In the right place at the right time: visualizing and understanding mRNA localization

PubMed Central

Buxbaum, Adina R.; Haimovich, Gal

2015-01-01

The spatial regulation of protein translation is an efficient way to create functional and structural asymmetries in cells. Recent research has furthered our understanding of how individual cells spatially organize protein synthesis, by applying innovative technology to characterize the relationship between mRNAs and their regulatory proteins, single-mRNA trafficking dynamics, physiological effects of abrogating mRNA localization in vivo and for endogenous mRNA labelling. The implementation of new imaging technologies has yielded valuable information on mRNA localization, for example, by observing single molecules in tissues. The emerging movements and localization patterns of mRNAs in morphologically distinct unicellular organisms and in neurons have illuminated shared and specialized mechanisms of mRNA localization, and this information is complemented by transgenic and biochemical techniques that reveal the biological consequences of mRNA mislocalization. PMID:25549890

Identification of Fat Mass and Obesity Associated (FTO) Protein Expression in Cardiomyocytes: Regulation by Leptin and Its Contribution to Leptin-Induced Hypertrophy

PubMed Central

Gan, Xiaohong Tracey; Zhao, Ganjian; Huang, Cathy X.; Rowe, Adrianna C.; Purdham, Daniel M.; Karmazyn, Morris

2013-01-01

The recently-identified fat mass and obesity-associated (FTO) protein is associated with various physiological functions including energy and body weight regulation. Ubiquitously expressed, FTO was identified in heart homogenates although its function is unknown. We studied whether FTO is specifically expressed within the cardiac myocyte and its potential role pertaining to the hypertrophic effect of the adipokine leptin. Most experiments were performed using cultured neonatal rat cardiomyocytes which showed nuclei-specific FTO expression. Leptin significantly increased FTO expression which was associated with myocyte hypertrophy although both events were abrogated by FTO knockdown with siRNA. Administration of a leptin receptor antibody to either normal or obese rats significant reduced myocardial FTO protein expression. Responses in cardiomyocytes were accompanied by JAK2/STAT3 activation whereas JAK2/STAT3 inhibition abolished these effects. Expression of the cut-like homeobox 1(CUX1) transcriptional factor was significantly increased by leptin although this was restricted to the cathepsin L-dependent, proteolytically-derived shorter p110CUX1 isoform whereas the longer p200CUX1 protein was not significantly affected. Cathepsin L expression and activity were both significantly increased by leptin whereas a cathepsin L peptide inhibitor or siRNA specific for CUX1 completely prevented the leptin-induced increase in FTO expression. The cathepsin L peptide inhibitor or siRNA-induced knockdown of either CUX1 or FTO abrogated the hypertrophic response to leptin. Two other pro-hypertrophic factors, endothelin-1 or angiotensin II had no effect on FTO expression and FTO knockdown did not alter the hypertrophic response to either agent. This study demonstrates leptin-induced FTO upregulation in cardiomyocytes via JAK2/STAT3- dependent CUX1 upregulation and suggests an FTO regulatory function of leptin. It also demonstrates for the first time a functional role of FTO in the cardiomyocyte. PMID:24019958

Identification of fat mass and obesity associated (FTO) protein expression in cardiomyocytes: regulation by leptin and its contribution to leptin-induced hypertrophy.

PubMed

Gan, Xiaohong Tracey; Zhao, Ganjian; Huang, Cathy X; Rowe, Adrianna C; Purdham, Daniel M; Karmazyn, Morris

2013-01-01

The recently-identified fat mass and obesity-associated (FTO) protein is associated with various physiological functions including energy and body weight regulation. Ubiquitously expressed, FTO was identified in heart homogenates although its function is unknown. We studied whether FTO is specifically expressed within the cardiac myocyte and its potential role pertaining to the hypertrophic effect of the adipokine leptin. Most experiments were performed using cultured neonatal rat cardiomyocytes which showed nuclei-specific FTO expression. Leptin significantly increased FTO expression which was associated with myocyte hypertrophy although both events were abrogated by FTO knockdown with siRNA. Administration of a leptin receptor antibody to either normal or obese rats significant reduced myocardial FTO protein expression. Responses in cardiomyocytes were accompanied by JAK2/STAT3 activation whereas JAK2/STAT3 inhibition abolished these effects. Expression of the cut-like homeobox 1(CUX1) transcriptional factor was significantly increased by leptin although this was restricted to the cathepsin L-dependent, proteolytically-derived shorter p110CUX1 isoform whereas the longer p200CUX1 protein was not significantly affected. Cathepsin L expression and activity were both significantly increased by leptin whereas a cathepsin L peptide inhibitor or siRNA specific for CUX1 completely prevented the leptin-induced increase in FTO expression. The cathepsin L peptide inhibitor or siRNA-induced knockdown of either CUX1 or FTO abrogated the hypertrophic response to leptin. Two other pro-hypertrophic factors, endothelin-1 or angiotensin II had no effect on FTO expression and FTO knockdown did not alter the hypertrophic response to either agent. This study demonstrates leptin-induced FTO upregulation in cardiomyocytes via JAK2/STAT3- dependent CUX1 upregulation and suggests an FTO regulatory function of leptin. It also demonstrates for the first time a functional role of FTO in the cardiomyocyte.

IFN-γ regulates human dental pulp stem cells behavior via NF-κB and MAPK signaling

PubMed Central

He, Xinyao; Jiang, Wenkai; Luo, Zhirong; Qu, Tiejun; Wang, Zhihua; Liu, Ningning; Zhang, Yaqing; Cooper, Paul R.; He, Wenxi

2017-01-01

During caries, dental pulp expresses a range of pro-inflammatory cytokines in response to the infectious challenge. Interferon gamma (IFN-γ) is a dimerized soluble cytokine, which is critical for immune responses. Previous study has demonstrated that IFN-γ at relative high concentration (100 ng/mL) treatment improved the impaired dentinogenic and immunosuppressive regulatory functions of disease-derived dental pulp stem cells (DPSCs). However, little is known about the regulatory effects of IFN-γ at relative low concentration on healthy DPSC behavior (including proliferation, migration, and multiple-potential differentiation). Here we demonstrate that IFN-γ at relatively low concentrations (0.5 ng/mL) promoted the proliferation and migration of DPSCs, but abrogated odonto/osteogenic differentiation. Additionally, we identified that NF-κB and MAPK signaling pathways are both involved in the process of IFN-γ-regulated odonto/osteogenic differentiation of DPSCs. DPSCs treated with IFN-γ and supplemented with pyrrolidine dithiocarbamate (PDTC, an NF-κB inhibitor) or SB203580 (a MAPK inhibitor) showed significantly improved potential for odonto/osteogenic differentiation of DPSCs both in vivo and in vitro. These data provide important insight into the regulatory effects of IFN-γ on the biological behavior of DPSCs and indicate a promising therapeutic strategy for dentin/pulp tissue engineering in future endodontic treatment. PMID:28098169

Osteopontin inhibits osteoblast responsiveness through the down-regulation of focal adhesion kinase mediated by the induction of low-molecular weight protein tyrosine phosphatase.

PubMed

Kusuyama, Joji; Bandow, Kenjiro; Ohnishi, Tomokazu; Hisadome, Mitsuhiro; Shima, Kaori; Semba, Ichiro; Matsuguchi, Tetsuya

2017-05-15

Osteopontin (OPN) is an osteogenic marker protein. Osteoblast functions are affected by inflammatory cytokines and pathological conditions. OPN is highly expressed in bone lesions such as those in rheumatoid arthritis. However, local regulatory effects of OPN on osteoblasts remain ambiguous. Here we examined how OPN influences osteoblast responses to mechanical stress and growth factors. Expression of NO synthase 1 ( Nos1 ) and Nos2 was increased by low-intensity pulsed ultrasound (LIPUS) in MC3T3-E1 cells and primary osteoblasts. The increase of Nos1/2 expression was abrogated by both exogenous OPN overexpression and recombinant OPN treatment, whereas it was promoted by OPN-specific siRNA and OPN antibody. Moreover, LIPUS-induced phosphorylation of focal adhesion kinase (FAK), a crucial regulator of mechanoresponses, was down-regulated by OPN treatments. OPN also attenuated hepatocyte growth factor-induced vitamin D receptor ( Vdr ) expression and platelet-derived growth factor-induced cell mobility through the repression of FAK activity. Of note, the expression of low-molecular weight protein tyrosine phosphatase (LMW-PTP), a FAK phosphatase, was increased in both OPN-treated and differentiated osteoblasts. CD44 was a specific OPN receptor for LWW-PTP induction. Consistently, the suppressive influence of OPN on osteoblast responsiveness was abrogated by LMW-PTP knockdown. Taken together, these results reveal novel functions of OPN in osteoblast physiology. © 2017 Kusuyama et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Identification of a cytoplasmic interaction partner of the large regulatory proteins Rep78/Rep68 of adeno-associated virus type 2 (AAV-2)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weger, Stefan; Hammer, Eva; Goetz, Anne

2007-05-25

Through yeast two-hybrid analysis and coimmunoprecipitation studies, we have identified a novel cellular AAV-2 Rep78/Rep68 interaction partner located predominantly in the cytoplasm. In public databases, it has been assigned as KCTD5, because of a region of high similarity to the cytoplasmic tetramerization domain of voltage-gated potassium channels. Whereas Rep/KCTD5 interaction relied on the region surrounding the Rep nuclear localization signal, nuclear accumulation of Rep was not required. Wildtype Rep78/Rep68 proteins induced the translocation of large portions of KCTD5 into the nucleus pointing to functional interactions both in the cytoplasm and the nucleus. In line with an anticipated functional interference inmore » the cytoplasm, KCTD5 overexpression completely abrogated Rep68-mediated posttranscriptional activation of a HIV-LTR driven luciferase reporter gene. Our study expands the panel of already identified nuclear Rep interaction partners to a cytoplasmic protein, which raises the awareness that important steps in the AAV life cycle may be regulated in this compartment.« less

Sea water acidification affects osmotic swelling, regulatory volume decrease and discharge in nematocytes of the jellyfish Pelagia noctiluca.

PubMed

Morabito, Rossana; Marino, Angela; Lauf, Peter K; Adragna, Norma C; La Spada, Giuseppa

2013-01-01

Increased acidification/PCO2 of sea water is a threat to the environment and affects the homeostasis of marine animals. In this study, the effect of sea water pH changes on the osmotic phase (OP), regulatory volume decrease (RVD) and discharge of the jellyfish Pelagia noctiluca (Cnidaria, Scyphozoa) nematocytes, collected from the Strait of Messina (Italy), was assessed. Isolated nematocytes, suspended in artificial sea water (ASW) with pH 7.65, 6.5 and 4.5, were exposed to hyposmotic ASW of the same pH values and their osmotic response and RVD measured optically in a special flow through chamber. Nematocyte discharge was analyzed in situ in ASW at all three pH values. At normal pH (7.65), nematocytes subjected to hyposmotic shock first expanded osmotically and then regulated their cell volume within 15 min. Exposure to hyposmotic ASW pH 6.5 and 4.5 compromised the OP and reduced or totally abrogated the ensuing RVD, respectively. Acidic pH also significantly reduced the nematocyte discharge response. Data indicate that the homeostasis and function of Cnidarians may be altered by environmental changes such as sea water acidification, thereby validating their use as novel bioindicators for the quality of the marine environment. © 2014 S. Karger AG, Basel.

Rational design of inducible CRISPR guide RNAs for de novo assembly of transcriptional programs

PubMed Central

Ferry, Quentin R. V.; Lyutova, Radostina; Fulga, Tudor A.

2017-01-01

CRISPR-based transcription regulators (CRISPR-TRs) have transformed the current synthetic biology landscape by allowing specific activation or repression of any target gene. Here we report a modular and versatile framework enabling rapid implementation of inducible CRISPR-TRs in mammalian cells. This strategy relies on the design of a spacer-blocking hairpin (SBH) structure at the 5′ end of the single guide RNA (sgRNA), which abrogates the function of CRISPR-transcriptional activators. By replacing the SBH loop with ligand-controlled RNA-cleaving units, we demonstrate conditional activation of quiescent sgRNAs programmed to respond to genetically encoded or externally delivered triggers. We use this system to couple multiple synthetic and endogenous target genes with specific inducers, and assemble gene regulatory modules demonstrating parallel and orthogonal transcriptional programs. We anticipate that this ‘plug and play' approach will be a valuable addition to the synthetic biology toolkit, facilitating the understanding of natural gene circuits and the design of cell-based therapeutic strategies. PMID:28256578

Planarian PTEN homologs regulate stem cells and regeneration through TOR signaling.

PubMed

Oviedo, Néstor J; Pearson, Bret J; Levin, Michael; Sánchez Alvarado, Alejandro

2008-01-01

We have identified two genes, Smed-PTEN-1 and Smed-PTEN-2, capable of regulating stem cell function in the planarian Schmidtea mediterranea. Both genes encode proteins homologous to the mammalian tumor suppressor, phosphatase and tensin homolog deleted on chromosome 10 (PTEN). Inactivation of Smed-PTEN-1 and -2 by RNA interference (RNAi) in planarians disrupts regeneration, and leads to abnormal outgrowths in both cut and uncut animals followed soon after by death (lysis). The resulting phenotype is characterized by hyperproliferation of neoblasts (planarian stem cells), tissue disorganization and a significant accumulation of postmitotic cells with impaired differentiation capacity. Further analyses revealed that rapamycin selectively prevented such accumulation without affecting the normal neoblast proliferation associated with physiological turnover and regeneration. In animals in which PTEN function is abrogated, we also detected a significant increase in the number of cells expressing the planarian Akt gene homolog (Smed-Akt). However, functional abrogation of Smed-Akt in Smed-PTEN RNAi-treated animals does not prevent cell overproliferation and lethality, indicating that functional abrogation of Smed-PTEN is sufficient to induce abnormal outgrowths. Altogether, our data reveal roles for PTEN in the regulation of planarian stem cells that are strikingly conserved to mammalian models. In addition, our results implicate this protein in the control of stem cell maintenance during the regeneration of complex structures in planarians.

Phosphorylation-mediated regulation of the Staphylococcus aureus secreted tyrosine phosphatase PtpA.

PubMed

Brelle, Solène; Baronian, Grégory; Huc-Brandt, Sylvaine; Zaki, Laila Gannoun; Cohen-Gonsaud, Martin; Bischoff, Markus; Molle, Virginie

2016-01-15

Due to the emergence of methicillin-resistant strains, Staphylococcus aureus has become as major public-health threat. Studies aimed at deciphering the molecular mechanism of virulence are thus required to identify new targets and develop efficient therapeutic agents. Protein phosphorylations are known to play key regulatory functions and their roles in pathogenesis are under intense scrutiny. Here we analyzed the protein tyrosine phosphatase PtpA of S. aureus, a member of the family of low molecular weight protein tyrosine phosphatases that are often secreted by pathogenic bacteria. We report for the first time that PtpA is phosphorylated in vitro by the S. aureus tyrosine kinase CapA1B2. A mass spectrometry approach allowed determining that Tyr122 and Tyr123 were the only two residues phosphorylated by this kinase. This result was confirmed by analysis of a double PtpA_Y122A/Y123A mutant that showed no phosphorylation by CapA1B2. Interestingly, PtpA phosphatase activity was abrogated in this mutant, suggesting a key regulatory function for these two tyrosine residues. This was further reinforced by the observation that CapA1B2-mediated phosphorylation significantly increased PtpA phosphatase activity. Moreover, we provide evidence that PtpA is secreted during growth of S. aureus. Together our results suggest that PtpA is an exported S. aureus signaling molecule controlled by tyrosine phosphorylation which may interfere with host cell signaling. Copyright © 2015 Elsevier Inc. All rights reserved.

Aurora A regulates the activity of HURP by controlling the accessibility of its microtubule-binding domain.

PubMed

Wong, Jim; Lerrigo, Robert; Jang, Chang-Young; Fang, Guowei

2008-05-01

HURP is a spindle-associated protein that mediates Ran-GTP-dependent assembly of the bipolar spindle and promotes chromosome congression and interkinetochore tension during mitosis. We report here a biochemical mechanism of HURP regulation by Aurora A, a key mitotic kinase that controls the assembly and function of the spindle. We found that HURP binds to microtubules through its N-terminal domain that hyperstabilizes spindle microtubules. Ectopic expression of this domain generates defects in spindle morphology and function that reduce the level of tension across sister kinetochores and activate the spindle checkpoint. Interestingly, the microtubule binding activity of this N-terminal domain is regulated by the C-terminal region of HURP: in its hypophosphorylated state, C-terminal HURP associates with the microtubule-binding domain, abrogating its affinity for microtubules. However, when the C-terminal domain is phosphorylated by Aurora A, it no longer binds to N-terminal HURP, thereby releasing the inhibition on its microtubule binding and stabilizing activity. In fact, ectopic expression of this C-terminal domain depletes endogenous HURP from the mitotic spindle in HeLa cells in trans, suggesting the physiological importance for this mode of regulation. We concluded that phosphorylation of HURP by Aurora A provides a regulatory mechanism for the control of spindle assembly and function.

Cytoprotective Polyphenols Against Chronological Skin Aging and Cutaneous Photodamage.

PubMed

Davinelli, Sergio; Bertoglio, Juan Carlos; Polimeni, Ascanio; Scapagnini, Giovanni

2018-01-01

Skin aging is a complex biological process influenced by a combination of intrinsic and extrinsic factors, leading to cumulative alterations of skin structure, function and appearance. Polyphenols, which are secondary plant metabolites, represent one of the largest classes of compounds used in dermatology and nutricosmetics to combat skin aging. The main objective is to provide an overview of the existing literature linking skin aging and the ability of polyphenols as regulatory elements able to maintain skin homeostasis. In this review, we discuss recent progress in understanding the molecular bases of skin aging, with specific emphasis on some well known and extensively studied polyphenols which have significant anti-aging influences and photoprotective effects. Although no relevant clinical data exist and standard delivery systems have not been established, promising results have been obtained in many in vitro and animal models. A wide variety of polyphenols may minimize mechanisms underlying the functional manifestations of photoaging and chronological skin aging. Polyphenols exert their influence mostly through their antioxidant and anti-inflammatory effects, thereby abrogating collagen degradation and/or increasing procollagen synthesis. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Donor CD4+ Foxp3+ regulatory T cells are necessary for posttransplantation cyclophosphamide-mediated protection against GVHD in mice

PubMed Central

Ganguly, Sudipto; Ross, Duncan B.; Panoskaltsis-Mortari, Angela; Kanakry, Christopher G.; Blazar, Bruce R.; Levy, Robert B.

2014-01-01

Posttransplantation cyclophosphamide (PTCy) is an effective prophylaxis against graft-versus-host disease (GVHD). However, it is unknown whether PTCy works singularly by eliminating alloreactive T cells via DNA alkylation or also by restoring the conventional (Tcon)/regulatory (Treg) T-cell balance. We studied the role of Tregs in PTCy-mediated GVHD prophylaxis in murine models of allogeneic blood or marrow transplantation (alloBMT). In 2 distinct MHC-matched alloBMT models, infusing Treg-depleted allografts abrogated the GVHD-prophylactic activity of PTCy. Using allografts in which Foxp3+ Tregs could be selectively depleted in vivo, either pre- or post-PTCy ablation of donor thymus–derived Tregs (tTregs) abolished PTCy protection against GVHD. PTCy treatment was associated with relative preservation of donor Tregs. Experiments using combinations of Foxp3– Tcons and Foxp3+ Tregs sorted from different Foxp3 reporter mice indicated that donor Treg persistence after PTCy treatment was predominantly caused by survival of functional tTregs that retained Treg-specific demethylation and also induction of peripherally derived Tregs. Finally, adoptive transfer of tTregs retrieved from PTCy-treated chimeras rescued PTCy-treated, Treg-depleted recipients from lethal GVHD. Our findings indicate that PTCy-mediated protection against GVHD is not singularly dependent on depletion of donor alloreactive T cells but also requires rapidly recovering donor Tregs to initiate and maintain alloimmune regulation. PMID:25139358

Hha has a defined regulatory role that is not dependent upon H-NS or StpA

PubMed Central

Solórzano, Carla; Srikumar, Shabarinath; Canals, Rocío; Juárez, Antonio; Paytubi, Sonia; Madrid, Cristina

2015-01-01

The Hha family of proteins is involved in the regulation of gene expression in enterobacteria by forming complexes with H-NS-like proteins. Whereas several amino acid residues of both proteins participate in the interaction, some of them play a key role. Residue D48 of Hha protein is essential for the interaction with H-NS, thus the D48N substitution in Hha protein abrogates H-NS/Hha interaction. Despite being a paralog of H-NS protein, StpA interacts with HhaD48N with higher affinity than with the wild type Hha protein. To analyze whether Hha is capable of acting independently of H-NS and StpA, we conducted transcriptomic analysis on the hha and stpA deletion strains and the hhaD48N substitution strain of Salmonella Typhimurium using a custom microarray. The results obtained allowed the identification of 120 genes regulated by Hha in an H-NS/StpA-independent manner, 38% of which are horizontally acquired genes. A significant number of the identified genes are involved in functions related to cell motility, iron uptake, and pathogenicity. Thus, motility assays, siderophore detection and intra-macrophage replication assays were performed to confirm the transcriptomic data. Our findings point out the importance of Hha protein as an independent regulator in S. Typhimurium, highlighting a regulatory role on virulence. PMID:26284052

IL-27 Production and STAT3-Dependent Upregulation of B7-H1 Mediate Immune Regulatory Functions of Liver Plasmacytoid DC1

PubMed Central

Matta, Benjamin M.; Raimondi, Giorgio; Rosborough, Brian R.; Sumpter, Tina L.; Thomson, Angus W.

2012-01-01

Plasmacytoid (p) dendritic cells (DC) are highly-specialized APC that, in addition to their well-recognized role in anti-viral immunity, also regulate immune responses. Liver-resident pDC are considerably less immunostimulatory than those from secondary lymphoid tissues and are equipped to promote immune tolerance/regulation through various mechanisms. IL-27 is an IL-12-family cytokine that regulates the function of both APC and T cells, although little is known about its role in pDC immunobiology. In this study, we show that mouse liver pDC express higher levels of IL-27p28 and EBV-induced protein (Ebi)3 compared to splenic pDC. Both populations of pDC express the IL-27Rα/WSX-1; however, only liver pDC significantly upregulate expression of the co-regulatory molecule B7 homolog-1 (B7-H1) in response to IL-27. Inhibition of STAT3 activation completely abrogates IL-27-induced upregulation of B7-H1 expression on liver pDC. Liver pDC treated with IL-27 increase the percentage of CD4+Foxp3+ T cells in MLR, which is dependent upon expression of B7-H1. pDC from Ebi3-deficient mice lacking functional IL-27, show increased capacity to stimulate allogeneic T cell proliferation and IFN-γ production in MLR. Liver but not spleen pDC suppress delayed-type hypersensitivity responses to OVA, an effect that is lost with Ebi3−/− and B7-H1−/− liver pDC compared to wild-type (WT) liver pDC. These data suggest that IL-27 signaling in pDC promotes their immunoregulatory function and that IL-27 produced by pDC contributes to their capacity to regulate immuneresponses in vitro and in vivo. PMID:22508931

SREBF1 Activity is Regulated by an AR/mTOR Nuclear Axis in Prostate Cancer.

PubMed

Audet-Walsh, Etienne; Vernier, Mathieu; Yee, Tracey; Laflamme, Chloe E; Li, Susan; Chen, Yonghong; Giguere, Vincent

2018-05-21

Reprogramming of cellular metabolism is an important feature of prostate cancer (PCa), including altered lipid metabolism. Recently, it was observed that the nuclear fraction of mTOR is essential for the androgen-mediated metabolic reprogramming of PCa cells. Herein, it is demonstrated that the androgen receptor (AR) and mTOR bind to regulatory regions of sterol regulatory element binding transcription factor 1 (SREBF1) to control its expression, while dual activation of these signaling pathways also promotes SREBF1 cleavage and its translocation to the nucleus. Consequently, SREBF1 recruitment to regulatory regions of its target genes is induced upon treatment with the synthetic androgen R1881, an effect abrogated upon inhibition of the mTOR signaling pathway. In turn, pharmacological and genetic inhibition of SREBF1 activity impairs the androgen-mediated induction of the key lipogenic genes fatty acid synthase (FASN) and stearoyl-CoA desaturase (SCD1). Consistent with these observations, the expression of SREBF1, FASN and SCD1 is significantly correlated in human PCa tumor clinical specimens. Functionally, blockade of SREBF1 activity reduces the androgen-driven lipid accumulation. Interestingly, decreased triglyceride accumulation observed upon SREBF1 inhibition is paralleled by an increase in mitochondrial respiration, indicating a potential rewiring of citrate metabolism in PCa cells. Altogether, these data define an AR/mTOR nuclear axis, in the context of PCa, as a novel pathway regulating SREBF1 activity and citrate metabolism. The finding that an AR/mTOR complex promotes SREBP expression and activity enhances our understanding of the metabolic adaptation necessary for prostate cancer cell growth and suggests novel therapeutic approaches to target metabolic vulnerabilities in tumors. Copyright ©2018, American Association for Cancer Research.

The role of JAK-3 in regulating TLR-mediated inflammatory cytokine production in innate immune cells.

PubMed

Wang, Huizhi; Brown, Jonathan; Gao, Shegan; Liang, Shuang; Jotwani, Ravi; Zhou, Huaxin; Suttles, Jill; Scott, David A; Lamont, Richard J

2013-08-01

The role of JAK-3 in TLR-mediated innate immune responses is poorly understood, although the suppressive function of JAK3 inhibition in adaptive immune response has been well studied. In this study, we found that JAK3 inhibition enhanced TLR-mediated immune responses by differentially regulating pro- and anti- inflammatory cytokine production in innate immune cells. Specifically, JAK3 inhibition by pharmacological inhibitors or specific small interfering RNA or JAK3 gene knockout resulted in an increase in TLR-mediated production of proinflammatory cytokines while concurrently decreasing the production of IL-10. Inhibition of JAK3 suppressed phosphorylation of PI3K downstream effectors including Akt, mammalian target of rapamycin complex 1, glycogen synthase kinase 3β (GSK3β), and CREB. Constitutive activation of Akt or inhibition of GSK3β abrogated the capability of JAK3 inhibition to enhance proinflammatory cytokines and suppress IL-10 production. In contrast, inhibition of PI3K enhanced this regulatory ability of JAK3 in LPS-stimulated monocytes. At the transcriptional level, JAK3 knockout lead to the increased phosphorylation of STATs that could be attenuated by neutralization of de novo inflammatory cytokines. JAK3 inhibition exhibited a GSK3 activity-dependent ability to enhance phosphorylation levels and DNA binding of NF-κB p65. Moreover, JAK3 inhibition correlated with an increased CD4(+) T cell response. Additionally, higher neutrophil infiltration, IL-17 expression, and intestinal epithelium erosion were observed in JAK3 knockout mice. These findings demonstrate the negative regulatory function of JAK3 and elucidate the signaling pathway by which JAK3 differentially regulates TLR-mediated inflammatory cytokine production in innate immune cells.

Critical role for TNF in the induction of human antigen-specific regulatory T cells by tolerogenic dendritic cells.

PubMed

Kleijwegt, Fleur S; Laban, Sandra; Duinkerken, Gaby; Joosten, Antoinette M; Zaldumbide, Arnaud; Nikolic, Tatjana; Roep, Bart O

2010-08-01

TNF is a pleiotropic cytokine with differential effects on immune cells and diseases. Anti-TNF therapy was shown to be effective in rheumatoid arthritis but proved inefficient or even detrimental in other autoimmune diseases. We studied the role of TNF in the induction of Ag-specific regulatory T cells (Tregs) by tolerogenic vitamin D3-modulated human dendritic cells (VD3-DCs), which previously were shown to release high amounts of soluble TNF (sTNF) upon maturation with LPS. First, production of TNF by modulated VD3-DCs was analyzed upon maturation with LPS or CD40L with respect to both secreted (cleaved) TNF (sTNF) and expression of the membrane-bound (uncleaved) form of TNF (mTNF). Next, TNF antagonists were tested for their effect on induction of Ag-specific Tregs by modulated DCs and the subsequent functionality of these Tregs. VD3-DCs expressed greater amounts of mTNF than did control DCs (nontreated DCs), independent of the maturation protocol. Inhibition of TNF with anti-TNF Ab (blocking both sTNF and mTNF) during the priming of Tregs with VD3-DCs prevented generation of Tregs and their suppression of proliferation of CD4(+) T cells. In contrast, sTNF receptor II (sTNFRII), mainly blocking sTNF, did not change the suppressive capacity of Tregs. Blocking of TNFRII by anti-CD120b Ab during Treg induction similarly abrogated their subsequent suppressive function. These data point to a specific role for mTNF on VD3-DCs in the induction of Ag-specific Tregs. Interaction between mTNF and TNFRII instructs the induction of suppressive Tregs by VD3-DCs. Anti-TNF therapy may therefore act adversely in different patients or disease pathways.

Modulation of mycobacterial-specific Th1 and Th17 cells in latent tuberculosis by coincident hookworm infection

PubMed Central

George, Parakkal Jovvian; Anuradha, Rajamanickam; Kumaran, Paramasivam Paul; Chandrasekaran, Vedachalam; Nutman, Thomas B.; Babu, Subash

2013-01-01

Hookworm infections and tuberculosis are co-endemic in many parts of the world. It has been suggested that infection with helminth parasites could suppress the predominant Th1 (IFN-γ-mediated) response needed to control Mycobacterium tuberculosis (Mtb) infection and enhance susceptibility to infection and/or disease. To determine the role of coincident hookworm infection on responses at steady state and on Mtb – specific immune responses in latent tuberculosis (TB), we examined the cellular responses in individuals with latent TB with or without concomitant hookworm infection. By analyzing the expression of Th1, Th2 and Th17 subsets of CD4+ T cells, we were able to demonstrate that the presence of coincident hookworm infection significantly diminished both spontaneously expressed and Mtb – specific mono – and dual – functional Th1 and Th17 cells. Hookworm infection, in contrast, was associated with expanded frequencies of mono – and dual – functional Th2 cells at both steady state and upon antigen – stimulation. This differential induction of CD4+ T cell subsets was abrogated upon mitogen stimulation. In addition, coincident hookworm infection was associated with increased adaptive T regulatory (aTreg) cells but not natural regulatory T cells (nTregs) in latent TB. Finally, the CD4+ T cell cytokine expression pattern was also associated with alterations in the systemic levels of Th1 and Th2 cytokines. Thus, coincident hookworm infection exerts a profound inhibitory effect on protective Th1 and Th17 responses in latent tuberculosis and may predispose toward the development of active tuberculosis in humans. PMID:23576678

Interaction between dendritic cells and natural killer cells during pregnancy in mice.

PubMed

Blois, Sandra M; Barrientos, Gabriela; Garcia, Mariana G; Orsal, Arif S; Tometten, Mareike; Cordo-Russo, Rosalia I; Klapp, Burghard F; Santoni, Angela; Fernández, Nelson; Terness, Peter; Arck, Petra C

2008-07-01

A complex regulation of innate and adaptive immune responses at the maternal fetal interface promotes tolerance of trophoblast cells carrying paternally derived antigens. Such regulatory functions involve uterine dendritic cells (uDC) and natural killer (uNK) cells. The existence of a NK and DC "cross talk" has been revealed in various experimental settings; its biological significance ranging from cooperative stimulation to cell lysis. Little is known about the presence or role of NK and DC cross talk at the maternal fetal interface. The present study shows that mouse NK and DC interactions are subject to modulation by trophoblast cells in vitro. This interaction promotes a tolerogenic microenvironment characterized by downregulation of the expression of activation markers on uNK cells and uDC and dominance of Th2 cytokines. NK and DC interactions would also influence uterine cell proliferation and this process would be strongly modulated by trophoblast-derived signals. Indeed; while low proliferation rates were observed upon regular coculture allowing direct contact between uterine cells and trophoblasts, incubation in a transwell culture system markedly increased uterine cell proliferation suggesting that soluble factors are key mediators in the molecular "dialog" between the mother and the conceptus during the establishment of mouse pregnancy. Our data further reveal that the regulatory functions of trophoblast cells associated with tolerance induction are impaired in high abortion murine matings. Interestingly, we observed that secretion of interleukin-12p70 by uDC is dramatically abrogated in the presence of uNK cells. Taken together, our results provide the first evidence that a delicate balance of interactions involving NK cells, DC, and trophoblasts at the mouse maternal fetal interface supports a successful pregnancy outcome.

Loss of Cbl-PI3K interaction in mice prevents significant bone loss following ovariectomy

PubMed Central

Adapala, Naga Suresh; Holland, Danielle; Piccuillo, Vanessa; Barbe, Mary F.; Langdon, Wallace Y.; Tsygankov, Alexander Y.; Lorenzo, Joseph A.; Sanjay, Archana

2014-01-01

Cbl and Cbl-b are E3 ubiquitin ligases and adaptor proteins, which perform regulatory roles in bone remodeling. Cbl−/− mice have delayed bone development due to decreased osteoclast migration. Cbl-b−/− mice are osteopenic due to increased bone resorbing activity of osteoclasts. Unique to Cbl, but not present in Cbl-b, is tyrosine 737 in the YEAM motif, which upon phosphorylation provides a binding site for the regulatory p85 subunit of PI3K. Substitution of tyrosine 737 with phenylalanine (Y737F, CblYF/YF mice) prevents Y737 phosphorylation and abrogates the Cbl-PI3K interaction. We have previously reported that CblYF/YF mice had increased bone volume due to defective bone resorption and increased bone formation. Here we show that the lumbar vertebra from CblYF/YF mice did not have significant bone loss following ovariectomy. Our data also suggests that abrogation of Cbl-PI3K interaction in mice results in the loss of coupling between bone resorption and formation, since ovariectomized CblYF/YF mice did not show significant changes in serum levels of c-terminal telopeptide (CTX), whereas the serum levels of pro-collagen type-1 amino-terminal pro-peptide (P1NP) were decreased. In contrast, following ovariectomy, Cbl−/− and Cbl-b−/− mice showed significant bone loss in tibiae and L2 vertebrae, concomitant with increased serum CTX and P1NP levels. These data indicate that while lack of Cbl or Cbl-b distinctly affects bone remodeling, only the loss of Cbl-PI3K interaction protects mice from significant bone loss following ovariectomy. PMID:24994594

Loss of Cbl-PI3K interaction in mice prevents significant bone loss following ovariectomy.

PubMed

Adapala, Naga Suresh; Holland, Danielle; Scanlon, Vanessa; Barbe, Mary F; Langdon, Wallace Y; Tsygankov, Alexander Y; Lorenzo, Joseph A; Sanjay, Archana

2014-10-01

Cbl and Cbl-b are E3 ubiquitin ligases and adaptor proteins, which perform regulatory roles in bone remodeling. Cbl-/- mice have delayed bone development due to decreased osteoclast migration. Cbl-b-/- mice are osteopenic due to increased bone resorbing activity of osteoclasts. Unique to Cbl, but not present in Cbl-b, is tyrosine 737 in the YEAM motif, which upon phosphorylation provides a binding site for the regulatory p85 subunit of PI3K. Substitution of tyrosine 737 with phenylalanine (Y737F, CblYF/YF mice) prevents Y737 phosphorylation and abrogates the Cbl-PI3K interaction. We have previously reported that CblYF/YF mice had increased bone volume due to defective bone resorption and increased bone formation. Here we show that the lumbar vertebra from CblYF/YF mice did not have significant bone loss following ovariectomy. Our data also suggests that abrogation of Cbl-PI3K interaction in mice results in the loss of coupling between bone resorption and formation, since ovariectomized CblYF/YF mice did not show significant changes in serum levels of c-terminal telopeptide (CTX), whereas the serum levels of pro-collagen type-1 amino-terminal pro-peptide (P1NP) were decreased. In contrast, following ovariectomy, Cbl-/- and Cbl-b-/- mice showed significant bone loss in the tibiae and L2 vertebrae, concomitant with increased serum CTX and P1NP levels. These data indicate that while lack of Cbl or Cbl-b distinctly affects bone remodeling, only the loss of Cbl-PI3K interaction protects mice from significant bone loss following ovariectomy. Copyright © 2014 Elsevier Inc. All rights reserved.

The novel HSP90 inhibitor NVP-AUY922 shows synergistic anti-leukemic activity with cytarabine in vivo.

PubMed

Wendel, Torunn; Zhen, Yan; Suo, Zenhe; Bruheim, Skjalg; Wiedlocha, Antoni

2016-01-15

HSP90 is a molecular chaperone essential for stability, activity and intracellular sorting of many proteins, including oncoproteins, such as tyrosine kinases, transcription factors and cell cycle regulatory proteins. Therefore, inhibitors of HSP90 are being investigated for their potential as anti-cancer drugs. Here we show that the HSP90 inhibitor NVP-AUY922 induced degradation of the fusion oncoprotein FOP2-FGFR1 in a human acute myeloid leukemia (AML) cell line, KG-1a. Concordantly, downstream signaling cascades, such as STAT1, STAT3 and PLCγ were abrogated. At concentrations that caused FOP2-FGFR1 degradation and signaling abrogation, NVP-AUY922 treatment caused significant cell death and inhibition of proliferation of KG-1a cells in vitro. In an animal model for AML, NVP-AUY922 administrated alone showed no anti-leukemic activity. However, when NVP-AUY922 was administered in combination with cytarabine, the two compounds showed significant synergistic anti-leukemic activity in vivo. Thus NVP-AUY922 and cytarabine combination therapy might be a prospective strategy for AML treatment. Copyright © 2016 Elsevier Inc. All rights reserved.

Highly conserved non-coding elements on either side of SOX9 associated with Pierre Robin sequence.

PubMed

Benko, Sabina; Fantes, Judy A; Amiel, Jeanne; Kleinjan, Dirk-Jan; Thomas, Sophie; Ramsay, Jacqueline; Jamshidi, Negar; Essafi, Abdelkader; Heaney, Simon; Gordon, Christopher T; McBride, David; Golzio, Christelle; Fisher, Malcolm; Perry, Paul; Abadie, Véronique; Ayuso, Carmen; Holder-Espinasse, Muriel; Kilpatrick, Nicky; Lees, Melissa M; Picard, Arnaud; Temple, I Karen; Thomas, Paul; Vazquez, Marie-Paule; Vekemans, Michel; Roest Crollius, Hugues; Hastie, Nicholas D; Munnich, Arnold; Etchevers, Heather C; Pelet, Anna; Farlie, Peter G; Fitzpatrick, David R; Lyonnet, Stanislas

2009-03-01

Pierre Robin sequence (PRS) is an important subgroup of cleft palate. We report several lines of evidence for the existence of a 17q24 locus underlying PRS, including linkage analysis results, a clustering of translocation breakpoints 1.06-1.23 Mb upstream of SOX9, and microdeletions both approximately 1.5 Mb centromeric and approximately 1.5 Mb telomeric of SOX9. We have also identified a heterozygous point mutation in an evolutionarily conserved region of DNA with in vitro and in vivo features of a developmental enhancer. This enhancer is centromeric to the breakpoint cluster and maps within one of the microdeletion regions. The mutation abrogates the in vitro enhancer function and alters binding of the transcription factor MSX1 as compared to the wild-type sequence. In the developing mouse mandible, the 3-Mb region bounded by the microdeletions shows a regionally specific chromatin decompaction in cells expressing Sox9. Some cases of PRS may thus result from developmental misexpression of SOX9 due to disruption of very-long-range cis-regulatory elements.

Potential Roles of GLUT12 for Glucose Sensing and Cellular Migration in MCF-7 Human Breast Cancer Cells Under High Glucose Conditions.

PubMed

Matsui, Chihiro; Takatani-Nakase, Tomoka; Maeda, Sachie; Nakase, Ikuhiko; Takahashi, Koichi

2017-12-01

Recent reports have indicated that hyperglycaemia is associated with breast cancer progression. High glucose conditions corresponding to hyperglycaemia significantly promote migration of MCF-7 human breast cancer cells, however, little is known about the mechanisms of glucose sensing for the acquisition of migratory properties by MCF-7 cells. This study investigated glucose sensing and mediation, which are responsible for the high motility of MCF-7 cells. We evaluated the migration of MCF-7 cells cultured in high glucose-containing medium and essential regulatory factors from the perspective of the glucose transport system. We demonstrated that glucose transporter 12 (GLUT12) protein level increased in MCF-7 cells and co-localized with actin organization under high glucose conditions. Moreover, GLUT12-knockdown completely abrogated high glucose-induced migration, indicating that GLUT12 functionally participates in sensing high glucose concentrations. GLUT12 plays a critical role in the model of breast cancer progression through high glucose concentrations. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

p53 mutations promote proteasomal activity.

PubMed

Oren, Moshe; Kotler, Eran

2016-07-27

p53 mutations occur very frequently in human cancer. Besides abrogating the tumour suppressive functions of wild-type p53, many of those mutations also acquire oncogenic gain-of-function activities. Augmentation of proteasome activity is now reported as a common gain-of-function mechanism shared by different p53 mutants, which promotes cancer resistance to proteasome inhibitors.

Toxoplasma gondii infection shifts dendritic cells into an amoeboid rapid migration mode encompassing podosome dissolution, secretion of TIMP-1, and reduced proteolysis of extracellular matrix.

PubMed

Ólafsson, Einar B; Varas-Godoy, Manuel; Barragan, Antonio

2018-03-01

Dendritic cells (DCs) infected by Toxoplasma gondii rapidly acquire a hypermigratory phenotype that promotes systemic parasite dissemination by a "Trojan horse" mechanism in mice. Recent paradigms of leukocyte migration have identified the amoeboid migration mode of DCs as particularly suited for rapid locomotion in extracellular matrix and tissues. Here, we have developed a microscopy-based high-throughput approach to assess motility and matrix degradation by Toxoplasma-challenged murine and human DCs. DCs challenged with T. gondii exhibited dependency on metalloproteinase activity for hypermotility and transmigration but, strikingly, also dramatically reduced pericellular proteolysis. Toxoplasma-challenged DCs up-regulated expression and secretion of tissue inhibitor of metalloproteinases-1 (TIMP-1) and their supernatants impaired matrix degradation by naïve DCs and by-stander DCs dose dependently. Gene silencing of TIMP-1 by short hairpin RNA restored matrix degradation activity in Toxoplasma-infected DCs. Additionally, dissolution of podosome structures in parasitised DCs coincided with abrogated matrix degradation. Toxoplasma lysates inhibited pericellular proteolysis in a MyD88-dependent fashion whereas abrogated proteolysis persevered in Toxoplasma-infected MyD88-deficient DCs. This indicated that both TLR/MyD88-dependent and TLR/MyD88-independent signalling pathways mediated podosome dissolution and the abrogated matrix degradation. We report that increased TIMP-1 secretion and cytoskeletal rearrangements encompassing podosome dissolution are features of Toxoplasma-induced hypermigration of DCs with an impact on matrix degradation. Jointly, the data highlight how an obligate intracellular parasite orchestrates key regulatory cellular processes consistent with non-proteolytic amoeboid migration of the vehicle cells that facilitate its dissemination. © 2017 John Wiley & Sons Ltd.

A NAP-Family Histone Chaperone Functions in Abiotic Stress Response and Adaptation1[OPEN

PubMed Central

Pareek, Ashwani; Singla-Pareek, Sneh Lata

2016-01-01

Modulation of gene expression is one of the most significant molecular mechanisms of abiotic stress response in plants. Via altering DNA accessibility, histone chaperones affect the transcriptional competence of genomic loci. However, in contrast to other factors affecting chromatin dynamics, the role of plant histone chaperones in abiotic stress response and adaptation remains elusive. Here, we studied the physiological function of a stress-responsive putative rice (Oryza sativa) histone chaperone of the NAP superfamily: OsNAPL6. We show that OsNAPL6 is a nuclear-localized H3/H4 histone chaperone capable of assembling a nucleosome-like structure. Utilizing overexpression and knockdown approaches, we found a positive correlation between OsNAPL6 expression levels and adaptation to multiple abiotic stresses. Results of comparative transcriptome profiling and promoter-recruitment studies indicate that OsNAPL6 functions during stress response via modulation of expression of various genes involved in diverse functions. For instance, we show that OsNAPL6 is recruited to OsRad51 promoter, activating its expression and leading to more efficient DNA repair and abrogation of programmed cell death under salinity and genotoxic stress conditions. These results suggest that the histone chaperone OsNAPL6 may serve a regulatory role in abiotic stress physiology possibly via modulating nucleosome dynamics at various stress-associated genomic loci. Taken together, our findings establish a hitherto unknown link between histone chaperones and abiotic stress response in plants. PMID:27342307

34 CFR 303.103 - Abrogation of State sovereign immunity.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 34 Education 2 2012-07-01 2012-07-01 false Abrogation of State sovereign immunity. 303.103 Section... System State Conformity with Part C of the Act and Abrogation of State Sovereign Immunity § 303.103 Abrogation of State sovereign immunity. (a) General. A State is not immune under the 11th amendment of the...

34 CFR 303.103 - Abrogation of State sovereign immunity.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 34 Education 2 2013-07-01 2013-07-01 false Abrogation of State sovereign immunity. 303.103 Section... System State Conformity with Part C of the Act and Abrogation of State Sovereign Immunity § 303.103 Abrogation of State sovereign immunity. (a) General. A State is not immune under the 11th amendment of the...

34 CFR 303.103 - Abrogation of State sovereign immunity.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 34 Education 2 2014-07-01 2013-07-01 true Abrogation of State sovereign immunity. 303.103 Section... System State Conformity with Part C of the Act and Abrogation of State Sovereign Immunity § 303.103 Abrogation of State sovereign immunity. (a) General. A State is not immune under the 11th amendment of the...

Upregulation of neurovascular communication through filamin abrogation promotes ectopic periventricular neurogenesis.

PubMed

Houlihan, Shauna L; Lanctot, Alison A; Guo, Yan; Feng, Yuanyi

2016-09-24

Neuronal fate-restricted intermediate progenitors (IPs) are derived from the multipotent radial glia (RGs) and serve as the direct precursors for cerebral cortical neurons, but factors that control their neurogenic plasticity remain elusive. Here we report that IPs' neuron production is enhanced by abrogating filamin function, leading to the generation of periventricular neurons independent of normal neocortical neurogenesis and neuronal migration. Loss of Flna in neural progenitor cells (NPCs) led RGs to undergo changes resembling epithelial-mesenchymal transition (EMT) along with exuberant angiogenesis that together changed the microenvironment and increased neurogenesis of IPs. We show that by collaborating with β-arrestin, Flna maintains the homeostatic signaling between the vasculature and NPCs, and loss of this function results in escalated Vegfa and Igf2 signaling, which exacerbates both EMT and angiogenesis to further potentiate IPs' neurogenesis. These results suggest that the neurogenic potential of IPs may be boosted in vivo by manipulating Flna-mediated neurovascular communication.

Immunodeficiency with thymoma: failure to induce Ig production in immunodeficient lymphocytes cocultured with normal T cells. [X radiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Litwin, S.D.

Blood mononuclear cells of two individuals having immunodeficiency with thymoma (ID-THY) were cocultured with normal mononuclear cells or treated mononuclear cell fractions in an attempt to correct an imbalance of regulatory cells postulated to be responsible for the failure of pokeweed mitogen-induced Ig synthesis in vitro. Treatment included abrogation of suppressor cell activity by irradiation or incubation with prednisolone in vitro. T cell help was provided by cocultivating lymphocytes of related and unrelated persons, and in some cases autologous treated cells. Ig secretion failed to be induced by any experimental maneuver suggesting that the primary problem in the above ID-THYmore » cells was related to defective or deficient B cells rather than an imbalance of T regulatory cells. Prednisolone treatment in vitro decreased suppressor cell activity in allogeneic cocultures of two ID-THY persons (S1 and S2) but not of an individual (S3) with variable immunodeficiency suggesting heterogeneity of suppressor cells.« less

A pivotal role for galectin-1 in fetomaternal tolerance.

PubMed

Blois, Sandra M; Ilarregui, Juan M; Tometten, Mareike; Garcia, Mariana; Orsal, Arif S; Cordo-Russo, Rosalia; Toscano, Marta A; Bianco, Germán A; Kobelt, Peter; Handjiski, Bori; Tirado, Irene; Markert, Udo R; Klapp, Burghard F; Poirier, Francoise; Szekeres-Bartho, Julia; Rabinovich, Gabriel A; Arck, Petra C

2007-12-01

A successful pregnancy requires synchronized adaptation of maternal immune-endocrine mechanisms to the fetus. Here we show that galectin-1 (Gal-1), an immunoregulatory glycan-binding protein, has a pivotal role in conferring fetomaternal tolerance. Consistently with a marked decrease in Gal-1 expression during failing pregnancies, Gal-1-deficient (Lgals1-/-) mice showed higher rates of fetal loss compared to wild-type mice in allogeneic matings, whereas fetal survival was unaffected in syngeneic matings. Treatment with recombinant Gal-1 prevented fetal loss and restored tolerance through multiple mechanisms, including the induction of tolerogenic dendritic cells, which in turn promoted the expansion of interleukin-10 (IL-10)-secreting regulatory T cells in vivo. Accordingly, Gal-1's protective effects were abrogated in mice depleted of regulatory T cells or deficient in IL-10. In addition, we provide evidence for synergy between Gal-1 and progesterone in the maintenance of pregnancy. Thus, Gal-1 is a pivotal regulator of fetomaternal tolerance that has potential therapeutic implications in threatened pregnancies.

Clusterin/ApoJ enhances central leptin signaling through Lrp2-mediated endocytosis.

PubMed

Byun, Kyunghee; Gil, So Young; Namkoong, Churl; Youn, Byung-Soo; Huang, Hu; Shin, Mi-Seon; Kang, Gil Myoung; Kim, Hyun-Kyong; Lee, Bonghee; Kim, Young-Bum; Kim, Min-Seon

2014-07-01

Hypothalamic leptin signaling plays a central role in maintaining body weight homeostasis. Here, we show that clusterin/ApoJ, recently identified as an anorexigenic neuropeptide, is an important regulator in the hypothalamic leptin signaling pathway. Coadministration of clusterin potentiates the anorexigenic effect of leptin and boosts leptin-induced hypothalamic Stat3 activation. In cultured neurons, clusterin enhances receptor binding and subsequent endocytosis of leptin. These effects are mainly mediated through the LDL receptor-related protein-2 (Lrp2). Notably, inhibition of hypothalamic clusterin, Lrp2 or endocytosis abrogates anorexia and hypothalamic Stat3 activation caused by leptin. These findings propose a novel regulatory mechanism in central leptin signaling pathways. © 2014 The Authors.

