Pierson, Duane

    2010-01-01

    Our previous studies on normal human lymphocytes have shown a five-fold increase (p less than 0.001) in angiogenic inducers such as Placental Induced Growth Factor (PIGf) in physiologically stressful environments such as modeled microgravity, a space analog. This suggests de-regulation of cardiovascular signalling pathways indicated by upregulation of PIGf. In the current study, we measured PIGf in the plasma of 33 patients with and without coronary artery disease (CAD) to investigate whether such disease is associated with increased levels of PIGf. A control consisting of 31 sex matched apparently healthy subjects was also included in the study. We observed that the levels of PIGf in CAD patients were significantly increased compared to those in healthy control subjects (p less than 0.001) and usually increased beyond the clinical threshold level (greater than 27ng/L). The mechanisms leading to up-regulation of angiogenic factors and the adaptation of organisms to stressful environments such as isolation, high altitude, hypoxia, ischemia, microgravity, increased radiation, etc are presently unknown and require further investigation in spaceflight and these other physiologically stressed environments.

  18. Regulation of erythropoiesis by hypoxia-inducible factors.

    PubMed

    Haase, Volker H

    2013-01-01

    A classic physiologic response to systemic hypoxia is the increase in red blood cell production. Hypoxia-inducible factors (HIFs) orchestrate this response by inducing cell-type specific gene expression changes that result in increased erythropoietin (EPO) production in kidney and liver, in enhanced iron uptake and utilization and in adjustments of the bone marrow microenvironment that facilitate erythroid progenitor maturation and proliferation. In particular HIF-2 has emerged as the transcription factor that regulates EPO synthesis in the kidney and liver and plays a critical role in the regulation of intestinal iron uptake. Its key function in the hypoxic regulation of erythropoiesis is underscored by genetic studies in human populations that live at high-altitude and by mutational analysis of patients with familial erythrocytosis. This review provides a perspective on recent insights into HIF-controlled erythropoiesis and iron metabolism, and examines cell types that have EPO-producing capability. Furthermore, the review summarizes clinical syndromes associated with mutations in the O(2)-sensing pathway and the genetic changes that occur in high altitude natives. The therapeutic potential of pharmacologic HIF activation for the treatment of anemia is discussed. PMID:23291219

  19. UV-B radiation induces macrophage migration inhibitory factor-mediated melanogenesis through activation of protease-activated receptor-2 and stem cell factor in keratinocytes.

    PubMed

    Enomoto, Akiko; Yoshihisa, Yoko; Yamakoshi, Takako; Ur Rehman, Mati; Norisugi, Osamu; Hara, Hiroshi; Matsunaga, Kenji; Makino, Teruhiko; Nishihira, Jun; Shimizu, Tadamichi

    2011-02-01

    UV radiation indirectly regulates melanogenesis in melanocytes through a paracrine regulatory mechanism involving keratinocytes. Protease-activated receptor (PAR)-2 activation induces melanosome transfer by increasing phagocytosis of melanosomes by keratinocytes. This study demonstrated that macrophage migration inhibitory factor (MIF) stimulated PAR-2 expression in human keratinocytes. In addition, we showed that MIF stimulated stem cell factor (SCF) release in keratinocytes; however, MIF had no effect on the release of endothelin-1 or prostaglandin E2 in keratinocytes. In addition, MIF had no direct effect on melanin and tyrosinase synthesis in cultured human melanocytes. The effect of MIF on melanogenesis was also examined using a three-dimensional reconstituted human epidermal culture model, which is a novel, commercially available, cultured human epidermis containing functional melanocytes. Migration inhibitory factor induced an increase in melanin content in the epidermis after a 9-day culture period. Moreover, melanin synthesis induced by UV-B stimulation was significantly down-regulated by anti-MIF antibody treatment. An in vivo study showed that the back skin of MIF transgenic mice had a higher melanin content than that of wild-type mice after 12 weeks of UV-B exposure. Therefore, MIF-mediated melanogenesis occurs mainly through the activation of PAR-2 and SCF expression in keratinocytes after exposure to UV-B radiation.

  20. Tumor Necrosis Factor Related Apoptosis Inducing Ligand (Trail) in endothelial response to biomechanical and biochemical stresses in arteries.

    PubMed

    D'Auria, F; Centurione, L; Centurione, M A; Angelini, A; Di Pietro, R

    2015-11-01

    Shear stress is determined by three physical components described in a famous triad: blood flow, blood viscosity and vessel geometry. Through the direct action on endothelium, shear stress is able to radically interfere with endothelial properties and the physiology of the vascular wall. Endothelial cells (ECs) have also to sustain biochemical stresses represented by chemokines, growth factors, cytokines, complement, hormones, nitric oxide (NO), oxygen and reactive oxygen species (ROS). Many growth factors, cytokines, chemokines, hormones, and chemical substances, like NO, act and regulate endothelium functions and homeostasis. Among these cytokines Tumor Necrosis Factor Related Apoptosis Inducing Ligand (TRAIL) has been assigned a regulatory role in ECs physiology and physiopathology. Thus, the aim of this review is to provide a general overview of the endothelial response pathways after different types of biomechanical and biochemical stress in in vitro models and to analyze the crucial role of TRAIL under pathological conditions of the cardiocirculatory system like atherosclerosis, coronary artery disease, and diabetes.