Rapid alterations of cell cycle control proteins in human T lymphocytes in microgravity

PubMed Central

2012-01-01

In our study we aimed to identify rapidly reacting gravity-responsive mechanisms in mammalian cells in order to understand if and how altered gravity is translated into a cellular response. In a combination of experiments using "functional weightlessness" provided by 2D-clinostats and real microgravity provided by several parabolic flight campaigns and compared to in-flight-1g-controls, we identified rapid gravity-responsive reactions inside the cell cycle regulatory machinery of human T lymphocytes. In response to 2D clinorotation, we detected an enhanced expression of p21 Waf1/Cip1 protein within minutes, less cdc25C protein expression and enhanced Ser147-phosphorylation of cyclinB1 after CD3/CD28 stimulation. Additionally, during 2D clinorotation, Tyr-15-phosphorylation occurred later and was shorter than in the 1 g controls. In CD3/CD28-stimulated primary human T cells, mRNA expression of the cell cycle arrest protein p21 increased 4.1-fold after 20s real microgravity in primary CD4+ T cells and 2.9-fold in Jurkat T cells, compared to 1 g in-flight controls after CD3/CD28 stimulation. The histone acetyltransferase (HAT) inhibitor curcumin was able to abrogate microgravity-induced p21 mRNA expression, whereas expression was enhanced by a histone deacetylase (HDAC) inhibitor. Therefore, we suppose that cell cycle progression in human T lymphocytes requires Earth gravity and that the disturbed expression of cell cycle regulatory proteins could contribute to the breakdown of the human immune system in space. PMID:22273506

GARP inhibits allergic airway inflammation in a humanized mouse model.

PubMed

Meyer-Martin, H; Hahn, S A; Beckert, H; Belz, C; Heinz, A; Jonuleit, H; Becker, C; Taube, C; Korn, S; Buhl, R; Reuter, S; Tuettenberg, A

2016-09-01

Regulatory T cells (Treg) represent a promising target for novel treatment strategies in patients with inflammatory/allergic diseases. A soluble derivate of the Treg surface molecule glycoprotein A repetitions predominant (sGARP) has strong anti-inflammatory and regulatory effects on human cells in vitro as well as in vivo through de novo induction of peripheral Treg. The aim of this study was to investigate the immunomodulatory function of sGARP and its possible role as a new therapeutic option in allergic diseases using a humanized mouse model. To analyze the therapeutic effects of sGARP, adult NOD/Scidγc(-/-) (NSG) mice received peripheral blood mononuclear cells (PBMC) derived from allergic patients with sensitization against birch allergen. Subsequently, allergic inflammation was induced in the presence of Treg alone or in combination with sGARP. In comparison with mice that received Treg alone, additional treatment with sGARP reduced airway hyperresponsiveness (AHR), influx of neutrophils and macrophages into the bronchoalveolar lavage (BAL), and human CD45(+) cells in the lungs. Furthermore, the numbers of mucus-producing goblet cells and inflammatory cell infiltrates were reduced. To elucidate whether the mechanism of action of sGARP involves the TGF-β receptor pathway, mice additionally received anti-TGF-β receptor II (TGF-βRII) antibodies. Blocking the signaling of TGF-β through TGF-βRII abrogated the anti-inflammatory effects of sGARP, confirming its essential role in inhibiting the allergic inflammation. Induction of peripheral tolerance via sGARP is a promising potential approach to treat allergic airway diseases. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Metabolomic profiling of the heart during acute ischemic preconditioning reveals a role for SIRT1 in rapid cardioprotective metabolic adaptation.

PubMed

Nadtochiy, Sergiy M; Urciuoli, William; Zhang, Jimmy; Schafer, Xenia; Munger, Joshua; Brookes, Paul S

2015-11-01

Ischemic preconditioning (IPC) protects tissues such as the heart from prolonged ischemia-reperfusion (IR) injury. We previously showed that the lysine deacetylase SIRT1 is required for acute IPC, and has numerous metabolic targets. While it is known that metabolism is altered during IPC, the underlying metabolic regulatory mechanisms are unknown, including the relative importance of SIRT1. Thus, we sought to test the hypothesis that some of the metabolic adaptations that occur in IPC may require SIRT1 as a regulatory mediator. Using both ex-vivo-perfused and in-vivo mouse hearts, LC-MS/MS based metabolomics and (13)C-labeled substrate tracing, we found that acute IPC altered several metabolic pathways including: (i) stimulation of glycolysis, (ii) increased synthesis of glycogen and several amino acids, (iii) increased reduced glutathione levels, (iv) elevation in the oncometabolite 2-hydroxyglutarate, and (v) inhibition of fatty-acid dependent respiration. The majority (83%) of metabolic alterations induced by IPC were ablated when SIRT1 was acutely inhibited with splitomicin, and a principal component analysis revealed that metabolic changes in response to IPC were fundamentally different in nature when SIRT1 was inhibited. Furthermore, the protective benefit of IPC was abrogated by eliminating glucose from perfusion media while sustaining normal cardiac function by burning fat, thus indicating that glucose dependency is required for acute IPC. Together, these data suggest that SIRT1 signaling is required for rapid cardioprotective metabolic adaptation in acute IPC. Copyright © 2015 Elsevier Ltd. All rights reserved.

Mutant p53 Promotes Tumor Cell Malignancy by Both Positive and Negative Regulation of the Transforming Growth Factor β (TGF-β) Pathway*

PubMed Central

Ji, Lei; Xu, Jinjin; Liu, Jian; Amjad, Ali; Zhang, Kun; Liu, Qingwu; Zhou, Lei; Xiao, Jianru; Li, Xiaotao

2015-01-01

Specific p53 mutations abrogate tumor-suppressive functions by gaining new abilities to promote tumorigenesis. Inactivation of p53 is known to distort TGF-β signaling, which paradoxically displays both tumor-suppressive and pro-oncogenic functions. The molecular mechanisms of how mutant p53 simultaneously antagonizes the tumor-suppressive and synergizes the tumor-promoting function of the TGF-β pathway remain elusive. Here we demonstrate that mutant p53 differentially regulates subsets of TGF-β target genes by enhanced binding to the MH2 domain in Smad3 upon the integration of ERK signaling, therefore disrupting Smad3/Smad4 complex formation. Silencing Smad2, inhibition of ERK, or introducing a phosphorylation-defective mutation at Ser-392 in p53 abrogates the R175H mutant p53-dependent regulation of these TGF-β target genes. Our study shows a mechanism to reconcile the seemingly contradictory observations that mutant p53 can both attenuate and cooperate with the TGF-β pathway to promote cancer cell malignancy in the same cell type. PMID:25767119

Codon-usage-based inhibition of HIV protein synthesis by human schlafen 11

PubMed Central

Li, Manqing; Kao, Elaine; Gao, Xia; Sandig, Hilary; Limmer, Kirsten; Pavon-Eternod, Mariana; Jones, Thomas E.; Landry, Sebastien; Pan, Tao; Weitzman, Matthew D.; David, Michael

2013-01-01

In mammals, one of the most pronounced consequences of viral infection is the induction of type I interferons, cytokines with potent antiviral activity. Schlafen (Slfn) genes are a subset of interferon-stimulated early response genes (ISGs) that are also induced directly by pathogens via the interferon regulatory factor 3 (IRF3) pathway1. However, many ISGs are of unknown or incompletely understood function. Here we show that human SLFN11 potently and specifically abrogates the production of retroviruses such as human immunodeficiency virus 1 (HIV-1). Our study revealed that SLFN11 has no effect on the early steps of the retroviral infection cycle, including reverse transcription, integration and transcription. Rather, SLFN11 acts at the late stage of virus production by selectively inhibiting the expression of viral proteins in a codon-usage-dependent manner. We further find that SLFN11 binds transfer RNA, and counteracts changes in the tRNA pool elicited by the presence of HIV. Our studies identified a novel antiviral mechanism within the innate immune response, in which SLFN11 selectively inhibits viral protein synthesis in HIV-infected cells by means of codon-bias discrimination. PMID:23000900

Codon-usage-based inhibition of HIV protein synthesis by human schlafen 11.

PubMed

Li, Manqing; Kao, Elaine; Gao, Xia; Sandig, Hilary; Limmer, Kirsten; Pavon-Eternod, Mariana; Jones, Thomas E; Landry, Sebastien; Pan, Tao; Weitzman, Matthew D; David, Michael

2012-11-01

In mammals, one of the most pronounced consequences of viral infection is the induction of type I interferons, cytokines with potent antiviral activity. Schlafen (Slfn) genes are a subset of interferon-stimulated early response genes (ISGs) that are also induced directly by pathogens via the interferon regulatory factor 3 (IRF3) pathway. However, many ISGs are of unknown or incompletely understood function. Here we show that human SLFN11 potently and specifically abrogates the production of retroviruses such as human immunodeficiency virus 1 (HIV-1). Our study revealed that SLFN11 has no effect on the early steps of the retroviral infection cycle, including reverse transcription, integration and transcription. Rather, SLFN11 acts at the late stage of virus production by selectively inhibiting the expression of viral proteins in a codon-usage-dependent manner. We further find that SLFN11 binds transfer RNA, and counteracts changes in the tRNA pool elicited by the presence of HIV. Our studies identified a novel antiviral mechanism within the innate immune response, in which SLFN11 selectively inhibits viral protein synthesis in HIV-infected cells by means of codon-bias discrimination.

Uveal Melanoma: Identifying Immunological and Chemotherapeutic Targets to Treat Metastases.

PubMed

Jager, Martine J; Dogrusöz, Mehmet; Woodman, Scott E

2017-01-01

Uveal melanoma is an intraocular malignancy that, depending on its size and genetic make-up, may lead to metastases in up to 50% of cases. Currently, no therapy has been proven to improve survival. However, new therapies exploiting immune responses against metastases are being developed. The primary tumor is well characterized: tumors at high risk of developing metastases often contain macrophages and lymphocytes. However, these lymphocytes are often regulatory T cells that may suppress immune response. Currently, immune checkpoint inhibitors have shown marked efficacy in multiple cancers (eg, cutaneous melanoma) but do not yet improve survival in uveal melanoma patients. More knowledge needs to be acquired regarding the function of T cells in uveal melanoma. Other therapeutic options are related to the biochemical pathways. Targeting the RAF-MEK-ERK pathway with small molecule MEK inhibitors abrogates the growth of UM cells harboring GNAQ/GNA11 Q209 mutations, suggesting that these aberrant G-protein oncogenes mediate, at least in part, their effect through this hallmark proliferation pathway. Other pathways are also implicated, such as those involving c-Jun and YAP. Further studies may show how interference in the different pathways may affect survival. Copyright 2017 Asia-Pacific Academy of Ophthalmology.

Telomeric noncoding RNA promotes mouse embryonic stem cell self-renewal through inhibition of TCF3 activity.

PubMed

Xu, Xiaojuan; Guo, Mengmeng; Zhang, Na; Ye, Shoudong

2018-06-01

Although long noncoding RNAs (lncRNAs) are emerging as new modulators in the fate decision of pluripotent stem cells, the functions of specific lncRNAs remain unclear. Here, we found that telomeric RNA (TERRA or TelRNA), one type of lncRNAs, is highly expressed in mouse embryonic stem cells (mESCs) but declines significantly upon differentiation. TERRA is induced by the Wnt/β-catenin signaling pathway and can reproduce its self-renewal-promoting effect when overexpressed. Further studies revealed that T cell factor 3 ( TCF3) is a potential downstream target of TERRA and mediates the effect of TERRA in mESC maintenance. TERRA inhibits TCF3 transcription, while enforced TCF3 expression abrogates the undifferentiated state of mESCs supported by TERRA. Accordingly, the transcripts of the pluripotency genes Esrrb, Tfcp2l1, and Klf2, repressed by TCF3 in mESCs, are increased in TERRA-overexpressing cells. Our study therefore highlights the important role of TERRA in mESC maintenance and also uncovers a mechanism by which TERRA promotes self-renewal. These data will expand our understanding of the pluripotent regulatory network of ESCs.

Evolving Strategies for Cancer and Autoimmunity: Back to the Future

PubMed Central

Lane, Peter J. L.; McConnell, Fiona M.; Anderson, Graham; Nawaf, Maher G.; Gaspal, Fabrina M.; Withers, David R.

2014-01-01

Although current thinking has focused on genetic variation between individuals and environmental influences as underpinning susceptibility to both autoimmunity and cancer, an alternative view is that human susceptibility to these diseases is a consequence of the way the immune system evolved. It is important to remember that the immunological genes that we inherit and the systems that they control were shaped by the drive for reproductive success rather than for individual survival. It is our view that human susceptibility to autoimmunity and cancer is the evolutionarily acceptable side effect of the immune adaptations that evolved in early placental mammals to accommodate a fundamental change in reproductive strategy. Studies of immune function in mammals show that high affinity antibodies and CD4 memory, along with its regulation, co-evolved with placentation. By dissection of the immunologically active genes and proteins that evolved to regulate this step change in the mammalian immune system, clues have emerged that may reveal ways of de-tuning both effector and regulatory arms of the immune system to abrogate autoimmune responses whilst preserving protection against infection. Paradoxically, it appears that such a detuned and deregulated immune system is much better equipped to mount anti-tumor immune responses against cancers. PMID:24782861

A cytoplasmic serine protein kinase binds and may regulate the Fanconi anemia protein FANCA.

PubMed

Yagasaki, H; Adachi, D; Oda, T; Garcia-Higuera, I; Tetteh, N; D'Andrea, A D; Futaki, M; Asano, S; Yamashita, T

2001-12-15

Fanconi anemia (FA) is an autosomal recessive disease with congenital anomalies, bone marrow failure, and susceptibility to leukemia. Patient cells show chromosome instability and hypersensitivity to DNA cross-linking agents. At least 8 complementation groups (A-G) have been identified and 6 FA genes (for subtypes A, C, D2, E, F, and G) have been cloned. Increasing evidence indicates that a protein complex assembly of multiple FA proteins, including FANCA and FANCG, plays a crucial role in the FA pathway. Previously, it was reported that FANCA was phosphorylated in lymphoblasts from normal controls, whereas the phosphorylation was defective in those derived from patients with FA of multiple complementation groups. The present study examined phosphorylation of FANCA ectopically expressed in FANCA(-) cells. Several patient-derived mutations abrogated in vivo phosphorylation of FANCA in this system, suggesting that FANCA phosphorylation is associated with its function. In vitro phosphorylation studies indicated that a physiologic protein kinase for FANCA (FANCA-PK) forms a complex with the substrate. Furthermore, at least a part of FANCA-PK as well as phosphorylated FANCA were included in the FANCA/FANCG complex. Thus, FANCA-PK appears to be another component of the FA protein complex and may regulate function of FANCA. FANCA-PK was characterized as a cytoplasmic serine kinase sensitive to wortmannin. Identification of the protein kinase is expected to elucidate regulatory mechanisms that control the FA pathway.

Characterization of New Otic Enhancers of the Pou3f4 Gene Reveal Distinct Signaling Pathway Regulation and Spatio-Temporal Patterns

PubMed Central

Robert-Moreno, Àlex; Naranjo, Silvia; de la Calle-Mustienes, Elisa; Gómez-Skarmeta, José Luis; Alsina, Berta

2010-01-01

POU3F4 is a member of the POU-homedomain transcription factor family with a prominent role in inner ear development. Mutations in the human POU3F4 coding unit leads to X-linked deafness type 3 (DFN3), characterized by conductive hearing loss and progressive sensorineural deafness. Microdeletions found 1 Mb 5′ upstream of the coding region also displayed the same phenotype, suggesting that cis-regulatory elements might be present in that region. Indeed, we and others have recently identified several enhancers at the 1 Mb 5′ upstream interval of the pou3f4 locus. Here we characterize the spatio-temporal patterns of these regulatory elements in zebrafish transgenic lines. We show that the most distal enhancer (HCNR 81675) is activated earlier and drives GFP reporter expression initially to a broad ear domain to progressively restrict to the sensory patches. The proximal enhancer (HCNR 82478) is switched later during development and promotes expression, among in other tissues, in sensory patches from its onset. The third enhancer (HCNR 81728) is also active at later stages in the otic mesenchyme and in the otic epithelium. We also characterize the signaling pathways regulating these enhancers. While HCNR 81675 is regulated by very early signals of retinoic acid, HCNR 82478 is regulated by Fgf activity at a later stage and the HCNR 81728 enhancer is under the control of Hh signaling. Finally, we show that Sox2 and Pax2 transcription factors are bound to HCNR 81675 genomic region during otic development and specific mutations to these transcription factor binding sites abrogates HCNR 81675 enhancer activity. Altogether, our results suggest that pou3f4 expression in inner ear might be under the control of distinct regulatory elements that fine-tune the spatio-temporal activity of this gene and provides novel data on the signaling mechanisms controlling pou3f4 function. PMID:21209840

Pulmonary CCR2+CD4+ T cells are immune regulatory and attenuate lung fibrosis development.

PubMed

Milger, Katrin; Yu, Yingyan; Brudy, Eva; Irmler, Martin; Skapenko, Alla; Mayinger, Michael; Lehmann, Mareike; Beckers, Johannes; Reichenberger, Frank; Behr, Jürgen; Eickelberg, Oliver; Königshoff, Melanie; Krauss-Etschmann, Susanne

2017-11-01

Animal models have suggested that CCR2-dependent signalling contributes to the pathogenesis of pulmonary fibrosis, but global blockade of CCL2 failed to improve the clinical course of patients with lung fibrosis. However, as levels of CCR2 + CD4 + T cells in paediatric lung fibrosis had previously been found to be increased, correlating with clinical symptoms, we hypothesised that distinct CCR2 + cell populations might either increase or decrease disease pathogenesis depending on their subtype. To investigate the role of CCR2 + CD4 + T cells in experimental lung fibrosis and in patients with idiopathic pulmonary fibrosis and other fibrosis. Pulmonary CCR2 + CD4 + T cells were analysed using flow cytometry and mRNA profiling, followed by in silico pathway analysis, in vitro assays and adoptive transfer experiments. Frequencies of CCR2 + CD4 + T cells were increased in experimental fibrosis-specifically the CD62L - CD44 + effector memory T cell phenotype, displaying a distinct chemokine receptor profile. mRNA profiling of isolated CCR2 + CD4 + T cells from fibrotic lungs suggested immune regulatory functions, a finding that was confirmed in vitro using suppressor assays. Importantly, adoptive transfer of CCR2 + CD4 + T cells attenuated fibrosis development. The results were partly corroborated in patients with lung fibrosis, by showing higher percentages of Foxp3 + CD25 + cells within bronchoalveolar lavage fluid CCR2 + CD4 + T cells as compared with CCR2 - CD4 + T cells. Pulmonary CCR2 + CD4 + T cells are immunosuppressive, and could attenuate lung inflammation and fibrosis. Therapeutic strategies completely abrogating CCR2-dependent signalling will therefore also eliminate cell populations with protective roles in fibrotic lung disease. This emphasises the need for a detailed understanding of the functions of immune cell subsets in fibrotic lung disease. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

The tail of the ParG DNA segregation protein remodels ParF polymers and enhances ATP hydrolysis via an arginine finger-like motif

PubMed Central

Barillà, Daniela; Carmelo, Emma; Hayes, Finbarr

2007-01-01

The ParF protein of plasmid TP228 belongs to the ubiquitous superfamily of ParA ATPases that drive DNA segregation in bacteria. ATP-bound ParF polymerizes into multistranded filaments. The partner protein ParG is dimeric, consisting of C-termini that interweave into a ribbon–helix–helix domain contacting the centromeric DNA and unstructured N-termini. ParG stimulates ATP hydrolysis by ParF ≈30-fold. Here, we establish that the mobile tails of ParG are crucial for this enhancement and that arginine R19 within the tail is absolutely required for activation of ParF nucleotide hydrolysis. R19 is part of an arginine finger-like loop in ParG that is predicted to intercalate into the ParF nucleotide-binding pocket thereby promoting ATP hydrolysis. Significantly, mutations of R19 abrogated DNA segregation in vivo, proving that intracellular stimulation of ATP hydrolysis by ParG is a key regulatory process for partitioning. Furthermore, ParG bundles ParF-ATP filaments as well as promoting nucleotide-independent polymerization. The N-terminal flexible tail is required for both activities, because N-terminal ΔParG polypeptides are defective in both functions. Strikingly, the critical arginine finger-like residue R19 is dispensable for ParG-mediated remodeling of ParF polymers, revealing that the ParG N-terminal tail possesses two separable activities in the interplay with ParF: a catalytic function during ATP hydrolysis and a mechanical role in modulation of polymerization. We speculate that activation of nucleotide hydrolysis via an arginine finger loop may be a conserved, regulatory mechanism of ParA family members and their partner proteins, including ParA-ParB and Soj-Spo0J that mediate DNA segregation and MinD-MinE that determine septum localization. PMID:17261809

The tail of the ParG DNA segregation protein remodels ParF polymers and enhances ATP hydrolysis via an arginine finger-like motif.

PubMed

Barillà, Daniela; Carmelo, Emma; Hayes, Finbarr

2007-02-06

The ParF protein of plasmid TP228 belongs to the ubiquitous superfamily of ParA ATPases that drive DNA segregation in bacteria. ATP-bound ParF polymerizes into multistranded filaments. The partner protein ParG is dimeric, consisting of C-termini that interweave into a ribbon-helix-helix domain contacting the centromeric DNA and unstructured N-termini. ParG stimulates ATP hydrolysis by ParF approximately 30-fold. Here, we establish that the mobile tails of ParG are crucial for this enhancement and that arginine R19 within the tail is absolutely required for activation of ParF nucleotide hydrolysis. R19 is part of an arginine finger-like loop in ParG that is predicted to intercalate into the ParF nucleotide-binding pocket thereby promoting ATP hydrolysis. Significantly, mutations of R19 abrogated DNA segregation in vivo, proving that intracellular stimulation of ATP hydrolysis by ParG is a key regulatory process for partitioning. Furthermore, ParG bundles ParF-ATP filaments as well as promoting nucleotide-independent polymerization. The N-terminal flexible tail is required for both activities, because N-terminal DeltaParG polypeptides are defective in both functions. Strikingly, the critical arginine finger-like residue R19 is dispensable for ParG-mediated remodeling of ParF polymers, revealing that the ParG N-terminal tail possesses two separable activities in the interplay with ParF: a catalytic function during ATP hydrolysis and a mechanical role in modulation of polymerization. We speculate that activation of nucleotide hydrolysis via an arginine finger loop may be a conserved, regulatory mechanism of ParA family members and their partner proteins, including ParA-ParB and Soj-Spo0J that mediate DNA segregation and MinD-MinE that determine septum localization.

Does helminth activation of toll-like receptors modulate immune response in multiple sclerosis patients?

PubMed Central

Correale, Jorge; Farez, Mauricio F.

2012-01-01

Multiple sclerosis (MS) is an inflammatory autoimmune demyelinating disease affecting the Central Nervous System (CNS), in which Th1 and Th17 cells appear to recognize and react against certain myelin sheath components. Epidemiological evidence has accumulated indicating steady increase in autoimmune disease incidence in developed countries. Reduced infectious disease prevalence in particular has been proposed as the cause. In agreement with this hypothesis, we recently demonstrated significantly better clinical and radiological outcome in helminth-infected MS patients, compared to uninfected ones. Parasite-driven protection was associated with regulatory T cell induction and anti-inflammatory cytokine secretion, including increased TGF-β and IL-10 levels. Interestingly, surface expression of TLR2, on both B cells and dendritic cells (DC) was significantly higher in infected MS patients. Moreover, stimulation of myelin-specific T cell lines with a TLR2 agonist induced inhibition of T cell proliferation, suppression of IFN-γ, IL-12, and IL-17 secretion, as well as increase in IL-10 production, suggesting the functional responses observed correlate with TLR2 expression patterns. Furthermore, parasite antigens were able to induce TLR2 expression on both B cells and DCs. All functional effects mediated by TLR2 were abrogated when MyD88 gene expression was silenced; indicating helminth-mediated signaling induced changes in cytokine secretion in a MyD88-dependent manner. In addition, helminth antigens significantly enhanced co-stimulatory molecule expression, effects not mediated by MyD88. Parasite antigens acting on MyD88 induced significant ERK kinase phosphorylation in DC. Addition of the ERK inhibitor U0126 was associated with dose-dependent IL-10 inhibition and reciprocal enhancement in IL-12, both correlating with ERK inhibition. Finally, cytokine effects and changes observed in co-stimulatory DC molecules after helminth antigen exposure were lost when TLR2 was silenced. Overall, the data described indicate that helminth molecules exert potent regulatory effects on both DCs and B cells from MS patients through TLR2 regulation. PMID:22937527

Optimal ex vivo expansion of neutrophils from PBSC CD34+ cells by a combination of SCF, Flt3-L and G-CSF and its inhibition by further addition of TPO.

PubMed

Tura, Olga; Barclay, G Robin; Roddie, Huw; Davies, John; Turner, Marc L

2007-10-30

Autologous mobilised peripheral blood stem cell (PBSC) transplantation is now a standard approach in the treatment of haematological diseases to reconstitute haematopoiesis following myeloablative chemotherapy. However, there remains a period of severe neutropenia and thrombocytopenia before haematopoietic reconstitution is achieved. Ex vivo expanded PBSC have been employed as an adjunct to unmanipulated HSC transplantation, but have tended to be produced using complex cytokine mixtures aimed at multilineage (neutrophil and megakaryocyte) progenitor expansion. These have been reported to reduce or abrogate neutropenia but have little major effect on thrombocytopenia. Selective megakaryocyte expansion has been to date ineffective in reducing thrombocytopenia. This study was implemented to evaluate neutrophil specific rather than multilineage ex vivo expansion of PBSC for specifically focusing on reduction or abrogation of neutropenia. CD34+ cells (PBSC) were enriched from peripheral blood mononuclear cells following G-CSF-mobilisation and cultured with different permutations of cytokines to determine optimal cytokine combinations and doses for expansion and functional differentiation and maturation of neutrophils and their progenitors. Results were assessed by cell number, morphology, phenotype and function. A simple cytokine combination, SCF + Flt3-L + G-CSF, synergised to optimally expand and mature neutrophil progenitors assessed by cell number, phenotype, morphology and function (superoxide respiratory burst measured by chemiluminescence). G-CSF appears mandatory for functional maturation. Addition of other commonly employed cytokines, IL-3 and IL-6, had no demonstrable additive effect on numbers or function compared to this optimal combination. Addition of TPO, commonly included in multilineage progenitor expansion for development of megakaryocytes, reduced the maturation of neutrophil progenitors as assessed by number, morphology and function (respiratory burst activity). Given that platelet transfusion support is available for autologous PBSC transplantation but granulocyte transfusion is generally lacking, and that multilineage expanded PBSC do not reduce thrombocytopenia, we suggest that instead of multilineage expansion selective neutrophil expansion based on this relatively simple cytokine combination might be prioritized for development for clinical use as an adjunct to unmanipulated PBSC transplantation to reduce or abrogate post-transplant neutropenia.

Ovarian function in mice results in abrogated skeletal muscle PPARdelta and FoxO1-mediated gene expression

USDA-ARS?s Scientific Manuscript database

Menopause, the age-related loss of ovarian hormone production, promotes increased adiposity and associated metabolic pathology, but molecular mechanisms remain unclear. We previously reported that estrogen increases skeletal muscle PPARDelta expression in vivo, and transgenic mice overexpressing mus...

Phosphoproteomics Reveals Regulatory T Cell-Mediated DEF6 Dephosphorylation That Affects Cytokine Expression in Human Conventional T Cells

PubMed Central

Joshi, Rubin N.; Binai, Nadine A.; Marabita, Francesco; Sui, Zhenhua; Altman, Amnon; Heck, Albert J. R.; Tegnér, Jesper; Schmidt, Angelika

2017-01-01

Regulatory T cells (Tregs) control key events of immune tolerance, primarily by suppression of effector T cells. We previously revealed that Tregs rapidly suppress T cell receptor (TCR)-induced calcium store depletion in conventional CD4+CD25− T cells (Tcons) independently of IP3 levels, consequently inhibiting NFAT signaling and effector cytokine expression. Here, we study Treg suppression mechanisms through unbiased phosphoproteomics of primary human Tcons upon TCR stimulation and Treg-mediated suppression, respectively. Tregs induced a state of overall decreased phosphorylation as opposed to TCR stimulation. We discovered novel phosphosites (T595_S597) in the DEF6 (SLAT) protein that were phosphorylated upon TCR stimulation and conversely dephosphorylated upon coculture with Tregs. Mutation of these DEF6 phosphosites abrogated interaction of DEF6 with the IP3 receptor and affected NFAT activation and cytokine transcription in primary Tcons. This novel mechanism and phosphoproteomics data resource may aid in modifying sensitivity of Tcons to Treg-mediated suppression in autoimmune disease or cancer. PMID:28993769

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Indukuri, Hemalatha; Castro, Shawn M.; Liao, S.-M.

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 withmore » 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.« less

Site-directed Nanotherapeutics to Abrogate RRMS and Promote Remyelination Repair (Rev)

DTIC Science & Technology

2013-03-01

in the labs additionally delayed the onset of the practical work.  Long lasting defect in freeze / dryer prevented a more in depth investigation of...drug content and reduced functionality of the peptides. Freezing of the NP without cryoprotection leads to disassembly; freezing with a cryoprotectant

Relaxed selection on the CBF/DREB1 regulatory genes and reduced freezing tolerance in the southern range of Arabidopsis thaliana.

PubMed

Zhen, Ying; Ungerer, Mark C

2008-12-01

Elucidating the molecular basis of adaptive phenotypic variation represents a central aim in evolutionary biology. Traits exhibiting patterns of clinal variation represent excellent models for studies of molecular adaptation, especially when variation in phenotype can be linked to organismal fitness in different environments. Natural accessions of the model plant species Arabidopsis thaliana exhibit clinal variation in freezing tolerance that follows a gradient of temperature variability across the species' native range (Zhen Y, Ungerer MC. 2008. Clinal variation in freezing tolerance among natural accessions of A. thaliana. New Phytol. 177:419-427). Here, we report that this pattern of variation is attributable, at least in part, to relaxed purifying selection on members of a small family of transcriptional activators (the CBF/DREB1s) in the species' southern range. These regulatory genes play a critical role in the ability of A. thaliana plants to undergo cold acclimation and thereby achieve maximum freezing tolerance. Relative to accessions from northern regions, accessions of A. thaliana from the southern part of their geographic range exhibit levels of nonsynonymous nucleotide polymorphism that are approximately 2.8-fold higher across this small gene subfamily. Relaxed selection on the CBF/DREB1s in southern accessions also has resulted in multiple mutations in regulatory regions resulting in abrogated expression of particular subfamily members in particular accessions. These coding-region and regulatory mutations compromise the ability of these genes to act as efficient transcriptional activators during the cold acclimation process, as determined by reductions in rates of induction and maximum levels of expression in the downstream genes they regulate. This study highlights the potential role of regulatory genes in underlying adaptive phenotypic variation in nature.

Ultraviolet B Exposure Inhibits Angiotensin II-Induced Abdominal Aortic Aneurysm Formation in Mice by Expanding CD4+Foxp3+ Regulatory T Cells.

PubMed

Hayashi, Tomohiro; Sasaki, Naoto; Yamashita, Tomoya; Mizoguchi, Taiji; Emoto, Takuo; Amin, Hilman Zulkifli; Yodoi, Keiko; Matsumoto, Takuya; Kasahara, Kazuyuki; Yoshida, Naofumi; Tabata, Tokiko; Kitano, Naoki; Fukunaga, Atsushi; Nishigori, Chikako; Rikitake, Yoshiyuki; Hirata, Ken-Ichi

2017-08-31

Pathogenic immune responses are known to play an important role in abdominal aortic aneurysm (AAA) development. Ultraviolet B (UVB) irradiation has been demonstrated to have therapeutic potential not only for cutaneous diseases but also for systemic inflammatory diseases in mice by suppressing immunoinflammatory responses. We investigated the effect of UVB irradiation on experimental AAA. We used an angiotensin II-induced AAA model in apolipoprotein E-deficient mice fed a high-cholesterol diet. Mice aged 10 weeks were irradiated with 5 kJ/m 2 UVB once weekly for 6 weeks (UVB-irradiated, n=38; nonirradiated, n=42) and were euthanized for evaluation of AAA formation at 16 weeks. Overall, 93% of angiotensin II-infused mice developed AAA, with 60% mortality possibly because of aneurysm rupture. UVB irradiation significantly decreased the incidence (66%) and mortality (29%) of AAA ( P =0.004 and P =0.006, respectively). UVB-irradiated mice had significantly smaller diameter AAA ( P =0.008) and fewer inflammatory cells in the aortic aneurysm tissue than nonirradiated mice, along with systemic expansion of CD4 + Foxp3 + regulatory T cells and decreased effector CD4 + CD44 high CD62L low T cells in para-aortic lymph nodes. Genetic depletion of regulatory T cells abrogated these beneficial effects of UVB treatment, demonstrating a critical role of regulatory T cells. Our data suggest that UVB-dependent expansion of regulatory T cells has beneficial effects on experimental AAA and may provide a novel strategy for the treatment of AAA. © 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.

Arabidopsis HOOKLESS1 Regulates Responses to Pathogens and Abscisic Acid through Interaction with MED18 and Acetylation of WRKY33 and ABI5 Chromatin

PubMed Central

Liao, Chao-Jan; Lee, Sanghun; Mengiste, Tesfaye

2016-01-01

Arabidopsis thaliana HOOKLESS1 (HLS1) encodes a putative histone acetyltransferase with known functions in seedling growth. Here, we show that HLS1 regulates plant responses to pathogens and abscisic acid (ABA) through histone acetylation at chromatin of target loci. The hls1 mutants show impaired responses to bacterial and fungal infection, accelerated senescence, and impaired responses to ABA. HLS1 modulates the expression of WRKY33 and ABA INSENSITIVE5 (ABI5), known regulators of pathogen and ABA responses, respectively, through direct association with these loci. Histone 3 acetylation (H3Ac), a positive mark of transcription, at WRKY33 and ABI5 requires HLS1 function. ABA treatment and pathogen infection enhance HLS1 recruitment and H3Ac at WRKY33. HLS1 associates with Mediator, a eukaryotic transcription coregulatory complex, through direct interaction with mediator subunit 18 (MED18), with which it shares multiple functions. HLS1 recruits MED18 to the WRKY33 promoter, boosting WKRY33 expression, suggesting the synergetic action of HLS1 and MED18. By contrast, MED18 recruitment to ABI5 and transcriptional activation are independent of HLS1. ABA-mediated priming of resistance to fungal infection was abrogated in hls1 and wrky33 mutants but correlated with ABA-induced HLS1 accumulation. In sum, HLS1 provides a regulatory node in pathogen and hormone response pathways through interaction with the Mediator complex and important transcription factors. PMID:27317674

Identification of the Intragenomic Promoter Controlling Hepatitis E Virus Subgenomic RNA Transcription.

PubMed

Ding, Qiang; Nimgaonkar, Ila; Archer, Nicholas F; Bram, Yaron; Heller, Brigitte; Schwartz, Robert E; Ploss, Alexander

2018-05-08

Approximately 20 million hepatitis E virus (HEV) infections occur annually in both developing and industrialized countries. Most infections are self-limiting, but they can lead to chronic infections and cirrhosis in immunocompromised patients, and death in pregnant women. The mechanisms of HEV replication remain incompletely understood due to scarcity of adequate experimental platforms. HEV undergoes asymmetric genome replication, but it produces an additional subgenomic (SG) RNA encoding the viral capsid and a viroporin in partially overlapping open reading frames. Using a novel transcomplementation system, we mapped the intragenomic subgenomic promoter regulating SG RNA synthesis. This cis -acting element is highly conserved across all eight HEV genotypes, and when the element is mutated, it abrogates particle assembly and release. Our work defines previously unappreciated viral regulatory elements and provides the first in-depth view of the intracellular genome dynamics of this emerging human pathogen. IMPORTANCE HEV is an emerging pathogen causing severe liver disease. The genetic information of HEV is encoded in RNA. The genomic RNA is initially copied into a complementary, antigenomic RNA that is a template for synthesis of more genomic RNA and for so-called subgenomic RNA. In this study, we identified the precise region within the HEV genome at which the synthesis of the subgenomic RNA is initiated. The nucleotides within this region are conserved across genetically distinct variants of HEV, highlighting the general importance of this segment for the virus. To identify this regulatory element, we developed a new experimental system that is a powerful tool with broad utility to mechanistically dissect many other poorly understood functional elements of HEV. Copyright © 2018 Ding et al.

Achieving donor-specific hyporesponsiveness is associated with FOXP3+ regulatory T cell recruitment in human renal allograft infiltrates.

PubMed

Bestard, Oriol; Cruzado, Josep M; Mestre, Mariona; Caldés, Anna; Bas, Jordi; Carrera, Marta; Torras, Joan; Rama, Inés; Moreso, Francesc; Serón, Daniel; Grinyó, Josep M

2007-10-01

Exploring new immunosuppressive strategies inducing donor-specific hyporesponsiveness is an important challenge in transplantation. For this purpose, a careful immune monitoring and graft histology assessment is mandatory. Here, we report the results of a pilot study conducted in twenty renal transplant recipients, analyzing the immunomodulatory effects of a protocol based on induction therapy with rabbit anti-thymocyte globulin low doses, sirolimus, and mofetil mycophenolate. Evolution of donor-specific cellular and humoral alloimmune response, peripheral blood lymphocyte subsets and apoptosis was evaluated. Six-month protocol biopsies were performed to assess histological lesions and presence of FOXP3+ regulatory T cells (Tregs) in interstitial infiltrates. After transplantation, there was an early and transient apoptotic effect, mainly within the CD8+ HLADR+ T cells, combined with a sustained enhancement of CD4+ CD25(+high) lymphocytes in peripheral blood. The incidence of acute rejection was 35%, all steroid sensitive. Importantly, only pretransplant donor-specific cellular alloreactivity could discriminate patients at risk to develop acute rejection. Two thirds of the patients became donor-specific hyporesponders at 6 and 24 mo, and the achievement of this immunologic state was not abrogated by prior acute rejection episodes. Remarkably, donor-specific hyporesponders had the better renal function and less chronic renal damage. Donor-specific hyporesponsiveness was inhibited by depleting CD4+ CD25(+high) T cells, which showed donor-Ag specificity. FOXP3+ CD4+ CD25(+high) Tregs both in peripheral blood and in renal infiltrates were higher in donor-specific hyporesponders than in nonhyporesponders, suggesting that the recruitment of Tregs in the allograft plays an important role for renal acceptance. In conclusion, reaching donor-specific hyporesponsiveness is feasible after renal transplantation and associated with Treg recruitment in the graft.

Negative regulation of NLRP3 inflammasome by SIRT1 in vascular endothelial cells.

PubMed

Li, Yanxiang; Yang, Xiaofeng; He, Yanhao; Wang, Weirong; Zhang, Jiye; Zhang, Wei; Jing, Ting; Wang, Bo; Lin, Rong

2017-03-01

NLRP3 inflammasome not only functions as a critical effector in innate immunity, but also triggers the production of proinflammatory cytokines involved in inflammation-associated diseases. Sirtuin 1 (SIRT1) plays an important role in the regulation of cellular inflammation. However, whether the activation of NLRP3 inflammasome is regulated by SIRT1 remains unknown. In this study, we investigated the regulatory effect of SIRT1 on NLRP3 inflammasome and the underlying mechanisms. We found that lipopolysaccharide (LPS) and adenosine triphosphate (ATP)-induced the activation of NLRP3 inflammasome in human umbilical vein endothelial cells (HUVECs). Activation of SIRT1 inhibited NLRP3 inflammasome activation and subsequent caspase-1 cleavage as well as interleukin (IL)-1β secretion, whereas SIRT1 knockdown obviously enhanced the activation of NLRP3 inflammasome in HUVECs. Importantly, gene silencing of SIRT1 abrogated the inhibitory effect of SIRT1 activator on NLRP3 inflammasome formation and IL-1β production in HUVECs stimulated with LPS plus ATP. Further study indicated that cluster of differentiation 40 (CD40) may be involved in the regulation of NLRP3 inflammasome by SIRT1. In vivo studies indicated that implantation of the periarterial carotid collar increased the arterial expression levels of CD40 and CD40 Ligand (CD40L), but inhibited arterial SIRT1 expression in the rabbits. Moreover, treatment with SIRT1 activator decreased CD40 and CD40L levels in collared arteries. Meanwhile, serum IL-1β level, the marker of inflammasome activation, was also inhibited by SIRT1 activation. Taken together, these findings revealed a novel regulatory mechanism of NLRP3 inflammasome by SIRT1, which may be related to suppression of CD40. Copyright © 2016 Elsevier GmbH. All rights reserved.

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

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. Copyright © 2012 Elsevier Ltd. All rights reserved.

Microgravity induces inhibition of osteoblastic differentiation and mineralization through abrogating primary cilia.

PubMed

Shi, Wengui; Xie, Yanfang; He, Jinpeng; Zhou, Jian; Gao, Yuhai; Wei, Wenjun; Ding, Nan; Ma, Huiping; Xian, Cory J; Chen, Keming; Wang, Jufang

2017-05-12

It is well documented that microgravity in space environment leads to bone loss in astronauts. These physiological changes have also been validated by human and animal studies and modeled in cell-based analogs. However, the underlying mechanisms are elusive. In the current study, we identified a novel phenomenon that primary cilia (key sensors and functioning organelles) of rat calvarial osteoblasts (ROBs) gradually shrank and disappeared almost completely after exposure to simulated microgravity generated by a random positioning machine (RPM). Along with the abrogation of primary cilia, the differentiation, maturation and mineralization of ROBs were inhibited. We also found that the disappearance of primary cilia was prevented by treating ROBs with cytochalasin D, but not with LiCl or dynein light chain Tctex-type 1 (Dynlt1) siRNA. The repression of the differentiation, maturation and mineralization of ROBs was effectively offset by cytochalasin D treatment in microgravity conditions. Blocking ciliogenesis using intraflagellar transport protein 88 (IFT88) siRNA knockdown inhibited the ability of cytochalasin D to counteract this reduction of osteogenesis. These results indicate that the abrogation of primary cilia may be responsible for the microgravity's inhibition on osteogenesis. Reconstruction of primary cilia may become a potential strategy against bone loss induced by microgravity.

N-Myc Interactor Inhibits Prototype Foamy Virus by Sequestering Viral Tas Protein in the Cytoplasm

PubMed Central

Hu, Xiaomei; Yang, Wei; Liu, Ruikang; Geng, Yunqi; Qiao, Wentao

2014-01-01

ABSTRACT Foamy viruses (FVs) are complex retroviruses that establish lifelong persistent infection without evident pathology. However, the roles of cellular factors in FV latency are poorly understood. This study revealed that N-Myc interactor (Nmi) could inhibit the replication of prototype foamy virus (PFV). Overexpression of Nmi reduced PFV replication, whereas its depletion by small interfering RNA increased PFV replication. The Nmi-mediated impairment of PFV replication resulted from the diminished transactivation by PFV Tas of the viral long terminal repeat (LTR) and an internal promoter (IP). Nmi was determined to interact with Tas and abrogate its function by sequestration in the cytoplasm. In addition, human and bovine Nmi proteins were found to inhibit the replication of bovine foamy virus (BFV) and PFV. Together, these results indicate that Nmi inhibits both human and bovine FVs by interfering with the transactivation function of Tas and may have a role in the host defense against FV infection. IMPORTANCE From this study, we report that the N-Myc interactor (Nmi), an interferon-induced protein, can interact with the regulatory protein Tas of the prototype foamy virus and sequester it in the cytoplasm. The results of this study suggest that Nmi plays an important role in maintaining foamy virus latency and may reveal a new pathway in the interferon-mediated antiviral barrier against viruses. These findings are important for understanding virus-host relationships not only with FVs but potentially for other retroviruses as well. PMID:24719420

Transcription of fgf8 is regulated by activating and repressive cis-elements at the midbrain-hindbrain boundary in zebrafish embryos.

PubMed

Inoue, Fumitaka; Parvin, Mst Shahnaj; Yamasu, Kyo

2008-04-15

Fgf8 is expressed in the isthmic region of the developing brain, serving an organizing function in vertebrate embryos. We previously identified S4.2 downstream to the zebrafish fgf8 gene as a regulatory region that drives transcription in the anterior hindbrain. Here, we investigated the mechanism of fgf8 regulation by the S4.2 region during development. Reporter analyses in embryos revealed that S4.2 closely recapitulates fgf8 expression in the anteriormost hindbrain during somitogenesis. This region contains a sequence highly conserved in fgf8 of diverse vertebrates. Further analyses of S4.2 revealed a 342-bp core region composed of three subregions (#2, #3, and #4). Regions #3 and #4 drove expression broadly in the brain from the midbrain to r5 of the hindbrain, whereas a 28-bp sequence in #2 repressed ectopic expression in the midbrain and in r2 to r5. The enhancer function of S4.2 was absent in pax2a mutant embryos, while it was activated ectopically by pax2a misexpression in the hindbrain. We identified two sites in the core region that are bound by Pax2a in vitro and in vivo, the disruption of which abrogated the S4.2 activity. Thus, fgf8 expression in the anteriormost hindbrain involves activation and repression, with Pax2a as a pivotal regulator.

High glucose disrupts oligosaccharide recognition function via competitive inhibition: a potential mechanism for immune dysregulation in diabetes mellitus.

PubMed

Ilyas, Rebecca; Wallis, Russell; Soilleux, Elizabeth J; Townsend, Paul; Zehnder, Daniel; Tan, Bee K; Sim, Robert B; Lehnert, Hendrik; Randeva, Harpal S; Mitchell, Daniel A

2011-01-01

Diabetic complications include infection and cardiovascular disease. Within the immune system, host-pathogen and regulatory host-host interactions operate through binding of oligosaccharides by C-type lectin. A number of C-type lectins recognise oligosaccharides rich in mannose and fucose - sugars with similar structures to glucose. This raises the possibility that high glucose conditions in diabetes affect protein-oligosaccharide interactions via competitive inhibition. Mannose-binding lectin, soluble DC-SIGN and DC-SIGNR, and surfactant protein D, were tested for carbohydrate binding in the presence of glucose concentrations typical of diabetes, via surface plasmon resonance and affinity chromatography. Complement activation assays were performed in high glucose. DC-SIGN and DC-SIGNR expression in adipose tissues was examined via immunohistochemistry. High glucose inhibited C-type lectin binding to high-mannose glycoprotein and binding of DC-SIGN to fucosylated ligand (blood group B) was abrogated in high glucose. Complement activation via the lectin pathway was inhibited in high glucose and also in high trehalose - a nonreducing sugar with glucoside stereochemistry. DC-SIGN staining was seen on cells with DC morphology within omental and subcutaneous adipose tissues. We conclude that high glucose disrupts C-type lectin function, potentially illuminating new perspectives on susceptibility to infectious and inflammatory disease in diabetes. Mechanisms involve competitive inhibition of carbohydrate binding within sets of defined proteins, in contrast to broadly indiscriminate, irreversible glycation of proteins. Copyright © 2010 Elsevier GmbH. All rights reserved.

Timely Degradation of Wip1 Phosphatase by APC/C Activator Protein Cdh1 is Necessary for Normal Mitotic Progression.

PubMed

Jeong, Ho-Chang; Gil, Na-Yeon; Lee, Ho-Soo; Cho, Seung-Ju; Kim, Kyungtae; Chun, Kwang-Hoon; Cho, Hyeseong; Cha, Hyuk-Jin

2015-08-01

Wip1 belongs to the protein phosphatase C (PP2C) family, of which expression is up-regulated by a number of external stresses, and serves as a stress modulator in normal physiological conditions. When overexpressed, premature dephosphorylation of stress-mediators by Wip1 results in abrogation of tumor surveillance, thus Wip1 acts as an oncogene. Previously, the functional regulation of Wip1 in cell-cycle progression by counteracting cellular G1 and G2/M checkpoint activity in response to DNA damage was reported. However, other than in stress conditions, the function and regulatory mechanism of Wip1 has not been fully determined. Herein, we demonstrated that protein regulation of Wip1 occurs in a cell cycle-dependent manner, which is directly governed by APC/C(Cdh1) at the end of mitosis. In particular, we also showed evidence that Wip1 phosphatase activity is closely associated with its own protein stability, suggesting that reduced phosphatase activity of Wip1 during mitosis could trigger its degradation. Furthermore, to verify the physiological role of its phosphatase activity during mitosis, we established doxycycline-inducible cell models, including a Wip1 wild type (WT) and phosphatase dead mutant (Wip1 DA). When ectopically expressing Wip1 WT, we observed a delay in the transition from metaphase to anaphase. In conclusion, these studies show that mitotic degradation of Wip1 by APC/C(Cdh1) is important for normal mitotic progression. © 2015 Wiley Periodicals, Inc.

Pivotal roles of CD4+ effector T cells in mediating agonistic anti-GITR mAb-induced-immune activation and tumor immunity in CT26 tumors.

PubMed

Zhou, Pengfei; L'italien, Lawrence; Hodges, Douglas; Schebye, Xiao Min

2007-12-01

Glucocorticoid-induced TNF receptor family related protein (GITR) is a member of the TNFR superfamily. Previous studies have shown that in vivo administration of a GITR agonistic Ab (DTA-1) is able to overcome tolerance and induce tumor rejection in several murine syngeneic tumor models. However, little is known about the in vivo targets and the mechanisms of how this tolerance is overcome in a tumor-bearing host, nor is much known about how the immune network is regulated to achieve this antitumor response. In this study, we demonstrate that the in vivo ligation of GITR on CD4(+) effector T cells renders them refractory to suppression by regulatory T (T(reg)) cells in the CT26 tumor-bearing mouse. GITR engagement on T(reg) cells does not appear to directly abrogate their suppressive function; rather, it increases the expansion of T(reg) cells and promotes IL-10 production, a cytokine important for their suppressive function. Moreover, CD4(+) effector T cells play a crucial role in mediating DTA-1-induced immune activation and expansion of CD8(+), NK, and B cells in the tumor-draining lymph nodes. This includes increased CD69 expression on all of these subsets. In addition, NK and tumor-specific CD8(+) T cells are generated that are cytolytic, which show increased intracellular IFN-gamma production and CD107a mobilization, the latter a hallmark of cytolytic activities that lead to tumor killing.

A ChIP-chip approach reveals a novel role for transcription factor IRF1 in the DNA damage response.

PubMed

Frontini, Mattia; Vijayakumar, Meeraa; Garvin, Alexander; Clarke, Nicole

2009-03-01

IRF1 is a transcription factor that regulates key processes in the immune system and in tumour suppression. To gain further insight into IRF1's role in these processes, we searched for new target genes by performing chromatin immunoprecipitation coupled to a CpG island microarray (ChIP-chip). Using this approach we identified 202 new IRF1-binding sites with high confidence. Functional categorization of the target genes revealed a surprising cadre of new roles that can be linked to IRF1. One of the major functional categories was the DNA damage response pathway. In order to further validate our findings, we show that IRF1 can regulate the mRNA expression of a number of the DNA damage response genes in our list. In particular, we demonstrate that the mRNA and protein levels of the DNA repair protein BRIP1 [Fanconi anemia gene J (FANC J)] are upregulated after IRF1 over-expression. We also demonstrate that knockdown of IRF1 by siRNA results in loss of BRIP1 expression, abrogation of BRIP1 foci after DNA interstrand crosslink (ICL) damage and hypersensitivity to the DNA crosslinking agent, melphalan; a characteristic phenotype of FANC J cells. Taken together, our data provides a more complete understanding of the regulatory networks controlled by IRF1 and reveals a novel role for IRF1 in regulating the ICL DNA damage response.

A ChIP–chip approach reveals a novel role for transcription factor IRF1 in the DNA damage response

PubMed Central

Frontini, Mattia; Vijayakumar, Meeraa; Garvin, Alexander; Clarke, Nicole

2009-01-01

IRF1 is a transcription factor that regulates key processes in the immune system and in tumour suppression. To gain further insight into IRF1's role in these processes, we searched for new target genes by performing chromatin immunoprecipitation coupled to a CpG island microarray (ChIP–chip). Using this approach we identified 202 new IRF1-binding sites with high confidence. Functional categorization of the target genes revealed a surprising cadre of new roles that can be linked to IRF1. One of the major functional categories was the DNA damage response pathway. In order to further validate our findings, we show that IRF1 can regulate the mRNA expression of a number of the DNA damage response genes in our list. In particular, we demonstrate that the mRNA and protein levels of the DNA repair protein BRIP1 [Fanconi anemia gene J (FANC J)] are upregulated after IRF1 over-expression. We also demonstrate that knockdown of IRF1 by siRNA results in loss of BRIP1 expression, abrogation of BRIP1 foci after DNA interstrand crosslink (ICL) damage and hypersensitivity to the DNA crosslinking agent, melphalan; a characteristic phenotype of FANC J cells. Taken together, our data provides a more complete understanding of the regulatory networks controlled by IRF1 and reveals a novel role for IRF1 in regulating the ICL DNA damage response. PMID:19129219

HIF-1α regulates epithelial inflammation by cell autonomous NFκB activation and paracrine stromal remodeling

PubMed Central

Scortegagna, Marzia; Cataisson, Christophe; Martin, Rebecca J.; Hicklin, Daniel J.; Schreiber, Robert D.; Yuspa, Stuart H.

2008-01-01

Hypoxia inducible factor-1 (HIF-1) is a master regulatory transcription factor controlling multiple cell-autonomous and non–cell-autonomous processes, such as metabolism, angiogenesis, matrix invasion, and cancer metastasis. Here we used a new line of transgenic mice with constitutive gain of HIF-1 function in basal keratinocytes and demonstrated a signaling pathway from HIF-1 to nuclear factor κ B (NFκB) activation to enhanced epithelial chemokine and cytokine elaboration. This pathway was responsible for a phenotypically silent accumulation of stromal inflammatory cells and a marked inflammatory hypersensitivity to a single 12-O-tetradecanoylphorbol-13-acetate (TPA) challenge. HIF-1–induced NFκB activation was composed of 2 elements, IκB hyperphosphorylation and phosphorylation of Ser276 on p65, enhancing p65 nuclear localization and transcriptional activity, respectively. NFκB transcriptional targets macrophage inflammatory protein-2 (MIP-2/CXCL2/3), keratinocyte chemokine (KC/CXCL1), and tumor necrosis factor [alfa] (TNFα) were constitutively up-regulated and further increased after TPA challenge both in cultured keratinocytes and in transgenic mice. Whole animal KC, MIP-2, or TNFα immunodepletion each abrogated TPA-induced inflammation, whereas blockade of either VEGF or placenta growth factor (PlGF) signaling did not affect transgenic inflammatory hyper-responsiveness. Thus, epithelial HIF-1 gain of function remodels the local environment by cell-autonomous NFκB-mediated chemokine and cytokine secretion, which may be another mechanism by which HIF-1 facilitates either inflammatory diseases or malignant progression. PMID:18199827

Blueberry and malvidin inhibit cell cycle progression and induce mitochondrial-mediated apoptosis by abrogating the JAK/STAT-3 signalling pathway.

PubMed

Baba, Abdul Basit; Nivetha, Ramesh; Chattopadhyay, Indranil; Nagini, Siddavaram

2017-11-01

Blueberries, a rich source of anthocyanins have attracted considerable attention as functional foods that confer immense health benefits including anticancer properties. Herein, we assessed the potential of blueberry and its major constituent malvidin to target STAT-3, a potentially druggable oncogenic transcription factor with high therapeutic index. We demonstrate that blueberry abrogates the JAK/STAT-3 pathway and modulates downstream targets that influence cell proliferation and apoptosis in a hamster model of oral oncogenesis. Further, we provide mechanistic evidence that blueberry and malvidin function as STAT-3 inhibitors in the oral cancer cell line SCC131. Blueberry and malvidin suppressed STAT-3 phosphorylation and nuclear translocation thereby inducing cell cycle arrest and mitochondrial-mediated apoptosis. However, the combination of blueberry and malvidin with the STAT-3 inhibitor S3I-201 was more efficacious in STAT-3 inhibition relative to single agents. The present study has provided leads for the development of novel combinations of compounds that can serve as inhibitors of STAT-mediated oncogenic signalling. Copyright © 2017 Elsevier Ltd. All rights reserved.

Helminth Infections Decrease Host Susceptibility to Immune-Mediated Diseases

PubMed Central

Weinstock, Joel V; Elliott, David E.

2014-01-01

Helminthic infection has become rare in highly industrialized nations. Concurrent with the decline in helminthic infection is an increase in prevalence of inflammatory disease. Removal of helminths from our environment and their powerful effects on host immunity may have contributed to this increase. Several different helminth species can abrogate disease in murine models of inflammatory bowel disease, type 1 diabetes, multiple sclerosis and other conditions. Helminths evoke immune regulatory pathways often involving dendritic cells, Tregs and macrophages that help control disease. Cytokines such as IL4, IL10 and TGFβ have a role. Notable is helminthic modulatory effect on innate immunity, which impedes development of aberrant adaptive immunity. Investigators are identifying key helminth-derived immune modulatory molecules that may have therapeutic utility in the control of inflammatory disease. PMID:25240019

SATB1 Expression Governs Epigenetic Repression of PD-1 in Tumor-Reactive T Cells.

PubMed

Stephen, Tom L; Payne, Kyle K; Chaurio, Ricardo A; Allegrezza, Michael J; Zhu, Hengrui; Perez-Sanz, Jairo; Perales-Puchalt, Alfredo; Nguyen, Jenny M; Vara-Ailor, Ana E; Eruslanov, Evgeniy B; Borowsky, Mark E; Zhang, Rugang; Laufer, Terri M; Conejo-Garcia, Jose R

2017-01-17

Despite the importance of programmed cell death-1 (PD-1) in inhibiting T cell effector activity, the mechanisms regulating its expression remain poorly defined. We found that the chromatin organizer special AT-rich sequence-binding protein-1 (Satb1) restrains PD-1 expression induced upon T cell activation by recruiting a nucleosome remodeling deacetylase (NuRD) complex to Pdcd1 regulatory regions. Satb1 deficienct T cells exhibited a 40-fold increase in PD-1 expression. Tumor-derived transforming growth factor β (Tgf-β) decreased Satb1 expression through binding of Smad proteins to the Satb1 promoter. Smad proteins also competed with the Satb1-NuRD complex for binding to Pdcd1 enhancers, releasing Pdcd1 expression from Satb1-mediated repression, Satb1-deficient tumor-reactive T cells lost effector activity more rapidly than wild-type lymphocytes at tumor beds expressing PD-1 ligand (CD274), and these differences were abrogated by sustained CD274 blockade. Our findings suggest that Satb1 functions to prevent premature T cell exhaustion by regulating Pdcd1 expression upon T cell activation. Dysregulation of this pathway in tumor-infiltrating T cells results in diminished anti-tumor immunity. Copyright © 2017 Elsevier Inc. All rights reserved.

MUC1 (CD227) interacts with lck tyrosine kinase in Jurkat lymphoma cells and normal T cells.

PubMed

Mukherjee, P; Tinder, T L; Basu, G D; Gendler, S J

2005-01-01

MUC1 (CD227) is a large transmembrane epithelial mucin glycoprotein, which is aberrantly overexpressed in most adenocarcinomas and is a target for immune therapy for epithelial tumors. Recently, MUC1 has been detected in a variety of hematopoietic cell malignancies including T and B cell lymphomas and myelomas; however, its function in these cells is not clearly defined. Using the Jurkat T cell lymphoma cell line and normal human T cells, we demonstrate that MUC1 is not only expressed in these cells but is also phosphorylated upon T cell receptor (TCR) ligation and associates with the Src-related T cell tyrosine kinase, p56lck. Upon TCR-mediated activation of Jurkat cells, MUC1 is found in the low-density membrane fractions, where linker of T cell activation is contained. Abrogation of MUC1 expression in Jurkat cells by MUC1-specific small interfering RNA resulted in defects in TCR-mediated downstream signaling events associated with T cell activation. These include reduction in Ca2+ influx and extracellular signal-regulated kinase 1/2 phosphorylation, leading to a decrease in CD69 expression, proliferation, and interleukin-2 production. These results suggest a regulatory role of MUC1 in modulating proximal signal transduction events through its interaction with proteins of the activation complex.

Embryonic demise caused by targeted disruption of a cysteine protease Dub-2.

PubMed

Baek, Kwang-Hyun; Lee, Heyjin; Yang, Sunmee; Lim, Soo-Bin; Lee, Wonwoo; Lee, Jeoung Eun; Lim, Jung-Jin; Jun, Kisun; Lee, Dong-Ryul; Chung, Young

2012-01-01

A plethora of biological metabolisms are regulated by the mechanisms of ubiquitination, wherein this process is balanced with the action of deubiquitination system. Dub-2 is an IL-2-inducible, immediate-early gene that encodes a deubiquitinating enzyme with growth regulatory activity. DUB-2 presumably removes ubiquitin from ubiquitin-conjugated target proteins regulating ubiquitin-mediated proteolysis, but its specific target proteins are unknown yet. To elucidate the functional role of Dub-2, we generated genetically modified mice by introducing neo cassette into the second exon of Dub-2 and then homologous recombination was done to completely abrogate the activity of DUB-2 proteins. We generated Dub-2+/- heterozygous mice showing a normal phenotype and are fertile, whereas new born mouse of Dub-2-/- homozygous alleles could not survive. In addition, Dub-2-/- embryo could not be seen between E6.5 and E12.5 stages. Furthermore, the number of embryos showing normal embryonic development for further stages is decreased in heterozygotes. Even embryonic stem cells from inner cell mass of Dub-2-/- embryos could not be established. Our study suggests that the targeted disruption of Dub-2 may cause embryonic lethality during early gestation, possibly due to the failure of cell proliferation during hatching process.

Role of Natural IgM Autoantibodies (IgM-NAA) and IgM Anti-Leukocyte Antibodies (IgM-ALA) in Regulating Inflammation.

PubMed

Lobo, Peter I

2017-01-01

Natural IgM autoantibodies (IgM-NAA) are rapidly produced to inhibit pathogens and abrogate inflammation mediated by invading microorganisms and host neoantigens. IgM-NAA achieve this difficult task by being polyreactive with low binding affinity but with high avidity, characteristics that allow these antibodies to bind antigenic determinants shared by pathogens and neoantigens. Hence the same clones of natural IgM can bind and mask host neoantigens as well as inhibit microorganisms. In addition, IgM-NAA regulate the inflammatory response via mechanisms involving binding of IgM to apoptotic cells to enhance their removal and binding of IgM to live leukocytes to regulate their function. Secondly, we review how natural IgM prevents autoimmune disorders arising from pathogenic IgG autoantibodies as well as by autoreactive B and T cells that have escaped tolerance mechanisms. Thirdly, using IgM knockout mice, we show that regulatory B and T cells require IgM to effectively regulate inflammation mediated by innate, adaptive and autoimmune mechanisms. It is therefore not surprising why the host positively selects such autoreactive B1 cells that generate protective IgM-NAA, which are also evolutionarily conserved. Fourthly, we show that IgM anti-leukocyte autoantibodies (IgM-ALA) levels and their repertoire can vary in normal humans and disease states and this variation may partly explain the observed differences in the inflammatory response after infection, ischemic injury or after a transplant. Finally we also show how protective IgM-NAA can be rendered pathogenic under non-physiological conditions. IgM-NAA have therapeutic potential. Polyclonal IgM infusions can be used to abrogate ongoing inflammation. Additionally, inflammation arising after ischemic kidney injury, e.g., during high-risk elective cardiac surgery or after allograft transplantation, can be prevented by pre-emptively infusing polyclonal IgM, or DC pretreated ex vivo with IgM, or by increasing in vivo IgM with a vaccine approach. Cell therapy with IgM pretreated cells, is appealing as less IgM will be required.

Making Aggressive Prostate Cancer Quiescent by Abrogating Cholesterol Esterification

DTIC Science & Technology

2015-10-01

AWARD NUMBER: W81XWH-14-1-0557 TITLE: Making Aggressive Prostate Cancer Quiescent by Abrogating Cholesterol Esterification PRINCIPAL...Aggressive Prostate Cancer Quiescent by Abrogating Cholesterol Esterification 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-14-1-0557 5c. PROGRAM...application is to establish the viability of a new strategy of treating late stage PCa through therapeutic targeting of cholesterol metabolism in vivo

Oxidative Stress Increases the Blood Brain Barrier Permeability Resulting in Increased Incidence of Brain Metastasis in BRCA Mutation Carriers

DTIC Science & Technology

2014-08-01

increase in ROS levels as compared to control, and this increased in ROS formation was abrogated by the antioxidant uric acid , UA (Table 1). Table 1...presence of UA antioxidant, uric acid , indicating the involvement of ROS in loss of the HBMEC integrity. The functional changes paralleled enhanced

Therapy-induced selective loss of leukemia-initiating activity in murine adult T cell leukemia

PubMed Central

El Hajj, Hiba; El-Sabban, Marwan; Hasegawa, Hideki; Zaatari, Ghazi; Ablain, Julien; Saab, Shahrazad T.; Janin, Anne; Mahfouz, Rami; Nasr, Rihab; Kfoury, Youmna; Nicot, Christophe; Hermine, Olivier; Hall, William

2010-01-01

Chronic HTLV-I (human T cell lymphotropic virus type I) infection may cause adult T cell leukemia/lymphoma (ATL), a disease with dismal long-term prognosis. The HTLV-I transactivator, Tax, initiates ATL in transgenic mice. In this study, we demonstrate that an As2O3 and IFN-α combination, known to trigger Tax proteolysis, cures Tax-driven ATL in mice. Unexpectedly, this combination therapy abrogated initial leukemia engraftment into secondary recipients, whereas the primary tumor bulk still grew in the primary hosts, only to ultimately abate later on. This loss of initial transplantability required proteasome function. A similar regimen recently yielded unprecedented disease control in human ATL. Our demonstration that this drug combination targeting Tax stability abrogates tumor cell immortality but not short-term growth may foretell a favorable long-term efficiency of this regimen in patients. PMID:21135137

Trimeric association of Hox and TALE homeodomain proteins mediates Hoxb2 hindbrain enhancer activity.

PubMed

Jacobs, Y; Schnabel, C A; Cleary, M L

1999-07-01

Pbx/exd proteins modulate the DNA binding affinities and specificities of Hox proteins and contribute to the execution of Hox-dependent developmental programs in arthropods and vertebrates. Pbx proteins also stably heterodimerize and bind DNA with Meis and Pknox1-Prep1, additional members of the TALE (three-amino-acid loop extension) superclass of homeodomain proteins that function on common genetic pathways with a subset of Hox proteins. In this study, we demonstrated that Pbx and Meis bind DNA as heterotrimeric complexes with Hoxb1 on a genetically defined Hoxb2 enhancer, r4, that mediates the cross-regulatory transcriptional effects of Hoxb1 in vivo. The DNA binding specificity of the heterotrimeric complex for r4 is mediated by a Pbx-Hox site in conjunction with a distal Meis site, which we showed to be required for ternary complex formation and Meis-enhanced transcription. Formation of heterotrimeric complexes in which all three homeodomains bind their cognate DNA sites is topologically facilitated by the ability of Pbx and Meis to interact through their amino termini and bind DNA without stringent half-site orientation and spacing requirements. Furthermore, Meis site mutation in the Hoxb2 enhancer phenocopies Pbx-Hox site mutation to abrogate enhancer-directed expression of a reporter transgene in the murine embryonic hindbrain, demonstrating that DNA binding by all three proteins is required for trimer function in vivo. Our data provide in vitro and in vivo evidence for the combinatorial regulation of Hox and TALE protein functions that are mediated, in part, by their interdependent DNA binding activities as ternary complexes. As a consequence, Hoxb1 employs Pbx and Meis-related proteins, as a pair of essential cofactors in a higher-order molecular complex, to mediate its transcriptional effects on an endogenous Hox response element.

Activity and subcellular compartmentalization of peroxisome proliferator-activated receptor alpha are altered by the centrosome-associated protein CAP350.

PubMed

Patel, Hansa; Truant, Ray; Rachubinski, Richard A; Capone, John P

2005-01-01

Peroxisome proliferator-activated nuclear hormone receptors (PPAR) are ligand-activated transcription factors that play pivotal roles in governing metabolic homeostasis and cell growth. PPARs are primarily in the nucleus but, under certain circumstances, can be found in the cytoplasm. We show here that PPAR(alpha) interacts with the centrosome-associated protein CAP350. CAP350 also interacts with PPAR(delta), PPAR(gamma) and liver-X-receptor alpha, but not with the 9-cis retinoic acid receptor, RXR(alpha). Immunofluorescence analysis indicated that PPAR(alpha) is diffusely distributed in the nucleus and excluded from the cytoplasm. However, in the presence of coexpressed CAP350, PPAR(alpha) colocalizes with CAP350 to discrete nuclear foci and to the centrosome, perinuclear region and intermediate filaments. In contrast, the subcellular distribution of RXR(alpha) or of thyroid hormone receptor alpha was not altered by coexpression of CAP350. An amino-terminal fragment of CAP350 was localized exclusively to nuclear foci and was sufficient to recruit PPAR(alpha) to these sites. Mutation of the single putative nuclear hormone receptor interacting signature motif LXXLL present in this fragment had no effect on its subnuclear localization but abrogated recruitment of PPAR(alpha) to nuclear foci. Surprisingly, mutation of the LXXLL motif in this CAP350 subfragment did not prevent its binding to PPAR(alpha) in vitro, suggesting that this motif serves some function other than PPAR(alpha) binding in recruiting PPAR(alpha) to nuclear spots. CAP350 inhibited PPAR(alpha)-mediated transactivation in an LXXLL-dependent manner, suggesting that CAP350 represses PPAR(alpha) function. Our findings implicate CAP350 in a dynamic process that recruits PPAR(alpha) to discrete nuclear and cytoplasmic compartments and suggest that altered intracellular compartmentalization represents a regulatory process that modulates PPAR function.

A close connection between the PERK and IRE arms of the UPR and the transcriptional regulation of autophagy.

PubMed

Deegan, Shane; Koryga, Izabela; Glynn, Sharon A; Gupta, Sanjeev; Gorman, Adrienne M; Samali, Afshin

2015-01-02

Endoplasmic reticulum (ER) stress is known to lead to activation of both the unfolded protein response (UPR) and autophagy. Although regulatory connections have been identified between the UPR and autophagy, it is still unclear to what extent the UPR regulates the genes involved at the different stages of the autophagy pathway. Here, we carried out a microarray analysis of HCT116 cells subjected to ER stress and observed the transcriptional upregulation of a large cohort of autophagy-related genes. Of particular interest, we identified the transcriptional upregulation of the autophagy receptor genes SQSTM1/p62, NBR1 and BNIP3L/NIX in response to ER stress and show that the inhibition of the UPR transmembrane receptors, PERK and IRE1, abrogates this upregulation. Copyright © 2014 Elsevier Inc. All rights reserved.

Mediation of mouse natural cytotoxic activity by tumour necrosis factor

NASA Astrophysics Data System (ADS)

Ortaldo, John R.; Mason, Llewellyn H.; Mathieson, Bonnie J.; Liang, Shu-Mei; Flick, David A.; Herberman, Ronald B.

1986-06-01

Natural cell-mediated cytotoxic activity in the mouse has been associated with two types of effector cells, the natural killer (NK) cell and the natural cytotoxic (NC) cell, which seem to differ with regard to their patterns of target selectivity, cell surface characteristics and susceptibility to regulatory factors1. During studies on the mechanism of action of cytotoxic molecules, it became evident that WEHI-164, the prototype NC target cell, was highly susceptible to direct lysis by both human and mouse recombinant tumour necrosis factor (TNF). Here we show that NC, but not NK activity mediated by normal splenocytes, is abrogated by rabbit antibodies to recombinant and natural TNF, respectively. Thus, the cell-mediated activity defined as NC is due to release of TNF by normal spleen cells and does not represent a unique natural effector mechanism.

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.

IL-23 (Interleukin-23)-Producing Conventional Dendritic Cells Control the Detrimental IL-17 (Interleukin-17) Response in Stroke.

PubMed

Gelderblom, Mathias; Gallizioli, Mattia; Ludewig, Peter; Thom, Vivien; Arunachalam, Priyadharshini; Rissiek, Björn; Bernreuther, Christian; Glatzel, Markus; Korn, Thomas; Arumugam, Thiruma Valavan; Sedlacik, Jan; Gerloff, Christian; Tolosa, Eva; Planas, Anna M; Magnus, Tim

2018-01-01

Inflammatory mechanisms can exacerbate ischemic tissue damage and worsen clinical outcome in patients with stroke. Both αβ and γδ T cells are established mediators of tissue damage in stroke, and the role of dendritic cells (DCs) in inducing the early events of T cell activation and differentiation in stroke is not well understood. In a murine model of experimental stroke, we defined the immune phenotype of infiltrating DC subsets based on flow cytometry of surface markers, the expression of ontogenetic markers, and cytokine levels. We used conditional DC depletion, bone marrow chimeric mice, and IL-23 (interleukin-23) receptor-deficient mice to further explore the functional role of DCs. We show that the ischemic brain was rapidly infiltrated by IRF4 + /CD172a + conventional type 2 DCs and that conventional type 2 DCs were the most abundant subset in comparison with all other DC subsets. Twenty-four hours after ischemia onset, conventional type 2 DCs became the major source of IL-23, promoting neutrophil infiltration by induction of IL-17 (interleukin-17) in γδ T cells. Functionally, the depletion of CD11c + cells or the genetic disruption of the IL-23 signaling abrogated both IL-17 production in γδ T cells and neutrophil infiltration. Interruption of the IL-23/IL-17 cascade decreased infarct size and improved neurological outcome after stroke. Our results suggest a central role for interferon regulatory factor 4-positive IL-23-producing conventional DCs in the IL-17-dependent secondary tissue damage in stroke. © 2017 American Heart Association, Inc.

Integrated microarray and ChIP analysis identifies multiple Foxa2 dependent target genes in the notochord.

PubMed

Tamplin, Owen J; Cox, Brian J; Rossant, Janet

2011-12-15

The node and notochord are key tissues required for patterning of the vertebrate body plan. Understanding the gene regulatory network that drives their formation and function is therefore important. Foxa2 is a key transcription factor at the top of this genetic hierarchy and finding its targets will help us to better understand node and notochord development. We performed an extensive microarray-based gene expression screen using sorted embryonic notochord cells to identify early notochord-enriched genes. We validated their specificity to the node and notochord by whole mount in situ hybridization. This provides the largest available resource of notochord-expressed genes, and therefore candidate Foxa2 target genes in the notochord. Using existing Foxa2 ChIP-seq data from adult liver, we were able to identify a set of genes expressed in the notochord that had associated regions of Foxa2-bound chromatin. Given that Foxa2 is a pioneer transcription factor, we reasoned that these sites might represent notochord-specific enhancers. Candidate Foxa2-bound regions were tested for notochord specific enhancer function in a zebrafish reporter assay and 7 novel notochord enhancers were identified. Importantly, sequence conservation or predictive models could not have readily identified these regions. Mutation of putative Foxa2 binding elements in two of these novel enhancers abrogated reporter expression and confirmed their Foxa2 dependence. The combination of highly specific gene expression profiling and genome-wide ChIP analysis is a powerful means of understanding developmental pathways, even for small cell populations such as the notochord. Copyright © 2011 Elsevier Inc. All rights reserved.

CCN5 attenuates profibrotic phenotypes of fibroblasts through the Smad6-CCN2 pathway: Potential role in epidural fibrosis.

PubMed

Xu, Honghai; Liu, Cong; Sun, Zhengming; Guo, Xiong; Zhang, Yuelin; Liu, Mengting; Li, Peng

2015-07-01

Epidural fibrosis is characterized by the development of dense and thick scar tissue adjacent to the dural mater and ranked as the major contributor for post-operative pain recurrence after laminectomy or discectomy. Recently, CCN5 exhibited an inhibitory effect on connective tissue growth factor (CTGF)/CCN2 (a critical regulator for fibrotic disease)‑mediated fibrogenesis. However, its function in epidural fibrosis and the underlying mechanisms involved remain to be determined. In this study, an obvious downregulation of CCN5 was observed in scar tissues from laminectomized rats, concomitant with a marked upregulation of CCN2, suggesting a potential negative regulatory role of CCN5 in fibrogenesis. Furthermore, CCN5 overexpression notably mitigated transforming growth factor‑β1-enhanced fibroblast viability and proliferation. Of note, CCN5 upregulation inhibited the switch of fibroblasts into myofibroblasts as its overexpression abrogated the expression of the myofibroblast marker, α-smooth muscle actin (α-SMA). CCN5 upregulation also reduced an increase in collagen type I, α1 (COL1A1) and total collagen concentrations. Additionally, CCN5 over-expression decreased CCN2 expression and increased Smad6 phosphorylation. Mechanism analysis revealed that blocking Smad6 signaling significantly ameliorated the inhibitory effect of CCN5 on the CCN2 levels, accompanied by the reduction in cell proliferation and collagen production. These results confirm that CCN5 exerts an anti-fibrotic function by regulating the Smad6-CCN2 pathway, thereby indicating a potential approach for ameliorating epidural fibrosis after laminectomy.

Serendipitous Discovery of an Immunoglobulin-Binding Autotransporter in Bordetella Species▿

PubMed Central

Williams, Corinne L.; Haines, Robert; Cotter, Peggy A.

2008-01-01

We describe the serendipitous discovery of BatB, a classical-type Bordetella autotransporter (AT) protein with an ∼180-kDa passenger domain that remains noncovalently associated with the outer membrane. Like genes encoding all characterized protein virulence factors in Bordetella species, batB transcription is positively regulated by the master virulence regulatory system BvgAS. BatB is predicted to share similarity with immunoglobulin A (IgA) proteases, and we showed that BatB binds Ig in vitro. In vivo, a Bordetella bronchiseptica ΔbatB mutant was unable to overcome innate immune defenses and was cleared from the lower respiratory tracts of mice more rapidly than wild-type B. bronchiseptica. This defect was abrogated in SCID mice, suggesting that BatB functions to resist clearance during the first week postinoculation in a manner dependent on B- and T-cell-mediated activities. Taken together with the previous demonstration that polymorphonuclear neutrophils (PMN) are critical for the control of B. bronchiseptica in mice, our data support the hypothesis that BatB prevents nonspecific antibodies from facilitating PMN-mediated clearance during the first few days postinoculation. Neither of the strictly human-adapted Bordetella subspecies produces a fully functional BatB protein; nucleotide differences within the putative promoter region prevent batB transcription in Bordetella pertussis, and although expressed, the batB gene of human-derived Bordetella parapertussis (B. parapertussishu) contains a large in-frame deletion relative to batB of B. bronchiseptica. Taken together, our data suggest that BatB played an important role in the evolution of virulence and host specificity among the mammalian-adapted bordetellae. PMID:18426869

Mutations Abrogating VP35 Interaction with Double-Stranded RNA Render Ebola Virus Avirulent in Guinea Pigs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Prins, Kathleen C.; Delpeut, Sebastien; Leung, Daisy W.

2010-10-11

Ebola virus (EBOV) protein VP35 is a double-stranded RNA (dsRNA) binding inhibitor of host interferon (IFN)-{alpha}/{beta} responses that also functions as a viral polymerase cofactor. Recent structural studies identified key features, including a central basic patch, required for VP35 dsRNA binding activity. To address the functional significance of these VP35 structural features for EBOV replication and pathogenesis, two point mutations, K319A/R322A, that abrogate VP35 dsRNA binding activity and severely impair its suppression of IFN-{alpha}/{beta} production were identified. Solution nuclear magnetic resonance (NMR) spectroscopy and X-ray crystallography reveal minimal structural perturbations in the K319A/R322A VP35 double mutant and suggest that lossmore » of basic charge leads to altered function. Recombinant EBOVs encoding the mutant VP35 exhibit, relative to wild-type VP35 viruses, minimal growth attenuation in IFN-defective Vero cells but severe impairment in IFN-competent cells. In guinea pigs, the VP35 mutant virus revealed a complete loss of virulence. Strikingly, the VP35 mutant virus effectively immunized animals against subsequent wild-type EBOV challenge. These in vivo studies, using recombinant EBOV viruses, combined with the accompanying biochemical and structural analyses directly correlate VP35 dsRNA binding and IFN inhibition functions with viral pathogenesis. Moreover, these studies provide a framework for the development of antivirals targeting this critical EBOV virulence factor.« less

Circulating tumor cells promote the metastatic colonization of disseminated carcinoma cells by inducing systemic inflammation

PubMed Central

Luo, Chao; Shu, Yu; Luo, Jing; Qin, Jian; Wang, Yu; Li, Dong; Wang, Shan-Shan; Chi, Gang; Guo, Fang; Zhang, Gui-Mei; Feng, Zuo-Hua

2017-01-01

Circulating tumor cells (CTCs) have been studied well in the prognosis for malignant diseases as liquid biopsy, but their contribution to tumor metastasis is not clearly defined. Here we report that CTCs could promote the metastatic colonization of disseminated carcinoma cells by inducing systemic inflammation and neutrophil recruitment to pre-metastatic organs. Depletion of neutrophils in vivo could effectively abrogate the promoting effect of CTCs on tumor cell metastasis. In the presence of CTCs, the pro-tumor function of neutrophils was augmented, whereas the antitumor function of neutrophils was suppressed. Mechanically, CTC-derived ligands for TLR2 and TLR4 (TLR2/4) induced the systemic inflammation, thus increasing the production of proinflammatory cytokines such as G-CSF and IL-6 that could induce the conversion of neutrophil function from tumor-suppressing to tumor-promoting. Moreover, CTCs induced the production of endogenous TLR2/4 ligands such as S100A8, S100A9, and SAA3, which may amplify the stimulating effect that induces the expression of proinflammatory cytokines. The promoting effect of CTCs on tumor cell metastasis could be abrogated by suppressing inflammatory response with IL-37, an anti-inflammatory cytokine, or blocking CTC-derived ligands for TLR2/4. Identification of the metastatic axis of CTCs/systemic inflammation/neutrophils may provide potential targets for preventing tumor cell metastasis. PMID:28415700

SIRT6 stabilizes DNA-dependent Protein Kinase at chromatin for DNA double-strand break repair

PubMed Central

McCord, Ronald A.; Michishita, Eriko; Hong, Tao; Berber, Elisabeth; Boxer, Lisa D.; Kusumoto, Rika; Guan, Shenheng; Shi, Xiaobing; Gozani, Or; Burlingame, Alma L.; Bohr, Vilhelm A.; Chua, Katrin F.

2009-01-01

The Sir2 chromatin regulatory factor links maintenance of genomic stability to life span extension in yeast. The mammalian Sir2 family member SIRT6 has been proposed to have analogous functions, because SIRT6-deficiency leads to shortened life span and an aging-like degenerative phenotype in mice, and SIRT6 knockout cells exhibit genomic instability and DNA damage hypersensitivity. However, the molecular mechanisms underlying these defects are not fully understood. Here, we show that SIRT6 forms a macromolecular complex with the DNA double-strand break (DSB) repair factor DNA-PK (DNA-dependent protein kinase) and promotes DNA DSB repair. In response to DSBs, SIRT6 associates dynamically with chromatin and is necessary for an acute decrease in global cellular acetylation levels on histone H3 Lysine 9. Moreover, SIRT6 is required for mobilization of the DNA-PK catalytic subunit (DNA-PKcs) to chromatin in response to DNA damage and stabilizes DNA-PKcs at chromatin adjacent to an induced site-specific DSB. Abrogation of these SIRT6 activities leads to impaired resolution of DSBs. Together, these findings elucidate a mechanism whereby regulation of dynamic interaction of a DNA repair factor with chromatin impacts on the efficiency of repair, and establish a link between chromatin regulation, DNA repair, and a mammalian Sir2 factor. PMID:20157594

Notch and PKC are involved in formation of the lateral region of the dorso-ventral axis in Drosophila embryos.

PubMed

Tremmel, Daniel M; Resad, Sedat; Little, Christopher J; Wesley, Cedric S

2013-01-01

The Notch gene encodes an evolutionarily conserved cell surface receptor that generates regulatory signals based on interactions between neighboring cells. In Drosophila embryos it is normally expressed at a low level due to strong negative regulation. When this negative regulation is abrogated neurogenesis in the ventral region is suppressed, the development of lateral epidermis is severely disrupted, and the dorsal aminoserosa is expanded. Of these phenotypes only the anti-neurogenic phenotype could be linked to excess canonical Notch signaling. The other phenotypes were linked to high levels of Notch protein expression at the surface of cells in the lateral regions indicating that a non-canonical Notch signaling activity normally functions in these regions. Results of our studies reported here provide evidence. They show that Notch activities are inextricably linked to that of Pkc98E, the homolog of mammalian PKCδ. Notch and Pkc98E up-regulate the levels of the phosphorylated form of IκBCactus, a negative regulator of Toll signaling, and Mothers against dpp (MAD), an effector of Dpp signaling. Our data suggest that in the lateral regions of the Drosophila embryos Notch activity, in conjunction with Pkc98E activity, is used to form the slopes of the opposing gradients of Toll and Dpp signaling that specify cell fates along the dorso-ventral axis.

Notch and PKC Are Involved in Formation of the Lateral Region of the Dorso-Ventral Axis in Drosophila Embryos

PubMed Central

Tremmel, Daniel M.; Resad, Sedat; Little, Christopher J.; Wesley, Cedric S.

2013-01-01

The Notch gene encodes an evolutionarily conserved cell surface receptor that generates regulatory signals based on interactions between neighboring cells. In Drosophila embryos it is normally expressed at a low level due to strong negative regulation. When this negative regulation is abrogated neurogenesis in the ventral region is suppressed, the development of lateral epidermis is severely disrupted, and the dorsal aminoserosa is expanded. Of these phenotypes only the anti-neurogenic phenotype could be linked to excess canonical Notch signaling. The other phenotypes were linked to high levels of Notch protein expression at the surface of cells in the lateral regions indicating that a non-canonical Notch signaling activity normally functions in these regions. Results of our studies reported here provide evidence. They show that Notch activities are inextricably linked to that of Pkc98E, the homolog of mammalian PKCδ. Notch and Pkc98E up-regulate the levels of the phosphorylated form of IκBCactus, a negative regulator of Toll signaling, and Mothers against dpp (MAD), an effector of Dpp signaling. Our data suggest that in the lateral regions of the Drosophila embryos Notch activity, in conjunction with Pkc98E activity, is used to form the slopes of the opposing gradients of Toll and Dpp signaling that specify cell fates along the dorso-ventral axis. PMID:23861806

Interleukin-6 increases matrix metalloproteinase-14 (MMP-14) levels via down-regulation of p53 to drive cancer progression.

PubMed

Cathcart, Jillian M; Banach, Anna; Liu, Alice; Chen, Jun; Goligorsky, Michael; Cao, Jian

2016-09-20

Matrix metalloproteinases (MMPs) play critical roles in cancer invasion and metastasis by digesting basement membrane and extracellular matrix (ECM). Much attention has focused on the enzymatic activities of MMPs; however, the regulatory mechanism of MMP expression remains elusive. By employing bioinformatics analysis, we identified a potential p53 response element within the MMP-14 promoter. Experimentally, we found that p53 can repress MMP-14 promoter activity, whereas deletion of this p53 response element abrogated this effect. Furthermore, we found that p53 expression decreases MMP-14 mRNA and protein levels and attenuates MMP-14-mediated cellular functions. Additional promoter analysis and chromatin immunoprecipitation studies identified a mechanism of regulation of MMP-14 expression by which p53 and transcription factor Sp1 competitively bind to the promoter. As the correlation between inflammation and cancer aggressiveness is well described, we next sought to evaluate if inflammatory cytokines could differentially affect p53 and MMP-14 levels. We demonstrate that interleukin-6 (IL-6) down-regulates p53 protein levels and thus results in a concomitant increase in MMP-14 expression, leading to enhanced cancer cell invasion and metastasis. Our data collectively indicate a novel mechanism of regulation of MMP-14 by a cascade of IL-6 and p53, demonstrating that the tumor microenvironment directly stimulates molecular changes in cancer cells to drive an invasive phenotype.

Mutual independence of alkaline‐ and calcium‐mediated signalling in Aspergillus fumigatus refutes the existence of a conserved druggable signalling nexus

PubMed Central

Loss, Omar; Bertuzzi, Margherita; Yan, Yu; Fedorova, Natalie; McCann, Bethany L.; Armstrong‐James, Darius; Espeso, Eduardo A.; Read, Nick D.; Nierman, William C.

2017-01-01

Summary Functional coupling of calcium‐ and alkaline responsive signalling occurs in multiple fungi to afford efficient cation homeostasis. Host microenvironments exert alkaline stress and potentially toxic concentrations of Ca2+, such that highly conserved regulators of both calcium‐ (Crz) and pH‐ (PacC/Rim101) responsive signalling are crucial for fungal pathogenicity. Drugs targeting calcineurin are potent antifungal agents but also perturb human immunity thereby negating their use as anti‐infectives, abrogation of alkaline signalling has, therefore, been postulated as an adjunctive antifungal strategy. We examined the interdependency of pH‐ and calcium‐mediated signalling in Aspergillus fumigatus and found that calcium chelation severely impedes hyphal growth indicating a critical requirement for this ion independently of ambient pH. Transcriptomic responses to alkaline pH or calcium excess exhibited minimal similarity. Mutants lacking calcineurin, or its client CrzA, displayed normal alkaline tolerance and nuclear translocation of CrzA was unaffected by ambient pH. Expression of a highly conserved, alkaline‐regulated, sodium ATPase was tolerant of genetic or chemical perturbations of calcium‐mediated signalling, but abolished in null mutants of the pH‐responsive transcription factor PacC, and PacC proteolytic processing occurred normally during calcium excess. Taken together our data demonstrate that in A. fumigatus the regulatory hierarchy governing alkaline tolerance circumvents calcineurin signalling. PMID:28922497

Tyrosine kinase oncogenes abrogate interleukin-3 dependence of murine myeloid cells through signaling pathways involving c-myc: conditional regulation of c-myc transcription by temperature-sensitive v-abl.

PubMed Central

Cleveland, J L; Dean, M; Rosenberg, N; Wang, J Y; Rapp, U R

1989-01-01

Retroviral expression vectors carrying the tyrosine kinase oncogenes abl, fms, src, and trk abrogate the requirements of murine myeloid FDC-P1 cells for interleukin-3 (IL-3). Factor-independent clones constitutively express c-myc in the absence of IL-3, whereas in parental cultures c-myc transcription requires the presence of the ligand. To directly test the effect of a tyrosine kinase oncogene on c-myc expression, retroviral constructs containing three different temperature-sensitive mutants of v-abl were introduced into myeloid IL-3-dependent FDC-P1 and 32D cells. At the permissive temperature, clones expressing temperature-sensitive abl behaved like wild-type abl-containing cells in their growth properties and expressed c-myc constitutively. Temperature shift experiments demonstrated that both IL-3 abrogation and the regulation of c-myc expression correlated with the presence of functional v-abl. Induction of c-myc expression by reactivation of temperature-sensitive v-abl mimicked c-myc induction by IL-3 in that it did not require protein synthesis and occurred at the level of transcription, with effects on both initiation and a transcription elongation block. However, v-abl-regulated FDC-P1 cell growth differed from IL-3-regulated growth in that c-fos and junB, which are normally induced by IL-3, were not induced by activation of v-abl. Images PMID:2555703

The role of a cell surface inhibitor in early signal transduction associated with the regulation of cell division and differentiation

NASA Technical Reports Server (NTRS)

Johnson, T. C.; Enebo, D. J.; Moos, P. J.; Fattaey, H. K.; Spooner, B. S. (Principal Investigator)

1992-01-01

Serum stimulation of quiescent human fibroblast cultures resulted in a hyperphosphorylation of the nuclear retinoblastoma gene susceptibility product (RB). However, serum stimulation in the presence of 9 x 10(-8) M of a purified bovine sialoglycopeptide (SGP) cell surface inhibitor abrogated the hyperphosphorylation of the RB protein and the subsequent progression of cells through the mitotic cycle. The experimental results suggest that the SGP mediated its cell cycle arrest at a site in the cell cycle that was at the time of RB phosphorylation or somewhat upstream of the modification of this regulatory protein of cell division. Both cells serum-deprived and serum stimulated in the presence of the SGP displayed only a hypophosphorylated RB protein, consistent with the SGP-mediated cell cycle arrest point being near the G1/S interface.

Janus-faced Acrolein prevents allergy but accelerates tumor growth by promoting immunoregulatory Foxp3+ cells: Mouse model for passive respiratory exposure.

PubMed

Roth-Walter, Franziska; Bergmayr, Cornelia; Meitz, Sarah; Buchleitner, Stefan; Stremnitzer, Caroline; Fazekas, Judit; Moskovskich, Anna; Müller, Mario A; Roth, Georg A; Manzano-Szalai, Krisztina; Dvorak, Zdenek; Neunkirchner, Alina; Jensen-Jarolim, Erika

2017-03-23

Acrolein, a highly reactive unsaturated aldehyde, is generated in large amounts during smoking and is best known for its genotoxic capacity. Here, we aimed to assess whether acrolein at concentrations relevant for smokers may also exert immunomodulatory effects that could be relevant in allergy or cancer. In a BALB/c allergy model repeated nasal exposure to acrolein abrogated allergen-specific antibody and cytokine formation, and led to a relative accumulation of regulatory T cells in the lungs. Only the acrolein-treated mice were protected from bronchial hyperreactivity as well as from anaphylactic reactions upon challenge with the specific allergen. Moreover, grafted D2F2 tumor cells grew faster and intratumoral Foxp3+ cell accumulation was observed in these mice compared to sham-treated controls. Results from reporter cell lines suggested that acrolein acts via the aryl-hydrocarbon receptor which could be inhibited by resveratrol and 3'-methoxy-4'-nitroflavone Acrolein- stimulation of human PBMCs increased Foxp3+ expression by T cells which could be antagonized by resveratrol. Our mouse and human data thus revealed that acrolein exerts systemic immunosuppression by promoting Foxp3+ regulatory cells. This provides a novel explanation why smokers have a lower allergy, but higher cancer risk.

An acidic polysaccharide of Panax ginseng ameliorates experimental autoimmune encephalomyelitis and induces regulatory T cells.

PubMed

Hwang, Insun; Ahn, Ginnae; Park, Eunjin; Ha, Danbee; Song, Jie-Young; Jee, Youngheun

2011-08-30

An acidic polysaccharide of Panax ginseng (APG), so called ginsan, is a purified polysaccharide. APG has multiple immunomodulatory effects of stimulating natural killer (NK) and T cells and producing a variety of cytokines that proved to diminish the proinflammatory response, and protect from septic lethality. To determine APG's role in the autoimmune demyelinating disease, we tested whether APG can regulate inflammatory and encephalitogenic response in experimental autoimmune encephalomyelitis (EAE), an animal model of human multiple sclerosis (MS). Here, we demonstrate the therapeutic efficacy of the APG which induces the suppression of an encephalitogenic response during EAE. APG significantly ameliorates the progression of EAE by inhibiting the proliferation of autoreactive T cells and the production of inflammatory cytokines such as IFN-γ, IL-1β and IL-17. More importantly, APG promotes the generation of immunosuppressive regulatory T cells (Tregs) through the activation of transcription factor, Foxp3. Furthermore, the depletion of CD25+ cells from APG-treated EAE mice abrogates the beneficial effects of EAE. The capacity of APG to induce clinically beneficial effects furthers our understanding of the basis for its therapeutic immunosuppression of EAE and, possibly, MS. Thus, our results suggest that APG may serve as an effective therapy for MS and other autoimmune diseases. Copyright © 2011 Elsevier B.V. All rights reserved.

Toward an Orofacial Gene Regulatory Network

PubMed Central

Kousa, Youssef A.; Schutte, Brian C.

2015-01-01

Orofacial clefting is a common birth defect with significant morbidity. A panoply of candidate genes have been discovered through synergy of animal models and human genetics. Among these, variants in Interferon Regulatory Factor 6 (IRF6) cause syndromic orofacial clefting and contribute risk toward isolated cleft lip and palate (1/700 live births). Rare variants in IRF6 can lead to Van der Woude Syndrome (1/35,000 live births) and Popliteal Pterygium Syndrome (1/300,000 live births). Furthermore, IRF6 regulates GRHL3 and rare variants in this downstream target can also lead to Van der Woude Syndrome. In addition, a common variant (rs642961) in the IRF6 locus is found in 30% of the world’s population and contributes risk for isolated orofacial clefting. Biochemical studies revealed that rs642961 abrogates one of four AP-2alpha binding sites. Like IRF6 and GRHL3, rare variants in TFAP2A can also lead to syndromic orofacial clefting with lip pits (Branchio-oculo-facial Syndrome). The literature suggests that AP-2alpha, IRF6 and GRHL3 are part of a pathway that is essential for lip and palate development. In addition to updating the pathways, players and pursuits, this review will highlight some of the current questions in the study of orofacial clefting. PMID:26332872

Hepatitis B Virus Lacks Immune Activating Capacity, but Actively Inhibits Plasmacytoid Dendritic Cell Function

PubMed Central

Woltman, Andrea M.; Shi, Cui C.; Janssen, Harry L. A.

2011-01-01

Chronic hepatitis B virus (HBV) infection is caused by inadequate anti-viral immunity. Activation of plasmacytoid dendritic cells (pDC) leading to IFNα production is important for effective anti-viral immunity. Hepatitis B virus (HBV) infection lacks IFNα induction in animal models and patients and chronic HBV patients display impaired IFNα production by pDC. Therefore, HBV and HBV-derived proteins were examined for their effect on human pDC in vitro. In addition, the in vitro findings were compared to the function of pDC derived from chronic HBV patients ex vivo. In contrast to other viruses, HBV did not activate pDC. Moreover, HBV and HBsAg abrogated CpG-A/TLR9-induced, but not Loxoribine/TLR7-induced, mTOR-mediated S6 phosphorylation, subsequent IRF7 phosphorylation and IFNα gene transcription. HBV/HBsAg also diminished upregulation of co-stimulatory molecules, production of TNFα, IP-10 and IL-6 and pDC-induced NK cell function, whereas TLR7-induced pDC function was hardly affected. In line, HBsAg preferentially bound to TLR9-triggered pDC demonstrating that once pDC are able to bind HBV/HBsAg, the virus exerts its immune regulatory effect. HBV not only directly interfered with pDC function, but also indirectly by interfering with monocyte-pDC interaction. Also HBeAg diminished pDC function to a certain extent, but via another unknown mechanism. Interestingly, patients with HBeAg-positive chronic hepatitis B displayed impaired CpG-induced IFNα production by pDC without significant alterations in Loxoribine-induced pDC function compared to HBeAg-negative patients and healthy controls. The lack of activation and the active inhibition of pDC by HBV may both contribute to HBV persistence. The finding that the interaction between pDC and HBV may change upon activation may aid in the identification of a scavenging receptor supporting immunosuppressive effects of HBV and also in the design of novel treatment strategies for chronic HBV. PMID:21246041

Characterization of noncoding regulatory DNA in the human genome.

PubMed

Elkon, Ran; Agami, Reuven

2017-08-08

Genetic variants associated with common diseases are usually located in noncoding parts of the human genome. Delineation of the full repertoire of functional noncoding elements, together with efficient methods for probing their biological roles, is therefore of crucial importance. Over the past decade, DNA accessibility and various epigenetic modifications have been associated with regulatory functions. Mapping these features across the genome has enabled researchers to begin to document the full complement of putative regulatory elements. High-throughput reporter assays to probe the functions of regulatory regions have also been developed but these methods separate putative regulatory elements from the chromosome so that any effects of chromatin context and long-range regulatory interactions are lost. Definitive assignment of function(s) to putative cis-regulatory elements requires perturbation of these elements. Genome-editing technologies are now transforming our ability to perturb regulatory elements across entire genomes. Interpretation of high-throughput genetic screens that incorporate genome editors might enable the construction of an unbiased map of functional noncoding elements in the human genome.

Transphosphorylation of Bruton's tyrosine kinase on tyrosine 551 is critical for B cell antigen receptor function.

PubMed

Kurosaki, T; Kurosaki, M

1997-06-20

Bruton's tyrosine kinase (Btk) is required for B cell development and B cell antigen receptor (BCR) function. Cross-linking of BCR induces phosphorylation of Btk at Tyr551 and Tyr223. However, the functional requirement of these phosphorylation for BCR signaling remains unclear. We demonstrate here that mutation of Tyr551, not Tyr223, abrogates the BCR-induced calcium mobilization. Not only Lyn, but also Syk was required for tyrosine phosphorylation of Btk in BCR signaling. These results suggest that transphosphorylation of Btk on Tyr551 is essential for BCR function and that this phosphorylation is mediated through the concerted actions of Lyn and Syk.

Hepatocyte-derived exosomes promote T follicular regulatory cell expansion during hepatitis C virus infection.