  1. Effects of estrogen on osteoprogenitor cells and cytokines/bone-regulatory factors in postmenopausal women.

    PubMed

    Mödder, Ulrike I; Roforth, Matthew M; Hoey, Kelley; McCready, Louise K; Peterson, James M; Monroe, David G; Oursler, Merry Jo; Khosla, Sundeep

    2011-08-01

    Decreases in estrogen levels contribute not only to early postmenopausal bone loss but also to bone loss with aging. While estrogen is critical for the maintenance of bone formation, the mechanism(s) of this effect remain unclear. Thus, we assessed the effects of 4months of transdermal estradiol treatment (0.05mg/day) of postmenopausal women as compared to no treatment (n=16 per group) on the expression of genes in pre-specified pathways in freshly isolated bone marrow osteoprogenitor cells (hematopoietic lineage [lin]-/Stro1+). We also evaluated whether estrogen treatment modulated peripheral blood or bone marrow plasma levels of the Wnt antagonists, sclerostin and DKK1, as well as serotonin, OPG, RANKL, adiponectin, oxytocin, and inflammatory cytokines (TNFα, IL-1β, and IL-6), as each of these molecules have recently been shown to play an important role in regulating osteoblast function and/or being responsive to estrogen. We observed a significant decrease in the expression of several proliferation markers (cyclin B1, cyclin E1, E2F1) and increase in adhesion molecules (N-cadherin) in bone marrow lin-/Stro1+ cells from estrogen-treated compared to control women. None of the peripheral blood or bone marrow plasma marker levels differed between the two groups, with the exception of sclerostin levels, which were significantly lower in the estrogen-treated as compared to the control women in peripheral serum (by 32%, P=0.009) and in bone marrow plasma (by 34%, P=0.017). There were significant differences in bone marrow versus peripheral plasma levels of several factors: sclerostin and OPG levels were higher in bone marrow as compared to peripheral plasma, whereas serotonin and adiponectin levels were higher in peripheral as compared to bone marrow plasma. In summary, our data directly assessing possible regulation by estrogen of osteoprogenitor cells in humans indicate that, consistent with previous studies in mice, estrogen suppresses the proliferation of human

  2. Sleep Loss as a Factor to Induce Cellular and Molecular Inflammatory Variations

    PubMed Central

    Hurtado-Alvarado, Gabriela; Castillo-García, Stephanie Ariadne; Hernández, María Eugenia; Domínguez-Salazar, Emilio; Velázquez-Moctezuma, Javier; Gómez-González, Beatriz

    2013-01-01

    A reduction in the amount of time spent sleeping occurs chronically in modern society. Clinical and experimental studies in humans and animal models have shown that immune function is impaired when sleep loss is experienced. Sleep loss exerts a strong regulatory influence on peripheral levels of inflammatory mediators of the immune response. An increasing number of research projects support the existence of reciprocal regulation between sleep and low-intensity inflammatory response. Recent studies show that sleep deficient humans and rodents exhibit a proinflammatory component; therefore, sleep loss is considered as a risk factor for developing cardiovascular, metabolic, and neurodegenerative diseases (e.g., diabetes, Alzheimer's disease, and multiple sclerosis). Circulating levels of proinflammatory mediators depend on the intensity and duration of the method employed to induce sleep loss. Recognizing the fact that the concentration of proinflammatory mediators is different between acute and chronic sleep-loss may expand the understanding of the relationship between sleep and the immune response. The aim of this review is to integrate data from recent published reports (2002–2013) on the effects of sleep loss on the immune response. This review may allow readers to have an integrated view of the mechanisms involved in central and peripheral deficits induced by sleep loss. PMID:24367384

  3. Basolateral Na+/HCO3– cotransport activity is regulated by the dissociable Na+/H+ exchanger regulatory factor

    PubMed Central

    Bernardo, Angelito A.; Kear, Felicidad T.; Santos, Anna V.P.; Ma, Jianfei; Steplock, Debra; Robey, R. Brooks; Weinman, Edward J.

    1999-01-01

    In the renal proximal tubule, the activities of the basolateral Na+/HCO3– cotransporter (NBC) and the apical Na+/H+ exchanger (NHE3) uniformly vary in parallel, suggesting that they are coordinately regulated. PKA-mediated inhibition of NHE3 is mediated by a PDZ motif–containing protein, the Na+/H+ exchanger regulatory factor (NHE-RF). Given the common inhibition of these transporters after protein kinase A (PKA) activation, we sought to determine whether NHE-RF also plays a role in PKA-regulated NBC activity. Renal cortex immunoblot analysis using anti-peptide antibodies directed against rabbit NHE-RF demonstrated the presence of this regulatory factor in both brush-border membranes (BBMs) and basolateral membranes (BLMs). Using a reconstitution assay, we found that limited trypsin digestion of detergent solubilized rabbit renal BLM preparations resulted in NBC activity that was unaffected by PKA activation. Co-reconstitution of these trypsinized preparations with a recombinant protein corresponding to wild-type rabbit NHE-RF restored the inhibitory effect of PKA on NBC activity in a concentration-dependent manner. NBC activity was inhibited 60% by 10–8M NHE-RF; this effect was not observed in the absence of PKA. Reconstitution with heat-denatured NHE-RF also failed to attenuate NBC activity. To establish further a physiologic role for NHE-RF in NBC regulation, the renal epithelial cell line B-SC-1, which lacks detectable endogenous NHE-RF expression, was engineered to express stably an NHE-RF transgene. NHE-RF–expressing B-SC-1 cells (B-SC-RF) exhibited markedly lower basal levels of NBC activity than did wild-type controls. Inhibition of NBC activity in B-SC-RF cells was enhanced after 10 μM of forskolin treatment, consistent with a postulated role for NHE-RF in mediating the inhibition of NBC activity by PKA. These findings not only suggest NHE-RF involvement in PKA-regulated NBC activity, but also provide a unique molecular mechanism whereby