PubMed

Cobb, Dustin A; Kim, Ok-Kyung; Golden-Mason, Lucy; Rosen, Hugo R; Hahn, Young S

2018-01-01

Hepatitis C virus (HCV) is a global health concern that can cause severe liver disease, such as cirrhosis and hepatocellular carcinoma. Control of HCV requires vigorous T-cell responses, yet CD4 + T cells in chronic HCV patients are dysfunctional. T follicular regulatory (Tfr) cells are a subset of regulatory T cells that suppress T follicular helper (Tfh) cells and the generation of high affinity antibody-producing B cells. In this study, we examined the accumulation of Tfr cells in the liver compartment during chronic HCV infection and defined the cellular and molecular mechanisms underlying their expansion. Our analysis revealed a substantial population of Tfr cells in livers of chronic HCV patients that is absent in liver tissues from nonviral hepatitis or healthy subjects. Coculture of PBMCs from healthy subjects with HCV-infected hepatoma cells resulted in preferential expansion of circulating Tfr cells, leading to suppression of Tfh cells. Additionally, coculture of tonsillar cells with infected hepatoma cells lead to an expansion of germinal center Tfr. Notably, expansion was mediated by transforming growth factor beta (TGF-β)-containing exosomes released from HCV-infected hepatocytes given that blockade of exosome-associated TGF-β or inhibition of exosome release abrogated Tfr expansion. These results show that liver-derived exosomes play a pivotal role in the accumulation of Tfr cells, likely leading to suppression of Tfh responses in HCV-infected patients. Our study identifies a novel pathway in which HCV infection in hepatocytes exacerbates Tfr cell responses to subvert antiviral immunity. (Hepatology 2018;67:71-85). © 2017 by the American Association for the Study of Liver Diseases.

Oligonucleotide-directed mutagenesis screen to identify pathogenic Lynch syndrome-associated MSH2 DNA mismatch repair gene variants

PubMed Central

Houlleberghs, Hellen; Dekker, Marleen; Lantermans, Hildo; Kleinendorst, Roos; Dubbink, Hendrikus Jan; Hofstra, Robert M. W.; Verhoef, Senno; te Riele, Hein

2016-01-01

Single-stranded DNA oligonucleotides can achieve targeted base-pair substitution with modest efficiency but high precision. We show that “oligo targeting” can be used effectively to study missense mutations in DNA mismatch repair (MMR) genes. Inherited inactivating mutations in DNA MMR genes are causative for the cancer predisposition Lynch syndrome (LS). Although overtly deleterious mutations in MMR genes can clearly be ascribed as the cause of LS, the functional implications of missense mutations are often unclear. We developed a genetic screen to determine the pathogenicity of these variants of uncertain significance (VUS), focusing on mutator S homolog 2 (MSH2). VUS were introduced into the endogenous Msh2 gene of mouse embryonic stem cells by oligo targeting. Subsequent selection for MMR-deficient cells using the guanine analog 6-thioguanine allowed the detection of MMR-abrogating VUS. The screen was able to distinguish weak and strong pathogenic variants from polymorphisms and was used to investigate 59 Msh2 VUS. Nineteen of the 59 VUS were identified as pathogenic. Functional assays revealed that 14 of the 19 detected variants fully abrogated MMR activity and that five of the detected variants attenuated MMR activity. Implementation of the screen in clinical practice allows proper counseling of mutation carriers and treatment of their tumors. PMID:26951660

Ergothioneine Biosynthesis and Functionality in the Opportunistic Fungal Pathogen, Aspergillus fumigatus.

PubMed

Sheridan, Kevin J; Lechner, Beatrix Elisabeth; Keeffe, Grainne O'; Keller, Markus A; Werner, Ernst R; Lindner, Herbert; Jones, Gary W; Haas, Hubertus; Doyle, Sean

2016-10-17

Ergothioneine (EGT; 2-mercaptohistidine trimethylbetaine) is a trimethylated and sulphurised histidine derivative which exhibits antioxidant properties. Here we report that deletion of Aspergillus fumigatus egtA (AFUA_2G15650), which encodes a trimodular enzyme, abrogated EGT biosynthesis in this opportunistic pathogen. EGT biosynthetic deficiency in A. fumigatus significantly reduced resistance to elevated H 2 O 2 and menadione, respectively, impaired gliotoxin production and resulted in attenuated conidiation. Quantitative proteomic analysis revealed substantial proteomic remodelling in ΔegtA compared to wild-type under both basal and ROS conditions, whereby the abundance of 290 proteins was altered. Specifically, the reciprocal differential abundance of cystathionine γ-synthase and β-lyase, respectively, influenced cystathionine availability to effect EGT biosynthesis. A combined deficiency in EGT biosynthesis and the oxidative stress response regulator Yap1, which led to extreme oxidative stress susceptibility, decreased resistance to heavy metals and production of the extracellular siderophore triacetylfusarinine C and increased accumulation of the intracellular siderophore ferricrocin. EGT dissipated H 2 O 2 in vitro, and elevated intracellular GSH levels accompanied abrogation of EGT biosynthesis. EGT deficiency only decreased resistance to high H 2 O 2 levels which suggests functionality as an auxiliary antioxidant, required for growth at elevated oxidative stress conditions. Combined, these data reveal new interactions between cellular redox homeostasis, secondary metabolism and metal ion homeostasis.

Functional Evolution of a cis-Regulatory Module

PubMed Central

Palsson, Arnar; Alekseeva, Elena; Bergman, Casey M; Nathan, Janaki; Kreitman, Martin

2005-01-01

Lack of knowledge about how regulatory regions evolve in relation to their structure–function may limit the utility of comparative sequence analysis in deciphering cis-regulatory sequences. To address this we applied reverse genetics to carry out a functional genetic complementation analysis of a eukaryotic cis-regulatory module—the even-skipped stripe 2 enhancer—from four Drosophila species. The evolution of this enhancer is non-clock-like, with important functional differences between closely related species and functional convergence between distantly related species. Functional divergence is attributable to differences in activation levels rather than spatiotemporal control of gene expression. Our findings have implications for understanding enhancer structure–function, mechanisms of speciation and computational identification of regulatory modules. PMID:15757364

MIR506 induces autophagy-related cell death in pancreatic cancer cells by targeting the STAT3 pathway.

PubMed

Sun, Longhao; Hu, Limei; Cogdell, David; Lu, Li; Gao, Chao; Tian, Weijun; Zhang, Zhixiang; Kang, Ya'an; Fleming, Jason B; Zhang, Wei

2017-04-03

Pancreatic ductal adenocarcinoma (PDAC) is the most aggressive and lethal cancer. The role of autophagy in the pathobiology of PDAC is intricate, with opposing functions manifested in different cellular contexts. MIR506 functions as a tumor suppressor in many cancer types through the regulation of multiple pathways. In this study, we hypothesized that MIR506 exerted a tumor suppression function in PDAC by inducing autophagy-related cell death. Our results provided evidence that downregulation of MIR506 expression was associated with disease progression in human PDAC. MIR506 triggered autophagic flux in PDAC cells, which led to autophagy-related cell death through direct targeting of the STAT3 (signal transducer and activator of transcription 3)-BCL2-BECN1 axis. Silencing and inhibiting STAT3 recapitulated the effects of MIR506, whereas forced expression of STAT3 abrogated the effects of MIR506. We propose that the apoptosis-inhibitory protein BCL2, which also inhibits induction of autophagy by blocking BECN1, was inhibited by MIR506 through targeting STAT3, thus augmenting BECN1 and promoting autophagy-related cell death. Silencing BECN1 and overexpression of BCL2 abrogated the effects of MIR506. These findings expand the known mechanisms of MIR506-mediated tumor suppression to activation of autophagy-related cell death and suggest a strategy for using MIR506 as an anti-STAT3 approach to PDAC treatment.

Andrographolide derivatives inhibit guanine nucleotide exchange and abrogate oncogenic Ras function

PubMed Central

Hocker, Harrison J.; Cho, Kwang-Jin; Chen, Chung-Ying K.; Rambahal, Nandini; Sagineedu, Sreenivasa Rao; Shaari, Khozirah; Stanslas, Johnson; Hancock, John F.; Gorfe, Alemayehu A.

2013-01-01

Aberrant signaling by oncogenic mutant rat sarcoma (Ras) proteins occurs in ∼15% of all human tumors, yet direct inhibition of Ras by small molecules has remained elusive. Recently, several small-molecule ligands have been discovered that directly bind Ras and inhibit its function by interfering with exchange factor binding. However, it is unclear whether, or how, these ligands could lead to drugs that act against constitutively active oncogenic mutant Ras. Using a dynamics-based pocket identification scheme, ensemble docking, and innovative cell-based assays, here we show that andrographolide (AGP)—a bicyclic diterpenoid lactone isolated from Andrographis paniculata—and its benzylidene derivatives bind to transient pockets on Kirsten-Ras (K-Ras) and inhibit GDP–GTP exchange. As expected for inhibitors of exchange factor binding, AGP derivatives reduced GTP loading of wild-type K-Ras in response to acute EGF stimulation with a concomitant reduction in MAPK activation. Remarkably, however, prolonged treatment with AGP derivatives also reduced GTP loading of, and signal transmission by, oncogenic mutant K-RasG12V. In sum, the combined analysis of our computational and cell biology results show that AGP derivatives directly bind Ras, block GDP–GTP exchange, and inhibit both wild-type and oncogenic K-Ras signaling. Importantly, our findings not only show that nucleotide exchange factors are required for oncogenic Ras signaling but also demonstrate that inhibiting nucleotide exchange is a valid approach to abrogating the function of oncogenic mutant Ras. PMID:23737504

In vitro effects of 4-hydroperoxycyclophosphamide on human immunoregulatory T subset function. I. Selective effects on lymphocyte function in T-B cell collaboration.

PubMed

Ozer, H; Cowens, J W; Colvin, M; Nussbaum-Blumenson, A; Sheedy, D

1982-01-01

The alkylating agent cyclophosphamide may suppress or enhance immune responses in vivo but is inactive in vitro unless metabolized by microsomal enzyme activation. 4-hydroperoxycyclophosphamide (4-HC) is a synthetic compound that is spontaneously converted in aqueous solution to the active metabolites. In this report, we examined the in vitro sensitivity of functional human T cell subsets to 4-HC in a polyclonal B cell differentiation assay and in the generation of mitogen-induced suppressor cells for effector B cell function. Con A-induced T suppression of B cell differentiation is completely abrogated by a 1-h pretreatment of T cells at very low concentrations of between 10(-2) and 20 nmol/ml, whereas inducer T cell function is sensitive only to concentrations in greater than 40 nmol/ml. The effects of 4-HC on suppressor T cells appear to occur at concentrations that do not result in DNA cross-linking or decreased blastogenesis. Con A-induced T suppressors are generated from within the OKT4+, OKT8- subset and are sensitive to low-dose 4-HC only before activation, whereas differentiated suppressor cells are resistant to concentrations in greater than 80 nmol/ml. Low-dose 4-HC pretreatment of the B cell population results in abrogation of immunoglobulin secretion when treated B cells are cocultured with unfractionated T cells, however, this effect is completely reversible if pretreated B cells are cocultured with T cells devoid of suppressor activity. These results demonstrate that human presuppressor cells for B-effector function differentiate in response to Con A from the OKT4+, OKT8- subset and are exquisitely sensitive to low concentrations of CYP whereas mature suppressor and inducer functions are resistant to all but very high concentrations in vitro. The differential sensitivity of functional T and B cell subsets to 4-HC in vitro can be a very useful probe in dissecting immunoregulatory interactions with man.

Characterisation and expression of a PP1 serine/threonine protein phosphatase (PfPP1) from the malaria parasite, Plasmodium falciparum: demonstration of its essential role using RNA interference

PubMed Central

Kumar, Rajinder; Adams, Brian; Oldenburg, Anja; Musiyenko, Alla; Barik, Sailen

2002-01-01

Background Reversible protein phosphorylation is relatively unexplored in the intracellular protozoa of the Apicomplexa family that includes the genus Plasmodium, to which belong the causative agents of malaria. Members of the PP1 family represent the most highly conserved protein phosphatase sequences in phylogeny and play essential regulatory roles in various cellular pathways. Previous evidence suggested a PP1-like activity in Plasmodium falciparum, not yet identified at the molecular level. Results We have identified a PP1 catalytic subunit from P. falciparum and named it PfPP1. The predicted primary structure of the 304-amino acid long protein was highly similar to PP1 sequences of other species, and showed conservation of all the signature motifs. The purified recombinant protein exhibited potent phosphatase activity in vitro. Its sensitivity to specific phosphatase inhibitors was characteristic of the PP1 class. The authenticity of the PfPP1 cDNA was further confirmed by mutational analysis of strategic amino acid residues important in catalysis. The protein was expressed in all erythrocytic stages of the parasite. Abrogation of PP1 expression by synthetic short interfering RNA (siRNA) led to inhibition of parasite DNA synthesis. Conclusions The high sequence similarity of PfPP1 with other PP1 members suggests conservation of function. Phenotypic gene knockdown studies using siRNA confirmed its essential role in the parasite. Detailed studies of PfPP1 and its regulation may unravel the role of reversible protein phosphorylation in the signalling pathways of the parasite, including glucose metabolism and parasitic cell division. The use of siRNA could be an important tool in the functional analysis of Apicomplexan genes. PMID:12057017

Arginine methylation of translocated in liposarcoma (TLS) inhibits its binding to long noncoding RNA, abrogating TLS-mediated repression of CBP/p300 activity.

PubMed

Cui, Wei; Yoneda, Ryoma; Ueda, Naomi; Kurokawa, Riki

2018-05-21

Translocated in liposarcoma (TLS) is an RNA-binding protein and a transcription-regulatory sensor of DNA damage. TLS binds promoter-associated noncoding RNA (pncRNA) and inhibits histone acetyltransferase (HAT) activity of CREB-binding protein (CBP)/E1A-binding protein P300 (p300) on the cyclin D1 (CCND1) gene. Although post-translational modifications of TLS, such as arginine methylation, are known to regulate TLS's nucleocytoplasmic shuttling and assembly in stress granules, its interactions with RNAs remain poorly characterized. Herein, using various biochemical assays, we confirmed the earlier observations that TLS is methylated by protein arginine methyltransferase 1 (PRMT1) in vitro. The arginine methylation of TLS disrupted binding to pncRNA and also prevented binding of TLS to and inhibition of CBP/p300. This result indicated that arginine methylation of TLS abrogates both binding to pncRNA and TLS-mediated inhibition of CBP/p300 HAT activities. We also report that an arginine residue within the Arg-Gly-Gly domain of TLS, Arg-476, serves as the major determinant for binding to pncRNA. Either methylation or mutation of Arg-476 of TLS significantly decreased pncRNA binding and thereby prevented a pncRNA-induced allosteric alteration in TLS that is required for its interaction with CBP/p300. Moreover, unlike wildtype TLS, an R476A TLS mutant did not inhibit CCND1 promoter activity in luciferase reporter assays. Taken together, we propose the hypothesis that arginine methylation of TLS regulates both TLS-nucleic acid and TLS-protein interactions and thereby participates in transcriptional regulation. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

The CCR4-NOT complex mediates deadenylation and degradation of stem cell mRNAs and promotes planarian stem cell differentiation.

PubMed

Solana, Jordi; Gamberi, Chiara; Mihaylova, Yuliana; Grosswendt, Stefanie; Chen, Chen; Lasko, Paul; Rajewsky, Nikolaus; Aboobaker, A Aziz

2013-01-01

Post-transcriptional regulatory mechanisms are of fundamental importance to form robust genetic networks, but their roles in stem cell pluripotency remain poorly understood. Here, we use freshwater planarians as a model system to investigate this and uncover a role for CCR4-NOT mediated deadenylation of mRNAs in stem cell differentiation. Planarian adult stem cells, the so-called neoblasts, drive the almost unlimited regenerative capabilities of planarians and allow their ongoing homeostatic tissue turnover. While many genes have been demonstrated to be required for these processes, currently almost no mechanistic insight is available into their regulation. We show that knockdown of planarian Not1, the CCR4-NOT deadenylating complex scaffolding subunit, abrogates regeneration and normal homeostasis. This abrogation is primarily due to severe impairment of their differentiation potential. We describe a stem cell specific increase in the mRNA levels of key neoblast genes after Smed-not1 knock down, consistent with a role of the CCR4-NOT complex in degradation of neoblast mRNAs upon the onset of differentiation. We also observe a stem cell specific increase in the frequency of longer poly(A) tails in these same mRNAs, showing that stem cells after Smed-not1 knock down fail to differentiate as they accumulate populations of transcripts with longer poly(A) tails. As other transcripts are unaffected our data hint at a targeted regulation of these key stem cell mRNAs by post-transcriptional regulators such as RNA-binding proteins or microRNAs. Together, our results show that the CCR4-NOT complex is crucial for stem cell differentiation and controls stem cell-specific degradation of mRNAs, thus providing clear mechanistic insight into this aspect of neoblast biology.

The CCR4-NOT Complex Mediates Deadenylation and Degradation of Stem Cell mRNAs and Promotes Planarian Stem Cell Differentiation

PubMed Central

Solana, Jordi; Gamberi, Chiara; Mihaylova, Yuliana; Grosswendt, Stefanie; Chen, Chen; Lasko, Paul; Rajewsky, Nikolaus; Aboobaker, A. Aziz

2013-01-01

Post-transcriptional regulatory mechanisms are of fundamental importance to form robust genetic networks, but their roles in stem cell pluripotency remain poorly understood. Here, we use freshwater planarians as a model system to investigate this and uncover a role for CCR4-NOT mediated deadenylation of mRNAs in stem cell differentiation. Planarian adult stem cells, the so-called neoblasts, drive the almost unlimited regenerative capabilities of planarians and allow their ongoing homeostatic tissue turnover. While many genes have been demonstrated to be required for these processes, currently almost no mechanistic insight is available into their regulation. We show that knockdown of planarian Not1, the CCR4-NOT deadenylating complex scaffolding subunit, abrogates regeneration and normal homeostasis. This abrogation is primarily due to severe impairment of their differentiation potential. We describe a stem cell specific increase in the mRNA levels of key neoblast genes after Smed-not1 knock down, consistent with a role of the CCR4-NOT complex in degradation of neoblast mRNAs upon the onset of differentiation. We also observe a stem cell specific increase in the frequency of longer poly(A) tails in these same mRNAs, showing that stem cells after Smed-not1 knock down fail to differentiate as they accumulate populations of transcripts with longer poly(A) tails. As other transcripts are unaffected our data hint at a targeted regulation of these key stem cell mRNAs by post-transcriptional regulators such as RNA-binding proteins or microRNAs. Together, our results show that the CCR4-NOT complex is crucial for stem cell differentiation and controls stem cell-specific degradation of mRNAs, thus providing clear mechanistic insight into this aspect of neoblast biology. PMID:24367277

Modulation of Human Allogeneic and Syngeneic Pluripotent Stem Cells and Immunological Implications for Transplantation

PubMed Central

Sackett, S.D.; Brown, M.E.; Tremmel, D.M.; Ellis, T.; Burlingham, W.J.; Odorico, J.S.

2016-01-01

Tissues derived from induced pluripotent stem cells (iPSCs) are a promising source of cells for building various regenerative medicine therapies; from simply transplanting cells to reseeding decellularized organs to reconstructing multicellular tissues. Although reprogramming strategies for producing iPSCs have improved, the clinical use of iPSCs is limited by the presence of unique human leukocyte antigen (HLA) genes, the main immunologic barrier to transplantation. In order to overcome the immunological hurdles associated with allogeneic tissues and organs, the generation of patient-histocompatible iPSCs (autologous or HLA-matched cells) provides an attractive platform for personalized medicine. However, concerns have been raised as to the fitness, safety and immunogenicity of iPSC derivatives because of variable differentiation potential of different lines and the identification of genetic and epigenetic aberrations that can occur during the reprogramming process. In addition, significant cost and regulatory barriers may deter commercialization of patient specific therapies in the short-term. Nonetheless, recent studies provide some evidence of immunological benefit for using autologous iPSCs. Yet, more studies are needed to evaluate the immunogenicity of various autologous and allogeneic human iPSC-derived cell types as well as test various methods to abrogate rejection. Here, we present perspectives of using allogeneic vs autologous iPSCs for transplantation therapies and the advantages and disadvantages of each related to differentiation potential, immunogenicity, genetic stability and tumorigenicity. We also review the current literature on the immunogenicity of syngeneic iPSCs and discuss evidence that questions the feasibility of HLA-matched iPSC banks. Finally, we will discuss emerging methods of abrogating or reducing host immune responses to PSC derivatives. PMID:26970668

Modulation of human allogeneic and syngeneic pluripotent stem cells and immunological implications for transplantation.

PubMed

Sackett, S D; Brown, M E; Tremmel, D M; Ellis, T; Burlingham, W J; Odorico, J S

2016-04-01

Tissues derived from induced pluripotent stem cells (iPSCs) are a promising source of cells for building various regenerative medicine therapies; from simply transplanting cells to reseeding decellularized organs to reconstructing multicellular tissues. Although reprogramming strategies for producing iPSCs have improved, the clinical use of iPSCs is limited by the presence of unique human leukocyte antigen (HLA) genes, the main immunologic barrier to transplantation. In order to overcome the immunological hurdles associated with allogeneic tissues and organs, the generation of patient-histocompatible iPSCs (autologous or HLA-matched cells) provides an attractive platform for personalized medicine. However, concerns have been raised as to the fitness, safety and immunogenicity of iPSC derivatives because of variable differentiation potential of different lines and the identification of genetic and epigenetic aberrations that can occur during the reprogramming process. In addition, significant cost and regulatory barriers may deter commercialization of patient specific therapies in the short-term. Nonetheless, recent studies provide some evidence of immunological benefit for using autologous iPSCs. Yet, more studies are needed to evaluate the immunogenicity of various autologous and allogeneic human iPSC-derived cell types as well as test various methods to abrogate rejection. Here, we present perspectives of using allogeneic vs. autologous iPSCs for transplantation therapies and the advantages and disadvantages of each related to differentiation potential, immunogenicity, genetic stability and tumorigenicity. We also review the current literature on the immunogenicity of syngeneic iPSCs and discuss evidence that questions the feasibility of HLA-matched iPSC banks. Finally, we will discuss emerging methods of abrogating or reducing host immune responses to PSC derivatives. Copyright © 2016 Elsevier Inc. All rights reserved.

Tregs: Where We Are and What Comes Next?

PubMed

Zhao, Hai; Liao, Xuelian; Kang, Yan

2017-01-01

Regulatory T cells are usually recognized as a specialized subset of CD4 + T cells functioning in establishment and maintenance of immune tolerance. Meanwhile, there is emerging evidence that regulatory T cells (Tregs) are also present in various non-lymphoid tissues, and that they have unique phenotypes credited with activities distinct from regulatory function. Their development and function have been described in plenty of manuscripts in the past two decades. However, with the deepening of research in recent years, emerging evidence revealed some novel mechanisms about how Tregs exert their activities. First, we discuss the expanding family of regulatory lymphocytes briefly and then, try to interpret how fork-head box P3 (Foxp3), a master regulator of the regulatory pathway in the development and function of regulatory T cells, functions. Subsequently, another part of our focus is varieties of tissue Tregs. Next, we primarily discuss recent research on how Tregs work and their faceted functions in terms of soluble mediators, functional proteins, and inhibitory receptors. In particular, unless otherwise noted, the term "Treg" is used here to refer specially to the "CD4 + CD25 + Foxp3 +" regulatory cells.

Studies of Cell-Mediated Immunity Against Immune Disorders Using Synthetic Peptides and Rotating Bioreactor System

NASA Technical Reports Server (NTRS)

Sastry, Jagannadha K.

1998-01-01

We conducted a series of experiments using mouse immune-precursor cells, and observed that bioreactor culturing results in the loss of antigen-specific cytotoxic T lymphocyte (CTL) function. The reason for the abrogation of CTL function is microgravity conditions in the bioreactor, but not the antigen per se or its MHC restriction. Similarly, we observed that allostimulation of human PBMC in the bioreactor, but not in the T flask, resulted in the blunting of both allo-CTL function and the NK activity, indicating that the microgravity-associated functional defects are not unique to the mouse system. These results provide further confirmation to the microgravity-associated immune dysfunction, and constitute ground-based confirmatory data for those related to space-travel.

miR-125/Pokemon auto-circuit contributes to the progression of hepatocellular carcinoma.

PubMed

Kong, Jing; Liu, Xiaoping; Li, Xiangqian; Wu, Jinsheng; Wu, Ning; Chen, Jun; Fang, Fang

2016-01-01

Hepatocellular carcinoma (HCC) is a type of human malignant tumor occurring in hepatic tissues with high mortality. Patients benefit little from current therapeutic modalities, at least partially due to the lack of complete elucidation of molecular network regulating HCC. miR-125 and Pokemon are well-recognized tumor suppressor and oncogenes for HCC, respectively. However, the underlying mechanism by which the two genes exert their functions and the relationship between miR-125 and Pokemon is still unexplored yet. In this study, we found that there is an inverse association between miR-125 and Pokemon expression levels in HCC specimen and cell lines. Online database mining indicated that there are three putative mRNA recognition elements (MREs) of miR-125 within 3' untranslated region (3'UTR) of Pokemon. MREs of miR-125 confer the expression of luciferase with a miR-125-dependent fashion. The alteration in miR-125 abundance regulates the expression of Pokemon at both protein and mRNA levels. Overexpression of Pokemon is able to abrogate the inhibitory effect of miR-125 on HCC progression. Further study showed that Pokemon inhibits the expression of miR-125 by binding of recognition sites within its promoter. In conclusion, we found that there is an auto-regulatory circuit consisting of miR-125 and Pokemon, which promotes the progression of HCC and may be a promising therapeutic target in clinical HCC treatment.

Advances in allergic skin disease, anaphylaxis, and hypersensitivity reactions to foods, drugs, and insects.

PubMed

Sicherer, Scott H; Leung, Donald Y M

2005-07-01

This review highlights some of the research advances in allergic skin disease, anaphylaxis, and hypersensitivity reactions to foods, drugs, and insects that were reported primarily in the Journal in 2004. Clinical observations included that gastrointestinal symptoms during anaphylaxis are associated with an increased risk for hypotension; recurrence of peanut allergy can occur for about 8% of children who pass an oral food challenge and is associated with continued avoidance of the food after the challenge; seafood allergy is reported by 2.3% of the US population; and determination of the time to resolution of childhood egg and milk allergy might be predictable by means of serial determination of food-specific IgE levels. The comorbid effects of atopic dermatitis (AD) on asthma and the role of topical calcineurin inhibitors in the therapy of AD were also addressed. Basic and translational research observations indicate that improved diagnosis and therapy might become possible on the basis of reported identification or characterization of allergens such as: lipid transfer proteins and birch pollen-related cross-reactive allergens in plant foods; proteins in scorpion venom that cross-react with proteins from imported fire ant; mosquito saliva proteins responsible for systemic anaphylaxis; and IgE binding to quinolones detectable with an in vitro immunoassay. In addition, advances in understanding immune regulation associated with abrogation of oral tolerance in food allergy and of dendritic cell function, modulation of regulatory T cells, and chemokine expression in AD have elucidated possible targets for future intervention.

Chaperone expression profiles correlate with distinct physiological states of Plasmodium falciparum in malaria patients

PubMed Central

2010-01-01

Background Molecular chaperones have been shown to be important in the growth of the malaria parasite Plasmodium falciparum and inhibition of chaperone function by pharmacological agents has been shown to abrogate parasite growth. A recent study has demonstrated that clinical isolates of the parasite have distinct physiological states, one of which resembles environmental stress response showing up-regulation of specific molecular chaperones. Methods Chaperone networks operational in the distinct physiological clusters in clinical malaria parasites were constructed using cytoscape by utilizing their clinical expression profiles. Results Molecular chaperones show distinct profiles in the previously defined physiologically distinct states. Further, expression profiles of the chaperones from different cellular compartments correlate with specific patient clusters. While cluster 1 parasites, representing a starvation response, show up-regulation of organellar chaperones, cluster 2 parasites, which resemble active growth based on glycolysis, show up-regulation of cytoplasmic chaperones. Interestingly, cytoplasmic Hsp90 and its co-chaperones, previously implicated as drug targets in malaria, cluster in the same group. Detailed analysis of chaperone expression in the patient cluster 2 reveals up-regulation of the entire Hsp90-dependent pro-survival circuitries. In addition, cluster 2 also shows up-regulation of Plasmodium export element (PEXEL)-containing Hsp40s thought to have regulatory and host remodeling roles in the infected erythrocyte. Conclusion In all, this study demonstrates an intimate involvement of parasite-encoded chaperones, PfHsp90 in particular, in defining pathogenesis of malaria. PMID:20719001

Interleukin-6 increases matrix metalloproteinase-14 (MMP-14) levels via down-regulation of p53 to drive cancer progression

PubMed Central

Cathcart, Jillian M.; Banach, Anna; Liu, Alice; Chen, Jun; Goligorsky, Michael; Cao, Jian

2016-01-01

Matrix metalloproteinases (MMPs) play critical roles in cancer invasion and metastasis by digesting basement membrane and extracellular matrix (ECM). Much attention has focused on the enzymatic activities of MMPs; however, the regulatory mechanism of MMP expression remains elusive. By employing bioinformatics analysis, we identified a potential p53 response element within the MMP-14 promoter. Experimentally, we found that p53 can repress MMP-14 promoter activity, whereas deletion of this p53 response element abrogated this effect. Furthermore, we found that p53 expression decreases MMP-14 mRNA and protein levels and attenuates MMP-14-mediated cellular functions. Additional promoter analysis and chromatin immunoprecipitation studies identified a mechanism of regulation of MMP-14 expression by which p53 and transcription factor Sp1 competitively bind to the promoter. As the correlation between inflammation and cancer aggressiveness is well described, we next sought to evaluate if inflammatory cytokines could differentially affect p53 and MMP-14 levels. We demonstrate that interleukin-6 (IL-6) down-regulates p53 protein levels and thus results in a concomitant increase in MMP-14 expression, leading to enhanced cancer cell invasion and metastasis. Our data collectively indicate a novel mechanism of regulation of MMP-14 by a cascade of IL-6 and p53, demonstrating that the tumor microenvironment directly stimulates molecular changes in cancer cells to drive an invasive phenotype. PMID:27531896

Mutual independence of alkaline- and calcium-mediated signalling in Aspergillus fumigatus refutes the existence of a conserved druggable signalling nexus.

PubMed

Loss, Omar; Bertuzzi, Margherita; Yan, Yu; Fedorova, Natalie; McCann, Bethany L; Armstrong-James, Darius; Espeso, Eduardo A; Read, Nick D; Nierman, William C; Bignell, Elaine M

2017-12-01

Functional coupling of calcium- and alkaline responsive signalling occurs in multiple fungi to afford efficient cation homeostasis. Host microenvironments exert alkaline stress and potentially toxic concentrations of Ca 2+ , such that highly conserved regulators of both calcium- (Crz) and pH- (PacC/Rim101) responsive signalling are crucial for fungal pathogenicity. Drugs targeting calcineurin are potent antifungal agents but also perturb human immunity thereby negating their use as anti-infectives, abrogation of alkaline signalling has, therefore, been postulated as an adjunctive antifungal strategy. We examined the interdependency of pH- and calcium-mediated signalling in Aspergillus fumigatus and found that calcium chelation severely impedes hyphal growth indicating a critical requirement for this ion independently of ambient pH. Transcriptomic responses to alkaline pH or calcium excess exhibited minimal similarity. Mutants lacking calcineurin, or its client CrzA, displayed normal alkaline tolerance and nuclear translocation of CrzA was unaffected by ambient pH. Expression of a highly conserved, alkaline-regulated, sodium ATPase was tolerant of genetic or chemical perturbations of calcium-mediated signalling, but abolished in null mutants of the pH-responsive transcription factor PacC, and PacC proteolytic processing occurred normally during calcium excess. Taken together our data demonstrate that in A. fumigatus the regulatory hierarchy governing alkaline tolerance circumvents calcineurin signalling. © 2017 The Authors. Molecular Microbiology Published by John Wiley & Sons Ltd.

PI3Kδ-selective and PI3Kα/δ-combinatorial inhibitors in clinical development for B-cell non-Hodgkin lymphoma.

PubMed

Lampson, Benjamin L; Brown, Jennifer R

2017-11-01

The efficacy of the prototypical phosphatidylinositol-3-kinase (PI3K) inhibitor idelalisib for the treatment of chronic lymphocytic leukemia (CLL) and indolent non-Hodgkin lymphoma (iNHL) has led to development of multiple compounds targeting this pathway. Areas Covered: We review the hypothesized therapeutic mechanisms of PI3K inhibitors, including abrogation of B cell receptor signaling, blockade of microenvironmental pro-survival signals, and enhancement of anti-tumor immunity. We examine toxicities of idelalisib, including bacterial infections (possibly secondary to drug-induced neutropenia), opportunistic infections (possibly attributable to on-target inhibition of T cell function), and organ toxicities such as transaminitis and enterocolitis (possibly autoimmune, secondary to on-target inhibition of p110δ in regulatory T cells). We evaluate PI3K inhibitors that have entered trials for the treatment of lymphoma, focusing on agents with selectivity for PI3Kα and PI3Kδ. Expert Opinion: PI3K inhibitors, particularly those that target p110δ, have robust efficacy in the treatment of CLL and iNHL. However, idelalisib has infectious and autoimmune toxicities that limit its use. Outside of trials, idelalisib should be restricted to CLL patients with progression on ibrutinib or iNHL patients with progression on two prior therapies. Whether newer PI3K inhibitors will demonstrate differentiated toxicity profiles in comparable patient populations while retaining efficacy remains to be seen.

Estrogen suppresses breast cancer proliferation through GPER / p38 MAPK axis during hypoxia.

PubMed

Sathya, S; Sudhagar, S; Lakshmi, B S

2015-12-05

Breast cancer cells frequently experience hypoxia which is associated with resistance to hormonal therapy and poor clinical prognosis, making it important to understand the function of estrogen under hypoxic condition. Here, we demonstrate that estrogen suppresses breast cancer cell growth under hypoxia, through inhibition at G1/S phase of cell cycle, by elevation of p21 expression. The involvement of GPER in estrogen mediated growth arrest was elucidated using specific ligands and siRNA. Although, estrogen was observed to activate both p44/42 and p38 MAPK signaling, pharmacological inhibition and silencing of p38 MAPK abrogated the induction of p21 expression and growth arrest, during hypoxia. The involvement of estrogen induced ROS in the p38 MAPK mediated p21 expression and cell growth arrest was established by observing that scavenging of ROS by NAC abrogated p38 MAPK activation and p21 expression during hypoxia. In conclusion, Estrogen suppresses breast cancer growth by inhibiting G1/S phase transition through GPER/ROS/p38 MAPK/p21 mediated signaling during hypoxic condition. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

TARGETED DELETION OF INDUCIBLE HEAT SHOCK PROTEIN 70 ABROGATES THE LATE INFARCT-SPARING EFFECT OF MYOCARDIAL ISCHEMIC PRECONDITIONING

EPA Science Inventory

Abstract submitted for 82nd annual meeting of the American Association for Thoracic Surgery, May 4-8, 2002 in Washington D.C.

Targeted Deletion of Inducible Heat Shock Protein 70 Abrogates the Late Infarct-Sparing Effect of Myocardial Ischemic Preconditioning

Craig...

Tregs: Where We Are and What Comes Next?

PubMed Central

Zhao, Hai; Liao, Xuelian; Kang, Yan

2017-01-01

Regulatory T cells are usually recognized as a specialized subset of CD4+ T cells functioning in establishment and maintenance of immune tolerance. Meanwhile, there is emerging evidence that regulatory T cells (Tregs) are also present in various non-lymphoid tissues, and that they have unique phenotypes credited with activities distinct from regulatory function. Their development and function have been described in plenty of manuscripts in the past two decades. However, with the deepening of research in recent years, emerging evidence revealed some novel mechanisms about how Tregs exert their activities. First, we discuss the expanding family of regulatory lymphocytes briefly and then, try to interpret how fork-head box P3 (Foxp3), a master regulator of the regulatory pathway in the development and function of regulatory T cells, functions. Subsequently, another part of our focus is varieties of tissue Tregs. Next, we primarily discuss recent research on how Tregs work and their faceted functions in terms of soluble mediators, functional proteins, and inhibitory receptors. In particular, unless otherwise noted, the term “Treg” is used here to refer specially to the “CD4+CD25+Foxp3+” regulatory cells. PMID:29225597

Trimeric Association of Hox and TALE Homeodomain Proteins Mediates Hoxb2 Hindbrain Enhancer Activity

PubMed Central

Jacobs, Yakop; Schnabel, Catherine A.; Cleary, Michael L.

1999-01-01

Pbx/exd proteins modulate the DNA binding affinities and specificities of Hox proteins and contribute to the execution of Hox-dependent developmental programs in arthropods and vertebrates. Pbx proteins also stably heterodimerize and bind DNA with Meis and Pknox1-Prep1, additional members of the TALE (three-amino-acid loop extension) superclass of homeodomain proteins that function on common genetic pathways with a subset of Hox proteins. In this study, we demonstrated that Pbx and Meis bind DNA as heterotrimeric complexes with Hoxb1 on a genetically defined Hoxb2 enhancer, r4, that mediates the cross-regulatory transcriptional effects of Hoxb1 in vivo. The DNA binding specificity of the heterotrimeric complex for r4 is mediated by a Pbx-Hox site in conjunction with a distal Meis site, which we showed to be required for ternary complex formation and Meis-enhanced transcription. Formation of heterotrimeric complexes in which all three homeodomains bind their cognate DNA sites is topologically facilitated by the ability of Pbx and Meis to interact through their amino termini and bind DNA without stringent half-site orientation and spacing requirements. Furthermore, Meis site mutation in the Hoxb2 enhancer phenocopies Pbx-Hox site mutation to abrogate enhancer-directed expression of a reporter transgene in the murine embryonic hindbrain, demonstrating that DNA binding by all three proteins is required for trimer function in vivo. Our data provide in vitro and in vivo evidence for the combinatorial regulation of Hox and TALE protein functions that are mediated, in part, by their interdependent DNA binding activities as ternary complexes. As a consequence, Hoxb1 employs Pbx and Meis-related proteins, as a pair of essential cofactors in a higher-order molecular complex, to mediate its transcriptional effects on an endogenous Hox response element. PMID:10373562

Differential effects of Denileukin Diftitox IL-2 immunotoxin on NK and regulatory T cells in non-human primates

PubMed Central

Yamada, Yohei; Aoyama, Akihiro; Tocco, Georges; Boskovic, Svjetlan; Nadazdin, Ognjenka; Alessandrini, Alessandro; Madsen, Joren C.; Cosimi, A. Benedict; Benichou, Gilles; Kawai, Tatsuo

2012-01-01

Denileukin Diftitox (DD), a fusion protein comprised of IL-2 and diphtheria toxin was initially expected to enhance anti-tumor immunity by selectively eliminating regulatory T cells (Tregs) displaying the high affinity IL-2R (α-β-γ trimers). While DD has been shown to deplete some Tregs in primates, its effects on NK cells (CD16+CD8+NKG2A+CD3−), which constitutively express the intermediate affinity IL-2R (β-γ dimers) and play a critical role in anti-tumor immunity, are still unknown. To address this question, cynomolgus monkeys were injected intravenously with two different doses of DD (8 or 18 μg/Kg). This treatment resulted in a rapid but short-term reduction in detectable peripheral blood resting Tregs (R-Tregs: CD4+CD45RA+Foxp3+) and a transient increase in the number of activated Tregs (A-Tregs: CD4+CD45RA−Foxp3high) followed by their partial depletion (50–60%). On the other hand, all NK cells were deleted immediately and durably after DD administration. This difference was not due to a higher binding or internalization of DD by NK cells as compared to Tregs. Co-administration of DD with IL-15, which binds to IL-2Rβ-γ, abrogated DD-induced NK cell deletion in vitro and in vivo while it did not affect Tregs elimination. Taken together, these results show that DD exerts a potent cytotoxic effect on NK cells, a phenomenon which might impair its anti-tumoral properties. However, co-administration of IL-15 with DD could alleviate this problem by selectively protecting potentially oncolytic NK cells while allowing the depletion of immunosuppressive regulatory T cells in cancer patients. PMID:22586034

Cross-Cohort Analysis Identifies a TEAD4-MYCN Positive Feedback Loop as the Core Regulatory Element of High-Risk Neuroblastoma.

PubMed

Rajbhandari, Presha; Lopez, Gonzalo; Capdevila, Claudia; Salvatori, Beatrice; Yu, Jiyang; Rodriguez-Barrueco, Ruth; Martinez, Daniel; Yarmarkovich, Mark; Weichert-Leahey, Nina; Abraham, Brian J; Alvarez, Mariano J; Iyer, Archana; Harenza, Jo Lynne; Oldridge, Derek; De Preter, Katleen; Koster, Jan; Asgharzadeh, Shahab; Seeger, Robert C; Wei, Jun S; Khan, Javed; Vandesompele, Jo; Mestdagh, Pieter; Versteeg, Rogier; Look, A Thomas; Young, Richard A; Iavarone, Antonio; Lasorella, Anna; Silva, Jose M; Maris, John M; Califano, Andrea

2018-05-01

High-risk neuroblastomas show a paucity of recurrent somatic mutations at diagnosis. As a result, the molecular basis for this aggressive phenotype remains elusive. Recent progress in regulatory network analysis helped us elucidate disease-driving mechanisms downstream of genomic alterations, including recurrent chromosomal alterations. Our analysis identified three molecular subtypes of high-risk neuroblastomas, consistent with chromosomal alterations, and identified subtype-specific master regulator proteins that were conserved across independent cohorts. A 10-protein transcriptional module-centered around a TEAD4-MYCN positive feedback loop-emerged as the regulatory driver of the high-risk subtype associated with MYCN amplification. Silencing of either gene collapsed MYCN -amplified ( MYCN Amp ) neuroblastoma transcriptional hallmarks and abrogated viability in vitro and in vivo Consistently, TEAD4 emerged as a robust prognostic marker of poor survival, with activity independent of the canonical Hippo pathway transcriptional coactivators YAP and TAZ. These results suggest novel therapeutic strategies for the large subset of MYCN-deregulated neuroblastomas. Significance: Despite progress in understanding of neuroblastoma genetics, little progress has been made toward personalized treatment. Here, we present a framework to determine the downstream effectors of the genetic alterations sustaining neuroblastoma subtypes, which can be easily extended to other tumor types. We show the critical effect of disrupting a 10-protein module centered around a YAP/TAZ-independent TEAD4-MYCN positive feedback loop in MYCN Amp neuroblastomas, nominating TEAD4 as a novel candidate for therapeutic intervention. Cancer Discov; 8(5); 582-99. ©2018 AACR. This article is highlighted in the In This Issue feature, p. 517 . ©2018 American Association for Cancer Research.

MiRNA-21 mediates the antiangiogenic activity of metformin through targeting PTEN and SMAD7 expression and PI3K/AKT pathway

PubMed Central

Luo, Mao; Tan, Xiaoyong; Mu, Lin; Luo, Yulin; Li, Rong; Deng, Xin; Chen, Ni; Ren, Meiping; Li, Yongjie; Wang, Liqun; Wu, Jianbo; Wan, Qin

2017-01-01

Metformin, an anti-diabetic drug commonly used for type 2 diabetes therapy, is associated with anti-angiogenic effects in conditions beyond diabetes. miR-21 has been reported to be involved in the process of angiogenesis. However, the precise regulatory mechanisms by which the metformin-induced endothelial suppression and its effects on miR-21-dependent pathways are still unclear. Bioinformatic analysis and identification of miR-21 and its targets and their effects on metformin-induced antiangiogenic activity were assessed using luciferase assays, quantitative real-time PCR, western blots, scratch assays, CCK-8 assays and tubule formation assays. In this study, miR-21 was strikingly downregulated by metformin in a time- and dose-dependent manner. miR-21 directly targeted the 3′-UTR of PTEN and SMAD7, and negatively regulated their expression. Overexpression of miR-21 abrogated the metformin-mediated inhibition of endothelial cells proliferation, migration, tubule formation and the TGF-β-induced AKT, SMAD- and ERK-dependent phosphorylations, and conversely, down-regulation of miR-21 aggravated metformin’s action and revealed significant promotion effects. Our study broadens our understanding of the regulatory mechanism of miR-21 mediating metformin-induced anti-angiogenic effects, providing important implications regarding the design of novel miRNA-based therapeutic strategies against angiogenesis. PMID:28230206

Suppression of OVA-alum induced allergy by Heligmosomoides polygyrus products is MyD88-, TRIF-, regulatory T- and B cell-independent, but is associated with reduced innate lymphoid cell activation.

PubMed

McSorley, Henry J; Blair, Natalie F; Robertson, Elaine; Maizels, Rick M

2015-11-01

The murine intestinal nematode Heligmosomoides polygyrus exerts multiple immunomodulatory effects in the host, including the suppression of allergic inflammation in mice sensitized to allergen presented with alum adjuvant. Similar suppression is attained by co-administration of H. polygyrus excretory/secretory products (HES) with the sensitizing dose of ovalbumin (OVA) in alum. We investigated the mechanism of suppression by HES in this model, and found it was maintained in MyD88xTRIF-deficient mice, implying no role for helminth- or host-derived TLR ligands, or IL-1 family cytokines that signal in a MyD88- or TRIF-dependent manner. We also found suppression was unchanged in µMT mice, which lack B2 B cells, and that suppression was not abrogated when regulatory T cells were depleted in Foxp3.LuciDTR-4 mice. However, reduced IL-5 production was seen in the first 12 h after injection of OVA-alum when HES was co-administered, associated with reduced activation of IL-5(+) and IL-13(+) group 2 innate lymphoid cells. Thus, the suppressive effects of HES on alum-mediated OVA sensitization are reflected in the very earliest innate response to allergen exposure in vivo. Copyright © 2015. Published by Elsevier Inc.

Cyclic Nucleotides Differentially Regulate Cx43 Gap Junction Function in Uterine Artery Endothelial Cells From Pregnant Ewes

PubMed Central

Ampey, Bryan C.; Ampey, Amanda C.; Lopez, Gladys E.; Bird, Ian M.

2017-01-01

Cell–cell communication is dependent on GJ (gap junction) proteins such as Cx43 (connexin 43). We previously demonstrated the importance of Cx43 function in establishing the enhanced pregnancy vasodilatory phenotype during pregnancy in uterine artery endothelial cells from pregnant (P-UAEC) ewes. Cx43 is regulated by elevating cAMP and PKA (protein kinase A)–dependent Cx43 S365 phosphorylation–associated trafficking and GJ open gating, which is opposed by PKC (protein kinase C)–dependent S368 phosphorylation-mediated GJ turnover and closed gating. However, the role of cyclic nucleotide-mediated signaling mechanisms that control Cx43 and GJ function in P-UAECs is unknown. We hypothesize that cAMP will mediate increases in S365 phosphorylation, thereby, enhancing GJ trafficking and open gating, while cGMP will stimulate S368, but not S365, phosphorylation to enhance GJ turnover and closed gating in P-UAECs. Treatment with 8-Bromo (8-Br)-cAMP signal significantly (P<0.05) increased nonphosphorylated S365 signal and total Cx43 phosphorylation, but not S368 phosphorylation, while 8-Br-cGMP significantly (P<0.05) increased Cx43 C-terminus-S365 signal, S368, and total Cx43 phosphorylation. Inhibition of PKA, but not PKG (protein kinase G), abrogated the 8-Br-cAMP–stimulated increase in nonphosphorylated S365 and total Cx43 phosphorylation and inhibited S368 below basal levels, whereas inhibition of PKG blocked (P<0.05) the 8-bromo-cGMP-stimulated rises in nonphosphorylated S365, total Cx43, and S368 phosphorylation levels in P-UAECs. Functional studies showed that 8-Br-cAMP increased dye transfer and sustained calcium bursts, while 8-Br-cGMP decreased both. Thus, in P-UAECs, only 8-Br-cAMP and not 8-Br-cGMP effectively enhances nonphosphorylated S365 and total Cx43 expression that correspondingly reduces S368 phosphorylation, allowing increased GJ communication. This provides new insights into the regulatory mechanisms behind Cx43 function and GJ communication. PMID:28559397

LncRNA ZFAS1 as a SERCA2a Inhibitor to Cause Intracellular Ca2+ Overload and Contractile Dysfunction in a Mouse Model of Myocardial Infarction.

PubMed

Zhang, Ying; Jiao, Lei; Sun, Lihua; Li, Yanru; Gao, Yuqiu; Xu, Chaoqian; Shao, Yingchun; Li, Mengmeng; Li, Chunyan; Lu, Yanjie; Pan, Zhenwei; Xuan, Lina; Zhang, Yiyuan; Li, Qingqi; Yang, Rui; Zhuang, Yuting; Zhang, Yong; Yang, Baofeng

2018-05-11

Ca 2+ homeostasis-a critical determinant of cardiac contractile function-is critically regulated by SERCA2a (sarcoplasmic reticulum Ca 2+ -ATPase 2a). Our previous study has identified ZFAS1 as a new lncRNA biomarker of acute myocardial infarction (MI). To evaluate the effects of ZFAS1 on SERCA2a and the associated Ca 2+ homeostasis and cardiac contractile function in the setting of MI. ZFAS1 expression was robustly increased in cytoplasm and sarcoplasmic reticulum in a mouse model of MI and a cellular model of hypoxia. Knockdown of endogenous ZFAS1 by virus-mediated silencing shRNA partially abrogated the ischemia-induced contractile dysfunction. Overexpression of ZFAS1 in otherwise normal mice created similar impairment of cardiac function as that observed in MI mice. Moreover, at the cellular level, ZFAS1 overexpression weakened the contractility of cardiac muscles. At the subcellular level, ZFAS1 deleteriously altered the Ca 2+ transient leading to intracellular Ca 2+ overload in cardiomyocytes. At the molecular level, ZFAS1 was found to directly bind SERCA2a protein and to limit its activity, as well as to repress its expression. The effects of ZFAS1 were readily reversible on knockdown of this lncRNA. Notably, a sequence domain of ZFAS1 gene that is conserved across species mimicked the effects of the full-length ZFAS1 . Mutation of this domain or application of an antisense fragment to this conserved region efficiently canceled out the deleterious actions of ZFAS1 . ZFAS1 had no significant effects on other Ca 2+ -handling regulatory proteins. ZFAS1 is an endogenous SERCA2a inhibitor, acting by binding to SERCA2a protein to limit its intracellular level and inhibit its activity, and a contributor to the impairment of cardiac contractile function in MI. Therefore, anti- ZFAS1 might be considered as a new therapeutic strategy for preserving SERCA2a activity and cardiac function under pathological conditions of the heart. © 2018 The Authors.