  4. Ganglioside GM1 Deficiency in Effector T Cells From NOD Mice Induces Resistance to Regulatory T-Cell Suppression

    PubMed Central

    Wu, Gusheng; Lu, Zi-Hua; Gabius, Hans-Joachim; Ledeen, Robert W.; Bleich, David

    2011-01-01

    OBJECTIVE To detect GM1 deficiency and determine its role in effector T cells (Teffs) from NOD mice in establishing resistance to regulatory T-cell (Treg) suppression. RESEARCH DESIGN AND METHODS CD4+ and CD8+ Teffs were isolated from spleens of prediabetic NOD mice for comparison with similar cells from Balb/c, C57BL/6, and NOR mice. GM1 was quantified with thin-layer chromatography for total cellular GM1 and flow cytometry for cell-surface GM1. Suppression of Teff proliferation was determined by application of GM1 cross-linking agents or coculturing with Tregs. Calcium influx in Teffs was quantified using fura-2. RESULTS Resting and activated CD4+ and CD8+ Teffs of NOD mice contained significantly less GM1 than Teffs from the other three mouse strains tested. After activation, NOD Teffs resisted suppression by Tregs or GM1 cross-linking agents in contrast to robust suppression of Balb/c Teffs; this was reversed by preincubation of NOD Teffs with GM1. NOD Teffs also showed attenuated Ca2+ influx via transient receptor potential channel 5 (TRPC5) channels induced by GM1 cross-linking, and this, too, was reversed by elevation of Teff GM1. CONCLUSIONS GM1 deficiency occurs in NOD Teffs and contributes importantly to failed suppression, which is rectified by increasing Teff GM1. Such elevation also reverses subthreshold Ca2+ influx via TRPC5 channels, an essential aspect of suppression. Our results also support a critical role for galectin-1 as a GM1 cross-linking counter-receptor that fittingly is upregulated and released by Tregs during activation. These findings suggest a novel mechanism by which pathogenic Teffs evade regulatory suppression, thereby leading to autoimmune β-cell destruction and type 1 diabetes. PMID:21788572

  5. Inducible nitric oxide synthase (NOS-2) in subarachnoid hemorrhage: Regulatory mechanisms and therapeutic implications

    PubMed Central

    Iqbal, Sana; Hayman, Erik G; Hong, Caron; Stokum, Jesse A; Kurland, David B; Gerzanich, Volodymyr; Simard, J Marc

    2016-01-01

    Aneurysmal subarachnoid hemorrhage (SAH) typically carries a poor prognosis. Growing evidence indicates that overabundant production of nitric oxide (NO) may be responsible for a large part of the secondary injury that follows SAH. Although SAH modulates the activity of all three isoforms of nitric oxide synthase (NOS), the inducible isoform, NOS-2, accounts for a majority of NO-mediated secondary injuries after SAH. Here, we review the indispensable physiological roles of NO that must be preserved, even while attempting to downmodulate the pathophysiologic effects of NO that are induced by SAH. We examine the effects of SAH on the function of the various NOS isoforms, with a particular focus on the pathological effects of NOS-2 and on the mechanisms responsible for its transcriptional upregulation. Finally, we review interventions to block NOS-2 upregulation or to counteract its effects, with an emphasis on the potential therapeutic strategies to improve outcomes in patients afflicted with SAH. There is still much to be learned regarding the apparently maladaptive response of NOS-2 and its harmful product NO in SAH. However, the available evidence points to crucial effects that, on balance, are adverse, making the NOS-2/NO/peroxynitrite axis an attractive therapeutic target in SAH. PMID:27774520

  6. Induced Disruption of the Iron-Regulatory Hormone Hepcidin Inhibits Acute Inflammatory Hypoferraemia.

    PubMed

    Armitage, Andrew E; Lim, Pei Jin; Frost, Joe N; Pasricha, Sant-Rayn; Soilleux, Elizabeth J; Evans, Emma; Morovat, Alireza; Santos, Ana; Diaz, Rebeca; Biggs, Daniel; Davies, Benjamin; Gileadi, Uzi; Robbins, Peter A; Lakhal-Littleton, Samira; Drakesmith, Hal