An exon 53 frameshift mutation in CUBN abrogates cubam function and causes Imerslund-Gräsbeck syndrome in dogs

PubMed Central

Fyfe, John C.; Hemker, Shelby L.; Venta, Patrick J.; Fitzgerald, Caitlin A.; Outerbridge, Catherine A.; Myers, Sherry L.; Giger, Urs

2013-01-01

Cobalamin malabsorption accompanied by selective proteinuria is an autosomal recessive disorder known as Imerslund-Gräsbeck syndrome in humans and was previously described in dogs due to amnionless (AMN) mutations. The resultant vitamin B12 deficiency causes dyshematopoiesis, lethargy, failure to thrive, and life-threatening metabolic disruption in the juvenile period. We studied 3 kindreds of border collies with cobalamin malabsorption and mapped the disease locus in affected dogs to a 2.9 Mb region of homozygosity on canine chromosome 2. The region included CUBN, the locus encoding cubilin, a peripheral membrane protein that in concert with AMN forms the functional intrinsic factor-cobalamin receptor expressed in ileum and a multi-ligand receptor in renal proximal tubules. Cobalamin malabsorption and proteinuria comprising CUBN ligands were demonstrated by radiolabeled cobalamin uptake studies and SDS-PAGE, respectively. CUBN mRNA and protein expression were reduced ~10 fold and ~20 fold, respectively, in both ileum and kidney of affected dogs. DNA sequencing demonstrated a single base deletion in exon 53 predicting a translational frameshift and early termination codon likely triggering nonsense mediated mRNA decay. The mutant allele segregated with disease in the border collie kindred. The border collie disorder indicates that a CUBN mutation far C-terminal from the intrinsic factor-cobalamin binding site can abrogate receptor expression and cause Imerslund-Gräsbeck syndrome. PMID:23746554

Surgical Stress Abrogates Pre-Existing Protective T Cell Mediated Anti-Tumor Immunity Leading to Postoperative Cancer Recurrence

PubMed Central

Lansdell, Casey; Alkayyal, Almohanad A.; Baxter, Katherine E.; Angka, Leonard; Zhang, Jiqing; Tanese de Souza, Christiano; Stephenson, Kyle B.; Parato, Kelley; Bramson, Jonathan L.; Bell, John C.; Lichty, Brian D.; Auer, Rebecca C.

2016-01-01

Anti-tumor CD8+ T cells are a key determinant for overall survival in patients following surgical resection for solid malignancies. Using a mouse model of cancer vaccination (adenovirus expressing melanoma tumor-associated antigen (TAA)—dopachrome tautomerase (AdDCT) and resection resulting in major surgical stress (abdominal nephrectomy), we demonstrate that surgical stress results in a reduction in the number of CD8+ T cell that produce cytokines (IFNγ, TNFα, Granzyme B) in response to TAA. This effect is secondary to both reduced proliferation and impaired T cell function following antigen binding. In a prophylactic model, surgical stress completely abrogates tumor protection conferred by vaccination in the immediate postoperative period. In a clinically relevant surgical resection model, vaccinated mice undergoing a positive margin resection with surgical stress had decreased survival compared to mice with positive margin resection alone. Preoperative immunotherapy with IFNα significantly extends survival in surgically stressed mice. Importantly, myeloid derived suppressor cell (MDSC) population numbers and functional impairment of TAA-specific CD8+ T cell were altered in surgically stressed mice. Our observations suggest that cancer progression may result from surgery-induced suppression of tumor-specific CD8+ T cells. Preoperative immunotherapies aimed at targeting the prometastatic effects of cancer surgery will reduce recurrence and improve survival in cancer surgery patients. PMID:27196057

Cell intrinsic abrogation of TGFβ signaling delays but does not prevent dysfunction of self/tumor specific CD8 T cells in a murine model of autochthonous prostate cancer

PubMed Central

Chou, Cassie K.; Schietinger, Andrea; Liggitt, H. Denny; Tan, Xiaoxia; Funk, Sarah; Freeman, Gordon J.; Ratliff, Timothy L.; Greenberg, Norman M.; Greenberg, Philip D.

2012-01-01

Adoptive T cell therapy (ACT) for the treatment of established cancers is actively being pursued in clinical trials. However, poor in vivo persistence and maintenance of anti-tumor activity of transferred T cells remain major problems. Transforming growth factor beta (TGFβ) is a potent immunosuppressive cytokine that is often expressed at high levels within the tumor microenvironment, potentially limiting T cell mediated anti-tumor activity. Here, we used a model of autochthonous murine prostate cancer to evaluate the effect of cell intrinsic abrogation of TGFβ signaling in self/tumor specific CD8 T cells used in ACT to target the tumor in situ. We found that persistence and anti-tumor activity of adoptively transferred effector T cells deficient in TGFβ signaling was significantly improved in the cancerous prostate. However, over time, despite persistence in peripheral lymphoid organs, the numbers of transferred cells in the prostate decreased and the residual prostate infiltrating T cells were no longer functional. These findings reveal that TGFβ negatively regulates the accumulation and effector function of transferred self/tumor specific CD8 T cells and highlight that, when targeting a tumor antigen that is also expressed as a self-protein, additional substantive obstacles are operative within the tumor microenvironment, potentially hampering the success of ACT for solid tumors. PMID:22984076

Ergothioneine Biosynthesis and Functionality in the Opportunistic Fungal Pathogen, Aspergillus fumigatus

PubMed Central

Sheridan, Kevin J.; Lechner, Beatrix Elisabeth; Keeffe, Grainne O’; Keller, Markus A.; Werner, Ernst R.; Lindner, Herbert; Jones, Gary W.; Haas, Hubertus; Doyle, Sean

2016-01-01

Ergothioneine (EGT; 2-mercaptohistidine trimethylbetaine) is a trimethylated and sulphurised histidine derivative which exhibits antioxidant properties. Here we report that deletion of Aspergillus fumigatus egtA (AFUA_2G15650), which encodes a trimodular enzyme, abrogated EGT biosynthesis in this opportunistic pathogen. EGT biosynthetic deficiency in A. fumigatus significantly reduced resistance to elevated H2O2 and menadione, respectively, impaired gliotoxin production and resulted in attenuated conidiation. Quantitative proteomic analysis revealed substantial proteomic remodelling in ΔegtA compared to wild-type under both basal and ROS conditions, whereby the abundance of 290 proteins was altered. Specifically, the reciprocal differential abundance of cystathionine γ-synthase and β-lyase, respectively, influenced cystathionine availability to effect EGT biosynthesis. A combined deficiency in EGT biosynthesis and the oxidative stress response regulator Yap1, which led to extreme oxidative stress susceptibility, decreased resistance to heavy metals and production of the extracellular siderophore triacetylfusarinine C and increased accumulation of the intracellular siderophore ferricrocin. EGT dissipated H2O2 in vitro, and elevated intracellular GSH levels accompanied abrogation of EGT biosynthesis. EGT deficiency only decreased resistance to high H2O2 levels which suggests functionality as an auxiliary antioxidant, required for growth at elevated oxidative stress conditions. Combined, these data reveal new interactions between cellular redox homeostasis, secondary metabolism and metal ion homeostasis. PMID:27748436

Inhibition of Oct 3/4 mitigates the cardiac progenitor-derived myocardial repair in infarcted myocardium.

PubMed

Zhao, Yu Tina; Du, Jianfeng; Chen, Youfang; Tang, Yaoliang; Qin, Gangjian; Lv, Guorong; Zhuang, Shougang; Zhao, Ting C

2015-12-24

Recent evidence has demonstrated that cardiac progenitor cells play an essential role in the induction of angiomyogenesis in infarcted myocardium. We and others have shown that engraftment of c-kit(+) cardiac stem cells (CSCs) into infarcted hearts led to myocardium regeneration and neovascularization, which was associated with an improvement of ventricular function. The purpose of this study is aimed at investigating the functional role of transcription factor (TF) Oct3/4 in facilitating CSCs to promote myocardium regeneration and preserve cardiac performance in the post-MI heart. c-kit(+) CSCs were isolated from adult hearts and re-introduced into the infarcted myocardium in which the mouse MI model was created by permanent ligation of the left anterior descending artery (LAD). The Oct3/4 of CSCs was inhibited by transfection of Oct3/4 siRNA, and transfection of CSCs with control siRNA serves as control groups. Myocardial functions were evaluated by echocardiographic measurement. Histological analysis was employed to assess newly formed cardiogenesis, neovascularization, and cell proliferations. Terminal deoxynucleotidyltransferase (TdT) nick-end labeling (TUNEL) was carried out to assess apoptotic cardiomyocytes. Real time polymerase chain reaction and Western blot were carried out to evaluate the level of Oct 3/4 in CSCs. Two weeks after engraftment, CSCs increased ventricular functional recovery as shown by a serial echocardiographic measurement, which is concomitant with the suppression of cardiac hypertrophy and attenuation of myocardial interstitial fibrosis. Suppression of Oct 3/4 of CSCs abrogated functional improvements and mitigated the hypertrophic response and cardiac remodeling. Transplantation of c-kit(+) CSCs into MI hearts promoted cardiac regeneration and neovascularization, which were abolished with the knockdown of Oct3/4. Additionally, suppression of Oct3/4 abrogated myocyte proliferation in the CSC-engrafted myocardium. Our results indicate that CSCs-derived cardiac regeneration improves the restoration of cardiac function and is mediated through Oct 3/4.

Alternative Mechanism by which IFN-γ Enhances Tumor Recognition: Active Release of Heat Shock Protein 721

PubMed Central

Bausero, Maria A.; Gastpar, Robert; Multhoff, Gabriele; Asea, Alexzander

2006-01-01

IFN-γ exhibits differential effects depending on the target and can induce cellular activation and enhance survival or mediate cell death via activation of apoptotic pathways. In this study, we demonstrate an alternative mechanism by which IFN-γ enhances tumor recognition, mediated by the active release of Hsp72. We demonstrate that stimulation of 4T1 breast adenocarcinoma cells and K562 erythroleukemic cells with IFN-γ triggers the cellular stress response, which results in the enhanced expression of total Hsp72 expression without a significant increase in cell death. Intracellular expression of Hsp72 was abrogated in cells stably transfected with a mutant hsf-1 gene. IFN-γ-induced Hsp72 expression correlated with enhanced surface expression and consequent release of Hsp72 into the culture medium. Pretreatment of tumors with compounds known to the block the classical protein transport pathway, including monensin, brefeldin A, tunicamycin, and thapsigargin, did not significantly block Hsp72 release. However, pretreatment with intracellular calcium chelator BAPTA-AM or disruption of lipid rafts using methyl β-cyclodextrin completely abrogated IFN-γ-induced Hsp72 release. Biochemical characterization revealed that Hsp72 is released within exosomes and has the ability to up-regulate CD83 expression and stimulate IL-12 release by naive dendritic cells. Pretreatment with neutralizing mAb or depletion of Hsp72 completely abrogated its chaperokine function. Taken together, these findings are indicative of an additional previously unknown mechanism by which IFN-γ promotes tumor surveillance and furthers our understanding of the central role of extracellular Hsp72 as an endogenous adjuvant and danger signal. PMID:16116176

Alternative mechanism by which IFN-gamma enhances tumor recognition: active release of heat shock protein 72.

PubMed

Bausero, Maria A; Gastpar, Robert; Multhoff, Gabriele; Asea, Alexzander

2005-09-01

IFN-gamma exhibits differential effects depending on the target and can induce cellular activation and enhance survival or mediate cell death via activation of apoptotic pathways. In this study, we demonstrate an alternative mechanism by which IFN-gamma enhances tumor recognition, mediated by the active release of Hsp72. We demonstrate that stimulation of 4T1 breast adenocarcinoma cells and K562 erythroleukemic cells with IFN-gamma triggers the cellular stress response, which results in the enhanced expression of total Hsp72 expression without a significant increase in cell death. Intracellular expression of Hsp72 was abrogated in cells stably transfected with a mutant hsf-1 gene. IFN-gamma-induced Hsp72 expression correlated with enhanced surface expression and consequent release of Hsp72 into the culture medium. Pretreatment of tumors with compounds known to the block the classical protein transport pathway, including monensin, brefeldin A, tunicamycin, and thapsigargin, did not significantly block Hsp72 release. However, pretreatment with intracellular calcium chelator BAPTA-AM or disruption of lipid rafts using methyl beta-cyclodextrin completely abrogated IFN-gamma-induced Hsp72 release. Biochemical characterization revealed that Hsp72 is released within exosomes and has the ability to up-regulate CD83 expression and stimulate IL-12 release by naive dendritic cells. Pretreatment with neutralizing mAb or depletion of Hsp72 completely abrogated its chaperokine function. Taken together, these findings are indicative of an additional previously unknown mechanism by which IFN-gamma promotes tumor surveillance and furthers our understanding of the central role of extracellular Hsp72 as an endogenous adjuvant and danger signal.

Preventing tissue fibrosis by local biomaterials interfacing of specific cryptic extracellular matrix information

PubMed Central

Horejs, Christine-Maria; St-Pierre, Jean-Philippe; Ojala, Juha R. M.; Steele, Joseph A. M.; da Silva, Patricia Barros; Rynne-Vidal, Angela; Maynard, Stephanie A.; Hansel, Catherine S.; Rodríguez-Fernández, Clara; Mazo, Manuel M.; You, Amanda Y. F.; Wang, Alex J.; von Erlach, Thomas; Tryggvason, Karl; López-Cabrera, Manuel; Stevens, Molly M.

2017-01-01

Matrix metalloproteinases (MMPs) contribute to the breakdown of tissue structures such as the basement membrane, promoting tissue fibrosis. Here we developed an electrospun membrane biofunctionalized with a fragment of the laminin β1-chain to modulate the expression of MMP2 in this context. We demonstrate that interfacing of the β1-fragment with the mesothelium of the peritoneal membrane via a biomaterial abrogates the release of active MMP2 in response to transforming growth factor β1 and rescues tissue integrity ex vivo and in vivo in a mouse model of peritoneal fibrosis. Importantly, our data demonstrate that the membrane inhibits MMP2 expression. Changes in the expression of epithelial-to-mesenchymal transition (EMT)-related molecules further point towards a contribution of the modulation of EMT. Biomaterial-based presentation of regulatory basement membrane signals directly addresses limitations of current therapeutic approaches by enabling a localized and specific method to counteract MMP2 release applicable to a broad range of therapeutic targets. PMID:28593951

Preventing tissue fibrosis by local biomaterials interfacing of specific cryptic extracellular matrix information

NASA Astrophysics Data System (ADS)

Horejs, Christine-Maria; St-Pierre, Jean-Philippe; Ojala, Juha R. M.; Steele, Joseph A. M.; da Silva, Patricia Barros; Rynne-Vidal, Angela; Maynard, Stephanie A.; Hansel, Catherine S.; Rodríguez-Fernández, Clara; Mazo, Manuel M.; You, Amanda Y. F.; Wang, Alex J.; von Erlach, Thomas; Tryggvason, Karl; López-Cabrera, Manuel; Stevens, Molly M.

2017-06-01

Matrix metalloproteinases (MMPs) contribute to the breakdown of tissue structures such as the basement membrane, promoting tissue fibrosis. Here we developed an electrospun membrane biofunctionalized with a fragment of the laminin β1-chain to modulate the expression of MMP2 in this context. We demonstrate that interfacing of the β1-fragment with the mesothelium of the peritoneal membrane via a biomaterial abrogates the release of active MMP2 in response to transforming growth factor β1 and rescues tissue integrity ex vivo and in vivo in a mouse model of peritoneal fibrosis. Importantly, our data demonstrate that the membrane inhibits MMP2 expression. Changes in the expression of epithelial-to-mesenchymal transition (EMT)-related molecules further point towards a contribution of the modulation of EMT. Biomaterial-based presentation of regulatory basement membrane signals directly addresses limitations of current therapeutic approaches by enabling a localized and specific method to counteract MMP2 release applicable to a broad range of therapeutic targets.

Stargazin regulates AMPA receptor trafficking through adaptor protein complexes during long-term depression

NASA Astrophysics Data System (ADS)

Matsuda, Shinji; Kakegawa, Wataru; Budisantoso, Timotheus; Nomura, Toshihiro; Kohda, Kazuhisa; Yuzaki, Michisuke

2013-11-01

Long-term depression (LTD) underlies learning and memory in various brain regions. Although postsynaptic AMPA receptor trafficking mediates LTD, its underlying molecular mechanisms remain largely unclear. Here we show that stargazin, a transmembrane AMPA receptor regulatory protein, forms a ternary complex with adaptor proteins AP-2 and AP-3A in hippocampal neurons, depending on its phosphorylation state. Inhibiting the stargazin-AP-2 interaction disrupts NMDA-induced AMPA receptor endocytosis, and inhibiting that of stargazin-AP-3A abrogates the late endosomal/lysosomal trafficking of AMPA receptors, thereby upregulating receptor recycling to the cell surface. Similarly, stargazin’s interaction with AP-2 or AP-3A is necessary for low-frequency stimulus-evoked LTD in CA1 hippocampal neurons. Thus, stargazin has a crucial role in NMDA-dependent LTD by regulating two trafficking pathways of AMPA receptors—transport from the cell surface to early endosomes and from early endosomes to late endosomes/lysosomes—through its sequential binding to AP-2 and AP-3A.

[CD4 + CD25 + regulatory T cells and their importance to human illnesses].

PubMed

Kelsen, Jens; Hvas, Christian Lodberg; Agnholt, Jørgen; Dahlerup, Jens F

2006-01-03

Regulatory T cells ensure a balanced immune response that is competent both to fight pathogens, at the same time, to recognize self-antigens and commensals as harmless. Regulatory mechanisms are essential in preventing autoimmune disorders but may also facilitate the progression of malignant diseases and the establishment of latent infections via suppression of the host immune response. Regulatory T cells arise in the thymus, and regulatory T cell function can be induced in the periphery, so-called infectious tolerance. An absolute or relative defect in regulatory T cell function may contribute to the development of autoimmune disorders such as rheumatoid arthritis, type 1 diabetes mellitus, multiple sclerosis and chronic inflammatory bowel disease. Regulatory T cell therapy is a tempting strategy for reestablishing the immune balance and thus preventing or reversing these disorders. Reestablishment of the immune balance may be accomplished by adoptive transfer of ex vivo-propagated regulatory T cells or by induction of regulatory functions locally in the organs, although such strategies are in their infancy in human research.

Sustained attention in infancy as a longitudinal predictor of self-regulatory functions.

PubMed

Johansson, Maria; Marciszko, Carin; Gredebäck, Gustaf; Nyström, Pär; Bohlin, Gunilla

2015-11-01

Previous literature suggests that attention processes such as sustained attention would constitute a developmental foundation for the self-regulatory functions executive functioning and effortful control (e.g., Garon, Bryson, & Smith, 2008; Rothbart, Derryberry, & Posner, 1994). Our main aim was to test this hypothesis by studying whether sustained attention at age 1 year can predict individual differences in self-regulatory functions at age 2 years. Longitudinal data from 66 infants and their parents were included in the study. Sustained attention was assessed during free play at age 1 year; executive functioning, measured using an eye-tracking version of the A-not-B task, and effortful control, measured using parental ratings, were assessed at both age 1 and age 2 years. The results did support a longitudinal prediction of individual differences in 2-year-olds' self-regulatory functions as a function of sustained attention at age 1 year. We also found significant improvement in both executive functioning and effortful control over time, and the two self-regulatory constructs were related in toddlerhood but not in infancy. The study helps increase our understanding of the early development of self-regulatory functions necessary for identifying developmental risks and, in the future, for developing new interventions. Copyright © 2015 Elsevier Inc. All rights reserved.

Nipah virus sequesters inactive STAT1 in the nucleus via a P gene-encoded mechanism.

PubMed

Ciancanelli, Michael J; Volchkova, Valentina A; Shaw, Megan L; Volchkov, Viktor E; Basler, Christopher F

2009-08-01

The Nipah virus (NiV) phosphoprotein (P) gene encodes the C, P, V, and W proteins. P, V, and W, have in common an amino-terminal domain sufficient to bind STAT1, inhibiting its interferon (IFN)-induced tyrosine phosphorylation. P is also essential for RNA-dependent RNA polymerase function. C is encoded by an alternate open reading frame (ORF) within the common amino-terminal domain. Mutations within residues 81 to 113 of P impaired its polymerase cofactor function, as assessed by a minireplicon assay, but these mutants retained STAT1 inhibitory function. Mutations within the residue 114 to 140 region were identified that abrogated interaction with and inhibition of STAT1 by P, V, and W without disrupting P polymerase cofactor function. Recombinant NiVs were then generated. A G121E mutation, which abrogated inhibition of STAT1, was introduced into a C protein knockout background (C(ko)) because the mutation would otherwise also alter the overlapping C ORF. In cell culture, relative to the wild-type virus, the C(ko) mutation proved attenuating but the G121E mutant virus replicated identically to the C(ko) virus. In cells infected with the wild-type and C(ko) viruses, STAT1 was nuclear despite the absence of tyrosine phosphorylation. This latter observation mirrors what has been seen in cells expressing NiV W. In the G121E mutant virus-infected cells, STAT1 was not phosphorylated and was cytoplasmic in the absence of IFN stimulation but became tyrosine phosphorylated and nuclear following IFN addition. These data demonstrate that the gene for NiV P encodes functions that sequester inactive STAT1 in the nucleus, preventing its activation and suggest that the W protein is the dominant inhibitor of STAT1 in NiV-infected cells.

Gene regulatory networks and the underlying biology of developmental toxicity

EPA Science Inventory

Embryonic cells are specified by large-scale networks of functionally linked regulatory genes. Knowledge of the relevant gene regulatory networks is essential for understanding phenotypic heterogeneity that emerges from disruption of molecular functions, cellular processes or sig...

Conserved and species-specific transcription factor co-binding patterns drive divergent gene regulation in human and mouse

PubMed Central

Diehl, Adam G

2018-01-01

Abstract The mouse is widely used as system to study human genetic mechanisms. However, extensive rewiring of transcriptional regulatory networks often confounds translation of findings between human and mouse. Site-specific gain and loss of individual transcription factor binding sites (TFBS) has caused functional divergence of orthologous regulatory loci, and so we must look beyond this positional conservation to understand common themes of regulatory control. Fortunately, transcription factor co-binding patterns shared across species often perform conserved regulatory functions. These can be compared to ‘regulatory sentences’ that retain the same meanings regardless of sequence and species context. By analyzing TFBS co-occupancy patterns observed in four human and mouse cell types, we learned a regulatory grammar: the rules by which TFBS are combined into meaningful regulatory sentences. Different parts of this grammar associate with specific sets of functional annotations regardless of sequence conservation and predict functional signatures more accurately than positional conservation. We further show that both species-specific and conserved portions of this grammar are involved in gene expression divergence and human disease risk. These findings expand our understanding of transcriptional regulatory mechanisms, suggesting that phenotypic divergence and disease risk are driven by a complex interplay between deeply conserved and species-specific transcriptional regulatory pathways. PMID:29361190

An exon 53 frameshift mutation in CUBN abrogates cubam function and causes Imerslund-Gräsbeck syndrome in dogs.

PubMed

Fyfe, John C; Hemker, Shelby L; Venta, Patrick J; Fitzgerald, Caitlin A; Outerbridge, Catherine A; Myers, Sherry L; Giger, Urs

2013-08-01

Cobalamin malabsorption accompanied by selective proteinuria is an autosomal recessive disorder known as Imerslund-Gräsbeck syndrome in humans and was previously described in dogs due to amnionless (AMN) mutations. The resultant vitamin B12 deficiency causes dyshematopoiesis, lethargy, failure to thrive, and life-threatening metabolic disruption in the juvenile period. We studied 3 kindreds of border collies with cobalamin malabsorption and mapped the disease locus in affected dogs to a 2.9Mb region of homozygosity on canine chromosome 2. The region included CUBN, the locus encoding cubilin, a peripheral membrane protein that in concert with AMN forms the functional intrinsic factor-cobalamin receptor expressed in ileum and a multi-ligand receptor in renal proximal tubules. Cobalamin malabsorption and proteinuria comprising CUBN ligands were demonstrated by radiolabeled cobalamin uptake studies and SDS-PAGE, respectively. CUBN mRNA and protein expression were reduced ~10 fold and ~20 fold, respectively, in both ileum and kidney of affected dogs. DNA sequencing demonstrated a single base deletion in exon 53 predicting a translational frameshift and early termination codon likely triggering nonsense mediated mRNA decay. The mutant allele segregated with the disease in the border collie kindred. The border collie disorder indicates that a CUBN mutation far C-terminal from the intrinsic factor-cobalamin binding site can abrogate receptor expression and cause Imerslund-Gräsbeck syndrome. Copyright © 2013 Elsevier Inc. All rights reserved.

Sulforaphane Potentiates the Efficacy of 17-Allylamino 17-Demethoxygeldanamycin Against Pancreatic Cancer Through Enhanced Abrogation of Hsp90 Chaperone Function

PubMed Central

Li, Yanyan; Zhang, Tao; Schwartz, Steven J.; Sun, Duxin

2013-01-01

Heat shock protein 90 (Hsp90), an essential molecular chaperone that regulates the stability of a wide range of oncogenic proteins, is a promising target for cancer therapeutics. We investigated the combination efficacy and potential mechanisms of sulforaphane, a dietary component from broccoli and broccoli sprouts, and 17-allylamino 17-demethoxygeldanamycin (17-AAG), an Hsp90 inhibitor, in pancreatic cancer. MTS assay demonstrated that sulforaphane sensitized pancreatic cancer cells to 17-AAG in vitro. Caspase-3 was activated to 6.4-fold in response to simultaneous treatment with sulforaphane and 17-AAG, whereas 17-AAG alone induced caspase-3 activity to 2-fold compared to control. ATP binding assay and coimmunoprecipitation revealed that sulforaphane disrupted Hsp90-p50Cdc37 interaction, whereas 17-AAG inhibited ATP binding to Hsp90. Concomitant use of sulforaphane and 17-AAG synergistically downregulated Hsp90 client proteins in Mia Paca-2 cells. Co-administration of sulforaphane and 17-AAG in pancreatic cancer xenograft model led to more than 70% inhibition of the tumor growth, whereas 17-AAG alone only suppressed the tumor growth by 50%. Our data suggest that sulforaphane potentiates the efficacy of 17-AAG against pancreatic cancer through enhanced abrogation of Hsp90 function. These findings provide a rationale for further evaluation of broccoli/broccoli sprout preparations combined with 17-AAG for better efficacy and lower dose-limiting toxicity in pancreatic cancer. PMID:21875325

Disturbed vesicular trafficking of membrane proteins in prion disease.

PubMed

Uchiyama, Keiji; Miyata, Hironori; Sakaguchi, Suehiro

2013-01-01

The pathogenic mechanism of prion diseases remains unknown. We recently reported that prion infection disturbs post-Golgi trafficking of certain types of membrane proteins to the cell surface, resulting in reduced surface expression of membrane proteins and abrogating the signal from the proteins. The surface expression of the membrane proteins was reduced in the brains of mice inoculated with prions, well before abnormal symptoms became evident. Prions or pathogenic prion proteins were mainly detected in endosomal compartments, being particularly abundant in recycling endosomes. Some newly synthesized membrane proteins are delivered to the surface from the Golgi apparatus through recycling endosomes, and some endocytosed membrane proteins are delivered back to the surface through recycling endosomes. These results suggest that prions might cause neuronal dysfunctions and cell loss by disturbing post-Golgi trafficking of membrane proteins via accumulation in recycling endosomes. Interestingly, it was recently shown that delivery of a calcium channel protein to the cell surface was impaired and its function was abrogated in a mouse model of hereditary prion disease. Taken together, these results suggest that impaired delivery of membrane proteins to the cell surface is a common pathogenic event in acquired and hereditary prion diseases.

The Role of Functional Foods in Cutaneous Anti-aging

PubMed Central

Cho, Soyun

2014-01-01

Oral supplementation of micronutrients, or functional foods, to prevent aging has gained much attention and popularity as society ages and becomes more affluent, and as science reveals the pathological mechanisms of aging. Aging of the skin combines biologic aging and extrinsic aging caused predominantly by sunlight and other environmental toxins. Anti-aging functional foods exert their influence mostly through their anti-oxidant and anti-inflammatory effects, thereby abrogating collagen degradation and/or increasing procollagen synthesis. Clinical evidence supporting a role in preventing cutaneous aging is available for oral supplements such as carotenoids, polyphenols, chlorophyll, aloe vera, vitamins C and E, red ginseng, squalene, and omega-3 fatty acids. Collagen peptides and proteoglycans are claimed to provide building blocks of the dermal matrix. This review summarizes the current study findings of these functional foods. PMID:26064850

Galectin-9 Signaling through TIM-3 Is Involved in Neutrophil-Mediated Gram-Negative Bacterial Killing: An Effect Abrogated within the Cystic Fibrosis Lung

PubMed Central

Vega-Carrascal, Isabel; Bergin, David A.; McElvaney, Oliver J.; McCarthy, Cormac; Banville, Nessa; Pohl, Kerstin; Hirashima, Mitsuomi; Kuchroo, Vijay K.; Reeves, Emer P.; McElvaney, Noel G.

2016-01-01

The T cell Ig and mucin domain–containing molecule (TIM) family of receptors have emerged as potential therapeutic targets to correct abnormal immune function in chronic inflammatory conditions. TIM-3 serves as a functional receptor in structural cells of the airways and via the ligand galectin-9 (Gal-9) can modulate the inflammatory response. The aim of this study was to investigate TIM-3 expression and function in neutrophils, focusing on its potential role in cystic fibrosis (CF) lung disease. Results revealed that TIM-3 mRNA and protein expression values of circulating neutrophils were equal between healthy controls (n = 20) and people with CF (n = 26). TIM-3 was detected on resting neutrophil membranes by FACS analysis, and expression levels significantly increased post IL-8 or TNF-α exposure (p < 0.05). Our data suggest a novel role for TIM-3/Gal-9 signaling involving modulation of cytosolic calcium levels. Via TIM-3 interaction, Gal-9 induced neutrophil degranulation and primed the cell for enhanced NADPH oxidase activity. Killing of Pseudomonas aeruginosa was significantly increased upon bacterial opsonization with Gal-9 (p < 0.05), an effect abrogated by blockade of TIM-3 receptors. This mechanism appeared to be Gram-negative bacteria specific and mediated via Gal-9/ LPS binding. Additionally, we have demonstrated that neutrophil TIM-3/Gal-9 signaling is perturbed in the CF airways due to proteolytic degradation of the receptor. In conclusion, results suggest a novel neutrophil defect potentially contributing to the defective bacterial clearance observed in the CF airways and suggest that manipulation of the TIM-3 signaling pathway may be of therapeutic value in CF, preferably in conjunction with antiprotease treatment. PMID:24477913

Inhibitory effect of CT domain of CCN3/NOV on proliferation and differentiation of osteogenic mesenchymal stem cells, Kusa-A1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Katsuki, Yuko; Oral Pathology, Graduate School of Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8549; Sakamoto, Kei

CCN3/NOV activates the Notch signal through the carboxyl terminal cysteine-rich (CT) domain. CCN3 transfection to Kusa-A1 inhibited osteogenic differentiation and cell proliferation, which is accompanied by upregulation of Hes/Hey, Notch downstream targets, and p21, a CDK inhibitor. Upregulation of Hes/Hey and p21 was abrogated by the deletion of CT domain. Anti-proliferative activity of CCN3 was also abrogated by CT domain deletion whereas anti-osteogenic activity was not completely abrogated. We found that CT domain-deleted CCN3 still possesses antagonistic effect on BMP-2. These results suggest that CCN3 employs Notch and BMP pathways in anti-osteogenic activity while it inhibits cell proliferation uniquely bymore » Notch/p21 pathway.« less

Predictive regulatory models in Drosophila melanogaster by integrative inference of transcriptional networks

PubMed Central

Marbach, Daniel; Roy, Sushmita; Ay, Ferhat; Meyer, Patrick E.; Candeias, Rogerio; Kahveci, Tamer; Bristow, Christopher A.; Kellis, Manolis

2012-01-01

Gaining insights on gene regulation from large-scale functional data sets is a grand challenge in systems biology. In this article, we develop and apply methods for transcriptional regulatory network inference from diverse functional genomics data sets and demonstrate their value for gene function and gene expression prediction. We formulate the network inference problem in a machine-learning framework and use both supervised and unsupervised methods to predict regulatory edges by integrating transcription factor (TF) binding, evolutionarily conserved sequence motifs, gene expression, and chromatin modification data sets as input features. Applying these methods to Drosophila melanogaster, we predict ∼300,000 regulatory edges in a network of ∼600 TFs and 12,000 target genes. We validate our predictions using known regulatory interactions, gene functional annotations, tissue-specific expression, protein–protein interactions, and three-dimensional maps of chromosome conformation. We use the inferred network to identify putative functions for hundreds of previously uncharacterized genes, including many in nervous system development, which are independently confirmed based on their tissue-specific expression patterns. Last, we use the regulatory network to predict target gene expression levels as a function of TF expression, and find significantly higher predictive power for integrative networks than for motif or ChIP-based networks. Our work reveals the complementarity between physical evidence of regulatory interactions (TF binding, motif conservation) and functional evidence (coordinated expression or chromatin patterns) and demonstrates the power of data integration for network inference and studies of gene regulation at the systems level. PMID:22456606

Constraint and Contingency in Multifunctional Gene Regulatory Circuits

PubMed Central

Payne, Joshua L.; Wagner, Andreas

2013-01-01

Gene regulatory circuits drive the development, physiology, and behavior of organisms from bacteria to humans. The phenotypes or functions of such circuits are embodied in the gene expression patterns they form. Regulatory circuits are typically multifunctional, forming distinct gene expression patterns in different embryonic stages, tissues, or physiological states. Any one circuit with a single function can be realized by many different regulatory genotypes. Multifunctionality presumably constrains this number, but we do not know to what extent. We here exhaustively characterize a genotype space harboring millions of model regulatory circuits and all their possible functions. As a circuit's number of functions increases, the number of genotypes with a given number of functions decreases exponentially but can remain very large for a modest number of functions. However, the sets of circuits that can form any one set of functions becomes increasingly fragmented. As a result, historical contingency becomes widespread in circuits with many functions. Whether a circuit can acquire an additional function in the course of its evolution becomes increasingly dependent on the function it already has. Circuits with many functions also become increasingly brittle and sensitive to mutation. These observations are generic properties of a broad class of circuits and independent of any one circuit genotype or phenotype. PMID:23762020

Dietary fiber prevents obesity-related liver lipotoxicity by modulating sterol-regulatory element binding protein pathway in C57BL/6J mice fed a high-fat/cholesterol diet.

PubMed

Han, Shufen; Jiao, Jun; Zhang, Wei; Xu, Jiaying; Wan, Zhongxiao; Zhang, Weiguo; Gao, Xiaoran; Qin, Liqiang

2015-10-29

Adequate intake of dietary fibers has proven metabolic and cardiovascular benefits, molecular mechanisms remain still limited. This study was aimed to investigate the effects of cereal dietary fiber on obesity-related liver lipotoxicity in C57BL/6J mice fed a high-fat/cholesterol (HFC) diet and underlying mechanism. Forty-eight adult male C57BL/6J mice were randomly given a reference chow diet, or a high fat/cholesterol (HFC) diet supplemented with or without oat fiber or wheat bran fiber for 24 weeks. Our results showed mice fed oat or wheat bran fiber exhibited lower weight gain, lipid profiles and insulin resistance, compared with HFC diet. The two cereal dietary fibers potently decreased protein expressions of sterol regulatory element binding protein-1 and key factors involved in lipogenesis, including fatty acid synthase and acetyl-CoA carboxylase in target tissues. At molecular level, the two cereal dietary fibers augmented protein expressions of peroxisome proliferator-activated receptor alpha and gamma, liver X receptor alpha, and ATP-binding cassette transporter A1 in target tissues. Our findings indicated that cereal dietary fiber supplementation abrogated obesity-related liver lipotoxicity and dyslipidemia in C57BL/6J mice fed a HFC diet. In addition, the efficacy of oat fiber is greater than wheat bran fiber in normalizing these metabolic disorders and pathological profiles.

Role of interferon regulatory factor-1 in lipopolysaccharide-induced mitochondrial damage and oxidative stress responses in macrophages

PubMed Central

Deng, Song-Yun; Zhang, Le-Meng; Ai, Yu-hang; Pan, Pin-Hua; Zhao, Shuang-Ping; Su, Xiao-Li; Wu, Dong-Dong; Tan, Hong-Yi; Zhang, Li-Na; Tsung, Allan

2017-01-01

Sepsis causes many early deaths; both macrophage mitochondrial damage and oxidative stress responses are key factors in its pathogenesis. Although the exact mechanisms responsible for sepsis-induced mitochondrial damage are unknown, the nuclear transcription factor, interferon regulatory factor-1 (IRF-1) has been reported to cause mitochondrial damage in several diseases. Previously, we reported that in addition to promoting systemic inflammation, IRF-1 promoted the apoptosis of and inhibited autophagy in macrophages. In the present study, we hypothesized that lipopolysaccharide (LPS)-induced IRF-1 activation in macrophages may promote mitochondrial damage and oxidative stress. In vitro, LPS was found to promote IRF-1 activation, reactive oxygen species (ROS) production, adenosine triphosphate (ATP) depletion, superoxide dismutase (SOD) consumption, malondialdehyde (MDA) accumulation and mitochondrial depolarization in macrophages in a time- and dose-dependent manner. These effects were abrogated in cells in which IRF-1 was knocked down. Furthermore, IRF-1 overexpression increased LPS-induced oxidative stress responses and mitochondrial damage. In vivo, peritoneal macrophages obtained from IRF-1 knockout (KO) mice produced less ROS and had less mitochondrial depolarization and damage following the administration of LPS, when compared to their wild-type (WT) counterparts. In addition, IRF-1 KO mice exhibited a decreased release of mitochondrial DNA (mtDNA) following the administration of LPS. Thus, IRF-1 may be a critical factor in augmenting LPS-induced oxidative stress and mitochondrial damage in macrophages. PMID:28849179

Targeting Antigens to Dec-205 on Dendritic Cells Induces Immune Protection in Experimental Colitis in Mice

PubMed Central

Wadwa, Munisch; Klopfleisch, Robert; Buer, Jan; Westendorf, Astrid M.

2016-01-01

The endocytotic c-type lectin receptor DEC-205 is highly expressed on immature dendritic cells. In previous studies, it was shown that antigen-targeting to DEC-205 is a useful tool for the induction of antigen-specific Foxp3+ regulatory T cells and thereby can prevent inflammatory processes. However, whether this approach is sufficient to mediate tolerance in mucosal tissues like the gut is unknown. In this study, we established a new mouse model in which the adoptive transfer of naive hemagglutinin (HA)-specific CD4+Foxp3– T cells into VILLIN-HA transgenic mice leads to severe colitis. To analyze if antigen-targeting to DEC-205 could protect against inflammation of the gut, VILLIN-HA transgenic mice were injected with an antibody–antigen complex consisting of the immunogenic HA110–120 peptide coupled to an α-DEC-205 antibody (DEC-HA) before adoptive T cell transfer. DEC-HA-treated mice showed significantly less signs of intestinal inflammation as was demonstrated by reduced loss of body weight and histopathology in the gut. Strikingly, abrogated intestinal inflammation was mediated via the conversion of naive HA-specific CD4+Foxp3– T cells into HA-specific CD4+Foxp3+ regulatory T cells. In this study, we provide evidence that antigen-targeting to DEC-205 can be utilized for the induction of tolerance in mucosal organs that are confronted with large numbers of exogenous antigens. PMID:27141310

Dietary fiber prevents obesity-related liver lipotoxicity by modulating sterol-regulatory element binding protein pathway in C57BL/6J mice fed a high-fat/cholesterol diet

PubMed Central

Han, Shufen; Jiao, Jun; Zhang, Wei; Xu, Jiaying; Wan, Zhongxiao; Zhang, Weiguo; Gao, Xiaoran; Qin, Liqiang

2015-01-01

Adequate intake of dietary fibers has proven metabolic and cardiovascular benefits, molecular mechanisms remain still limited. This study was aimed to investigate the effects of cereal dietary fiber on obesity-related liver lipotoxicity in C57BL/6J mice fed a high-fat/cholesterol (HFC) diet and underlying mechanism. Forty-eight adult male C57BL/6J mice were randomly given a reference chow diet, or a high fat/choleserol (HFC) diet supplemented with or without oat fiber or wheat bran fiber for 24 weeks. Our results showed mice fed oat or wheat bran fiber exhibtied lower weight gain, lipid profiles and insulin resistance, compared with HFC diet. The two cereal dietary fibers potently decreased protein expressions of sterol regulatory element binding protein-1 and key factors involved in lipogenesis, including fatty acid synthase and acetyl-CoA carboxylase in target tissues. At molecular level, the two cereal dietary fibers augmented protein expressions of peroxisome proliferator-activated receptor alpha and gamma, liver X receptor alpha, and ATP-binding cassette transporter A1 in target tissues. Our findings indicated that cereal dietary fiber supplementation abrogated obesity-related liver lipotoxicity and dyslipidemia in C57BL/6J mice fed a HFC diet. In addition, the efficacy of oat fiber is greater than wheat bran fiber in normalizing these metabolic disorders and pathological profiles. PMID:26510459

Blockade of Treg Cell Differentiation and Function by the Interleukin-21-Mechanistic Target of Rapamycin Axis Via Suppression of Autophagy in Patients With Systemic Lupus Erythematosus.

PubMed

Kato, Hiroshi; Perl, Andras

2018-03-01

The mechanistic target of rapamycin (mTOR) has become a therapeutic target in systemic lupus erythematosus (SLE). In T cells, mTOR plays a central role in lineage specification, including development of regulatory cells (Treg cells). This study sought to investigate whether mTOR is activated within Treg cells and whether this contributes to the depletion and dysfunction of Treg cells in patients with SLE. Activities of mTOR complexes 1 (mTORC1) and 2 (mTORC2) were examined by quantifying phosphorylation of translation initiation factor 4E-binding protein 1, S6 kinase, and Akt in SLE patients relative to age- and sex-matched female healthy control subjects. Polarization of Treg cells from naive CD4+ T cells was assessed in the presence of interleukin-6 (IL-6), IL-17, and IL-21. The suppressor function of sorted CD4+CD25+ Treg cells was measured by determining their impact on the proliferation of autologous CD4+CD25- responder T cells. Treg cell expression of FoxP3, GATA-3, and CTLA-4 was monitored by flow cytometry. Autophagy was assessed using immunoblotting of light chain 3 lipidation. The effect of mTOR blockade was evaluated by testing the impact of rapamycin treatment on Treg cell function. SLE Treg cells exhibited increased activities of mTORC1 and mTORC2, whereas autophagy, the expression of GATA-3 and CTLA-4, and the suppressor function of Treg cells were diminished. IL-21, but not IL-6 or IL-17, blocked the development of Treg cells. IL-21 stimulated mTORC1 and mTORC2, and it abrogated the autophagy, differentiation, and function of Treg cells. Moreover, IL-21 constrained the expression of GATA-3 and CTLA-4 selectively in Treg cells. In turn, blockade of mTORC1 by 3-day rapamycin treatment enhanced transforming growth factor β production, while dual blockade of mTORC1 and mTORC2 by 4-week rapamycin treatment induced autophagy, restored the expression of GATA-3 and CTLA-4, and corrected Treg cell function. IL-21-driven mTOR activation is a pharmacologically targetable checkpoint of the deficient autophagy that underlies Treg cell dysfunction in SLE. © 2017, American College of Rheumatology.

Genome-wide analysis of the regulatory function mediated by the small regulatory psm-mec RNA of methicillin-resistant Staphylococcus aureus.

PubMed

Cheung, Gordon Y C; Villaruz, Amer E; Joo, Hwang-Soo; Duong, Anthony C; Yeh, Anthony J; Nguyen, Thuan H; Sturdevant, Daniel E; Queck, S Y; Otto, M

2014-07-01

Several methicillin resistance (SCCmec) clusters characteristic of hospital-associated methicillin-resistant Staphylococcus aureus (MRSA) strains harbor the psm-mec locus. In addition to encoding the cytolysin, phenol-soluble modulin (PSM)-mec, this locus has been attributed gene regulatory functions. Here we employed genome-wide transcriptional profiling to define the regulatory function of the psm-mec locus. The immune evasion factor protein A emerged as the primary conserved and strongly regulated target of psm-mec, an effect we show is mediated by the psm-mec RNA. Furthermore, the psm-mec locus exerted regulatory effects that were more moderate in extent. For example, expression of PSM-mec limited expression of mecA, thereby decreasing methicillin resistance. Our study shows that the psm-mec locus has a rare dual regulatory RNA and encoded cytolysin function. Furthermore, our findings reveal a specific mechanism underscoring the recently emerging concept that S. aureus strains balance pronounced virulence and high expression of antibiotic resistance. Published by Elsevier GmbH.

Prolonged Neutrophil Dysfunction Following Plasmodium falciparum Malaria is Related to Hemolysis and Heme Oxygenase-1 Induction1

PubMed Central

Cunnington, Aubrey J.; Njie, Madi; Correa, Simon; Takem, Ebako N.; Riley, Eleanor M.; Walther, Michael

2012-01-01

It is not known why people are more susceptible to bacterial infections such as non-Typhoid Salmonella (NTS) during and after a malaria infection but, in mice, malarial hemolysis impairs resistance to NTS by impairing the neutrophil oxidative burst. This acquired neutrophil dysfunction is a consequence of induction of the cytoprotective, heme degrading enzyme heme oxygenase-1 (HO-1) in neutrophil progenitors in bone marrow. In this study, we assessed whether neutrophil dysfunction occurs in humans with malaria and how this relates to hemolysis. We evaluated neutrophil function in 58 Gambian children with Plasmodium falciparum malaria (55 (95%) with uncomplicated disease), and examined associations with erythrocyte count, haptoglobin, hemopexin, plasma heme, expression of receptors for heme uptake, and HO-1 induction. Malaria caused the appearance of a dominant population of neutrophils with reduced oxidative burst activity, which gradually normalized over 8 weeks of follow-up. The degree of neutrophil impairment correlated significantly with markers of hemolysis and HO-1 induction. HO-1 expression was increased in blood during acute malaria, but at a cellular level HO-1 expression was modulated by changes in surface expression of the haptoglobin receptor (CD163). These findings demonstrate that neutrophil dysfunction occurs in P. falciparum malaria and support the relevance of the mechanistic studies in mice. Furthermore, they suggest the presence of a regulatory pathway to limit HO-1 induction by hemolysis in the context of infection, and indicate new targets for therapeutic intervention to abrogate the susceptibility to bacterial infection in the context of hemolysis in humans. PMID:23100518

Proresolving lipid mediators resolvin D1, resolvin D2, and maresin 1 are critical in modulating T cell responses.

PubMed

Chiurchiù, Valerio; Leuti, Alessandro; Dalli, Jesmond; Jacobsson, Anders; Battistini, Luca; Maccarrone, Mauro; Serhan, Charles N

2016-08-24

Resolution of inflammation is a finely regulated process mediated by specialized proresolving lipid mediators (SPMs), including docosahexaenoic acid (DHA)-derived resolvins and maresins. The immunomodulatory role of SPMs in adaptive immune cells is of interest. We report that D-series resolvins (resolvin D1 and resolvin D2) and maresin 1 modulate adaptive immune responses in human peripheral blood lymphocytes. These lipid mediators reduce cytokine production by activated CD8(+) T cells and CD4(+) T helper 1 (TH1) and TH17 cells but do not modulate T cell inhibitory receptors or abrogate their capacity to proliferate. Moreover, these SPMs prevented naïve CD4(+) T cell differentiation into TH1 and TH17 by down-regulating their signature transcription factors, T-bet and Rorc, in a mechanism mediated by the GPR32 and ALX/FPR2 receptors; they concomitantly enhanced de novo generation and function of Foxp3(+) regulatory T (Treg) cells via the GPR32 receptor. These results were also supported in vivo in a mouse deficient for DHA synthesis (Elovl2(-/-)) that showed an increase in TH1/TH17 cells and a decrease in Treg cells compared to wild-type mice. Additionally, either DHA supplementation in Elovl2(-/-) mice or in vivo administration of resolvin D1 significantly reduced cytokine production upon specific stimulation of T cells. These findings demonstrate actions of specific SPMs on adaptive immunity and provide a new avenue for SPM-based approaches to modulate chronic inflammation. Copyright © 2016, American Association for the Advancement of Science.

Genetic deficiency of indoleamine 2,3-dioxygenase promotes gut microbiota-mediated metabolic health.

PubMed

Laurans, Ludivine; Venteclef, Nicolas; Haddad, Yacine; Chajadine, Mouna; Alzaid, Fawaz; Metghalchi, Sarvenaz; Sovran, Bruno; Denis, Raphael G P; Dairou, Julien; Cardellini, Marina; Moreno-Navarrete, Jose-Maria; Straub, Marjolene; Jegou, Sarah; McQuitty, Claire; Viel, Thomas; Esposito, Bruno; Tavitian, Bertrand; Callebert, Jacques; Luquet, Serge H; Federici, Massimo; Fernandez-Real, José Manuel; Burcelin, Remy; Launay, Jean-Marie; Tedgui, Alain; Mallat, Ziad; Sokol, Harry; Taleb, Soraya

2018-06-25

The association between altered gut microbiota, intestinal permeability, inflammation and cardiometabolic diseases is becoming increasingly clear but remains poorly understood 1,2 . Indoleamine 2,3-dioxygenase is an enzyme induced in many types of immune cells, including macrophages in response to inflammatory stimuli, and catalyzes the degradation of tryptophan along the kynurenine pathway. Indoleamine 2,3-dioxygenase activity is better known for its suppression of effector T cell immunity and its activation of regulatory T cells 3,4 . However, high indoleamine 2,3-dioxygenase activity predicts worse cardiovascular outcome 5-9 and may promote atherosclerosis and vascular inflammation 6 , suggesting a more complex role in chronic inflammatory settings. Indoleamine 2,3-dioxygenase activity is also increased in obesity 10-13 , yet its role in metabolic disease is still unexplored. Here, we show that obesity is associated with an increase of intestinal indoleamine 2,3-dioxygenase activity, which shifts tryptophan metabolism from indole derivative and interleukin-22 production toward kynurenine production. Indoleamine 2,3-dioxygenase deletion or inhibition improves insulin sensitivity, preserves the gut mucosal barrier, decreases endotoxemia and chronic inflammation, and regulates lipid metabolism in liver and adipose tissues. These beneficial effects are due to rewiring of tryptophan metabolism toward a microbiota-dependent production of interleukin-22 and are abrogated after treatment with a neutralizing anti-interleukin-22 antibody. In summary, we identify an unexpected function of indoleamine 2,3-dioxygenase in the fine tuning of intestinal tryptophan metabolism with major consequences on microbiota-dependent control of metabolic disease, which suggests indoleamine 2,3-dioxygenase as a potential therapeutic target.

Par-4-mediated recruitment of Amida to the actin cytoskeleton leads to the induction of apoptosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boosen, Meike; Vetterkind, Susanne; Koplin, Ansgar

Par-4 (prostate apoptosis response-4) sensitizes cells to apoptotic stimuli, but the exact mechanisms are still poorly understood. Using Par-4 as bait in a yeast two-hybrid screen, we identified Amida as a novel interaction partner, a ubiquitously expressed protein which has been suggested to be involved in apoptotic processes. Complex formation of Par-4 and Amida occurs in vitro and in vivo and is mediated via the C-termini of both proteins, involving the leucine zipper of Par-4. Amida resides mainly in the nucleus but displays nucleo-cytoplasmic shuttling in heterokaryons. Upon coexpression with Par-4 in REF52.2 cells, Amida translocates to the cytoplasm andmore » is recruited to actin filaments by Par-4, resulting in enhanced induction of apoptosis. The synergistic effect of Amida/Par-4 complexes on the induction of apoptosis is abrogated when either Amida/Par-4 complex formation or association of these complexes with the actin cytoskeleton is impaired, indicating that the Par-4-mediated relocation of Amida to the actin cytoskeleton is crucial for the pro-apoptotic function of Par-4/Amida complexes in REF52.2 cells. The latter results in enhanced phosphorylation of the regulatory light chain of myosin II (MLC) as has previously been shown for Par-4-mediated recruitment of DAP-like kinase (Dlk), suggesting that the recruitment of nuclear proteins involved in the regulation of apoptotic processes to the actin filament system by Par-4 represents a potent mechanism how Par-4 can trigger apoptosis.« less

[The Influence of the Functioning of Brain Regulatory Systems onto the Voluntary Regulation of Cognitive Performance in Children. Report 2. Neuropsychological and Electrophysiological Assessment of Brain Regulatory Functions in Children Aged 10-12 with Learning Difficulties].

PubMed

Semenova, O A; Machinskaya, R I

2015-01-01

A total number of 172 children aged 10-12 were electrophysiologically and neuropsychologically assessed in order to analyze the influence of the functioning of brain regulatory systems onto the voluntary regulation of cognitive performance during the preteen years. EEG patterns associated with the nonoptimal functioning of brain regulatory systems, particularly fronto-thalamic, limbic and fronto-striatal structures were significantly more often observed in children with learning and behavioral difficulties, as compared to the control group. Neuropsychological assessment showed that the nonoptimal functioning of different brain regulatory systems specifically affect the voluntary regulation of cognitive performance. Children with EEG patterns of fronto-thalamic nonoptimal functioning demonstrated poor voluntary regulation such as impulsiveness and difficulties in continuing the same algorithms. Children with EEG patterns of limbic nonoptimal functioning showed a less pronounced executive dysfunction manifested only in poor switching between program units within a task. Children with EEG patterns of fronto-striatal nonoptimal functioning struggled with such executive dysfunctions as motor and tactile perseverations and emotional-motivational deviations such as poor motivation and communicative skills.

The regulatory network analysis of long noncoding RNAs in human colorectal cancer.

PubMed

Zhang, Yuwei; Tao, Yang; Li, Yang; Zhao, Jinshun; Zhang, Lina; Zhang, Xiaohong; Dong, Changzheng; Xie, Yangyang; Dai, Xiaoyu; Zhang, Xinjun; Liao, Qi

2018-05-01

Colorectal cancer (CRC) is among one of the most prevalent and lethiferous diseases worldwide. Long noncoding RNAs (lncRNAs) are commonly accepted to function as a key regulatory factor in human cancer, but the potential regulatory mechanisms of CRC-associated lncRNA are largely obscure. Here, we integrated several expression profiles to obtain 55 differentially expressed (DE) lncRNAs. We first detected lncRNA interactions with transcription factors, microRNAs, mRNAs, and RNA-binding proteins to construct a regulatory network and then create functional enrichment analyses for them using bioinformatics approaches. We found the upregulated genes in the regulatory network are enriched in cell cycle and DNA damage response, while the downregulated genes are enriched in cell differentiation, cellular response, and cell signaling. We then employed module-based methods to mine several intriguing modules from the overall network, which helps to classify the functions of genes more specifically. Next, we confirmed the validity of our network by comparisons with a randomized network using computational method. Finally, we attempted to annotate lncRNA functions based on the regulatory network, which indicated its potential application. Our study of the lncRNA regulatory network provided significant clues to unveil lncRNAs potential regulatory mechanisms in CRC and laid a foundation for further experimental investigation.

MEF2 Cooperates With Forskolin/cAMP and GATA4 to Regulate Star Gene Expression in Mouse MA-10 Leydig Cells.

PubMed

Daems, Caroline; Di-Luoffo, Mickaël; Paradis, Élise; Tremblay, Jacques J

2015-07-01

In Leydig cells, steroidogenic acute regulatory protein (STAR) participates in cholesterol shuttling from the outer to the inner mitochondrial membrane, the rate-limiting step in steroidogenesis. Steroid hormone biosynthesis and steroidogenic gene expression are regulated by LH, which activates various signaling pathways and transcription factors, including cAMP/Ca(2+)/CAMK (Ca(2+)/calmodulin-dependent kinase)-myocyte enhancer factor 2 (MEF2). The 4 MEF2 transcription factors are essential regulators of cell differentiation and organogenesis in numerous tissues. Recently, MEF2 was identified in Sertoli and Leydig cells of the testis. Here, we report that MEF2 regulates steroidogenesis in mouse MA-10 Leydig cells by acting on the Star gene. In MA-10 cells depleted of MEF2 using siRNAs (small interfering RNAs), STAR protein levels, Star mRNA levels, and promoter activity were significantly decreased. On its own, MEF2 did not activate the mouse Star promoter but was found to cooperate with forskolin/cAMP. By chromatin immunoprecipitation and DNA precipitation assays, we confirmed MEF2 binding to a consensus element located at -232 bp of the Star promoter. Mutation or deletion of the MEF2 element reduced but did not abrogate the MEF2/cAMP cooperation, indicating that MEF2 cooperates with other DNA-bound transcription factor(s). We identified GATA4 (GATA binding protein 4) as a partner for MEF2 in Leydig cells, because mutation of the GATA element abrogated the MEF2/cAMP cooperation on a reporter lacking a MEF2 element. MEF2 and GATA4 interact as revealed by coimmunoprecipitation, and MEF2 and GATA4 transcriptionally cooperate on the Star promoter. Altogether, our results define MEF2 as a novel regulator of steroidogenesis and Star transcription in Leydig cells and identify GATA4 as a key partner for MEF2-mediated action.

A rapid, PPAR-gamma-dependent effect of pioglitazone on the phosphorylation of MYPT.

PubMed

Atkins, Kevin B; Irey, Brittany; Xiang, Nan; Brosius, Frank C

2009-05-01

Peroxisome proliferator-activated receptor (PPAR)-gamma ligands, thiazolidinediones, have been demonstrated to regulate vascular reactivity. We examined the effect of pioglitazone (PIO; 20 muM) in rat primary cultured aortic smooth muscle cells on constitutive phosphorylation of the regulatory subunit of myosin phosphatase (MYPT). PIO decreased the phosphorylation of Thr(697) on MYPT within 15 min, and the inhibition was maintained up to 6 h. The PPAR-gamma antagonist GW-9662 (5 microM) abrogated the inhibition of Thr(697) phosphorylation mediated by PIO. Because longer-term PIO treatment inhibits RhoA/Rho kinase (ROCK) signaling and Thr(697) phosphorylation, we tested the effect of the ROCK inhibitor Y-27632 (10 muM) on the inhibition of Thr(697) phosphorylation by PIO. Y-27632 alone inhibited Thr(697) phosphorylation, and there was an additive effect with PIO. In addition, up to 1 h of PIO treatment did not affect RhoA localization or decrease ROCK-dependent phosphorylation of Thr(855). These results suggest that the effect of PIO is independent of inhibition of RhoA/ROCK. PIO increased the phosphorylation of Ser(696) in the same time course as its effect on Thr(697). Ser(696) has been shown to be phosphorylated by PKA and PKG. PKA inhibitor H-89 (10 microM) and PKG inhibitor KT-5823 (0.5 microM) abrogated the effect of PIO on both Thr(697) and Ser(696) phosphorylation. The constitutive turnover of phosphorylation of Thr(697) is rapid, suggesting that the decreased phosphorylation of Thr(697) by PIO is due to enhanced phosphorylation of Ser(696). This is supported by the finding that PIO blocks ANG II-stimulated phosphorylation of Thr(697) but not ANG II-stimulated RhoA translocation. Therefore, the effect of shorter-term PIO apparently is to increase myosin light chain phosphatase activity, thereby desensitizing the vascular smooth muscle to agonist signaling.

Downregulation of HIF-1a sensitizes U251 glioma cells to the temozolomide (TMZ) treatment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tang, Jun-Hai; Ma, Zhi-Xiong; Huang, Guo-Hao

Purpose: The aim of this study was to investigate the effect of downregulation of HIF-1α gene on human U251 glioma cells and examine the consequent changes of TMZ induced effects and explore the molecular mechanisms. Methods: U251 cell line stably expressing HIF-1α shRNA was acquired via lentiviral vector transfection. The mRNA and protein expression alterations of genes involved in our study were determined respectively by qRT-PCR and Western blot. Cell proliferation was measured by MTT assay and colony formation assay, cell invasion/migration capacity was determined by transwell invasion assay/wound healing assay, and cell apoptosis was detected by flow cytometry. Results:more » We successfully established a U251 cell line with highly efficient HIF-1α knockdown. HIF-1a downregulation sensitized U251 cells to TMZ treatment and enhanced the proliferation-inhibiting, invasion/migration-suppressing, apoptosis-inducing and differentiation-promoting effects exerted by TMZ. The related molecular mechanisms demonstrated that expression of O{sup 6}-methylguanine DNA methyltransferase gene (MGMT) and genes of Notch1 pathway were significantly upregulated by TMZ treatment. However, this upregulation was abrogated by HIF-1α knockdown. We further confirmed important regulatory roles of HIF-1α in the expression of MGMT and activation of Notch1 pathways. Conclusion: HIF-1α downregulation sensitizes U251 glioma cells to the temozolomide treatment via inhibiting MGMT expression and Notch1 pathway activation. - Highlights: • TMZ caused more significant proliferation inhibition and apoptosis in U251 cells after downregulating HIF-1α. • Under TMZ treatment, HIF-1 downregulated U251 cells exhibited weaker mobility and more differentiated state. • TMZ caused MGMT over-expression and Notch1 pathway activation, which could be abrogated by HIF-1α downregulation.« less

Chrysin alleviates testicular dysfunction in adjuvant arthritic rats via suppression of inflammation and apoptosis: Comparison with celecoxib

DOE Office of Scientific and Technical Information (OSTI.GOV)

Darwish, Hebatallah A.; Arab, Hany H., E-mail: hany.arab@pharma.cu.edu.eg; Abdelsalam, Rania M.

Long standing rheumatoid arthritis (RA) is associated with testicular dysfunction and subfertility. Few studies have addressed the pathogenesis of testicular injury in RA and its modulation by effective agents. Thus, the current study aimed at evaluating the effects of two testosterone boosting agents; chrysin, a natural flavone and celecoxib, a selective COX-2 inhibitor, in testicular impairment in rats with adjuvant arthritis, an experimental model of RA. Chrysin (25 and 50 mg/kg) and celecoxib (5 mg/kg) were orally administered to Wistar rats once daily for 21 days starting 1 h before arthritis induction. Chrysin suppressed paw edema with comparable efficacy tomore » celecoxib. More important, chrysin, dose-dependently and celecoxib attenuated the testicular injury via reversing lowered gonadosomatic index and histopathologic alterations with preservation of spermatogenesis. Both agents upregulated steroidogenic acute regulatory (StAR) mRNA expression and serum testosterone with concomitant restoration of LH and FSH. Furthermore, they suppressed inflammation via abrogation of myeloperoxidase, TNF-α and protein expression of COX-2 and iNOS besides elevation of IL-10. Alleviation of the testicular impairment was accompanied with suppression of oxidative stress via lowering testicular lipid peroxides and nitric oxide. With respect to apoptosis, both agents downregulated FasL mRNA expression and caspase-3 activity in favor of cell survival. For the first time, these findings highlight the protective effects of chrysin and celecoxib against testicular dysfunction in experimental RA which were mediated via boosting testosterone in addition to attenuation of testicular inflammation, oxidative stress and apoptosis. Generally, the 50 mg/kg dose of chrysin exerted comparable protective actions to celecoxib. - Highlights: • Chrysin and celecoxib alleviated testicular suppression in adjuvant arthritis. • They attenuated histopathological damage and preserved spermatogenesis. • They upregulated StAR expression and testosterone with restoration of LH and FSH. • They abrogated myeloperoxidase, TNF-α and COX-2 and iNOS expression. • They suppressed oxidative stress and FasL-associated apoptosis.« less

Regulatory networks and connected components of the neutral space. A look at functional islands

NASA Astrophysics Data System (ADS)

Boldhaus, G.; Klemm, K.

2010-09-01

The functioning of a living cell is largely determined by the structure of its regulatory network, comprising non-linear interactions between regulatory genes. An important factor for the stability and evolvability of such regulatory systems is neutrality - typically a large number of alternative network structures give rise to the necessary dynamics. Here we study the discretized regulatory dynamics of the yeast cell cycle [Li et al., PNAS, 2004] and the set of networks capable of reproducing it, which we call functional. Among these, the empirical yeast wildtype network is close to optimal with respect to sparse wiring. Under point mutations, which establish or delete single interactions, the neutral space of functional networks is fragmented into ≈ 4.7 × 108 components. One of the smaller ones contains the wildtype network. On average, functional networks reachable from the wildtype by mutations are sparser, have higher noise resilience and fewer fixed point attractors as compared with networks outside of this wildtype component.

Regulatory T cells in human disease and their potential for therapeutic manipulation

PubMed Central

Taams, Leonie S; Palmer, Donald B; Akbar, Arne N; Robinson, Douglas S; Brown, Zarin; Hawrylowicz, Catherine M

2006-01-01

Regulatory T cells are proposed to play a central role in the maintenance of immunological tolerance in the periphery, and studies in many animal models demonstrate their capacity to inhibit inflammatory pathologies in vivo. At a recent meeting [Clinical Application of Regulatory T Cells, 7–8 April 2005, Horsham, UK, organized by the authors of this review, in collaboration with the British Society for Immunology and Novartis] evidence was discussed that certain human autoimmune, infectious and allergic diseases are associated with impaired regulatory T-cell function. In contrast, evidence from several human cancer studies and some infections indicates that regulatory T cells may impair the development of protective immunity. Importantly, certain therapies, both those that act non-specifically to reduce inflammation and antigen-specific immunotherapies, may induce or enhance regulatory T-cell function. The purpose of this review was to summarize current knowledge on regulatory T-cell function in human disease, and to assess critically how this can be tailored to suit the therapeutic manipulation of immunity. PMID:16630018

Regulatory states in the developmental control of gene expression.

PubMed

Peter, Isabelle S

2017-09-01

A growing body of evidence shows that gene expression in multicellular organisms is controlled by the combinatorial function of multiple transcription factors. This indicates that not the individual transcription factors or signaling molecules, but the combination of expressed regulatory molecules, the regulatory state, should be viewed as the functional unit in gene regulation. Here, I discuss the concept of the regulatory state and its proposed role in the genome-wide control of gene expression. Recent analyses of regulatory gene expression in sea urchin embryos have been instrumental for solving the genomic control of cell fate specification in this system. Some of the approaches that were used to determine the expression of regulatory states during sea urchin embryogenesis are reviewed. Significant developmental changes in regulatory state expression leading to the distinct specification of cell fates are regulated by gene regulatory network circuits. How these regulatory state transitions are encoded in the genome is illuminated using the sea urchin endoderm-mesoderms cell fate decision circuit as an example. These observations highlight the importance of considering developmental gene regulation, and the function of individual transcription factors, in the context of regulatory states. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Dynamics and function of distal regulatory elements during neurogenesis and neuroplasticity

PubMed Central

Thakurela, Sudhir; Sahu, Sanjeeb Kumar; Garding, Angela; Tiwari, Vijay K.

2015-01-01

Gene regulation in mammals involves a complex interplay between promoters and distal regulatory elements that function in concert to drive precise spatiotemporal gene expression programs. However, the dynamics of the distal gene regulatory landscape and its function in the transcriptional reprogramming that underlies neurogenesis and neuronal activity remain largely unknown. Here, we performed a combinatorial analysis of genome-wide data sets for chromatin accessibility (FAIRE-seq) and the enhancer mark H3K27ac, revealing the highly dynamic nature of distal gene regulation during neurogenesis, which gets progressively restricted to distinct genomic regions as neurons acquire a post-mitotic, terminally differentiated state. We further find that the distal accessible and active regions serve as target sites for distinct transcription factors that function in a stage-specific manner to contribute to the transcriptional program underlying neuronal commitment and maturation. Mature neurons respond to a sustained activity of NMDA receptors by epigenetic reprogramming at a large number of distal regulatory regions as well as dramatic reorganization of super-enhancers. Such massive remodeling of the distal regulatory landscape in turn results in a transcriptome that confers a transient loss of neuronal identity and gain of cellular plasticity. Furthermore, NMDA receptor activity also induces many novel prosurvival genes that function in neuroprotective pathways. Taken together, these findings reveal the dynamics of the distal regulatory landscape during neurogenesis and uncover novel regulatory elements that function in concert with epigenetic mechanisms and transcription factors to generate the transcriptome underlying neuronal development and activity. PMID:26170447

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

Arabidopsis ensemble reverse-engineered gene regulatory network discloses interconnected transcription factors in oxidative stress.

PubMed

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

2014-12-01

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. © 2014 American Society of Plant Biologists. All rights reserved.

The Association between Infants' Self-Regulatory Behavior and MAOA Gene Polymorphism

ERIC Educational Resources Information Center

Zhang, Minghao; Chen, Xinyin; Way, Niobe; Yoshikawa, Hirokazu; Deng, Huihua; Ke, Xiaoyan; Yu, Weiwei; Chen, Ping; He, Chuan; Chi, Xia; Lu, Zuhong

2011-01-01

Self-regulatory behavior in early childhood is an important characteristic that has considerable implications for the development of adaptive and maladaptive functioning. The present study investigated the relations between a functional polymorphism in the upstream region of monoamine oxidase A gene (MAOA) and self-regulatory behavior in a sample…

Galectin-9-CD44 interaction enhances stability and function of adaptive regulatory T cells | Center for Cancer Research

Cancer.gov

The β-galactoside-binding protein galectin-9 is critical in regulating the immune response, but the mechanism by which it functions remains unclear. We have demonstrated that galectin-9 is highly expressed by induced regulatory T cells (iTreg) and was crucial for the generation and function of iTreg cells, but not natural regulatory T (nTreg) cells. Galectin-9 expression

The Drosophila TGF-beta/Activin-like ligands Dawdle and Myoglianin appear to modulate adult lifespan through regulation of 26S proteasome function in adult muscle

PubMed Central

Langerak, Shaughna; Kim, Myung-Jun; Lamberg, Hannah; Godinez, Michael; Main, Mackenzie; Winslow, Lindsey; O'Connor, Michael B.

2018-01-01

ABSTRACT The Drosophila Activin signaling pathway employs at least three separate ligands – Activin-β (Actβ), Dawdle (Daw), and Myoglianin (Myo) – to regulate several general aspects of fruit fly larval development, including cell proliferation, neuronal remodeling, and metabolism. Here we provide experimental evidence indicating that both Daw and Myo are anti-ageing factors in adult fruit flies. Knockdown of Myo or Daw in adult fruit flies reduced mean lifespan, while overexpression of either ligand in adult muscle tissues but not in adipose tissues enhanced mean lifespan. An examination of ubiquitinated protein aggregates in adult muscles revealed a strong inverse correlation between Myo- or Daw-initiated Activin signaling and the amount of ubiquitinated protein aggregates. We show that this correlation has important functional implications by demonstrating that the lifespan extension effect caused by overexpression of wild-type Daw or Myo in adult muscle tissues can be completely abrogated by knockdown of a 26S proteasome regulatory subunit Rpn1 in adult fly muscle, and that the prolonged lifespan caused by overexpression of Daw or Myo in adult muscle could be due to enhanced protein levels of the key subunits of 26S proteasome. Overall, our data suggest that Activin signaling initiated by Myo and Daw in adult Drosophila muscles influences lifespan, in part, by modulation of protein homeostasis through either direct or indirect regulation of the 26S proteasome levels. Since Myo is closely related to the vertebrate muscle mass regulator Myostatin (GDF8) and the Myostatin paralog GDF11, our observations may offer a new experimental model for probing the roles of GDF11/8 in ageing regulation in vertebrates. This article has an associated First Person interview with the first author of the paper. PMID:29615416

CK2 phosphorylates and inhibits TAp73 tumor suppressor function to promote expression of cancer stem cell genes and phenotype in head and neck cancer.

PubMed

Lu, Hai; Yan, Carol; Quan, Xin Xin; Yang, Xinping; Zhang, Jialing; Bian, Yansong; Chen, Zhong; Van Waes, Carter

2014-10-01

Cancer stem cells (CSC) and genes have been linked to cancer development and therapeutic resistance, but the signaling mechanisms regulating CSC genes and phenotype are incompletely understood. CK2 has emerged as a key signal serine/threonine kinase that modulates diverse signal cascades regulating cell fate and growth. We previously showed that CK2 is often aberrantly expressed and activated in head and neck squamous cell carcinomas (HNSCC), concomitantly with mutant (mt) tumor suppressor TP53, and inactivation of its family member, TAp73. Unexpectedly, we observed that classical stem cell genes Nanog, Sox2, and Oct4, are overexpressed in HNSCC with inactivated TAp73 and mtTP53. However, the potential relationship between CK2, TAp73 inactivation, and CSC phenotype is unknown. We reveal that inhibition of CK2 by pharmacologic inhibitors or siRNA inhibits the expression of CSC genes and side population (SP), while enhancing TAp73 mRNA and protein expression. Conversely, CK2 inhibitor attenuation of CSC protein expression and the SP by was abrogated by TAp73 siRNA. Bioinformatic analysis uncovered a single predicted CK2 threonine phosphorylation site (T27) within the N-terminal transactivation domain of TAp73. Nuclear CK2 and TAp73 interaction, confirmed by co-immunoprecipitation, was attenuated by CK2 inhibitor, or a T27A point-mutation of this predicted CK2 threonine phospho-acceptor site of TAp73. Further, T27A mutation attenuated phosphorylation, while enhancing TAp73 function in repressing CSC gene expression and SP cells. A new CK2 inhibitor, CX-4945, inhibited CSC related SP cells, clonogenic survival, and spheroid formation. Our study unveils a novel regulatory mechanism whereby aberrant CK2 signaling inhibits TAp73 to promote the expression of CSC genes and phenotype.

Signal Transducer and Activator of Transcription 1 Plays a Pivotal Role in RET/PTC3 Oncogene-induced Expression of Indoleamine 2,3-Dioxygenase 1.

PubMed

Moretti, Sonia; Menicali, Elisa; Nucci, Nicole; Voce, Pasquale; Colella, Renato; Melillo, Rosa Marina; Liotti, Federica; Morelli, Silvia; Fallarino, Francesca; Macchiarulo, Antonio; Santoro, Massimo; Avenia, Nicola; Puxeddu, Efisio

2017-02-03

Indoleamine 2,3-dioxygenase 1 (IDO1) is a single chain oxidoreductase that catalyzes tryptophan degradation to kynurenine. In cancer, it exerts an immunosuppressive function as part of an acquired mechanism of immune escape. Recently, we demonstrated that IDO1 expression is significantly higher in all thyroid cancer histotypes compared with normal thyroid and that its expression levels correlate with T regulatory (Treg) lymphocyte densities in the tumor microenvironment. BRAF V600E - and RET/PTC3-expressing PcCL3 cells were used as cellular models for the evaluation of IDO1 expression in thyroid carcinoma cells and for the study of involved signal transduction pathways. BRAF V600E -expressing PcCL3 cells did not show IDO1 expression. Conversely, RET/PTC3-expressing cells were characterized by a high IDO1 expression. Moreover, we found that, the STAT1-IRF1 pathway was instrumental for IDO1 expression in RET/PTC3 expressing cells. In detail, RET/PTC3 induced STAT1 overexpression and phosphorylation at Ser-727 and Tyr-701. STAT1 transcriptional regulation appeared to require activation of the canonical NF-κB pathway. Conversely, activation of the MAPK and PI3K-AKT pathways primarily regulated Ser-727 phosphorylation, whereas a physical interaction between RET/PTC3 and STAT1, followed by a direct tyrosine phosphorylation event, was necessary for STAT1 Tyr-701 phosphorylation. These data provide the first evidence of a direct link between IDO1 expression and the oncogenic activation of RET in thyroid carcinoma and describe the involved signal transduction pathways. Moreover, they suggest possible novel molecular targets for the abrogation of tumor microenvironment immunosuppression. The detection of those targets is becoming increasingly important to yield the full function of novel immune checkpoint inhibitors. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

nAChRs-ERK1/2-Egr-1 signaling participates in the developmental toxicity of nicotine by epigenetically down-regulating placental 11β-HSD2.

PubMed

Zhou, Jin; Liu, Fulin; Yu, Luting; Xu, Dan; Li, Bin; Zhang, Guohui; Huang, Wen; Li, Lu; Zhang, Yuanzhen; Zhang, Wei; Wang, Hui

2018-04-01

Impaired placental 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) activity which inactivates maternal glucocorticoids is associated with poor fetal growth and a higher risk of chronic diseases in adulthood. This study aimed to elucidate the epigenetically regulatory mechanism of nicotine on placental 11β-HSD2 expression. Pregnant Wistar rats were administered 1.0 mg/kg nicotine subcutaneously twice a day from gestational day 9 to 20. The results showed that prenatal nicotine exposure increased corticosterone levels in the placenta and fetal serum, disrupted placental morphology and endocrine function, and reduced fetal bodyweight. Meanwhile, histone modification abnormalities (decreased acetylation and increased di-methylation of histone 3 Lysine 9) on the HSD11B2 promoter and lower-expression of 11β-HSD2 were observed. Furthermore, the expression of nicotinic acetylcholine receptor (nAChR) α4/β2, the phosphorylation of extracellular regulated kinase 1/2 (ERK1/2) and Ets-like protein-1 (Elk-1), and the expression of early growth response-1 (Egr-1) were increased in the nicotine groups. In human BeWo cells, nicotine decreased 11β-HSD2 expression, increased nAChRα9 expression, and activated ERK1/2/Elk-1/Egr-1 signaling in the concentration (0.1-10 μM)-dependent manner. Antagonism of nAChRs, inhibition of ERK1/2 and Egr-1 knockdown by siRNA were able to block/abrogate the effects of nicotine on histone modification and expression of 11β-HSD2. Taken together, nicotine can impair placental structure and function, and induce fetal developmental toxicity. The underlying mechanism involves histone modifications and down-regulation of 11β-HSD2 through nAChRs/ERK1/2/Elk-1/Egr-1 signaling, which increases active glucocorticoids levels in the placenta and fetus, and eventually inhibits the fetal development. Copyright © 2018 Elsevier Inc. All rights reserved.

The dual role of poly(ADP-ribose) polymerase-1 in modulating parthanatos and autophagy under oxidative stress in rat cochlear marginal cells of the stria vascularis.

PubMed

Jiang, Hong-Yan; Yang, Yang; Zhang, Yuan-Yuan; Xie, Zhen; Zhao, Xue-Yan; Sun, Yu; Kong, Wei-Jia

2018-04-01

Oxidative stress is reported to regulate several apoptotic and necrotic cell death pathways in auditory tissues. Poly(ADP-ribose) polymerase-1 (PARP-1) can be activated under oxidative stress, which is the hallmark of parthanatos. Autophagy, which serves either a pro-survival or pro-death function, can also be stimulated by oxidative stress, but the role of autophagy and its relationship with parthanatos underlying this activation in the inner ear remains unknown. In this study, we established an oxidative stress model in vitro by glucose oxidase/glucose (GO/G), which could continuously generate low concentrations of H 2 O 2 to mimic continuous exposure to H 2 O 2 in physiological conditions, for investigation of oxidative stress-induced cell death mechanisms and the regulatory role of PARP-1 in this process. We observed that GO/G induced stria marginal cells (MCs) death via upregulation of PARP-1 expression, accumulation of polyADP-ribose (PAR) polymers, decline of mitochondrial membrane potential (MMP) and nuclear translocation of apoptosis-inducing factor (AIF), which all are biochemical features of parthanatos. PARP-1 knockdown rescued GO/G-induced MCs death, as well as abrogated downstream molecular events of PARP-1 activation. In addition, we demonstrated that GO/G stimulated autophagy and PARP-1 knockdown suppressed GO/G-induced autophagy in MCs. Interestingly, autophagy suppression by 3-Methyladenine (3-MA) accelerated GO/G-induced parthanatos, indicating a pro-survival function of autophagy in GO/G-induced MCs death. Taken together, these data suggested that PARP-1 played dual roles by modulating parthanatos and autophagy in oxidative stress-induced MCs death, which may be considered as a promising therapeutic target for ameliorating oxidative stress-related hearing disorders. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

TIRAP, an Adaptor Protein for TLR2/4, Transduces a Signal from RAGE Phosphorylated upon Ligand Binding

PubMed Central

Sakaguchi, Masakiyo; Murata, Hitoshi; Yamamoto, Ken-ichi; Ono, Tomoyuki; Sakaguchi, Yoshihiko; Motoyama, Akira; Hibino, Toshihiko; Kataoka, Ken; Huh, Nam-ho

2011-01-01

The receptor for advanced glycation end products (RAGE) is thought to be involved in the pathogenesis of a broad range of inflammatory, degenerative and hyperproliferative diseases. It binds to diverse ligands and activates multiple intracellular signaling pathways. Despite these pivotal functions, molecular events just downstream of ligand-activated RAGE have been surprisingly unknown. Here we show that the cytoplasmic domain of RAGE is phosphorylated at Ser391 by PKCζ upon binding of ligands. TIRAP and MyD88, which are known to be adaptor proteins for Toll-like receptor-2 and -4 (TLR2/4), bound to the phosphorylated RAGE and transduced a signal to downstream molecules. Blocking of the function of TIRAP and MyD88 largely abrogated intracellular signaling from ligand-activated RAGE. Our findings indicate that functional interaction between RAGE and TLRs coordinately regulates inflammation, immune response and other cellular functions. PMID:21829704

Functional role of EMMPRIN in the formation and mineralisation of dental matrix in mouse molars.

PubMed

Xie, Ming; Xing, Guofang; Hou, Liwen; Bao, Jing; Chen, Yuqing; Jiao, Ting; Zhang, Fuqiang

2015-02-01

Our previous research has shown that the extracellular matrix metalloproteinase inducer (EMMPRIN) is expressed during and may function in the early development of tooth germs. In the present study, we observed the specific expression of EMMPRIN in ameloblasts and odontoblasts during the middle and late stages of tooth germ development using immunohistochemistry. Furthermore, to extend our understanding of the function of EMMPRIN in odontogenesis, we used an anti-EMMPRIN function-blocking antibody to remove EMMPRIN activity in tooth germ culture in vitro. Both the formation and mineralisation of dental hard tissues were suppressed in the tooth germ culture after the abrogation of EMMPRIN. Meanwhile, significant reductions in VEGF, MMP-9, ALPL, ameloblastin, amelogenin and enamelin expression were observed in antibody-treated tooth germ explants compared to control and normal serum-treated explants. The current results illustrate that EMMPRIN may play a critical role in the processing and maturation of the dental matrix.

Ipilimumab (Yervoy) and the TGN1412 catastrophe.

PubMed

Bakacs, Tibor; Mehrishi, Jitendra N; Moss, Ralph W

2012-06-01

The development of the anti-CTLA-4 antibody (ipilimumab; marketed as Yervoy) immune regulatory therapy was based on the premise that "Abrogation of the function of CTLA-4 would permit CD28 to function unopposed and might swing the balance in favor of immune stimulation, tolerance breakdown and tumor eradication…" (Weber, 2009). By now, the vast majority of data collected from more than 4000 patients proves that this prediction was entirely correct. Paradoxically, the successful blockade of immune checkpoints raises the question whether an anti-CTLA-4 antibody could ever become an important therapy against cancer. T cells lost their ability to discriminate between self and non-self. Thus, tolerance to self tissues was broken in ∼70% of the patients. In the recent industry-sponsored phase III clinical trial of ipilimumab, 147 (38.7%) of the patients experienced severe adverse events and 6.8% suffered dose-limiting events (8.4%, in the ipilimumab-alone group). There were 14 deaths related to the study drugs and 7 of these were associated with immune-related adverse events. In contrast, the complete response rate was only 0.2%, in one patient out of 403 who received ipilimumab plus a peptide vaccine. Promoters of ipilimumab appear to be unmindful of the clinical trial catastrophe in London. Then, a humanized "superagonist" anti-CD28 monoclonal antibody, TGN1412, which "preferentially" activated regulatory T cells, at a higher dose, also activated all CD28 positive T cells. This precipitated a "cytokine storm" leading to life-threatening multiple organ failure in the six healthy human volunteers. Neither anti-CD28 nor anti-CTLA-4 therapies rely on antigen-specificity. Both release free antibody into the body against common molecular targets that are expressed on the targeted as well as on the non-targeted T cells. At lower antibody doses specific T cells are preferentially activated. With increasing antibody dose, however, the kinetics of the interaction is pushed in favor of widespread non-specific T cell expansion. Using the law of mass action we calculated that the vast majority of the CTLA-4 receptors on all activated T cells (including melanoma specific T cells) in the phase III clinical trial of ipilimumab will have been saturated. This would explain the runaway immune response observed. The conclusions drawn by the authors of the ipilimumab trial paper could bear an independent inspection and reassessment concerning the validation of the blockade of immune checkpoints as an important therapeutic strategy against cancer. Copyright © 2011 Elsevier GmbH. All rights reserved.

In silico evolution of the Drosophila gap gene regulatory sequence under elevated mutational pressure.

PubMed

Chertkova, Aleksandra A; Schiffman, Joshua S; Nuzhdin, Sergey V; Kozlov, Konstantin N; Samsonova, Maria G; Gursky, Vitaly V

2017-02-07

Cis-regulatory sequences are often composed of many low-affinity transcription factor binding sites (TFBSs). Determining the evolutionary and functional importance of regulatory sequence composition is impeded without a detailed knowledge of the genotype-phenotype map. We simulate the evolution of regulatory sequences involved in Drosophila melanogaster embryo segmentation during early development. Natural selection evaluates gene expression dynamics produced by a computational model of the developmental network. We observe a dramatic decrease in the total number of transcription factor binding sites through the course of evolution. Despite a decrease in average sequence binding energies through time, the regulatory sequences tend towards organisations containing increased high affinity transcription factor binding sites. Additionally, the binding energies of separate sequence segments demonstrate ubiquitous mutual correlations through time. Fewer than 10% of initial TFBSs are maintained throughout the entire simulation, deemed 'core' sites. These sites have increased functional importance as assessed under wild-type conditions and their binding energy distributions are highly conserved. Furthermore, TFBSs within close proximity of core sites exhibit increased longevity, reflecting functional regulatory interactions with core sites. In response to elevated mutational pressure, evolution tends to sample regulatory sequence organisations with fewer, albeit on average, stronger functional transcription factor binding sites. These organisations are also shaped by the regulatory interactions among core binding sites with sites in their local vicinity.

A Tyrosine Residue on the TSH Receptor Stabilizes Multimer Formation

PubMed Central

Latif, Rauf; Michalek, Krzysztof; Morshed, Syed Ahmed; Davies, Terry F.

2010-01-01

Background The thyrotropin stimulating hormone receptor (TSHR) is a G protein coupled receptor (GPCR) with a large ectodomain. The ligand, TSH, acting via this receptor regulates thyroid growth and thyroid hormone production and secretion. The TSH receptor (TSHR) undergoes complex post –translational modifications including intramolecular cleavage and receptor multimerization. Since monomeric and multimeric receptors coexist in cells, understanding the functional role of just the TSHR multimers is difficult. Therefore, to help understand the physiological significance of receptor multimerization, it will be necessary to abrogate multimer formation, which requires identifying the ectodomain and endodomain interaction sites on the TSHR. Here, we have examined the contribution of the ectodomain to constitutive multimerization of the TSHR and determined the possible residue(s) that may be involved in this interaction. Methodology/Principal Findings We studied ectodomain multimer formation by expressing the extracellular domain of the TSHR linked to a glycophosphotidyl (GPI) anchor in both stable and transient expression systems. Using co-immunoprecipitation and FRET of tagged receptors, we established that the TSH receptor ectodomain was capable of multimerization even when totally devoid of the transmembrane domain. Further, we studied the effect of two residues that likely made critical contact points in this interaction. We showed that a conserved tyrosine residue (Y116) on the convex surface of the LRR3 was a critical residue in ectodomain multimer formation since mutation of this residue to serine totally abrogated ectodomain multimers. This abrogation was not seen with the mutation of cysteine 176 on the inner side of the LRR5, demonstrating that inter-receptor disulfide bonding was not involved in ectodomain multimer formation. Additionally, the Y116 mutation in the intact wild type receptor enhanced receptor degradation. Conclusions/Significance These data establish the TSH receptor ectodomain as one site of multimerization, independent of the transmembrane region, and that this interaction was primarily via a conserved tyrosine residue in LRR3. PMID:20195479

Quorum-sensing inhibition abrogates the deleterious impact of Pseudomonas aeruginosa on airway epithelial repair.

PubMed

Ruffin, Manon; Bilodeau, Claudia; Maillé, Émilie; LaFayette, Shantelle L; McKay, Geoffrey A; Trinh, Nguyen Thu Ngan; Beaudoin, Trevor; Desrosiers, Martin-Yvon; Rousseau, Simon; Nguyen, Dao; Brochiero, Emmanuelle

2016-09-01

Chronic Pseudomonas aeruginosa lung infections are associated with progressive epithelial damage and lung function decline. In addition to its role in tissue injury, the persistent presence of P. aeruginosa-secreted products may also affect epithelial repair ability, raising the need for new antivirulence therapies. The purpose of our study was to better understand the outcomes of P. aeruginosa exoproducts exposure on airway epithelial repair processes to identify a strategy to counteract their deleterious effect. We found that P. aeruginosa exoproducts significantly decreased wound healing, migration, and proliferation rates, and impaired the ability of directional migration of primary non-cystic fibrosis (CF) human airway epithelial cells. Impact of exoproducts was inhibited after mutations in P. aeruginosa genes that encoded for the quorum-sensing (QS) transcriptional regulator, LasR, and the elastase, LasB, whereas impact was restored by LasB induction in ΔlasR mutants. P. aeruginosa purified elastase also induced a significant decrease in non-CF epithelial repair, whereas protease inhibition with phosphoramidon prevented the effect of P. aeruginosa exoproducts. Furthermore, treatment of P. aeruginosa cultures with 4-hydroxy-2,5-dimethyl-3(2H)-furanone, a QS inhibitor, abrogated the negative impact of P. aeruginosa exoproducts on airway epithelial repair. Finally, we confirmed our findings in human airway epithelial cells from patients with CF, a disease featuring P. aeruginosa chronic respiratory infection. These data demonstrate that secreted proteases under the control of the LasR QS system impair airway epithelial repair and that QS inhibitors could be of benefit to counteract the deleterious effect of P. aeruginosa in infected patients.-Ruffin, M., Bilodeau, C., Maillé, É., LaFayette, S. L., McKay, G. A., Trinh, N. T. N., Beaudoin, T., Desrosiers, M.-Y., Rousseau, S., Nguyen, D., Brochiero, E. Quorum-sensing inhibition abrogates the deleterious impact of Pseudomonas aeruginosa on airway epithelial repair. © FASEB.

GARP: a surface molecule of regulatory T cells that is involved in the regulatory function and TGF-β releasing.

PubMed

Sun, Liping; Jin, Hao; Li, Hui

2016-07-05

There are many molecules that define regulatory T cells (Tregs) phenotypically and functionally. Glycoprotein A repetitions predominant (GARP) is a transmembrane protein containing leucine rich repeats. Recently, GARP is found to express highly on the surface of activated Tregs. The combination of GARP and other surface molecules isolates Tregs with higher purity. Besides, GARP is a cell surface molecule of Tregs that maintains their regulatory function and homeosatsis. GARP has also been proved to promote the activation and secretion of transforming growth factor β (TGF-β). Moreover, its potential value in cancer immunotherapy is also discussed in this work.

Docosahexaneoic acid (22:6,n-3) regulates rat hepatocyte SREBP-1 nuclear abundance by Erk- and 26S proteasome-dependent pathways

PubMed Central

Botolin, Daniela; Wang, Yun; Christian, Barbara; Jump, Donald B.

2009-01-01

Insulin induces and dietary n-3 PUFAs suppress hepatic de novo lipogenesis by controlling sterol-regulatory element binding protein-1 nuclear abundance (nSREBP-1). Our goal was to define the mechanisms involved in this regulatory process. Insulin treatment of rat primary hepatocytes rapidly augments nSREBP-1 and mRNASREBP-1c while suppressing mRNAInsig-2 but not mRNAInsig-1. These events are preceded by rapid but transient increases in Akt and Erk phosphorylation. Removal of insulin from hepatocytes leads to a rapid decline in nSREBP-1 [half-time (T1/2) ~ 10 h] that is abrogated by inhibitors of 26S proteasomal degradation. 22:6,n-3, the major n-3 PUFA accumulating in livers of fish oil-fed rats, suppresses hepatocyte levels of nSREBP-1, mRNASREBP-1c, and mRNAInsig-2 but modestly and transiently induces mRNAInsig-1. More importantly, 22:6,n-3 accelerates the disappearance of hepatocyte nSREBP-1 (T1/2 ~ 4 h) through a 26S proteasome-dependent process. 22:6,n-3 has minimal effects on microsomal SREBP-1 and sterol-regulatory element binding protein cleavage-activating protein or nuclear SREBP-2. 22:6,n-3 transiently inhibits insulin-induced Akt phosphorylation but induces Erk phosphorylation. Inhibitors of Erk phosphorylation, but not overexpressed constitutively active Akt, rapidly attenuate 22:6,n-3 suppression of nSREBP-1. Thus, 22:6,n-3 suppresses hepatocyte nSREBP-1 through 26S proteasome- and Erk-dependent pathways. These studies reveal a novel mechanism for n-3 PUFA regulation of hepatocyte nSREBP-1 and lipid metabolism.—Botolin, D., Y. Wang, B. Christian, and D. B. Jump. Docosahexaneoic acid (22:6,n-3) regulates rat hepatocyte SREBP-1 nuclear abundance by Erk- and 26S proteasome-dependent pathways. PMID:16222032

Gut Microbiota-Derived Tryptophan Metabolites Modulate Inflammatory Response in Hepatocytes and Macrophages.

PubMed

Krishnan, Smitha; Ding, Yufang; Saedi, Nima; Choi, Maria; Sridharan, Gautham V; Sherr, David H; Yarmush, Martin L; Alaniz, Robert C; Jayaraman, Arul; Lee, Kyongbum

2018-04-24

The gut microbiota plays a significant role in the progression of fatty liver disease; however, the mediators and their mechanisms remain to be elucidated. Comparing metabolite profile differences between germ-free and conventionally raised mice against differences between mice fed a low- and high-fat diet (HFD), we identified tryptamine and indole-3-acetate (I3A) as metabolites that depend on the microbiota and are depleted under a HFD. Both metabolites reduced fatty-acid- and LPS-stimulated production of pro-inflammatory cytokines in macrophages and inhibited the migration of cells toward a chemokine, with I3A exhibiting greater potency. In hepatocytes, I3A attenuated inflammatory responses under lipid loading and reduced the expression of fatty acid synthase and sterol regulatory element-binding protein-1c. These effects were abrogated in the presence of an aryl-hydrocarbon receptor (AhR) antagonist, indicating that the effects are AhR dependent. Our results suggest that gut microbiota could influence inflammatory responses in the liver through metabolites engaging host receptors. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

DFR1-Mediated Inhibition of Proline Degradation Pathway Regulates Drought and Freezing Tolerance in Arabidopsis.

PubMed

Ren, Yongbing; Miao, Min; Meng, Yun; Cao, Jiasheng; Fan, Tingting; Yue, Junyang; Xiao, Fangming; Liu, Yongsheng; Cao, Shuqing

2018-06-26

Proline accumulation is one of the most important adaptation mechanisms for plants to cope with environmental stresses, such as drought and freezing. However, the molecular mechanism of proline homeostasis under these stresses is largely unknown. Here, we identified a mitochondrial protein, DFR1, involved in the inhibition of proline degradation in Arabidopsis. DFR1 was strongly induced by drought and cold stresses. The dfr1 knockdown mutants showed hypersensitivity to drought and freezing stresses, whereas the DFR1 overexpression plants exhibited enhanced tolerance, which was positively correlated with proline levels. DFR1 interacts with proline degradation enzymes PDH1/2 and P5CDH and compromises their activities. Genetic analysis showed that DFR1 acts upstream of PDH1/2 and P5CDH to positively regulate proline accumulation. Our results demonstrate a regulatory mechanism by which, under drought and freezing stresses, DFR1 interacts with PDH1/2 and P5CDH to abrogate their activities to maintain proline homeostasis, thereby conferring drought and freezing tolerance. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Loss of immunological tolerance in Gimap5-deficient mice is associated with loss of Foxo in CD4+ T cells

PubMed Central

Aksoylar, H. Ibrahim; Lampe, Kristin; Barnes, Michael J.; Plas, David R.; Hoebe, Kasper

2011-01-01

Previously, we reported the abrogation of quiescence and reduced survival in lymphocytes from Gimap5sph/sph mice, an ENU germline mutant with a missense mutation in the GTPase of immunity-associated nucleotide binding protein 5 (Gimap5). These mice showed a progressive loss of peripheral lymphocyte populations and developed spontaneous colitis, resulting in early mortality. Here, we identify the molecular pathways that contribute to the onset of colitis in Gimap5sph/sph mice. We show that CD4+ T cells become Th1/Th17-polarized and are critically important for the development of colitis. Concomitantly, Treg cells become reduced in frequency in the peripheral tissues and their immune-suppressive capacity becomes impaired. Most importantly, these progressive changes in CD4+ T cells are associated with the loss of Foxo1, Foxo3 and Foxo4 expression. Our data establish a novel link between Gimap5 and Foxo expression and provide evidence for a regulatory mechanism that controls Foxo protein expression and may help maintain immunological tolerance. PMID:22106000

Red wine consumption may affect sperm biology: the effects of different concentrations of the phytoestrogen myricetin on human male gamete function.

PubMed

Aquila, Saveria; Santoro, Marta; De Amicis, Francesca; Guido, Carmela; Bonofiglio, Daniela; Lanzino, Marilena; Cesario, Maria Grazia; Perrotta, Ida; Sisci, Diego; Morelli, Catia

2013-02-01

Myricetin is a natural flavonoid, particularly enriched in red wines, whose occurrence is widespread among plants. Despite extensive research, the beneficial effects of Myricetin on human health are still controversial. Here, we tested the estrogen-like effect of the phytoestrogen Myricetin on human ejaculated sperm biology. To this aim, human normozoospermic samples were exposed to increasing concentrations (10 nM, 100 nM, and 1 µM) of Myricetin. Motility, viability, capacitation-associated biochemical changes (i.e., cholesterol efflux and tyrosine phosphorylation), acrosin activity, as well as glucose utilization and fatty-acid oxidation (i.e., glucose and lipid metabolism) were all significantly increased by low doses of Myricetin. Importantly, both estrogen receptors α and β (ERs) and phosphatidylinositol-3-OH kinase (PI3K)/AKT signaling are activated in the presence of Myricetin since these were both abrogated by specific inhibitors of each pathway. Our results show how Myricetin, through ERs and PI3K/AKT signalings, potentiates sperm function. This effect is dose-dependent at low concentrations of Myricetin (up to 100 nM), whereas higher amounts do not seem to improve any further sperm motility, viability, or other tested features, and, in some cases, they reduced or even abrogated the efficacy exerted by lower doses. Further studies are needed to elucidate if high levels of Myricetin, which could be attained even with moderate wine consumption, could synergize with endogenous estrogens in the female reproductive tract, interfering with the physiological sperm fertilization process. Copyright © 2012 Wiley Periodicals, Inc.

The effect of regulatory mode on procrastination: Bi-stable parahippocampus connectivity with dorsal anterior cingulate and anterior prefrontal cortex.

PubMed

Zhang, Chenyan; Ni, Yan; Feng, Tingyong

2017-06-30

Previous research has elucidated that procrastination can be influenced by regulatory mode orientations. However, the neural mechanism of regulatory modes affecting procrastination is not well understood. To address this question, we employed resting-state functional magnetic resonance imaging (RS-fMRI) to test the influence of two regulatory modes (assessment and locomotion) on procrastination. The behavioral results showed that procrastination was positively correlated with assessment orientation but negatively correlated with locomotion orientation. Neuroimaging results indicated that the functional connectivity between parahippocampal cortex (PHC) and dorsal anterior cingulate (dACC) was negatively correlated with assessment scores, while the functional connectivity between anterior prefrontal cortex (aPFC) and parahippocampal cortex (PHC) was negatively correlated with locomotion scores. Critically, mediation analysis showed that the different effects of two distinct regulatory modes on procrastination were mediated by PHC-dACC and aPFC-PHC functional connectivity respectively. These results suggested that people's procrastination could be predicted by regulatory mode orientations, which is mediated by PHC connectivity with dACC and aPFC respectively. The present study extends our knowledge on procrastination and provides neural mechanism for understanding the link between regulatory mode orientations and procrastination. Copyright © 2017. Published by Elsevier B.V.

Team structure and regulatory focus: the impact of regulatory fit on team dynamic.

PubMed

Dimotakis, Nikolaos; Davison, Robert B; Hollenbeck, John R

2012-03-01

We report a within-teams experiment testing the effects of fit between team structure and regulatory task demands on task performance and satisfaction through average team member positive affect and helping behaviors. We used a completely crossed repeated-observations design in which 21 teams enacted 2 tasks with different regulatory focus characteristics (prevention and promotion) in 2 organizational structures (functional and divisional), resulting in 84 observations. Results suggested that salient regulatory demands inherent in the task interacted with structure to determine objective and subjective team-level outcomes, such that functional structures were best suited to (i.e., had best fit with) tasks with a prevention regulatory focus and divisional structures were best suited to tasks with a promotion regulatory focus. This contingency finding integrates regulatory focus and structural contingency theories, and extends them to the team level with implications for models of performance, satisfaction, and team dynamics.