    2016-01-01

    Withdrawal of iron from serum (hypoferraemia) is a conserved innate immune antimicrobial strategy that can withhold this critical nutrient from invading pathogens, impairing their growth. Hepcidin (Hamp1) is the master regulator of iron and its expression is induced by inflammation. Mice lacking Hamp1 from birth rapidly accumulate iron and are susceptible to infection by blood-dwelling siderophilic bacteria such as Vibrio vulnificus. In order to study the innate immune role of hepcidin against a background of normal iron status, we developed a transgenic mouse model of tamoxifen-sensitive conditional Hamp1 deletion (termed iHamp1-KO mice). These mice attain adulthood with an iron status indistinguishable from littermate controls. Hamp1 disruption and the consequent decline of serum hepcidin concentrations occurred within hours of a single tamoxifen dose. We found that the TLR ligands LPS and Pam3CSK4 and heat-killed Brucella abortus caused an equivalent induction of inflammation in control and iHamp1-KO mice. Pam3CSK4 and B. abortus only caused a drop in serum iron in control mice, while hypoferraemia due to LPS was evident but substantially blunted in iHamp1-KO mice. Our results characterise a powerful new model of rapidly inducible hepcidin disruption, and demonstrate the critical contribution of hepcidin to the hypoferraemia of inflammation. PMID:27423740

  7. Induced regulatory T cells in allograft tolerance via transient mixed chimerism

    PubMed Central

    Hotta, Kiyohiko; Aoyama, Akihiro; Oura, Tetsu; Yamada, Yohei; Tonsho, Makoto; Huh, Kyu Ha; Kawai, Kento; Schoenfeld, David; Allan, James S.; Madsen, Joren C.; Benichou, Gilles; Smith, Rex-Neal; Colvin, Robert B.; Sachs, David H.; Cosimi, A. Benedict

    2016-01-01

    Successful induction of allograft tolerance has been achieved in nonhuman primates (NHPs) and humans via induction of transient hematopoietic chimerism. Since allograft tolerance was achieved in these recipients without durable chimerism, peripheral mechanisms are postulated to play a major role. Here, we report our studies of T cell immunity in NHP recipients that achieved long-term tolerance versus those that rejected the allograft (AR). All kidney, heart, and lung transplant recipients underwent simultaneous or delayed donor bone marrow transplantation (DBMT) following conditioning with a nonmyeloablative regimen. After DBMT, mixed lymphocyte culture with CFSE consistently revealed donor-specific loss of CD8+ T cell responses in tolerant (TOL) recipients, while marked CD4+ T cell proliferation in response to donor antigens was found to persist. Interestingly, a significant proportion of the proliferated CD4+ cells were FOXP3+ in TOL recipients, but not in AR or naive NHPs. In TOL recipients, CD4+FOXP3+ cell proliferation against donor antigens was greater than that observed against third-party antigens. Finally, the expanded Tregs appeared to be induced Tregs (iTregs) that were converted from non-Tregs. These data provide support for the hypothesis that specific induction of iTregs by donor antigens is key to long-term allograft tolerance induced by transient mixed chimerism. PMID:27446989

  8. The effect of nutritional status and myogenic satellite cell age on turkey satellite cell proliferation, differentiation, and expression of myogenic transcriptional regulatory factors and heparan sulfate proteoglycans syndecan-4 and glypican-1.

    PubMed

    Harthan, Laura B; McFarland, Douglas C; Velleman, Sandra G

    2014-01-01

    Posthatch satellite cell mitotic activity is a critical component of muscle development and growth. Satellite cells are myogenic stem cells that can be induced by nutrition to follow other cellular developmental pathways, and whose mitotic activity declines with age. The objective of the current study was to determine the effect of restricting protein synthesis on the proliferation and differentiation, expression of myogenic transcriptional regulatory factors myogenic determination factor 1, myogenin, and myogenic regulatory factor 4, and expression of the heparan sulfate proteoglycans syndecan-4 and glypican-1 in satellite cells isolated from 1-d-, 7-wk-, and 16-wk-old turkey pectoralis major muscle (1 d, 7 wk, and 16 wk cells, respectively) by using variable concentrations of Met and Cys. Four Met concentrations-30 (control), 7.5, 3, or 0 mg/L with 3.2 mg/L of Cys per 1 mg/L of Met-were used for culture of satellite cells to determine the effect of nutrition and age on satellite cell behavior during proliferation and differentiation. Proliferation was reduced by lower Met and Cys concentrations in all ages at 96 h of proliferation. Differentiation was increased in the 1 d Met-restricted cells, whereas the 7 wk cells treated with 3 mg/L of Met had decreased differentiation. Reduced Met and Cys levels from the control did not significantly affect the 16 wk cells at 72 h of differentiation. However, medium with no Met or Cys suppressed differentiation at all ages. The expression of myogenic determination factor 1, myogenin, myogenic regulatory factor 4, syndecan-4, and glypican-1 was differentially affected by age and Met or Cys treatment. These data demonstrate the age-specific manner in which turkey pectoralis major muscle satellite cells respond to nutritional availability and the importance of defining optimal nutrition to maximize satellite cell proliferation and differentiation for subsequent muscle mass accretion.

  9. Discovery of transcription factors and regulatory regions driving in vivo tumor development by ATAC-seq and FAIRE-seq open chromatin profiling.