Further characterization of loss of heterozygosity enhanced by p53 abrogation in human lymphoblastoid TK6 cells: disappearance of endpoint hotspots.

PubMed

Yatagai, Fumio; Morimoto, Shigeko; Kato, Takesi; Honma, Masamitsu

2004-06-13

Loss of heterozygosity (LOH) is the predominant mechanism of spontaneous mutagenesis at the heterozygous thymindine kinase locus (tk) in TK6 cells. LOH events detected in spontaneous TK(-) mutants (110 clones from p53 wild-type cells TK6-20C and 117 clones from p53-abrogated cells TK6-E6) were analyzed using 13 microsatellite markers spanning the whole of chromosome 17. Our analysis indicated an approximately 60-fold higher frequency of terminal deletions in p53-abrogated cells TK6-E6 compared to p53 wild-type cells TK6-20C whereas frequencies of point mutations (non-LOH events), interstitial deletions, and crossing over events were found to increase only less than twofold by such p53 abrogation. We then made use of an additional 17 microsatellite markers which provided an average map-interval of 1.6Mb to map various LOH endpoints on the 45Mb portion of chromosome 17q corresponding to the maximum length of LOH tracts (i.e. from the distal marker D17S932 to the terminal end). There appeared to be four prominent peaks (I-IV) in the distribution of LOH endpoints/Mb of Tk6-20C cells that were not evident in p53-abrogated cells TK6-E6, where they appeared to be rather broadly distributed along the 15-20Mb length (D17S1807 to D17S1607) surrounding two of the peaks that we detected in TK6-20C cells (peaks II and III). We suggest that the chromosomal instability that is so evident in TK6-E6 cells may be due to DNA double-strand break repair occurring through non homologous end-joining rather than allelic recombination.

17 CFR 1.59 - Activities of self-regulatory organization employees, governing board members, committee members...

Code of Federal Regulations, 2012 CFR

2012-04-01

... COMMODITY EXCHANGE ACT Miscellaneous § 1.59 Activities of self-regulatory organization employees, governing...) Self-regulatory organization means “self-regulatory organization,” as defined in Commission regulation... governors of a self-regulatory organization. (3) Committee member means a member, or functional equivalent...

17 CFR 1.59 - Activities of self-regulatory organization employees, governing board members, committee members...

Code of Federal Regulations, 2011 CFR

2011-04-01

... COMMODITY EXCHANGE ACT Miscellaneous § 1.59 Activities of self-regulatory organization employees, governing...) Self-regulatory organization means “self-regulatory organization,” as defined in Commission regulation... governors of a self-regulatory organization. (3) Committee member means a member, or functional equivalent...

17 CFR 1.59 - Activities of self-regulatory organization employees, governing board members, committee members...

Code of Federal Regulations, 2010 CFR

2010-04-01

... COMMODITY EXCHANGE ACT Miscellaneous § 1.59 Activities of self-regulatory organization employees, governing...) Self-regulatory organization means “self-regulatory organization,” as defined in Commission regulation... governors of a self-regulatory organization. (3) Committee member means a member, or functional equivalent...

Amyloid Precursor Protein (APP) Mediated Regulation of Ganglioside Homeostasis Linking Alzheimer's Disease Pathology with Ganglioside Metabolism

PubMed Central

Grimm, Marcus O. W.; Zinser, Eva G.; Grösgen, Sven; Hundsdörfer, Benjamin; Rothhaar, Tatjana L.; Burg, Verena K.; Kaestner, Lars; Bayer, Thomas A.; Lipp, Peter; Müller, Ulrike; Grimm, Heike S.; Hartmann, Tobias

2012-01-01

Gangliosides are important players for controlling neuronal function and are directly involved in AD pathology. They are among the most potent stimulators of Aβ production, are enriched in amyloid plaques and bind amyloid beta (Aβ). However, the molecular mechanisms linking gangliosides with AD are unknown. Here we identified the previously unknown function of the amyloid precursor protein (APP), specifically its cleavage products Aβ and the APP intracellular domain (AICD), of regulating GD3-synthase (GD3S). Since GD3S is the key enzyme converting a- to b-series gangliosides, it therefore plays a major role in controlling the levels of major brain gangliosides. This regulation occurs by two separate and additive mechanisms. The first mechanism directly targets the enzymatic activity of GD3S: Upon binding of Aβ to the ganglioside GM3, the immediate substrate of the GD3S, enzymatic turnover of GM3 by GD3S was strongly reduced. The second mechanism targets GD3S expression. APP cleavage results, in addition to Aβ release, in the release of AICD, a known candidate for gene transcriptional regulation. AICD strongly down regulated GD3S transcription and knock-in of an AICD deletion mutant of APP in vivo, or knock-down of Fe65 in neuroblastoma cells, was sufficient to abrogate normal GD3S functionality. Equally, knock-out of the presenilin genes, presenilin 1 and presenilin 2, essential for Aβ and AICD production, or of APP itself, increased GD3S activity and expression and consequently resulted in a major shift of a- to b-series gangliosides. In addition to GD3S regulation by APP processing, gangliosides in turn altered APP cleavage. GM3 decreased, whereas the ganglioside GD3, the GD3S product, increased Aβ production, resulting in a regulatory feedback cycle, directly linking ganglioside metabolism with APP processing and Aβ generation. A central aspect of this homeostatic control is the reduction of GD3S activity via an Aβ-GM3 complex and AICD-mediated repression of GD3S transcription. PMID:22470521

The three-dimensional structure of TrmB, a transcriptional regulator of dual function in the hyperthermophilic archaeon Pyrococcus furiosus in complex with sucrose

PubMed Central

Krug, Michael; Lee, Sung-Jae; Boos, Winfried; Diederichs, Kay; Welte, Wolfram

2013-01-01

TrmB is a repressor that binds maltose, maltotriose, and sucrose, as well as other α-glucosides. It recognizes two different operator sequences controlling the TM (Trehalose/Maltose) and the MD (Maltodextrin) operon encoding the respective ABC transporters and sugar-degrading enzymes. Binding of maltose to TrmB abrogates repression of the TM operon but maintains the repression of the MD operon. On the other hand, binding of sucrose abrogates repression of the MD operon but maintains repression of the TM operon. The three-dimensional structure of TrmB in complex with sucrose was solved and refined to a resolution of 3.0 Å. The structure shows the N-terminal DNA binding domain containing a winged-helix-turn-helix (wHTH) domain followed by an amphipathic helix with a coiled-coil motif. The latter promotes dimerization and places the symmetry mates of the putative recognition helix in the wHTH motif about 30 Å apart suggesting a canonical binding to two successive major grooves of duplex palindromic DNA. This suggests that the structure resembles the conformation of TrmB recognizing the pseudopalindromic TM promoter but not the conformation recognizing the nonpalindromic MD promoter. PMID:23576322

Complex pattern of immune evasion in MSI colorectal cancer.

PubMed

Ozcan, Mine; Janikovits, Jonas; von Knebel Doeberitz, Magnus; Kloor, Matthias

2018-01-01

Mismatch repair (MMR)-deficient cancers accumulate multiple insertion/deletion mutations at coding microsatellites (cMS), which give rise to frameshift peptide neoantigens. The high mutational neoantigen load of MMR-deficient cancers is reflected by pronounced anti-tumoral immune responses of the host and high responsiveness towards immune checkpoint blockade. However, immune evasion mechanisms can interfere with the immune response against MMR-deficient tumors. We here performed a comprehensive analysis of immune evasion in MMR-deficient colorectal cancers, focusing on HLA class I-mediated antigen presentation. 72% of MMR-deficient colorectal cancers of the DFCI database harbored alterations affecting genes involved in HLA class I-mediated antigen presentation, and 54% of these mutations were predicted to abrogate function. Mutations affecting the HLA class I transactivator NLRC5 were observed as a potential new immune evasion mechanism in 26% (6% abrogating) of the analyzed tumors. NLRC5 mutations in MMR-deficient cancers were associated with decreased levels of HLA class I antigen expression. In summary, the majority of MMR-deficient cancers display mutations interfering with HLA class I antigen presentation that reflect active immune surveillance and immunoselection during tumor development. Clinical studies focusing on immune checkpoint blockade in MSI cancer should account for the broad variety of immune evasion mechanisms as potential biomarkers of therapy success.

Cloning and characterization of GETS-1, a goldfish Ets family member that functions as a transcriptional repressor in muscle.

PubMed

Goldman, D; Sapru, M K; Stewart, S; Plotkin, J; Libermann, T A; Wasylyk, B; Guan, K

1998-10-15

An Ets transcription factor family member, GETS-1, was cloned from a goldfish retina cDNA library. GETS-1 contains a conserved Ets DNA-binding domain at its N-terminus and is most similar to ternary complex factor (TCF) serum-response-factor protein-1a (SAP-1a). GETS-1 is expressed in many tissues, but is enriched in retina and brain. As with the TCFs SAP-1a and ets-related protein (ERP), overexpression of the GETS-1 promoter suppresses nicotinic acetylcholine receptor epsilon-subunit gene expression in cultured muscle cells. A consensus Ets binding site sequence in the promoter of the epsilon-subunit gene is required for GETS-1-mediated repression. GETS-1 repressor activity is abrogated by overexpression of an activated Ras/mitogen-activated protein kinase (MAP kinase) or by mutation of Ser-405, a MAP kinase phosphorylation site in GETS-1. Fusion proteins created between GETS-1 and the Gal4 DNA-binding domain show that, like other TCFs, GETS-1 contains a C-terminal activation domain that is activated by a Ras/MAP kinase signalling cascade. Interestingly, mutation of Ser-405 located within this activation domain abrogated transcriptional activation of the fusion protein.

Expression of PKA inhibitor (PKI) gene abolishes cAMP-mediated protection to endothelial barrier dysfunction.

PubMed

Lum, H; Jaffe, H A; Schulz, I T; Masood, A; RayChaudhury, A; Green, R D

1999-09-01

We investigated the hypothesis that cAMP-dependent protein kinase (PKA) protects against endothelial barrier dysfunction in response to proinflammatory mediators. An E1-, E3-, replication-deficient adenovirus (Ad) vector was constructed containing the complete sequence of PKA inhibitor (PKI) gene (AdPKI). Infection of human microvascular endothelial cells (HMEC) with AdPKI resulted in overexpression of PKI. Treatment with 0.5 microM thrombin increased transendothelial albumin clearance rate (0.012 +/- 0.003 and 0.035 +/- 0.005 microl/min for control and thrombin, respectively); the increase was prevented with forskolin + 3-isobutyl-1-methylxanthine (F + I) treatment. Overexpression of PKI resulted in abrogation of the F + I-induced inhibition of the permeability increase. However, with HMEC infected with ultraviolet-inactivated AdPKI, the F + I-induced inhibition was present. Also, F + I treatment of HMEC transfected with reporter plasmid containing the cAMP response element-directed transcription of the luciferase gene resulted in an almost threefold increase in luciferase activity. Overexpression of PKI inhibited this induction of luciferase activity. The results show that Ad-mediated overexpression of PKI in endothelial cells abrogated the cAMP-mediated protection against increased endothelial permeability, providing direct evidence that cAMP-dependent protein kinase promotes endothelial barrier function.

Ionizing radiation abrogates the pro-tumorigenic capacity of cancer-associated fibroblasts co-implanted in xenografts.

PubMed

Grinde, Maria Tunset; Vik, Jørg; Camilio, Ketil André; Martinez-Zubiaurre, Inigo; Hellevik, Turid

2017-04-25

Cancer-associated fibroblasts (CAFs) are abundantly present in solid tumors and affect tumorigenesis and therapeutic responses. In the context of clinical radiotherapy, the impact of irradiated CAFs to treatment outcomes is largely unexplored. Aiming at improving radiotherapy efficacy, we have here explored the effect of radiation on the inherent pro-tumorigenic capacity of CAFs in animals. Ionizing radiation was delivered to cultured CAFs as single-high or fractionated doses. Tumor development was compared in mice receiving A549 lung tumor cells admixed with irradiated or control CAFs. Biological mechanisms behind tumor growth regulation were investigated by quantitative histology and immunohistochemistry. Viability assessments confirmed that irradiated CAFs are fully functional prior to implantation. However, the enhanced tumorigenic effect observed in tumors co-implanted with control CAFs was abrogated in tumors established with irradiated CAFs. Experiments to ascertain fate of implanted fibroblasts showed that exogenously administered CAFs reside at the implantation site for few days, suggesting that tumor growth regulation from admixed CAFs take place during initial tumor formation. Our work demonstrate that irradiated CAFs lose their pro-tumorigenic potential in vivo, affecting angiogenesis and tumor engraftment. This finding propose a previously unknown advantageous effect induced by radiotherapy, adding to the direct cytotoxic effects on transformed epithelial cells.

Sociotechnical systems as a framework for regulatory system design and evaluation: Using Work Domain Analysis to examine a new regulatory system.

PubMed

Carden, Tony; Goode, Natassia; Read, Gemma J M; Salmon, Paul M

2017-03-15

Like most work systems, the domain of adventure activities has seen a series of serious incidents and subsequent calls to improve regulation. Safety regulation systems aim to promote safety and reduce accidents. However, there is scant evidence they have led to improved safety outcomes. In fact there is some evidence that the poor integration of regulatory system components has led to adverse safety outcomes in some contexts. Despite this, there is an absence of methods for evaluating regulatory and compliance systems. This article argues that sociotechnical systems theory and methods provide a suitable framework for evaluating regulatory systems. This is demonstrated through an analysis of a recently introduced set of adventure activity regulations. Work Domain Analysis (WDA) was used to describe the regulatory system in terms of its functional purposes, values and priority measures, purpose-related functions, object-related processes and cognitive objects. This allowed judgement to be made on the nature of the new regulatory system and on the constraints that may impact its efficacy following implementation. Importantly, the analysis suggests that the new system's functional purpose of ensuring safe activities is not fully supported in terms of the functions and objects available to fulfil them. Potential improvements to the design of the system are discussed along with the implications for regulatory system design and evaluation across the safety critical domains generally. Copyright © 2017 Elsevier Ltd. All rights reserved.

THE ROLES OF METAL IONS IN REGULATION BY RIBOSWITCHES

PubMed Central

2012-01-01

Metal ions are required by all organisms in order to execute an array of essential molecular functions. They play a critical role in many catalytic mechanisms and structural properties. Proper homeostasis of ions is critical; levels that are aberrantly low or high are deleterious to cellular physiology. To maintain stable intracellular pools, metal ion-sensing regulatory (metalloregulatory) proteins couple metal ion concentration fluctuations with expression of genes encoding for cation transport or sequestration. However, these transcriptional-based regulatory strategies are not the only mechanisms by which organisms coordinate metal ions with gene expression. Intriguingly, a few classes of signal-responsive RNA elements have also been discovered to function as metalloregulatory agents. This suggests that RNA-based regulatory strategies can be precisely tuned to intracellular metal ion pools, functionally akin to metalloregulatory proteins. In addition to these metal-sensing regulatory RNAs, there is a yet broader role for metal ions in directly assisting the structural integrity of other signal-responsive regulatory RNA elements. In this chapter, we discuss how the intimate physicochemical relationship between metal ions and nucleic acids is important for the structure and function of metal ion- and metabolite-sensing regulatory RNAs. PMID:22010271

17 CFR 1.59 - Activities of self-regulatory organization employees, governing board members, committee members...

Code of Federal Regulations, 2014 CFR

2014-04-01

... COMMODITY EXCHANGE ACT Miscellaneous § 1.59 Activities of self-regulatory organization employees, governing...) Self-regulatory organization means “self-regulatory organization,” as defined in § 1.3(ee), and... member, or functional equivalent thereof, of the board of governors of a self-regulatory organization. (3...

17 CFR 1.59 - Activities of self-regulatory organization employees, governing board members, committee members...

Code of Federal Regulations, 2013 CFR

2013-04-01

... COMMODITY EXCHANGE ACT Miscellaneous § 1.59 Activities of self-regulatory organization employees, governing...) Self-regulatory organization means “self-regulatory organization,” as defined in § 1.3(ee), and... member, or functional equivalent thereof, of the board of governors of a self-regulatory organization. (3...

Cigarette smoke-induced cell cycle arrest in spermatocytes [GC-2spd(ts)] is mediated through crosstalk between Ahr-Nrf2 pathway and MAPK signaling.

PubMed

Esakky, Prabagaran; Hansen, Deborah A; Drury, Andrea M; Moley, Kelle H

2015-02-01

Our earlier studies have demonstrated that the cigarette smoke in the form of cigarette smoke condensate (CSC) causes growth arrest of a mouse spermatocyte cell line [GC-2spd(ts)] through activation of the AHR-NRF2 pathway. The present study demonstrates the CSC-activated p38 and ERK MAPK signaling in GC-2spd(ts) via arylhydrocarbon receptor (AHR). Pharmacological inhibition by using AHR-antagonist, or p38 MAPK and ERK (MEK1) inhibitors significantly abrogates CSC-induced growth arrest by AHR and MAPK inactivation. QRT-PCR, western blot, and immunofluorescence of Ahr-target of Nrf2, and stress-inducible growth suppressive Atf3 and E2f4 following treatments indicate a crosstalk among these pathways. Regulation of Atf3 by Nrf2 and Ahr through RNA interference suggests the existence of a cross-regulatory loop between the targets. CSC induction of E2f4 via Atf3 and its regulation by pharmacological inhibitors reveal a possible regulatory mechanism of growth inhibitory CSC. SiRNA silencing of Ahr, Nrf2, Atf3, and E2f4 genes and downregulation of cyclins by CSC corroborate the growth inhibitory effect of cigarette smoke. Thus, the data obtained suggest that the CSC-mediated MAPKs and AHR-NRF2 crosstalks lay the molecular basis for the growth arrest and cell death of spermatocytes. © The Author (2014). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved.

Sost, independent of the non-coding enhancer ECR5, is required for bone mechanoadaptation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Robling, Alexander G.; Kang, Kyung Shin; Bullock, Whitney A.

Here, sclerostin ( Sost) is a negative regulator of bone formation that acts upon the Wnt signaling pathway. Sost is mechanically regulated at both mRNA and protein level such that loading represses and unloading enhances Sost expression, in osteocytes and in circulation. The non-coding evolutionarily conserved enhancer ECR5 has been previously reported as a transcriptional regulatory element required for modulating Sost expression in osteocytes. Here we explored the mechanisms by which ECR5, or several other putative transcriptional enhancers regulate Sost expression, in response to mechanical stimulation. We found that in vivo ulna loading is equally osteoanabolic in wildtype and Sostmore » –/– mice, although Sost is required for proper distribution of load-induced bone formation to regions of high strain. Using Luciferase reporters carrying the ECR5 non-coding enhancer and heterologous or homologous h SOST promoters, we found that ECR5 is mechanosensitive in vitro and that ECR5-driven Luciferase activity decreases in osteoblasts exposed to oscillatory fluid flow. Yet, ECR5–/– mice showed similar magnitude of load-induced bone formation and similar periosteal distribution of bone formation to high-strain regions compared to wildtype mice. Further, we found that in contrast to Sost–/– mice, which are resistant to disuse-induced bone loss, ECR5–/– mice lose bone upon unloading to a degree similar to wildtype control mice. ECR5 deletion did not abrogate positive effects of unloading on Sost, suggesting that additional transcriptional regulators and regulatory elements contribute to load-induced regulation of Sost.« less

Cadmium-induced malignant transformation of rat liver cells: Potential key role and regulatory mechanism of altered apolipoprotein E expression in enhanced invasiveness.

PubMed

Suzuki, Masayo; Takeda, Shuso; Teraoka-Nishitani, Noriko; Yamagata, Akane; Tanaka, Takahiro; Sasaki, Marika; Yasuda, Natsuki; Oda, Makiko; Okano, Tatsuji; Yamahira, Kazuhiro; Nakamura, Yuta; Kobayashi, Takanobu; Kino, Katsuhito; Miyazawa, Hiroshi; Waalkes, Michael P; Takiguchi, Masufumi

2017-05-01

Cadmium is a transition metal that is classified as human carcinogen by the International Agency for Research on Cancer (IARC) with multiple target sites. Many studies using various model systems provide evidence of cadmium-induced malignancy formation in vivo or malignant cell transformation in vitro. Nonetheless, further studies are needed to completely understand the mechanisms of cadmium carcinogenicity. Our prior studies have utilized a rat liver epithelial cell line (TRL 1215) as a model for cadmium-induced malignant transformation. In the present study, we focused on the molecular mechanisms of this malignant transformation, especially with regard to hyper-invasiveness stimulated by cadmium transformation. By performing a series of biochemical analyses on cadmium transformed cells, it was determined that cadmium had significantly down-regulated the expression of apolipoprotein E (ApoE). ApoE was recently established as a suppressor of cell invasion. A key factor in the suppression of ApoE by cadmium appeared to be that the metal evoked a 5-aza-2'-deoxycytidine-sensitive hypermethylation of the regulatory region of ApoE, coupled with interference of the action of liver X receptor α (LXRα), a transcriptional regulator for ApoE. Furthermore, the expression of LXRα itself was suppressed by cadmium-mediated epigenetic modification. Re-expression of ApoE clearly abrogated the cell invasion stimulated by cadmium-induced malignant transformation. Together, the current results suggest that the cadmium-mediated enhanced cell invasion is linked to down-regulation of ApoE during malignant transformation these liver cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Sost, independent of the non-coding enhancer ECR5, is required for bone mechanoadaptation

DOE PAGES

Robling, Alexander G.; Kang, Kyung Shin; Bullock, Whitney A.; ...

2016-09-04

Here, sclerostin ( Sost) is a negative regulator of bone formation that acts upon the Wnt signaling pathway. Sost is mechanically regulated at both mRNA and protein level such that loading represses and unloading enhances Sost expression, in osteocytes and in circulation. The non-coding evolutionarily conserved enhancer ECR5 has been previously reported as a transcriptional regulatory element required for modulating Sost expression in osteocytes. Here we explored the mechanisms by which ECR5, or several other putative transcriptional enhancers regulate Sost expression, in response to mechanical stimulation. We found that in vivo ulna loading is equally osteoanabolic in wildtype and Sostmore » –/– mice, although Sost is required for proper distribution of load-induced bone formation to regions of high strain. Using Luciferase reporters carrying the ECR5 non-coding enhancer and heterologous or homologous h SOST promoters, we found that ECR5 is mechanosensitive in vitro and that ECR5-driven Luciferase activity decreases in osteoblasts exposed to oscillatory fluid flow. Yet, ECR5–/– mice showed similar magnitude of load-induced bone formation and similar periosteal distribution of bone formation to high-strain regions compared to wildtype mice. Further, we found that in contrast to Sost–/– mice, which are resistant to disuse-induced bone loss, ECR5–/– mice lose bone upon unloading to a degree similar to wildtype control mice. ECR5 deletion did not abrogate positive effects of unloading on Sost, suggesting that additional transcriptional regulators and regulatory elements contribute to load-induced regulation of Sost.« less

Fox smell abrogates the effect of herbal odor to prolong mouse cardiac allograft survival.

PubMed

Jin, Xiangyuan; Uchiyama, Masateru; Zhang, Qi; Niimi, Masanori

2014-05-09

Herbal medicines have unique odors, and the act of smelling may have modulatory effects on the immune system. We investigated the effect of olfactory exposure to Tokishakuyaku-san (TJ-23), a Japanese herbal medicine, on alloimmune responses in a murine model of cardiac allograft transplantation. Naïve or olfactory-dysfunctional CBA mice underwent transplantation of a C57BL/6 heart and were exposed to the odor of TJ-23 until rejection. Some naïve CBA recipients of an allograft were given olfactory exposure to Sairei-to (TJ-114), trimethylthiazoline (TMT), individual components of TJ-23, or a TJ-23 preparation lacking one component. Adoptive transfer studies were performed to determine whether regulatory cells were generated. Untreated CBA mice rejected their C57BL/6 allografts acutely, as did olfactory-dysfunctional CBA mice exposed to the odor of TJ-23. CBA recipients of a C57BL/6 heart given olfactory exposure to TJ-23 had significantly prolonged allograft survival, whereas those exposed to the odor of TJ-114, TMT, one component of TJ-23, or TJ-23 lacking a component did not. Secondary allograft recipients that were given, at 30 days after transplantation, either whole splenocytes, CD4+ cells, or CD4+CD25+ cells from primary recipients exposed to the odor of TJ-23 had indefinitely prolonged allograft survival. Prolonged survival of cardiac allografts and generation of regulatory cells was associated with exposure to the odor of TJ-23 in our model. The olfactory area of the brain may have a role in the modulation of immune responses.

Cinnamon Ameliorates Experimental Allergic Encephalomyelitis in Mice via Regulatory T Cells: Implications for Multiple Sclerosis Therapy

PubMed Central

Mondal, Susanta; Pahan, Kalipada

2015-01-01

Upregulation and/or maintenance of regulatory T cells (Tregs) during an autoimmune insult may have therapeutic efficacy in autoimmune diseases. Although several immunomodulatory drugs and molecules are available, most present significant side effects over long-term use. Cinnamon is a commonly used natural spice and flavoring material used for centuries throughout the world. Here, we have explored a novel use of cinnamon powder in protecting Tregs and treating the disease process of experimental allergic encephalomyelitis (EAE), an animal model of MS. Oral feeding of cinnamon (Cinnamonum verum) powder suppresses clinical symptoms of relapsing-remitting EAE in female PLP-TCR transgenic mice and adoptive transfer mouse model. Cinnamon also inhibited clinical symptoms of chronic EAE in male C57/BL6 mice. Dose-dependent study shows that cinnamon powder at a dose of 50 mg/kg body wt/d or higher significantly suppresses clinical symptoms of EAE in mice. Accordingly, oral administration of cinnamon also inhibited perivascular cuffing, maintained the integrity of blood-brain barrier and blood-spinal cord barrier, suppressed inflammation, normalized the expression of myelin genes, and blocked demyelination in the central nervous system of EAE mice. Interestingly, cinnamon treatment upregulated Tregs via reduction of nitric oxide production. Furthermore, we demonstrate that blocking of Tregs by neutralizing antibodies against CD25 abrogates cinnamon-mediated protection of EAE. Taken together, our results suggest that oral administration of cinnamon powder may be beneficial in MS patients and that no other existing anti-MS therapies could be so economical and trouble-free as this approach. PMID:25569428

Histone Deacetylase-1 Is Enriched at the Platelet-derived Growth Factor-D Promoter in Response to Interleukin-1β and Forms a Cytokine-inducible Gene-silencing Complex with NF-κB p65 and Interferon Regulatory Factor-1*

PubMed Central

Liu, Mary Y.; Khachigian, Levon M.

2009-01-01

Understanding the mechanisms governing cytokine control of growth factor expression in smooth muscle cells would provide invaluable insight into the molecular regulation of vascular phenotypes and create future opportunities for therapeutic intervention. Here, we report that the proinflammatory cytokine interleukin (IL)-1β suppresses platelet-derived growth factor (PDGF)-D promoter activity and mRNA and protein expression in smooth muscle cells. NF-κB p65, induced by IL-1β, interacts with a novel element in the PDGF-D promoter and inhibits PDGF-D transcription. Interferon regulatory factor-1 (IRF-1) is also induced by IL-1β and binds to a different element upstream in the promoter. Immunoprecipitation and chromatin immunoprecipitation experiments showed that IL-1β stimulates p65 interaction with IRF-1 and the accumulation of both factors at the PDGF-D promoter. Mutation of the IRF-1 and p65 DNA-binding elements relieved the promoter from IL-1β-mediated repression. PDGF-D repression by IL-1β involves histone deacetylation and interaction of HDAC-1 with IRF-1 and p65. HDAC-1 small interfering RNA ablates complex formation with IRF-1 and p65 and abrogates IRF-1 and p65 occupancy of the PDGF-D promoter. Thus, HDAC-1 is enriched at the PDGF-D promoter in cells exposed to IL-1β and forms a cytokine-inducible gene-silencing complex with p65 and IRF-1. PMID:19843519

Phosphodiesterases regulate airway smooth muscle function in health and disease.

PubMed

Krymskaya, Vera P; Panettieri, Reynold A

2007-01-01

On the basis of structure, regulation, and kinetic properties, phosphodiesterases (PDEs) represent a superfamily of enzymes divided into 11 subfamilies that catalyze cytosolic levels of 3',5'-cyclic adenosine monophosphate (cAMP) or 3',5'-cyclic guanosine monophosphate (cGMP) to 5'-AMP or 5'-GMP, respectively. PDE4 represents the major PDE expressed in inflammatory cells as well as airway smooth muscle (ASM), and selective PDE4 inhibitors provide a broad spectrum of anti-inflammatory effects such as abrogating cytokine and chemokine release from inflammatory cells and inhibiting inflammatory cell trafficking. Due to cell- and tissue-specific gene expression and regulation, PDEs modulate unique organ-based functions. New tools or compounds that selectively inhibit PDE subfamilies and genetically engineered mice deficient in selective isoforms have greatly enhanced our understanding of PDE function in airway inflammation and resident cell function. This chapter will focus on recent advances in our understanding of the role of PDE in regulating ASM function.

Actin dynamics regulate immediate PAR-2-dependent responses to acute epidermal permeability barrier abrogation.

PubMed

Roelandt, Truus; Heughebaert, Carol; Verween, Gunther; Giddelo, Christina; Verbeken, Gilbert; Pirnay, Jean-Paul; Devos, Daniel; Crumrine, Debra; Roseeuw, Diane; Elias, Peter M; Hachem, Jean-Pierre

2011-02-01

Lamellar body (LB) secretion and terminal differentiation of stratum granulosum (SG) cells are signaled by both protease activated receptor-2 (PAR-2) and caveolin-1 (cav-1). To address the early dynamics of LB secretion, we examined cytoskeletal remodeling of keratinocytes in 3 mouse models following acute barrier abrogation: hairless mice, PAR-2 knockout (-/-) and cav-1 -/-. Under basal conditions, globular (G)-actin accumulates in SG cells cytosol, while filamentous (F)-actin is restricted to peri-membrane domains. Barrier abrogation induces the apical movement of F-actin and the retreat of the SG-G-actin front, paralleled by upstream cytoskeletal kinases activation. This phenomenon was both enhanced by PAR-2 agonist, and inhibited by cytochalasin-D and in PAR-2 knockout mice. We found that plasma membrane conformational changes causing LB secretion are controlled by PAR-2-dependent cytoskeletal rearrangements. We next addressed the interaction dynamics between cytoskeleton and plasma membrane following PAR-2-induced actin stress fiber formation in both cav-1 -/- and wildtype cells. Actin stress fiber formation is increased in cav-1 -/- cells prior to and following PAR-2 agonist peptide-treatment, while absence of cav-1 inhibits E-cadherin-mediated cell-to-cell adhesion. PAR-2 drives cytoskeletal/plasma membrane dynamics that regulate early LB secretion following barrier abrogation, stress fiber formation and keratinocyte adhesion. Copyright © 2010 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

Thymic Stromal Lymphopoietin Attenuates the Development of Atherosclerosis in ApoE−/− Mice

PubMed Central

Yu, Kunwu; Zhu, Pengfei; Dong, Qian; Zhong, Yucheng; Zhu, Zhengfeng; Lin, Yingzhong; Huang, Ying; Meng, Kai; Ji, Qingwei; Yi, Guiwen; Zhang, Wei; Wu, Bangwei; Mao, Yi; Cheng, Peng; Zhao, Xiaoqi; Mao, Xiaobo; Zeng, Qiutang

2013-01-01

Background Thymic stromal lymphopoietin (TSLP) is a cytokine with multiple effects on the body. For one thing, TSLP induces Th2 immunoreaction and facilitates allergic reaction; for another, it promotes the differentiation of naturally occurring CD4+CD25+Foxp3+ regulatory T cells (nTregs) and maintains immune tolerance. However, the exact role of TSLP in atherosclerosis remains unknown. Methods and Results In vitro, we examined the phenotype of TSLP‐conditioned bone marrow dendritic cells (TSLP‐DCs) of apolipoprotein E–deficient (ApoE−/−) mice and their capacity to induce the differentiation of Tregs. Our results indicated that TSLP‐DCs obtained the characteristics of tolerogenic dendritic cells and increased a generation of CD4+ latency‐associated peptide (LAP)+ Tregs and nTregs when cocultured with naive T cells. In addition, the functional relevance of TSLP and TSLP‐DCs in the development of atherosclerosis was also determined. Interestingly, we found that TSLP was almost absent in cardiovascular tissue of ApoE−/− mice, and TSLP administration increased the levels of antioxidized low‐density lipoprotein IgM and IgG1, but decreased the levels of IgG2a in plasma. Furthermore, mice treated with TSLP and TSLP‐DCs developed significantly fewer (32.6% and 28.2%, respectively) atherosclerotic plaques in the aortic root compared with controls, along with increased numbers of CD4+LAP+ Tregs and nTregs in the spleen and decreased inflammation in the aorta, which could be abrogated by anti‐TGF‐β antibody. Conclusions Our results revealed a protective role for TSLP in atherosclerosis that is possibly mediated by reestablishing a tolerogenic immune response, which may represent a novel possibility for treatment or prevention of atherosclerosis. PMID:23985377

Protein kinase-A-dependent osteoprotegerin production on interleukin-1 stimulation in human gingival fibroblasts is distinct from periodontal ligament fibroblasts

PubMed Central

Hormdee, D; Nagasawa, T; Kiji, M; Yashiro, R; Kobayashi, H; Koshy, G; Noguchi, K; Nitta, H; Ishikawa, I

2005-01-01

Periodontitis, a chronic inflammatory disease, is characterized by increased expression of interleukin (IL)-1 and other inflammatory mediators resulting in extensive osteoclast formation and bone loss. Expression of receptor activator of nuclear factor kappa B ligand (RANKL) and its decoy receptor, osteoprotegerin (OPG), by osteoblasts is important to regulate osteoclast differentiation. The aim of the present study was to investigate the regulatory effects of IL-1 on RANKL and OPG production by mesenchymal fibroblasts in periodontal tissue. Human gingival fibroblasts (HGF) and periodontal ligament fibroblasts (PDL) were stimulated with IL-1α with or without protein synthesis inhibitor cycloheximide (CHX), protein kinase A (PKA) inhibitors, protein kinase C (PKC) inhibitors and prostaglandin E2 (PGE2) inhibitor. In some experiments, the cultured cells were directly stimulated with either PKA or PKC activators. In HGF, IL-1α-stimulated OPG mRNA expression was high and could be reduced by CHX. PKA inhibitor completely abrogated IL-1α-induced OPG mRNA expression and OPG production. Endogenous PGE2 further enhanced IL-1α-induced OPG production in HGF. In PDL, RANKL mRNA expression was greatly augmented by IL-1α. IL-1α induced OPG mRNA expression and protein production. PKC inhibitor partially reduced IL-1α-induced OPG production and PKC activator enhanced OPG production in PDL. The IL-1α-stimulated OPG mRNA expression in HGF was greater than PDL. These results provide new evidence for the possible osteoclastogenesis-inhibitory function of HGF through PKA activity pathway. PDL utilized PKC for OPG production. Thus, we emphasize that HGF and PDL have different characteristics of host defence mechanism against inflammatory process. PMID:16297161

Implication of matrix metalloproteinases 2 and 9 in ceramide 1-phosphate-stimulated macrophage migration.

PubMed

Ordoñez, Marta; Rivera, Io-Guané; Presa, Natalia; Gomez-Muñoz, Antonio

2016-08-01

Cell migration is a complex biological function involved in both physiologic and pathologic processes. Although this is a subject of intense investigation, the mechanisms by which cell migration is regulated are not completely understood. In this study we show that the bioactive sphingolipid ceramide 1-phosphate (C1P), which is involved in inflammatory responses, causes upregulation of metalloproteinases (MMP) -2 and -9 in J774A.1 macrophages. This effect was shown to be dependent on stimulation of phosphatidylinositol 3-kinase (PI3K) and extracellularly regulated kinases 1-2 (ERK1-2) as demonstrated by treating the cells with specific siRNA to knockdown the p85 regulatory subunit of PI3K, or ERK1-2. Inhibition of MMP-2 or MMP-9 pharmacologically or with specific siRNA to silence the genes encoding these MMPs abrogated C1P-stimulated macrophage migration. Also, C1P induced actin polymerization and potently increased phosphorylation of the focal adhesion protein paxillin, which are essential factors in the regulation of cell migration. As expected, blockade of paxillin activation with specific siRNA significantly reduced actin polymerization. In addition, inhibition of actin polymerization with cytochalasin D completely blocked C1P-induced MMP-2 and -9 expression as well as C1P-stimulated macrophage migration. It was also observed that pertussis toxin (Ptx) inhibited Akt, ERK1-2, and paxillin phosphorylation, and completely blocked cell migration. The latter findings support the notion that C1P-stimulated macrophage migration is a receptor mediated effect, and point to MMP-2 and -9 as possible therapeutic targets to control inflammation. Copyright © 2016 Elsevier Inc. All rights reserved.

Met receptor tyrosine kinase signals through a cortactin–Gab1 scaffold complex, to mediate invadopodia

PubMed Central

Rajadurai, Charles V.; Havrylov, Serhiy; Zaoui, Kossay; Vaillancourt, Richard; Stuible, Matthew; Naujokas, Monica; Zuo, Dongmei; Tremblay, Michel L.; Park, Morag

2012-01-01

Summary Invasive carcinoma cells form actin-rich matrix-degrading protrusions called invadopodia. These structures resemble podosomes produced by some normal cells and play a crucial role in extracellular matrix remodeling. In cancer, formation of invadopodia is strongly associated with invasive potential. Although deregulated signals from the receptor tyrosine kinase Met (also known as hepatocyte growth factor are linked to cancer metastasis and poor prognosis, its role in invadopodia formation is not known. Here we show that stimulation of breast cancer cells with the ligand for Met, hepatocyte growth factor, promotes invadopodia formation, and in aggressive gastric tumor cells where Met is amplified, invadopodia formation is dependent on Met activity. Using both GRB2-associated-binding protein 1 (Gab1)-null fibroblasts and specific knockdown of Gab1 in tumor cells we show that Met-mediated invadopodia formation and cell invasion requires the scaffold protein Gab1. By a structure–function approach, we demonstrate that two proline-rich motifs (P4/5) within Gab1 are essential for invadopodia formation. We identify the actin regulatory protein, cortactin, as a direct interaction partner for Gab1 and show that a Gab1–cortactin interaction is dependent on the SH3 domain of cortactin and the integrity of the P4/5 region of Gab1. Both cortactin and Gab1 localize to invadopodia rosettes in Met-transformed cells and the specific uncoupling of cortactin from Gab1 abrogates invadopodia biogenesis and cell invasion downstream from the Met receptor tyrosine kinase. Met localizes to invadopodia along with cortactin and promotes phosphorylation of cortactin. These findings provide insights into the molecular mechanisms of invadopodia formation and identify Gab1 as a scaffold protein involved in this process. PMID:22366451

A stable transcription factor complex nucleated by oligomeric AML1–ETO controls leukaemogenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sun, Xiao-Jian; Wang, Zhanxin; Wang, Lan

2013-06-30

Transcription factors are frequently altered in leukaemia through chromosomal translocation, mutation or aberrant expression. AML1–ETO, a fusion protein generated by the t(8;21) translocation in acute myeloid leukaemia, is a transcription factor implicated in both gene repression and activation. AML1–ETO oligomerization, mediated by the NHR2 domain, is critical for leukaemogenesis, making it important to identify co-regulatory factors that ‘read’ the NHR2 oligomerization and contribute to leukaemogenesis. Here we show that, in human leukaemic cells, AML1–ETO resides in and functions through a stable AML1–ETO-containing transcription factor complex (AETFC) that contains several haematopoietic transcription (co)factors. These AETFC components stabilize the complex through multivalentmore » interactions, provide multiple DNA-binding domains for diverse target genes, co-localize genome wide, cooperatively regulate gene expression, and contribute to leukaemogenesis. Within the AETFC complex, AML1–ETO oligomerization is required for a specific interaction between the oligomerized NHR2 domain and a novel NHR2-binding (N2B) motif in E proteins. Crystallographic analysis of the NHR2–N2B complex reveals a unique interaction pattern in which an N2B peptide makes direct contact with side chains of two NHR2 domains as a dimer, providing a novel model of how dimeric/oligomeric transcription factors create a new protein-binding interface through dimerization/oligomerization. Intriguingly, disruption of this interaction by point mutations abrogates AML1–ETO-induced haematopoietic stem/progenitor cell self-renewal and leukaemogenesis. These results reveal new mechanisms of action of AML1–ETO, and provide a potential therapeutic target in t(8;21)-positive acute myeloid leukaemia.« less

Functional analysis of regulatory single-nucleotide polymorphisms.

PubMed

Pampín, Sandra; Rodríguez-Rey, José C

2007-04-01

The identification of regulatory polymorphisms has become a key problem in human genetics. In the past few years there has been a conceptual change in the way in which regulatory single-nucleotide polymorphisms are studied. We revise the new approaches and discuss how gene expression studies can contribute to a better knowledge of the genetics of common diseases. New techniques for the association of single-nucleotide polymorphisms with changes in gene expression have been recently developed. This, together with a more comprehensive use of the old in-vitro methods, has produced a great amount of genetic information. When added to current databases, it will help to design better tools for the detection of regulatory single-nucleotide polymorphisms. The identification of functional regulatory single-nucleotide polymorphisms cannot be done by the simple inspection of DNA sequence. In-vivo techniques, based on primer-extension, and the more recently developed 'haploChIP' allow the association of gene variants to changes in gene expression. Gene expression analysis by conventional in-vitro techniques is the only way to identify the functional consequences of regulatory single-nucleotide polymorphisms. The amount of information produced in the last few years will help to refine the tools for the future analysis of regulatory gene variants.

7 CFR 1700.32 - Program Accounting and Regulatory Analysis.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 7 Agriculture 11 2014-01-01 2014-01-01 false Program Accounting and Regulatory Analysis. 1700.32... SERVICE, DEPARTMENT OF AGRICULTURE GENERAL INFORMATION Agency Organization and Functions § 1700.32 Program Accounting and Regulatory Analysis. RUS, through Program Accounting and Regulatory Analysis, monitors and...

7 CFR 1700.32 - Program Accounting and Regulatory Analysis.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 7 Agriculture 11 2013-01-01 2013-01-01 false Program Accounting and Regulatory Analysis. 1700.32... SERVICE, DEPARTMENT OF AGRICULTURE GENERAL INFORMATION Agency Organization and Functions § 1700.32 Program Accounting and Regulatory Analysis. RUS, through Program Accounting and Regulatory Analysis, monitors and...

Self-Regulatory Processes and Exercise Adherence in Older Adults

PubMed Central

McAuley, Edward; Mullen, Sean P.; Szabo, Amanda N.; White, Siobhan M.; Wójcicki, Thomas R.; Mailey, Emily L.; Gothe, Neha P.; Olson, Erin A.; Voss, Michelle; Erickson, Kirk; Prakash, Ruchika; Kramer, Arthur F.

2011-01-01

Background Self-efficacy and the use of self-regulatory strategies are consistently associated with physical activity behavior. Similarly, behavioral inhibition and cognitive resource allocation, indices of executive control function, have also been associated with this health behavior. Purpose The purpose of this study was to examine the hypothesis that self-efficacy mediates the relationship between self-regulatory processes, such as executive function, and sustained exercise behavior. Methods Older adults (N = 177, mean age = 66.44 years) completed measures of executive function, self-reported use of self-regulatory strategies and self-efficacy prior to and during the first month of a 12-month exercise intervention. Percentage of exercise classes attended over the following 11 months was used to represent adherence. Data were collected from 2007 to 2010 and analyzed in 2010–2011. Structural equation models were tested examining the effect of executive function and strategy use on adherence via efficacy. Results As hypothesized, results showed significant direct effects of two elements of executive function and of strategy use on self-efficacy and of efficacy on adherence. In addition, there were significant indirect effects of strategy use and executive function on adherence via self-efficacy. Conclusions Higher levels of executive function and use of self-regulatory strategies at the start of an exercise program enhance beliefs in exercise capabilities, which in turn leads to greater adherence. PMID:21855742

T cell-B cell interactions in primary immunodeficiencies.

PubMed

Tangye, Stuart G; Deenick, Elissa K; Palendira, Umaimainthan; Ma, Cindy S

2012-02-01

Regulated interactions between cells of the immune system facilitate the generation of successful immune responses, thereby enabling efficient neutralization and clearance of pathogens and the establishment of both cell- and humoral-mediated immunological memory. The corollary of this is that impediments to efficient cell-cell interactions, normally necessary for differentiation and effector functions of immune cells, underly the clinical features and disease pathogenesis of primary immunodeficiencies. In affected individuals, these defects manifest as impaired long-term humoral immunity and susceptibility to infection by specific pathogens. In this review, we discuss the importance of, and requirements for, effective interactions between B cells and T cells during the formation of CD4(+) T follicular helper cells and the elicitation of cytotoxic function of virus-specific CD8(+) T cells, as well as how these processes are abrogated in primary immunodeficiencies due to loss-of-function mutations in defined genes. © 2012 New York Academy of Sciences.

A Web-Accessible Protein Structure Prediction Pipeline

DTIC Science & Technology

2009-06-01

Abstract Proteins are the molecular basis of nearly all structural, catalytic, sensory, and regulatory functions in living organisms. The biological...sensory, and regulatory functions in living organisms. The structure of a protein is essential in understanding its function at the molecular level...Characterizing sequence-structure and structure-function relationships have been the goals of molecular biology for more than three decades

Self-regulatory processes and exercise adherence in older adults: executive function and self-efficacy effects.

PubMed

McAuley, Edward; Mullen, Sean P; Szabo, Amanda N; White, Siobhan M; Wójcicki, Thomas R; Mailey, Emily L; Gothe, Neha P; Olson, Erin A; Voss, Michelle; Erickson, Kirk; Prakash, Ruchika; Kramer, Arthur F

2011-09-01

Self-efficacy and the use of self-regulatory strategies are consistently associated with physical activity behavior. Similarly, behavioral inhibition and cognitive resource allocation-indices of executive control function-have also been associated with this health behavior. The purpose of this study was to examine the hypothesis that self-efficacy mediates the relationship between self-regulatory processes, such as executive function, and sustained exercise behavior. Older adults (N=177, mean age=66.44 years) completed measures of executive function, self-reported use of self-regulatory strategies, and self-efficacy prior to and during the first month of a 12-month exercise intervention. Percentage of exercise classes attended over the following 11 months was used to represent adherence. Data were collected from 2007 to 2010 and analyzed in 2010-2011. Structural equation models were tested examining the effect of executive function and strategy use on adherence via efficacy. As hypothesized, results showed significant direct effects of two elements of executive function and of strategy use on self-efficacy and of efficacy on adherence. In addition, there were significant indirect effects of strategy use and executive function on adherence via self-efficacy. Higher levels of executive function and use of self-regulatory strategies at the start of an exercise program enhance beliefs in exercise capabilities, which in turn leads to greater adherence. Copyright © 2011 American Journal of Preventive Medicine. Published by Elsevier Inc. All rights reserved.

Health Risks and Adverse Reactions to Functional Foods.

PubMed

Ameratunga, Rohan; Crooks, Christine; Simmons, Greg; Woon, See-Tarn

2016-01-01

Functional foods have become increasingly popular with consumers anxious to mitigate the effects of an unhealthy lifestyle or aging. In spite of attractive health claims, these products do not have legal or regulatory status in most countries and are regulated through their health claims. Regulation of functional foods by health claims does not address health risks and adverse effects of these products. In this essay regulatory aspects of functional foods are reviewed along with adverse effects published in the peer-reviewed literature. We detail why the lack of an internationally accepted definition of functional foods places consumers at risk of adverse outcomes. Our review will assist regulatory agencies, manufacturers and consumer groups to assess the benefits and reduce the risks associated with these products.

Betaine supplementation mitigates cisplatin-induced nephrotoxicity by abrogation of oxidative/nitrosative stress and suppression of inflammation and apoptosis in rats.

PubMed

Hagar, Hanan; Medany, Azza El; Salam, Reem; Medany, Gamila El; Nayal, Omina A

2015-02-01

Cisplatin is one of the most potent chemotherapeutic antitumor drugs used in the treatment of a wide range of solid tumors. Its primary dose-limiting side effect is nephrotoxicity. This study aims to investigate the effect of betaine supplementation on cisplatin-induced nephrotoxicity. A single intraperitoneal injection of cisplatin (5mg/kg) deteriorated the kidney functions as reflected by elevated blood urea nitrogen and serum creatinine levels. Oxidative/nitrosative stress was evident in cisplatin group by increased renal thiobarbituric acid-reactive substances (TBARS), an indicator of lipid peroxidation, reduced renal total antioxidant status and increased renal nitrite concentration. Cisplatin resulted in a decline in the concentrations of reduced glutathione, glutathione peroxidase, catalase, and superoxide dismutase in renal tissues. Renal tumor necrosis factor-α (TNF-α) was also elevated. Expressions of nuclear factor-kappa B (NF-κB) and caspase-3 were up-regulated in renal tissues as indicated by immunohistochemical analysis. Histopathological changes were observed in cisplatin group. Betaine supplementation (250 mg/kg/day) orally via gavage for 21 days prior to cisplatin injection was able to protect against deterioration in kidney function, abrogate the decline in antioxidants enzymes and suppressed the increase in TBARS, nitrite and TNF-α concentrations. Moreover, betaine inhibited NF-κB and caspase-3 activation and improved the histological changes induced by cisplatin. Thus, the present study demonstrated the renoprotective nature of betaine by attenuating the pro-inflammatory and apoptotic mediators and improving antioxidant competence in kidney tissues of cisplatin treated rats. Betaine could be a beneficial dietary supplement to attenuate cisplatin nephrotoxicity. Copyright © 2014 Elsevier GmbH. All rights reserved.

Novel Mps1 Kinase Inhibitors with Potent Antitumor Activity.