    PubMed

    Davie, Kristofer; Jacobs, Jelle; Atkins, Mardelle; Potier, Delphine; Christiaens, Valerie; Halder, Georg; Aerts, Stein

    2015-02-01

    Genomic enhancers regulate spatio-temporal gene expression by recruiting specific combinations of transcription factors (TFs). When TFs are bound to active regulatory regions, they displace canonical nucleosomes, making these regions biochemically detectable as nucleosome-depleted regions or accessible/open chromatin. Here we ask whether open chromatin profiling can be used to identify the entire repertoire of active promoters and enhancers underlying tissue-specific gene expression during normal development and oncogenesis in vivo. To this end, we first compare two different approaches to detect open chromatin in vivo using the Drosophila eye primordium as a model system: FAIRE-seq, based on physical separation of open versus closed chromatin; and ATAC-seq, based on preferential integration of a transposon into open chromatin. We find that both methods reproducibly capture the tissue-specific chromatin activity of regulatory regions, including promoters, enhancers, and insulators. Using both techniques, we screened for regulatory regions that become ectopically active during Ras-dependent oncogenesis, and identified 3778 regions that become (over-)activated during tumor development. Next, we applied motif discovery to search for candidate transcription factors that could bind these regions and identified AP-1 and Stat92E as key regulators. We validated the importance of Stat92E in the development of the tumors by introducing a loss of function Stat92E mutant, which was sufficient to rescue the tumor phenotype. Additionally we tested if the predicted Stat92E responsive regulatory regions are genuine, using ectopic induction of JAK/STAT signaling in developing eye discs, and observed that similar chromatin changes indeed occurred. Finally, we determine that these are functionally significant regulatory changes, as nearby target genes are up- or down-regulated. In conclusion, we show that FAIRE-seq and ATAC-seq based open chromatin profiling, combined with motif

  10. Non-small-cell lung cancer-induced immunosuppression by increased human regulatory T cells via Foxp3 promoter demethylation.

    PubMed

    Ke, Xing; Zhang, Shuping; Xu, Jian; Liu, Genyan; Zhang, Lixia; Xie, Erfu; Gao, Li; Li, Daqian; Sun, Ruihong; Wang, Fang; Pan, Shiyang

    2016-05-01

    Patients with non-small-cell lung cancer (NSCLC) have immune defects that are poorly understood. Forkhead box protein P3 (Foxp3) is crucial for immunosuppression by CD4(+) regulatory T cells (Tregs). It is not well known how NSCLC induces Foxp3 expression and causes immunosuppression in tumor-bearing patients. Our study found a higher percentage of CD4(+) Tregs in the peripheral blood of NSCLC compared with healthy donors. NSCLC patients showed demethylation of eight CpG sites within the Foxp3 promoter with methylation ratios negatively correlated with CD4(+)CD25(+)Foxp3(+) T levels. Foxp3 expression in CD4(+) Tregs was directly regulated by Foxp3 promoter demethylation and was involved in immunosuppression by NSCLC. To verify the effect of tumor cells on the phenotype and function of CD4(+) Tregs, we established a coculture system using NSCLC cell line and healthy CD4(+) T cells and showed that SPC-A1 induced IL-10 and TGF-β1 secretion by affecting the function of CD4(+) Tregs. The activity of DNA methyltransferases from CD4(+) T was decreased during this process. Furthermore, eight CpG sites within the Foxp3 promoter also appeared to have undergone demethylation. Foxp3 is highly expressed in CD4(+) T cells, and this may be caused by gene promoter demethylation. These induced Tregs are highly immunosuppressive and dramatically inhibit the proliferative activity of naïve CD4(+) T cells. Our study provides one possible mechanism describing Foxp3 promoter demethylation changes by which NSCLC down-regulates immune responses and contributes to tumor progression. Foxp3 represents an important target for NSCLC anti-tumor immunotherapy.

  11. Tumor necrosis factor alpha induces the expression of transforming growth factor alpha and the epidermal growth factor receptor in human pancreatic cancer cells.

    PubMed Central

    Schmiegel, W; Roeder, C; Schmielau, J; Rodeck, U; Kalthoff, H

    1993-01-01

    Recombinant human tumor necrosis factor (TNF)-alpha increased the expression of epidermal growth factor receptor (EGFR) mRNA and protein in all of six human pancreatic carcinoma cell lines tested. In addition, TNF-alpha increased the expression of an EGFR ligand, transforming growth factor (TGF)-alpha, at the mRNA and protein level in all cell lines. Increased expression of EGFR protein was associated with elevated steady-state EGFR mRNA levels. Nuclear run-on analysis showed that increase in EGFR mRNA was due to an increased rate of transcription. Induction of EGFR mRNA expression by TNF-alpha was abrogated by cycloheximide but occurred independently of TNF-alpha-induced production of TGF-alpha protein. Protein kinase A or Gi-type guanine nucleotide-binding proteins were not involved in this process as assessed by using appropriate stimulators and inhibitors of these signal transduction pathways. By contrast, staurosporine, an inhibitor of protein kinase C, partially inhibited, and 4-bromophenacyl bromide, a phospholipase inhibitor, completely inhibited TNF-alpha-dependent EGFR mRNA expression. The phospholipase C-specific inhibitor tricyclodecan-9-yl xanthogenate did not alter TNF-alpha-dependent EGFR mRNA expression, suggesting that phospholipase A2 is involved in the modulation of EGFR expression by TNF-alpha. The simultaneous induction of a ligand/receptor system by TNF-alpha suggests that this cytokine modulates autocrine growth-regulatory pathways in pancreatic cancer cells. Images PMID:8430098

  12. Growth Factors and Tension-Induced Skeletal Muscle Growth

    NASA Technical Reports Server (NTRS)

    Vandenburgh, Herman H.

    1994-01-01

    The project investigated biochemical mechanisms to enhance skeletal muscle growth, and developed a computer based mechanical cell stimulator system. The biochemicals investigated in this study were insulin/(Insulin like Growth Factor) IGF-1 and Steroids. In order to analyze which growth factors are essential for stretch-induced muscle growth in vitro, we developed a defined, serum-free medium in which the differentiated, cultured avian muscle fibers could be maintained for extended periods of time. The defined medium (muscle maintenance medium, MM medium) maintains the nitrogen balance of the myofibers for 3 to 7 days, based on myofiber diameter measurements and myosin heavy chain content. Insulin and IGF-1, but not IGF-2, induced pronounced myofiber hypertrophy when added to this medium. In 5 to 7 days, muscle fiber diameters increase by 71 % to 98% compared to untreated controls. Mechanical stimulation of the avian muscle fibers in MM medium increased the sensitivity of the cells to insulin and IGF-1, based on a leftward shift of the insulin dose/response curve for protein synthesis rates. (54). We developed a ligand binding assay for IGF-1 binding proteins and found that the avian skeletal muscle cultures produced three major species of 31, 36 and 43 kD molecular weight (54) Stretch of the myofibers was found to have no significant effect on the efflux of IGF-1 binding proteins, but addition of exogenous collagen stimulated IGF-1 binding protein production 1.5 to 5 fold. Steroid hormones have a profound effect on muscle protein turnover rates in vivo, with the stress-related glucocorticoids inducing rapid skeletal muscle atrophy while androgenic steroids induce skeletal muscle growth. Exercise in humans and animals reduces the catabolic effects of glucocorticoids and may enhance the anabolic effects of androgenic steroids on skeletal muscle. In our continuing work on the involvement of exogenrus growth factors in stretch-induced avian skeletal muscle growth, we

  13. Self-regulatory role of 4-hydroxynonenal in signaling for stress-induced programmed cell death.

    PubMed

    Awasthi, Yogesh C; Sharma, Rajendra; Sharma, Abha; Yadav, Sushma; Singhal, Sharad S; Chaudhary, Pankaj; Awasthi, Sanjay

    2008-07-15

    Within the last two decades, 4-hydroxynonenal has emerged as an important second messenger involved in the regulation of various cellular processes. Our recent studies suggest that HNE can induce apoptosis in various cells through the death receptor Fas (CD95)-mediated extrinsic pathway as well as through the p53-dependent intrinsic pathway. Interestingly, through its interaction with the nuclear protein Daxx, HNE can self-limit its apoptotic role by translocating Daxx to cytoplasm where it binds to Fas and inhibits Fas-mediated apoptosis. In this paper, after briefly describing recent studies on various biological activities of HNE, based on its interactions with Fas, Daxx, and p53, we speculate on possible mechanisms through which HNE may affect a multitude of cellular processes and draw a parallel between signaling roles of H(2)O(2) and HNE.

  14. Role of IL-10-producing regulatory B cells in modulating T-helper cell immune responses during silica-induced lung inflammation and fibrosis

    PubMed Central

    Liu, Fangwei; Dai, Wujing; Li, Chao; Lu, Xiaowei; Chen, Ying; Weng, Dong; Chen, Jie

    2016-01-01

    Silicosis is characterized by chronic lung inflammation and fibrosis, which are seriously harmful to human health. Previous research demonstrated that uncontrolled T-helper (Th) cell immune responses were involved in the pathogenesis of silicosis. Lymphocytes also are reported to have important roles. Existing studies on lymphocyte regulation of Th immune responses were limited to T cells, such as the regulatory T (Treg) cell, which could negatively regulate inflammation and promote the process of silicosis. However, other regulatory subsets in silicosis have not been investigated in detail, and the mechanism of immune homeostasis modulation needs further exploration. Another regulatory lymphocyte, the regulatory B cell, has recently drawn increasing attention. In this study, we comprehensively showed the role of IL-10-producing regulatory B cell (B10) in a silicosis model of mice. B10 was inducible by silica instillation. Insufficient B10 amplified inflammation and attenuated lung fibrosis by promoting the Th1 immune response. Insufficient B10 clearly inhibited Treg and decreased the level of IL-10. Our study indicated that B10 could control lung inflammation and exacerbate lung fibrosis by inhibiting Th1 response and modulating the Th balance. The regulatory function of B10 could be associated with Treg induction and IL-10 secretion. PMID:27354007

  15. Regulatory regions that control expression of two chloramphenicol-inducible cat genes cloned in Bacillus subtilis.

    PubMed Central

    Duvall, E J; Williams, D M; Mongkolsuk, S; Lovett, P S

    1984-01-01

    Plasmid pPL603 is a promoter cloning vector for Bacillus subtilis and consists of a 1.1-kilobase fragment of Bacillus pumilus DNA inserted between the EcoRI and BamHI sites of pUB110. The gene cat-86, specifying chloramphenicol-inducible chloramphenicol acetyltransferase, is located on the 1.1-kilobase cloned DNA. When pPL603 is present in B. subtilis, cat-86 is unexpressed during vegetative growth but expressed during sporulation. The regulation of cat-86 in pPL603 is due to sequences within two restriction fragments, designated P1 and R1, that precede the main coding portion of the gene. The P1 fragment promotes transcription of cat-86 only during sporulation, whereas the adjacent R1 fragment lacks promoter function but contains sequences essential to chloramphenicol inducibility. A second B. pumilus gene, cat-66, was cloned in B. subtilis and is expressed throughout the vegetative growth and sporulation cycle. The cat-66 coding region is preceded by two adjacent restriction fragments designated as P2 and R2. P1 and P2 are identical in size and share 95% conservation of base sequence. R1 and R2 are also identical in size and share 91% conservation of base sequence. Fragment substitution experiments demonstrate that R2 can functionally replace R1. The substitution of P2 for P1 promotes cat-86 expression throughout vegetative growth and sporulation. Analysis of a derivative of pPL603 in which P2 has replaced P1 demonstrates that P2 promotes transcription of cat-86 during vegetative growth and that P2 contains the start site for transcription of cat-86. Thus, P1 and P2 differ strikingly in vegetative promoter function, yet they differ by single-base substitutions at only 11 positions of 203. Images PMID:6327638

  16. Overexpression of steroidogenic acute regulatory protein in rat aortic endothelial cells attenuates palmitic acid-induced inflammation and reduction in nitric oxide bioavailability

    PubMed Central

    2012-01-01

    Background Endothelial dysfunction is a well documented evidence for the onset of atherosclerosis and other cardiovascular diseases. Lipids disorder is among the main risk factors for endothelial dysfunction in these diseases. Steroidogenic acute regulatory protein (StAR), one of the cholesterol transporters, plays an important role in the maintenance of intracellular lipid homeostasis. However, the effect of StAR on endothelial dysfunction is not well understood. Palmitic acid (PA) has been shown to decrease eNOS activity and induce inflammation, both are the causes of endothelial dysfunction, in an endothelial cell culture model. Methods StAR gene was introduced into primary rat aortic endothelial cells by adenovirus infection. Real-time PCR and Western blotting were performed to determine the relative genes and proteins expression level to elucidate the underlying mechanism. The free fatty acid and cholesterol quantification kits were used to detect total cellular free fatty acid and cholesterol. The levels of inflammatory factors and nitric oxide were determined by ELISA and classic Griess reagent methods respectively. Results We successfully overexpressed StAR in primary rat aortic endothelial cells. Following StAR overexpression, mRNA levels of IL-1β, TNFα, IL6 and VCAM-1 and protein levels of IL-1β, , TNFα and IL-6 in culture supernatant were significantly decreased, which duing to blocke NFκB nuclear translocation and activation. Moreover, StAR overexpression attenuated the PA-induced reduction of nitric oxide bioavailability by protecting the bioactivity of pAkt/peNOS/NO pathway. Furthermore, the key genes involved in lipid metabolism were greatly reduced following StAR overexpression. In order to investigate the underlying mechanism, cerulenin and lovastatin, the inhibitor of fatty acid and cholesterol synthase, were added prior to PA treatment. The results showed that both cerulenin and lovastatin had a similar effect as StAR overexpression. On the

  17. Regulatory SNPs and transcriptional factor binding sites in ADRBK1, AKT3, ATF3, DIO2, TBXA2R and VEGFA

    PubMed Central

    Buroker, Norman E

    2014-01-01

    Abstract Regulatory single nucleotide polymorphisms (rSNPs) which change the transcriptional factor binding sites (TFBS) for transcriptional factors (TFs) to bind DNA were reviewed for the ADRBK1 (GRK2), AKT3, ATF3, DIO2, TBXA2R and VEGFA genes. Changes in the TFBS where TFs attach to regulate these genes may result in human sickness and disease. The highlights of this previous work were reviewed for these genes. PMID:25483406

  18. Auxin Response Factor SlARF2 Is an Essential Component of the Regulatory Mechanism Controlling Fruit Ripening in Tomato.

    PubMed

    Hao, Yanwei; Hu, Guojian; Breitel, Dario; Liu, Mingchun; Mila, Isabelle; Frasse, Pierre; Fu, Yongyao; Aharoni, Asaph; Bouzayen, Mondher; Zouine, Mohamed

    2015-12-01

    Ethylene is the main regulator of climacteric fruit ripening, by contrast the putative role of other phytohormones in this process remains poorly understood. The present study brings auxin signaling components into the mechanism regulating tomato fruit ripening through the functional characterization of Auxin Response Factor2 (SlARF2) which encodes a downstream component of auxin signaling. Two paralogs, SlARF2A and SlARF2B, are found in the tomato genome, both displaying a marked ripening-associated expression but distinct responsiveness to ethylene and auxin. Down-regulation of either SlARF2A or SlARF2B resulted in ripening defects while simultaneous silencing of both genes led to severe ripening inhibition suggesting a functional redundancy among the two ARFs. Tomato fruits under-expressing SlARF2 produced less climacteric ethylene and exhibited a dramatic down-regulation of the key ripening regulators RIN, CNR, NOR and TAGL1. Ethylene treatment failed to reverse the non-ripening phenotype and the expression of ethylene signaling and biosynthesis genes was strongly altered in SlARF2 down-regulated fruits. Although both SlARF proteins are transcriptional repressors the data indicate they work as positive regulators of tomato fruit ripening. Altogether, the study defines SlARF2 as a new component of the regulatory network controlling the ripening process in tomato. PMID:26716451

  19. The PDZ Protein Na+/H+ Exchanger Regulatory Factor-1 (NHERF1) Regulates Planar Cell Polarity and Motile Cilia Organization

    PubMed Central

    Stolz, Donna B.; Tsang, Michael; Friedman, Peter A.; Romero, Guillermo

    2016-01-01

    Directional flow of the cerebrospinal fluid requires coordinated movement of the motile cilia of the ependymal epithelium that lines the cerebral ventricles. Here we report that mice lacking the Na+/H+ Exchanger Regulatory Factor 1 (NHERF1/Slc9a3r1, also known as EBP50) develop profound communicating hydrocephalus associated with fewer and disorganized ependymal cilia. Knockdown of NHERF1/slc9a3r1 in zebrafish embryos also causes severe hydrocephalus of the hindbrain and impaired ciliogenesis in the otic vesicle. Ultrastructural analysis did not reveal defects in the shape or organization of individual cilia. Similar phenotypes have been described in animals with deficiencies in Wnt signaling and the Planar Cell Polarity (PCP) pathway. We show that NHERF1 binds the PCP core genes Frizzled (Fzd) and Vangl. We further show that NHERF1 assembles a ternary complex with Fzd4 and Vangl2 and promotes translocation of Vangl2 to the plasma membrane, in particular to the apical surface of ependymal cells. Taken together, these results strongly support an important role for NHERF1 in the regulation of PCP signaling and the development of functional motile cilia. PMID:27055101

  20. Auxin Response Factor SlARF2 Is an Essential Component of the Regulatory Mechanism Controlling Fruit Ripening in Tomato

    PubMed Central

    Hao, Yanwei; Hu, Guojian; Breitel, Dario; Liu, Mingchun; Mila, Isabelle; Frasse, Pierre; Fu, Yongyao; Aharoni, Asaph; Bouzayen, Mondher; Zouine, Mohamed

    2015-01-01

    Ethylene is the main regulator of climacteric fruit ripening, by contrast the putative role of other phytohormones in this process remains poorly understood. The present study brings auxin signaling components into the mechanism regulating tomato fruit ripening through the functional characterization of Auxin Response Factor2 (SlARF2) which encodes a downstream component of auxin signaling. Two paralogs, SlARF2A and SlARF2B, are found in the tomato genome, both displaying a marked ripening-associated expression but distinct responsiveness to ethylene and auxin. Down-regulation of either SlARF2A or SlARF2B resulted in ripening defects while simultaneous silencing of both genes led to severe ripening inhibition suggesting a functional redundancy among the two ARFs. Tomato fruits under-expressing SlARF2 produced less climacteric ethylene and exhibited a dramatic down-regulation of the key ripening regulators RIN, CNR, NOR and TAGL1. Ethylene treatment failed to reverse the non-ripening phenotype and the expression of ethylene signaling and biosynthesis genes was strongly altered in SlARF2 down-regulated fruits. Although both SlARF proteins are transcriptional repressors the data indicate they work as positive regulators of tomato fruit ripening. Altogether, the study defines SlARF2 as a new component of the regulatory network controlling the ripening process in tomato. PMID:26716451

  1. Virulence factor regulation and regulatory networks in Streptococcus pyogenes and their impact on pathogen-host interactions.

    PubMed

    Kreikemeyer, Bernd; McIver, Kevin S; Podbielski, Andreas

    2003-05-01

    Streptococcus pyogenes (group A streptococcus, GAS) is a very important human pathogen with remarkable adaptation capabilities. Survival within the harsh host surroundings requires sensing potential on the bacterial side, which leads in particular to coordinately regulated virulence factor expression. GAS 'stand-alone' response regulators (RRs) and two-component signal transduction systems (TCSs) link the signals from the host environment with adaptive responses of the bacterial cell. Numerous putative regulatory systems emerged from GAS genome sequences. Only three RRs [Mga, RofA-like protein (RALP) and Rgg/RopB] and three TCSs (CsrRS/CovRS, FasBCAX and Ihk/Irr) have been studied in some detail with respect to their growth-phase-dependent activity and their influence on GAS-host cell interaction. In particular, the Mga-, RALP- and Rgg/RopB-regulated pathways display interconnected activities that appear to influence GAS colonization, persistence and spreading mechanisms, in a growth-phase-related fashion. Here, we have summarized our current knowledge about these RRs and TCSs to highlight the questions that should be addressed in future research on GAS pathogenicity.