PubMed

Wengner, Antje M; Siemeister, Gerhard; Koppitz, Marcus; Schulze, Volker; Kosemund, Dirk; Klar, Ulrich; Stoeckigt, Detlef; Neuhaus, Roland; Lienau, Philip; Bader, Benjamin; Prechtl, Stefan; Raschke, Marian; Frisk, Anna-Lena; von Ahsen, Oliver; Michels, Martin; Kreft, Bertolt; von Nussbaum, Franz; Brands, Michael; Mumberg, Dominik; Ziegelbauer, Karl

2016-04-01

Monopolar spindle 1 (Mps1) has been shown to function as the key kinase that activates the spindle assembly checkpoint (SAC) to secure proper distribution of chromosomes to daughter cells. Here, we report the structure and functional characterization of two novel selective Mps1 inhibitors, BAY 1161909 and BAY 1217389, derived from structurally distinct chemical classes. BAY 1161909 and BAY 1217389 inhibited Mps1 kinase activity with IC50 values below 10 nmol/L while showing an excellent selectivity profile. In cellular mechanistic assays, both Mps1 inhibitors abrogated nocodazole-induced SAC activity and induced premature exit from mitosis ("mitotic breakthrough"), resulting in multinuclearity and tumor cell death. Both compounds efficiently inhibited tumor cell proliferation in vitro (IC50 nmol/L range). In vivo, BAY 1161909 and BAY 1217389 achieved moderate efficacy in monotherapy in tumor xenograft studies. However, in line with its unique mode of action, when combined with paclitaxel, low doses of Mps1 inhibitor reduced paclitaxel-induced mitotic arrest by the weakening of SAC activity. As a result, combination therapy strongly improved efficacy over paclitaxel or Mps1 inhibitor monotreatment at the respective MTDs in a broad range of xenograft models, including those showing acquired or intrinsic paclitaxel resistance. Both Mps1 inhibitors showed good tolerability without adding toxicity to paclitaxel monotherapy. These preclinical findings validate the innovative concept of SAC abrogation for cancer therapy and justify clinical proof-of-concept studies evaluating the Mps1 inhibitors BAY 1161909 and BAY 1217389 in combination with antimitotic cancer drugs to enhance their efficacy and potentially overcome resistance. Mol Cancer Ther; 15(4); 583-92. ©2016 AACR. ©2016 American Association for Cancer Research.

Endothelial microparticle-mediated transfer of MicroRNA-126 promotes vascular endothelial cell repair via SPRED1 and is abrogated in glucose-damaged endothelial microparticles.

PubMed

Jansen, Felix; Yang, Xiaoyan; Hoelscher, Marion; Cattelan, Arianna; Schmitz, Theresa; Proebsting, Sebastian; Wenzel, Daniela; Vosen, Sarah; Franklin, Bernardo S; Fleischmann, Bernd K; Nickenig, Georg; Werner, Nikos

2013-10-29

Repair of the endothelium after vascular injury is crucial for preserving endothelial integrity and preventing the development of vascular disease. The underlying mechanisms of endothelial cell repair are largely unknown. We sought to investigate whether endothelial microparticles (EMPs), released from apoptotic endothelial cells (ECs), influence EC repair. Systemic treatment of mice with EMPs after electric denudation of the endothelium accelerated reendothelialization in vivo. In vitro experiments revealed that EMP uptake in ECs promotes EC migration and proliferation, both critical steps in endothelial repair. To dissect the underlying mechanisms, Taqman microRNA array was performed, and microRNA (miR)-126 was identified as the predominantly expressed miR in EMPs. The following experiments demonstrated that miR-126 was transported into recipient human coronary artery endothelial cells by EMPs and functionally regulated the target protein sprouty-related, EVH1 domain-containing protein 1 (SPRED1). Knockdown of miR-126 in EMPs abrogated EMP-mediated effects on human coronary artery endothelial cell migration and proliferation in vitro and reendothelialization in vivo. Interestingly, after simulating diabetic conditions, EMPs derived from glucose-treated ECs contained significantly lower amounts of miR-126 and showed reduced endothelial repair capacity in vitro and in vivo. Finally, expression analysis of miR-126 in circulating microparticles from 176 patients with stable coronary artery disease with and without diabetes mellitus revealed a significantly reduced miR-126 expression in circulating microparticles from diabetic patients. Endothelial microparticles promote vascular endothelial repair by delivering functional miR-126 into recipient cells. In pathological hyperglycemic conditions, EMP-mediated miR-126-induced EC repair is altered.

Predictive computation of genomic logic processing functions in embryonic development

PubMed Central

Peter, Isabelle S.; Faure, Emmanuel; Davidson, Eric H.

2012-01-01

Gene regulatory networks (GRNs) control the dynamic spatial patterns of regulatory gene expression in development. Thus, in principle, GRN models may provide system-level, causal explanations of developmental process. To test this assertion, we have transformed a relatively well-established GRN model into a predictive, dynamic Boolean computational model. This Boolean model computes spatial and temporal gene expression according to the regulatory logic and gene interactions specified in a GRN model for embryonic development in the sea urchin. Additional information input into the model included the progressive embryonic geometry and gene expression kinetics. The resulting model predicted gene expression patterns for a large number of individual regulatory genes each hour up to gastrulation (30 h) in four different spatial domains of the embryo. Direct comparison with experimental observations showed that the model predictively computed these patterns with remarkable spatial and temporal accuracy. In addition, we used this model to carry out in silico perturbations of regulatory functions and of embryonic spatial organization. The model computationally reproduced the altered developmental functions observed experimentally. Two major conclusions are that the starting GRN model contains sufficiently complete regulatory information to permit explanation of a complex developmental process of gene expression solely in terms of genomic regulatory code, and that the Boolean model provides a tool with which to test in silico regulatory circuitry and developmental perturbations. PMID:22927416

Human epidermal growth factor receptor bispecific ligand trap RB200: abrogation of collagen-induced arthritis in combination with tumour necrosis factor blockade

PubMed Central

2011-01-01

Introduction Rheumatoid arthritis (RA) is a chronic disease associated with inflammation and destruction of bone and cartilage. Although inhibition of TNFα is widely used to treat RA, a significant number of patients do not respond to TNFα blockade, and therefore there is a compelling need to continue to identify alternative therapeutic strategies for treating chronic inflammatory diseases such as RA. The anti-epidermal growth factor (anti-EGF) receptor antibody trastuzumab has revolutionised the treatment of patients with EGF receptor-positive breast cancer. Expression of EGF ligands and receptors (known as HER) has also been documented in RA. The highly unique compound RB200 is a bispecific ligand trap that is composed of full-length extracellular domains of HER1 and HER3 EGF receptors. Because of its pan-HER specificity, RB200 inhibits responses mediated by HER1, HER2 and HER3 in vitro and in vivo. The objective of this study was to assess the effect of RB200 combined with TNF blockade in a murine collagen-induced arthritis (CIA) model of RA. Methods Arthritic mice were treated with RB200 alone or in combination with the TNF receptor fusion protein etanercept. We performed immunohistochemistry to assess CD31 and in vivo fluorescent imaging using anti-E-selectin antibody labelled with fluorescent dye to elucidate the effect of RB200 on the vasculature in CIA. Results RB200 significantly abrogated CIA by reducing paw swelling and clinical scores. Importantly, low-dose RB200 combined with a suboptimal dose of etanercept led to complete abrogation of arthritis. Moreover, the combination of RB200 with etanercept abrogated the intensity of the E-selectin-targeted signal to the level seen in control animals not immunised to CIA. Conclusions The human pan-EGF receptor bispecific ligand trap RB200, when combined with low-dose etanercept, abrogates CIA, suggesting that inhibition of events downstream of EGF receptor activation, in combination with TNFα inhibitors, may hold promise as a future therapy for patients with RA. PMID:21982514

Deleterious impact of hyperglycemia on cystic fibrosis airway ion transport and epithelial repair.

PubMed

Bilodeau, Claudia; Bardou, Olivier; Maillé, Émilie; Berthiaume, Yves; Brochiero, Emmanuelle

2016-01-01

Cystic fibrosis (CF)-related diabetes (CFRD) is associated with faster pulmonary function decline. Thus, we evaluated the impact of hyperglycemia on airway epithelial repair and transepithelial ion transport, which are critical in maintaining lung integrity and function. Non-CF and CF airway epithelial cells were exposed to low (LG) or high (HG) glucose before ion current and wound repair rate measurements. CFTR and K+ currents decreased after HG treatments. HG also reduced the wound healing rates of non-CF and CF cell monolayers. Although CFTR correction with VRT-325 accelerated the healing rates of CF cells monolayers under LG conditions, this improvement was significantly abrogated under HG conditions. Our data highlights a deleterious impact of hyperglycemia on ion transport and epithelial repair functions, which could contribute to the deterioration in lung function in CFRD patients. HG may also interfere with the beneficial effects of CFTR rescue on airway epithelial repair. Copyright © 2015 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

The Role of Diacylglycerol Acyltransferase (DGAT) 1 and 2 in Cardiac Metabolism and Function.

PubMed

Roe, Nathan D; Handzlik, Michal K; Li, Tao; Tian, Rong

2018-03-21

It is increasingly recognized that synthesis and turnover of cardiac triglyceride (TG) play a pivotal role in the regulation of lipid metabolism and function of the heart. The last step in TG synthesis is catalyzed by diacylglycerol:acyltransferase (DGAT) which esterifies the diacylglycerol with a fatty acid. Mammalian heart has two DGAT isoforms, DGAT1 and DGAT2, yet their roles in cardiac metabolism and function remain poorly defined. Here, we show that inactivation of DGAT1 or DGAT2 in adult mouse heart results in a moderate suppression of TG synthesis and turnover. Partial inhibition of DGAT activity increases cardiac fatty acid oxidation without affecting PPARα signaling, myocardial energetics or contractile function. Moreover, coinhibition of DGAT1/2 in the heart abrogates TG turnover and protects the heart against high fat diet-induced lipid accumulation with no adverse effects on basal or dobutamine-stimulated cardiac function. Thus, the two DGAT isoforms in the heart have partially redundant function, and pharmacological inhibition of one DGAT isoform is well tolerated in adult hearts.

Short-lived non-coding transcripts (SLiTs): Clues to regulatory long non-coding RNA.

PubMed

Tani, Hidenori

2017-03-22

Whole transcriptome analyses have revealed a large number of novel long non-coding RNAs (lncRNAs). Although the importance of lncRNAs has been documented in previous reports, the biological and physiological functions of lncRNAs remain largely unknown. The role of lncRNAs seems an elusive problem. Here, I propose a clue to the identification of regulatory lncRNAs. The key point is RNA half-life. RNAs with a long half-life (t 1/2 > 4 h) contain a significant proportion of ncRNAs, as well as mRNAs involved in housekeeping functions, whereas RNAs with a short half-life (t 1/2 < 4 h) include known regulatory ncRNAs and regulatory mRNAs. This novel class of ncRNAs with a short half-life can be categorized as Short-Lived non-coding Transcripts (SLiTs). I consider that SLiTs are likely to be rich in functionally uncharacterized regulatory RNAs. This review describes recent progress in research into SLiTs.

Synthetic Oligodeoxynucleotides (ODN) Containing Suppressive TTAGGG Motifs Inhibit AIM2 Inflammasome Activation

PubMed Central

Kaminski, John J.; Schattgen, Stefan A.; Tzeng, Te-Chen; Bode, Christian; Klinman, Dennis M.; Fitzgerald, Katherine A.

2013-01-01

Synthetic oligodeoxynucleotides comprised of the immunosuppressive motif TTAGGG block TLR9 signaling, prevent STAT1 and STAT4 phosphorylation and attenuate a variety of inflammatory responses in vivo. Here, we demonstrate that such suppressive oligodeoxynucleotides (sup ODN) abrogate activation of cytosolic nucleic acid sensing pathways. Pretreatment of dendritic cells and macrophages with the suppressive ODN-A151 abrogated type I IFN, TNFα and ISG induction in response to cytosolic dsDNA. In addition, A151 abrogated caspase-1-dependent IL-1β and IL-18 maturation in dendritic cells stimulated with dsDNA and murine cytomegalovirus (MCMV). Inhibition was dependent on A151’s phosphorothioate backbone while substitution of the guanosine residues for adenosine negatively affected potency. A151 mediates these effects by binding to AIM2 in a manner that is competitive with immune-stimulatory DNA and as a consequence prevents AIM2 inflammasome complex formation. Collectively, these findings reveal a new route by which suppressive ODNs modulate the immune system and unveil novel applications for suppressive ODNs in the treatment of infectious and autoimmune diseases. PMID:23986531

FOXP3: required but not sufficient. the role of GARP (LRRC32) as a safeguard of the regulatory phenotype.

PubMed

Probst-Kepper, M; Balling, R; Buer, J

2010-08-01

FOXP3 is essential for the development and function of regulatory CD4(+)CD25(hi) T (T(reg)) cells. However, recent evidence suggests that FOXP3 alone is not sufficient to completely explain the regulatory phenotype of these key players in autoimmunity and inflammation: after being activated, conventional human CD4(+) T cells transiently up-regulate FOXP3 without acquiring a regulatory function. Researchers have recently found that glycoprotein A repetitions predominant (GARP, or LRRC32) is a T(reg)-specific receptor that binds latent TGF-beta and dominantly controls FOXP3 and the regulatory phenotype via a positive feedback loop. This finding provides a missing link in our molecular understanding of FOXP3 in T(reg) cells. This viewpoint focuses on GARP as safeguard of FOXP3 and the regulatory phenotype.

Isolation of CD4+CD25+ regulatory T cells for clinical trials.

PubMed

Hoffmann, Petra; Boeld, Tina J; Eder, Ruediger; Albrecht, Julia; Doser, Kristina; Piseshka, Biserka; Dada, Ashraf; Niemand, Claudia; Assenmacher, Mario; Orsó, Evelyn; Andreesen, Reinhard; Holler, Ernst; Edinger, Matthias

2006-03-01

The adoptive transfer of donor CD4+CD25+ regulatory T cells has been shown to protect from lethal graft-versus-host disease after allogeneic bone marrow transplantation in murine disease models. Efficient isolation strategies that comply with good manufacturing practice (GMP) guidelines are prerequisites for the clinical application of human CD4+CD25+ regulatory T cells. Here we describe the isolation of CD4+CD25+ T cells with regulatory function from standard leukapheresis products by using a 2-step magnetic cell-separation protocol performed under GMP conditions. The generated cell products contained on average 49.5% CD4+CD25high T cells that phenotypically and functionally represented natural CD4+CD25+ regulatory T cells and showed a suppressive activity comparable to that of CD4+CD25+ regulatory T-cell preparations purified by non-GMP-approved fluorescence-activated cell sorting.

Loss of p53 induces M-phase retardation following G2 DNA damage checkpoint abrogation.

PubMed

Minemoto, Yuzuru; Uchida, Sanae; Ohtsubo, Motoaki; Shimura, Mari; Sasagawa, Toshiyuki; Hirata, Masato; Nakagama, Hitoshi; Ishizaka, Yukihito; Yamashita, Katsumi

2003-04-01

Most cell lines that lack functional p53 protein are arrested in the G2 phase of the cell cycle due to DNA damage. When the G2 checkpoint is abrogated, these cells are forced into mitotic catastrophe. A549 lung adenocarcinoma cells, in which p53 was eliminated with the HPV16 E6 gene, exhibited efficient arrest in the G2 phase when treated with adriamycin. Administration of caffeine to G2-arrested cells induced a drastic change in cell phenotype, the nature of which depended on the status of p53. Flow cytometric and microscopic observations revealed that cells that either contained or lacked p53 resumed their cell cycles and entered mitosis upon caffeine treatment. However, transit to the M phase was slower in p53-negative cells than in p53-positive cells. Consistent with these observations, CDK1 activity was maintained at high levels, along with stable cyclin B1, in p53-negative cells. The addition of butyrolactone I, which is an inhibitor of CDK1 and CDK2, to the p53-negative cells reduced the floating round cell population and induced the disappearance of cyclin B1. These results suggest a relationship between the p53 pathway and the ubiquitin-mediated degradation of mitotic cyclins and possible cross-talk between the G2-DNA damage checkpoint and the mitotic checkpoint.

Molecular Mapping of the RNA Cap 2′-O-Methyltransferase Activation Interface between Severe Acute Respiratory Syndrome Coronavirus nsp10 and nsp16*

PubMed Central

Lugari, Adrien; Betzi, Stephane; Decroly, Etienne; Bonnaud, Emmanuel; Hermant, Aurélie; Guillemot, Jean-Claude; Debarnot, Claire; Borg, Jean-Paul; Bouvet, Mickaël; Canard, Bruno; Morelli, Xavier; Lécine, Patrick

2010-01-01

Several protein-protein interactions within the SARS-CoV proteome have been identified, one of them being between non-structural proteins nsp10 and nsp16. In this work, we have mapped key residues on the nsp10 surface involved in this interaction. Alanine-scanning mutagenesis, bioinformatics, and molecular modeling were used to identify several “hot spots,” such as Val42, Met44, Ala71, Lys93, Gly94, and Tyr96, forming a continuous protein-protein surface of about 830 Å2, bearing very conserved amino acids among coronaviruses. Because nsp16 carries RNA cap 2′-O-methyltransferase (2′O-MTase) activity only in the presence of its interacting partner nsp10 (Bouvet, M., Debarnot, C., Imbert, I., Selisko, B., Snijder, E. J., Canard, B., and Decroly, E. (2010) PLoS Pathog. 6, e1000863), functional consequences of mutations on this surface were evaluated biochemically. Most changes that disrupted the nsp10-nsp16 interaction without structural perturbations were shown to abrogate stimulation of nsp16 RNA cap 2′O-MTase activity. More strikingly, the Y96A mutation abrogates stimulation of nsp16 2′O-MTase activity, whereas Y96F overstimulates it. Thus, the nsp10-nsp16 interface may represent an attractive target for antivirals against human and animal pathogenic coronaviruses. PMID:20699222

Whole-genome duplication increases tumor cell sensitivity to MPS1 inhibition.

PubMed

Jemaà, Mohamed; Manic, Gwenola; Lledo, Gwendaline; Lissa, Delphine; Reynes, Christelle; Morin, Nathalie; Chibon, Frédéric; Sistigu, Antonella; Castedo, Maria; Vitale, Ilio; Kroemer, Guido; Abrieu, Ariane

2016-01-05

Several lines of evidence indicate that whole-genome duplication resulting in tetraploidy facilitates carcinogenesis by providing an intermediate and metastable state more prone to generate oncogenic aneuploidy. Here, we report a novel strategy to preferentially kill tetraploid cells based on the abrogation of the spindle assembly checkpoint (SAC) via the targeting of TTK protein kinase (better known as monopolar spindle 1, MPS1). The pharmacological inhibition as well as the knockdown of MPS1 kills more efficiently tetraploid cells than their diploid counterparts. By using time-lapse videomicroscopy, we show that tetraploid cells do not survive the aborted mitosis due to SAC abrogation upon MPS1 depletion. On the contrary diploid cells are able to survive up to at least two more cell cycles upon the same treatment. This effect might reflect the enhanced difficulty of cells with whole-genome doubling to tolerate a further increase in ploidy and/or an elevated level of chromosome instability in the absence of SAC functions. We further show that MPS1-inhibited tetraploid cells promote mitotic catastrophe executed by the intrinsic pathway of apoptosis, as indicated by the loss of mitochondrial potential, the release of the pro-apoptotic cytochrome c from mitochondria, and the activation of caspases. Altogether, our results suggest that MPS1 inhibition could be used as a therapeutic strategy for targeting tetraploid cancer cells.

Specific inhibition of NLRP3 in chikungunya disease reveals a role for inflammasomes in alphavirus-induced inflammation.

PubMed

Chen, Weiqiang; Foo, Suan-Sin; Zaid, Ali; Teng, Terk-Shin; Herrero, Lara J; Wolf, Stefan; Tharmarajah, Kothila; Vu, Luan D; van Vreden, Caryn; Taylor, Adam; Freitas, Joseph R; Li, Rachel W; Woodruff, Trent M; Gordon, Richard; Ojcius, David M; Nakaya, Helder I; Kanneganti, Thirumala-Devi; O'Neill, Luke A J; Robertson, Avril A B; King, Nicholas J; Suhrbier, Andreas; Cooper, Matthew A; Ng, Lisa F P; Mahalingam, Suresh

2017-10-01

Mosquito-borne viruses can cause severe inflammatory diseases and there are limited therapeutic solutions targeted specifically at virus-induced inflammation. Chikungunya virus (CHIKV), a re-emerging alphavirus responsible for several outbreaks worldwide in the past decade, causes debilitating joint inflammation and severe pain. Here, we show that CHIKV infection activates the NLRP3 inflammasome in humans and mice. Peripheral blood mononuclear cells isolated from CHIKV-infected patients showed elevated NLRP3, caspase-1 and interleukin-18 messenger RNA expression and, using a mouse model of CHIKV infection, we found that high NLRP3 expression was associated with peak inflammatory symptoms. Inhibition of NLRP3 activation using the small-molecule inhibitor MCC950 resulted in reduced CHIKV-induced inflammation and abrogated osteoclastogenic bone loss and myositis, but did not affect in vivo viral replication. Mice treated with MCC950 displayed lower expression levels of the cytokines interleukin-6, chemokine ligand 2 and tumour necrosis factor in joint tissue. Interestingly, MCC950 treatment abrogated disease signs in mice infected with a related arthritogenic alphavirus, Ross River virus, but not in mice infected with West Nile virus-a flavivirus. Here, using mouse models of alphavirus-induced musculoskeletal disease, we demonstrate that NLRP3 inhibition in vivo can reduce inflammatory pathology and that further development of therapeutic solutions targeting inflammasome function could help treat arboviral diseases.

Up-regulation of Ciliary Neurotrophic Factor in Astrocytes by Aspirin

PubMed Central

Modi, Khushbu K.; Sendtner, Michael; Pahan, Kalipada

2013-01-01

Ciliary neurotrophic factor (CNTF) is a promyelinating trophic factor, and the mechanisms by which CNTF expression could be increased in the brain are poorly understood. Acetylsalicylic acid (aspirin) is one of the most widely used analgesics. Interestingly, aspirin increased mRNA and protein expression of CNTF in primary mouse and human astrocytes in a dose- and time-dependent manner. Aspirin induced the activation of protein kinase A (PKA) but not protein kinase C (PKC). H-89, an inhibitor of PKA, abrogated aspirin-induced expression of CNTF. The activation of cAMP-response element-binding protein (CREB), but not NF-κB, by aspirin, the abrogation of aspirin-induced expression of CNTF by siRNA knockdown of CREB, the presence of a consensus cAMP-response element in the promoter of CNTF, and the recruitment of CREB and CREB-binding protein to the CNTF promoter by aspirin suggest that aspirin increases the expression of the Cntf gene via the activation of CREB. Furthermore, we demonstrate that aspirin-induced astroglial CNTF was also functionally active and that supernatants of aspirin-treated astrocytes of wild type, but not Cntf null, mice increased myelin-associated proteins in oligodendrocytes and protected oligodendrocytes from TNF-α insult. These results highlight a new and novel myelinogenic property of aspirin, which may be of benefit for multiple sclerosis and other demyelinating disorders. PMID:23653362

Comparative genomics of metabolic capacities of regulons controlled by cis-regulatory RNA motifs in bacteria.

PubMed

Sun, Eric I; Leyn, Semen A; Kazanov, Marat D; Saier, Milton H; Novichkov, Pavel S; Rodionov, Dmitry A

2013-09-02

In silico comparative genomics approaches have been efficiently used for functional prediction and reconstruction of metabolic and regulatory networks. Riboswitches are metabolite-sensing structures often found in bacterial mRNA leaders controlling gene expression on transcriptional or translational levels.An increasing number of riboswitches and other cis-regulatory RNAs have been recently classified into numerous RNA families in the Rfam database. High conservation of these RNA motifs provides a unique advantage for their genomic identification and comparative analysis. A comparative genomics approach implemented in the RegPredict tool was used for reconstruction and functional annotation of regulons controlled by RNAs from 43 Rfam families in diverse taxonomic groups of Bacteria. The inferred regulons include ~5200 cis-regulatory RNAs and more than 12000 target genes in 255 microbial genomes. All predicted RNA-regulated genes were classified into specific and overall functional categories. Analysis of taxonomic distribution of these categories allowed us to establish major functional preferences for each analyzed cis-regulatory RNA motif family. Overall, most RNA motif regulons showed predictable functional content in accordance with their experimentally established effector ligands. Our results suggest that some RNA motifs (including thiamin pyrophosphate and cobalamin riboswitches that control the cofactor metabolism) are widespread and likely originated from the last common ancestor of all bacteria. However, many more analyzed RNA motifs are restricted to a narrow taxonomic group of bacteria and likely represent more recent evolutionary innovations. The reconstructed regulatory networks for major known RNA motifs substantially expand the existing knowledge of transcriptional regulation in bacteria. The inferred regulons can be used for genetic experiments, functional annotations of genes, metabolic reconstruction and evolutionary analysis. The obtained genome-wide collection of reference RNA motif regulons is available in the RegPrecise database (http://regprecise.lbl.gov/).

Abasy Atlas: a comprehensive inventory of systems, global network properties and systems-level elements across bacteria

PubMed Central

Ibarra-Arellano, Miguel A.; Campos-González, Adrián I.; Treviño-Quintanilla, Luis G.; Tauch, Andreas; Freyre-González, Julio A.

2016-01-01

The availability of databases electronically encoding curated regulatory networks and of high-throughput technologies and methods to discover regulatory interactions provides an invaluable source of data to understand the principles underpinning the organization and evolution of these networks responsible for cellular regulation. Nevertheless, data on these sources never goes beyond the regulon level despite the fact that regulatory networks are complex hierarchical-modular structures still challenging our understanding. This brings the necessity for an inventory of systems across a large range of organisms, a key step to rendering feasible comparative systems biology approaches. In this work, we take the first step towards a global understanding of the regulatory networks organization by making a cartography of the functional architectures of diverse bacteria. Abasy (Across-bacteria systems) Atlas provides a comprehensive inventory of annotated functional systems, global network properties and systems-level elements (global regulators, modular genes shaping functional systems, basal machinery genes and intermodular genes) predicted by the natural decomposition approach for reconstructed and meta-curated regulatory networks across a large range of bacteria, including pathogenically and biotechnologically relevant organisms. The meta-curation of regulatory datasets provides the most complete and reliable set of regulatory interactions currently available, which can even be projected into subsets by considering the force or weight of evidence supporting them or the systems that they belong to. Besides, Abasy Atlas provides data enabling large-scale comparative systems biology studies aimed at understanding the common principles and particular lifestyle adaptions of systems across bacteria. Abasy Atlas contains systems and system-level elements for 50 regulatory networks comprising 78 649 regulatory interactions covering 42 bacteria in nine taxa, containing 3708 regulons and 1776 systems. All this brings together a large corpus of data that will surely inspire studies to generate hypothesis regarding the principles governing the evolution and organization of systems and the functional architectures controlling them. Database URL: http://abasy.ccg.unam.mx PMID:27242034

Iodide Protects Heart Tissue from Reperfusion Injury

PubMed Central

Iwata, Akiko; Morrison, Michael L.; Roth, Mark B.

2014-01-01

Iodine is an elemental nutrient that is essential for mammals. Here we provide evidence for an acute therapeutic role for iodine in ischemia reperfusion injury. Infusion of the reduced form, iodide, but not the oxidized form iodate, reduces heart damage by as much as 75% when delivered intravenously following temporary loss of blood flow but prior to reperfusion of the heart in a mouse model of acute myocardial infarction. Normal thyroid function may be required because loss of thyroid activity abrogates the iodide benefit. Given the high degree of protection and the high degree of safety, iodide should be explored further as a therapy for reperfusion injury. PMID:25379708

Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) and IPEX-related disorders: an evolving web of heritable autoimmune diseases.

PubMed

Verbsky, James W; Chatila, Talal A

2013-12-01

To summarize recent progress in our understanding of immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) and IPEX-related disorders. A number of Mendelian disorders of immune dysregulation and autoimmunity have been noted to result from defects in T regulatory cell, development and function. The best characterized of these is IPEX, resulting from mutations affecting FOXP3. A number of other gene defects that affect T regulatory cell function also give rise to IPEX-related phenotypes, including loss-of-function mutations in CD25, STAT5b and ITCH. Recent progress includes the identification of gain-of-function mutations in STAT1 as a cause of an IPEX-like disease, emerging FOXP3 genotype/phenotype relationships in IPEX, and the elucidation of a role for the microbiota in the immune dysregulation associated with regulatory T cell deficiency. An expanding spectrum of genetic defects that compromise T regulatory cell function underlies human disorders of immune dysregulation and autoimmunity. Collectively, these disorders offer novel insights into pathways of peripheral tolerance and their disruption in autoimmunity.

Experimental asthma persists in IL-33 receptor knockout mice because of the emergence of thymic stromal lymphopoietin-driven IL-9+ and IL-13+ type 2 innate lymphoid cell subpopulations.

PubMed

Verma, Mukesh; Liu, Sucai; Michalec, Lidia; Sripada, Anand; Gorska, Magdalena M; Alam, Rafeul

2017-11-10

IL-33 plays an important role in the development of experimental asthma. We sought to study the role of the IL-33 receptor suppressor of tumorigenicity 2 (ST2) in the persistence of asthma in a mouse model. We studied allergen-induced experimental asthma in ST2 knockout (KO) and wild-type control mice. We measured airway hyperresponsiveness by using flexiVent; inflammatory indices by using ELISA, histology, and real-time PCR; and type 2 innate lymphoid cells (ILC2s) in lung single-cell preparations by using flow cytometry. Airway hyperresponsiveness was increased in allergen-treated ST2 KO mice and comparable with that in allergen-treated wild-type control mice. Peribronchial and perivascular inflammation and mucus production were largely similar in both groups. Persistence of experimental asthma in ST2 KO mice was associated with an increase in levels of thymic stromal lymphopoietin (TSLP), IL-9, and IL-13, but not IL-5, in bronchoalveolar lavage fluid. Expectedly, ST2 deletion caused a reduction in IL-13 + CD4 T cells, forkhead box P3-positive regulatory T cells, and IL-5 + ILC2s. Unexpectedly, ST2 deletion led to an overall increase in innate lymphoid cells (CD45 + lin - CD25 + cells) and IL-13 + ILC2s, emergence of a TSLP receptor-positive IL-9 + ILC2 population, and an increase in intraepithelial mast cell numbers in the lung. An anti-TSLP antibody abrogated airway hyperresponsiveness, inflammation, and mucus production in allergen-treated ST2 KO mice. It also caused a reduction in innate lymphoid cell, ILC2, and IL-9 + and IL-13 + ILC2 numbers in the lung. Genetic deletion of the IL-33 receptor paradoxically increases TSLP production, which stimulates the emergence of IL-9 + and IL-13 + ILC2s and mast cells and leads to development of chronic experimental asthma. An anti-TSLP antibody abrogates all pathologic features of asthma in this model. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Identification and characterization of a silencer regulatory element in the 3'-flanking region of the murine CD46 gene.

PubMed Central

Nomura, M; Tsujimura, A; Begum, N A; Matsumoto, M; Wabiko, H; Toyoshima, K; Seya, T

2000-01-01

The murine membrane cofactor protein (CD46) gene is expressed exclusively in testis, in contrast to human CD46, which is expressed ubiquitously. To elucidate the mechanism of differential CD46 gene expression among species, we cloned entire murine CD46 genomic DNA and possible regulatory regions were placed in the flanking region of the luciferase reporter gene. The reporter gene assay revealed a silencing activity not in the promoter, but in the 3'-flanking region of the gene and the silencer-like element was identified within a 0.2-kb region between 0.6 and 0.8 kb downstream of the stop codon. This silencer-like element was highly similar to that of the pig MHC class-I gene. The introduction of a mutation into this putative silencer element of murine CD46 resulted in an abrogation of the silencing effect. Electrophoretic mobility-shift assay indicated the presence of the binding molecule(s) for this silencer sequence in murine cell lines and tissues. A size difference of the protein-silencer-element complex was observed depending upon the solubilizers used for preparation of the nuclear extracts. A mutated silencer sequence failed to interact with the binding molecules. The level of the binding factor was lower in the testicular germ cells compared with other organs. Thus the silencer element and its binding factor may play a role in transcriptional regulation of murine CD46 gene expression. These results imply that the effects of the CD46 silencer element encompass the innate immune and reproductive systems, and in mice may determine the testicular germ-cell-dominant expression of CD46. PMID:11023821

Female-specific down-regulation of tissue-PMN drives impaired Treg and amplified effector T cell responses in autoimmune dry eye disease1

PubMed Central

Gao, Yuan; Min, Kyungji; Zhang, Yibing; Su, John; Greenwood, Matthew; Gronert, Karsten

2015-01-01

Immune-driven dry eye disease primarily affects women; the cause for this sex-specific prevalence is unknown. PMN have distinct phenotypes that drive inflammation but also regulate lymphocytes and are the rate-limiting cell for generating anti-inflammatory lipoxin A4 (LXA4). Estrogen regulates the LXA4 circuit to induce delayed female-specific wound healing in the cornea. However, the role of PMN in dry eye disease remains unexplored. We discovered a LXA4-producing tissue-PMN population in the corneal limbus, lacrimal glands and cervical lymph nodes of healthy male and female mice. These tissue-PMN, unlike inflammatory-PMN, expressed a highly amplified LXA4 circuit and were sex-specifically regulated during immune-driven dry eye disease. Desiccating stress in females, unlike in males, triggered a remarkable decrease in lymph node PMN and LXA4 formation that remained depressed during dry eye disease. Depressed lymph node PMN and LXA4 in females correlated with an increase in T effector cells (TH1 and TH17), a decrease in regulatory T cells (Treg) and increased dry eye pathogenesis. Antibody depletion of tissue-PMN abrogated LXA4 formation in lymph nodes, caused a marked increase in TH1 and TH17 and decrease in Treg cells. To establish an immune regulatory role for PMN-derived LXA4 in dry eye females were treated with LXA4. LXA4 treatment markedly inhibited TH1 and TH17 and amplified Treg cells in draining lymph nodes, while reducing dry eye pathogenesis. These results identify female-specific regulation of LXA4-producing tissue-PMN as a potential key factor in aberrant T effector cell activation and initiation of immune-driven dry eye disease. PMID:26324767

Indigo Naturalis ameliorates murine dextran sodium sulfate-induced colitis via aryl hydrocarbon receptor activation.

PubMed

Kawai, Shoichiro; Iijima, Hideki; Shinzaki, Shinichiro; Hiyama, Satoshi; Yamaguchi, Toshio; Araki, Manabu; Iwatani, Shuko; Shiraishi, Eri; Mukai, Akira; Inoue, Takahiro; Hayashi, Yoshito; Tsujii, Masahiko; Motooka, Daisuke; Nakamura, Shota; Iida, Tetsuya; Takehara, Tetsuo

2017-08-01

Indigo Naturalis (IN) is used as a traditional herbal medicine for ulcerative colitis (UC). However, the mechanisms of action of IN have not been clarified. We aimed to evaluate the efficacy of IN for ameliorating colonic inflammation. We further investigated the mechanisms of action of IN. Colitis severity was assessed in dextran sodium sulfate-induced colitis and trinitrobenzene sulfonic acid-induced colitis models with or without the oral administration of IN or indigo, which is a known major component of IN. Colonic lamina propria (LP) mononuclear cells isolated from IN-treated mice were analyzed with quantitative reverse transcription polymerase chain reaction (qRT-PCR) and flow cytometry. LP and splenic mononuclear cells cultured in vitro with IN or indigo were also analyzed. The role of the candidate receptor for indigo, the aryl hydrocarbon receptor (AhR), was analyzed using Ahr-deficient mice. Colitis severity was significantly ameliorated in the IN and indigo treatment groups compared with the control group. The mRNA expression levels of interleukin (Il)-10 and Il-22 in the LP lymphocytes were increased by IN treatment. The treatment of splenocytes with IN or indigo increased the expression of anti-inflammatory cytokines and resulted in the expansion of IL-10-producing CD4 + T cells and IL-22-producing CD3 - RORγt + cells, but not CD4 + Foxp3 + regulatory T cells. The amelioration of colitis by IN or indigo was abrogated in Ahr-deficient mice, in association with diminished regulatory cytokine production. IN and indigo ameliorated murine colitis through AhR signaling activation, suggesting that AhR could be a promising therapeutic target for UC.

Pro-apoptotic signaling induced by Retinoic acid and dsRNA is under the control of Interferon Regulatory Factor-3 in breast cancer cells.

PubMed

Bernardo, Ana R; Cosgaya, José M; Aranda, Ana; Jiménez-Lara, Ana M

2017-07-01

Breast cancer is one of the most lethal malignancies for women. Retinoic acid (RA) and double-stranded RNA (dsRNA) are considered signaling molecules with potential anticancer activity. RA, co-administered with the dsRNA mimic polyinosinic-polycytidylic acid (poly(I:C)), synergizes to induce a TRAIL (Tumor-Necrosis-Factor Related Apoptosis-Inducing Ligand)- dependent apoptotic program in breast cancer cells. Here, we report that RA/poly(I:C) co-treatment, synergically, induce the activation of Interferon Regulatory Factor-3 (IRF3) in breast cancer cells. IRF3 activation is mediated by a member of the pathogen recognition receptors, Toll-like receptor-3 (TLR3), since its depletion abrogates IRF3 activation by RA/poly(I:C) co-treatment. Besides induction of TRAIL, apoptosis induced by RA/poly(I:C) correlates with the increased expression of pro-apoptotic TRAIL receptors, TRAIL-R1/2, and the inhibition of the antagonistic receptors TRAIL-R3/4. IRF3 plays an important role in RA/poly(I:C)-induced apoptosis since IRF3 depletion suppresses caspase-8 and caspase-3 activation, TRAIL expression upregulation and apoptosis. Interestingly, RA/poly(I:C) combination synergizes to induce a bioactive autocrine/paracrine loop of type-I Interferons (IFNs) which is ultimately responsible for TRAIL and TRAIL-R1/2 expression upregulation, while inhibition of TRAIL-R3/4 expression is type-I IFN-independent. Our results highlight the importance of IRF3 and type-I IFNs signaling for the pro-apoptotic effects induced by RA and synthetic dsRNA in breast cancer cells.

3C Protease of Enterovirus D68 Inhibits Cellular Defense Mediated by Interferon Regulatory Factor 7

PubMed Central

Xiang, Zichun; Liu, Lulu; Lei, Xiaobo; Zhou, Zhuo

2015-01-01

ABSTRACT Human enterovirus 68 (EV-D68) is a member of the EV-D species, which belongs to the EV genus of the Picornaviridae family. Over the past several years, clusters of EV-D68 infections have occurred worldwide. A recent outbreak in the United States is the largest one associated with severe respiratory illness and neurological complication. Although clinical symptoms are recognized, the virus remains poorly understood. Here we report that EV-D68 inhibits innate antiviral immunity by downregulation of interferon regulatory factor 7 (IRF7), an immune factor with a pivotal role in viral pathogenesis. This process depends on 3Cpro, an EV-D68-encoded protease, to mediate IRF7 cleavage. When expressed in host cells, 3Cpro targets Q167 and Q189 within the constitutive activation domain, resulting in cleavage of IRF7. Accordingly, wild-type IRF7 is fully active. However, IRF7 cleavage abrogated its capacity to activate type I interferon expression and limit replication of EV-D68. Notably, IRF7 cleavage strictly requires the protease activity of 3Cpro. Together, these results suggest that a dynamic interplay between 3Cpro and IRF7 may determine the outcome of EV-D68 infection. IMPORTANCE EV-D68 is a globally emerging pathogen, but the molecular basis of EV-D68 pathogenesis is unclear. Here we report that EV-D68 inhibits innate immune responses by targeting an immune factor, IRF7. This involves the 3C protease encoded by EV-D68, which mediates the cleavage of IRF7. These observations suggest that the 3Cpro-IRF7 interaction may represent an interface that dictates EV-D68 infection. PMID:26608321

Nicotine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone induce cyclooxygenase-2 activity in human gastric cancer cells: Involvement of nicotinic acetylcholine receptor (nAChR) and {beta}-adrenergic receptor signaling pathways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shin, Vivian Yvonne; Jin, H.C.; Ng, Enders K.O.

Induction of cyclooxygenase-2 (COX-2) associates with cigarette smoke exposure in many malignancies. Nicotine and its derivative, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), are the two important components in cigarette smoke that contributes to cancer development. However, the molecular mechanism(s) by which nicotine or NNK promotes gastric carcinogenesis remains largely unknown. We found that nicotine and NNK significantly enhanced cell proliferation in AGS cells that expressed both alpha7 nicotinic acetylcholine receptor ({alpha}7 nAChR) and {beta}-adrenergic receptors. Treatment of cells with {alpha}-bungarotoxin ({alpha}-BTX, {alpha}7nAChR antagonist) or propranolol ({beta}-adrenergic receptor antagonist) blocked NNK-induced COX-2/PGE{sub 2} and cell proliferation, while nicotine-mediated cell growth and COX-2/PGE{sub 2} induction canmore » only be suppressed by propranolol, but not {alpha}-BTX. Moreover, in contrast to the dependence of growth promoting effect of nicotine on Erk activation, inhibitor of p38 mitogen-activated protein kinase (MAPK) repressed NNK-induced COX-2 upregulation and resulted in suppression of cell growth. In addition, nicotine and NNK mediated COX-2 induction via different receptors to modulate several G1/S transition regulatory proteins and promote gastric cancer cell growth. Selective COX-2 inhibitor (SC-236) caused G1 arrest and abrogated nicotine/NNK-induced cell proliferation. Aberrant expression of cyclin D1 and other G1 regulatory proteins are reversed by blockade of COX-2. These results pointed to the importance of adrenergic and nicotinic receptors in gastric tumor growth through MAPK/COX-2 activation, which may perhaps provide a chemoprevention strategy for cigarette smoke-related gastric carcinogenesis.« less

EVI1 Interferes with Myeloid Maturation via Transcriptional Repression of Cebpa, via Binding to Two Far Downstream Regulatory Elements*

PubMed Central

Wilson, Michael; Tsakraklides, Vasiliki; Tran, Minh; Xiao, Ying-Yi; Zhang, Yi; Perkins, Archibald S.

2016-01-01

One mechanism by which oncoproteins work is through perturbation of cellular maturation; understanding the mechanisms by which this occurs can lead to the development of targeted therapies. EVI1 is a zinc finger oncoprotein involved in the development of acute myeloid leukemia; previous work has shown it to interfere with the maturation of granulocytes from immature precursors. Here we investigate the mechanism by which that occurs, using an immortalized hematopoietic progenitor cell line, EML-C1, as a model system. We document that overexpression of EVI1 abrogates retinoic acid-induced maturation of EML cells into committed myeloid cells, a process that can be documented by the down-regulation of stem cell antigen-1 and acquisition of responsiveness to granulocyte-macrophage colony-stimulating factor. We show that this requires DNA binding capacity of EVI1, suggesting that downstream target genes are involved. We identify the myeloid regulator Cebpa as a target gene and identify two EVI1 binding regions within evolutionarily conserved enhancer elements at +35 and +37 kb relative to the gene. EVI1 can strongly suppress Cebpa transcription, and add-back of Cebpa into EVI1-expressing EML cells partially corrects the block in maturation. We identify the DNA sequences to which EVI1 binds at +35 and +37 kb and show that mutation of one of these releases Cebpa from EVI1-induced suppression. We observe a more complex picture in primary bone marrow cells, where EVI1 suppresses Cebpa in stem cells but not in more committed progenitors. Our data thus identify a regulatory node by which EVI1 contributes to leukemia, and this represents a possible therapeutic target for treatment of EVI1-expressing leukemia. PMID:27129260

Hepatocyte‐induced CD4+ T cell alloresponse is associated with major histocompatibility complex class II up‐regulation on hepatocytes and suppressible by regulatory T cells

PubMed Central

DeTemple, Daphne E.; Oldhafer, Felix; Falk, Christine S.; Chen‐Wacker, Chen; Figueiredo, Constanca; Kleine, Moritz; Ramackers, Wolf; Timrott, Kai; Lehner, Frank; Klempnauer, Juergen; Bock, Michael

2018-01-01

Hepatocyte transplantation is a promising therapeutic approach for various liver diseases. Despite the liver's tolerogenic potential, early immune‐mediated loss of transplanted cells is observed, and longterm acceptance has not been achieved yet. Patients deemed tolerant after liver transplantation presented an increased frequency of regulatory T cells (Tregs), which therefore also might enable reduction of posttransplant cell loss and enhance longterm allograft acceptance. We hence characterized hepatocyte‐induced immune reactions and evaluated the immunomodulatory potential of Tregs applying mixed lymphocyte cultures and mixed lymphocyte hepatocyte cultures. These were set up using peripheral blood mononuclear cells and primary human hepatocytes, respectively. Polyclonally expanded CD4+CD25highCD127low Tregs were added to cocultures in single‐/trans‐well setups with/without supplementation of anti‐interferon γ (IFNγ) antibodies. Hepatocyte‐induced alloresponses were then analyzed by multicolor flow cytometry. Measurements indicated that T cell response upon stimulation was associated with IFNγ‐induced major histocompatibility complex (MHC) class II up‐regulation on hepatocytes and mediated by CD4+ T cells. An indirect route of antigen presentation could be ruled out by use of fragmented hepatocytes and culture supernatants of hepatocytes. Allospecific proliferation was accompanied by inflammatory cytokine secretion. CD8+ T cells showed early up‐regulation of CD69 despite lack of cell proliferation in the course of coculture. Supplementation of Tregs effectively abrogated hepatocyte‐induced alloresponses and was primarily cell contact dependent. In conclusion, human hepatocytes induce a CD4+ T cell alloresponse in vitro, which is associated with MHC class II up‐regulation on hepatocytes and is susceptible to suppression by Tregs. Liver Transplantation 24 407–419 2018 AASLD. PMID:29365365

Therapeutic action of ghrelin in a mouse model of colitis.

PubMed

Gonzalez-Rey, Elena; Chorny, Alejo; Delgado, Mario

2006-05-01

Ghrelin is a novel growth hormone-releasing peptide with potential endogenous anti-inflammatory activities ameliorating some pathologic inflammatory conditions. Crohn's disease is a chronic debilitating disease characterized by severe T helper cell (Th)1-driven inflammation of the colon. The aim of this study was to investigate the therapeutic effect of ghrelin in a murine model of colitis. We examined the anti-inflammatory action of ghrelin in the colitis induced by intracolonic administration of trinitrobenzene sulfonic acid. Diverse clinical signs of the disease were evaluated, including weight loss, diarrhea, colitis, and histopathology. We also investigated the mechanisms involved in the potential therapeutic effect of ghrelin, such as inflammatory cytokines and chemokines, Th1-type response, and regulatory factors. Ghrelin ameliorated significantly the clinical and histopathologic severity of the trinitrobenzene sulfonic acid-induced colitis; abrogating body weight loss, diarrhea, and inflammation; and increasing survival. The therapeutic effect was associated with down-regulation of both inflammatory and Th1-driven autoimmune response through the regulation of a wide spectrum of inflammatory mediators. In addition, a partial involvement of interluekin-10/transforming growth factor-beta1-secreting regulatory T cells in this therapeutic effect was demonstrated. Importantly, the ghrelin treatment was therapeutically effective in established colitis and avoided the recurrence of the disease. Our data demonstrate novel anti-inflammatory actions for ghrelin in the gastrointestinal tract, ie, the capacity to deactivate the intestinal inflammatory response and to restore mucosal immune tolerance at multiple levels. Consequently, ghrelin administration represents a novel possible therapeutic approach for the treatment of Crohn's disease and other Th1-mediated inflammatory diseases, such as rheumatoid arthritis and multiple sclerosis.

Iron-Regulated Expression of Alginate Production, Mucoid Phenotype, and Biofilm Formation by Pseudomonas aeruginosa

PubMed Central

Wiens, Jacinta R.; Vasil, Adriana I.; Schurr, Michael J.; Vasil, Michael L.

2014-01-01

ABSTRACT Pseudomonas aeruginosa strains of non-cystic fibrosis (non-CF) origin do not produce significant amounts of extracellular alginate and are nonmucoid. In CF, such isolates can become mucoid through mutation of one of the genes (mucA, mucB, mucC, or mucD) that produce regulatory factors that sequester AlgU, required for increased expression of alginate genes. Mutation of the muc genes in the nonmucoid PAO1, PA14, PAKS-1, and Ps388 strains led to increased levels of extracellular alginate and an obvious mucoid phenotype, but only under iron-limiting growth conditions (≤5 µM), not under iron-replete conditions (≥10 µM). In contrast, >50% of P. aeruginosa isolates from chronic CF pulmonary infections expressed increased levels of alginate and mucoidy both under iron-limiting and iron-replete conditions (i.e., iron-constitutive phenotype). No single iron regulatory factor (e.g., Fur, PvdS) was associated with this loss of iron-regulated alginate expression and mucoidy in these CF isolates. However, the loss of only pyoverdine production, or its uptake, abrogated the ability of P. aeruginosa to produce a robust biofilm that represents the Psl-type of biofilm. In contrast, we show that mutation of the pyoverdine and pyochelin biosynthesis genes and the pyoverdine receptor (FpvA) lead to iron-constitutive expression of the key alginate biosynthesis gene, algD, and an explicitly mucoid phenotype in both iron-limiting and iron-replete conditions. These data indicate that alginate production and mucoidy, in contrast to other types of biofilms produced by P. aeruginosa, are substantially enhanced under iron limitation. These results also have compelling implications in relation to the use of iron chelators in the treatment of P. aeruginosa CF infections. PMID:24496793

Yeast PAH1-encoded phosphatidate phosphatase controls the expression of CHO1-encoded phosphatidylserine synthase for membrane phospholipid synthesis.

PubMed

Han, Gil-Soo; Carman, George M

2017-08-11

The PAH1 -encoded phosphatidate phosphatase (PAP), which catalyzes the committed step for the synthesis of triacylglycerol in Saccharomyces cerevisiae , exerts a negative regulatory effect on the level of phosphatidate used for the de novo synthesis of membrane phospholipids. This raises the question whether PAP thereby affects the expression and activity of enzymes involved in phospholipid synthesis. Here, we examined the PAP-mediated regulation of CHO1 -encoded phosphatidylserine synthase (PSS), which catalyzes the committed step for the synthesis of major phospholipids via the CDP-diacylglycerol pathway. The lack of PAP in the pah1 Δ mutant highly elevated PSS activity, exhibiting a growth-dependent up-regulation from the exponential to the stationary phase of growth. Immunoblot analysis showed that the elevation of PSS activity results from an increase in the level of the enzyme encoded by CHO1 Truncation analysis and site-directed mutagenesis of the CHO1 promoter indicated that Cho1 expression in the pah1 Δ mutant is induced through the inositol-sensitive upstream activation sequence (UAS INO ), a cis -acting element for the phosphatidate-controlled Henry (Ino2-Ino4/Opi1) regulatory circuit. The abrogation of Cho1 induction and PSS activity by a CHO1 UAS INO mutation suppressed pah1 Δ effects on lipid synthesis, nuclear/endoplasmic reticulum membrane morphology, and lipid droplet formation, but not on growth at elevated temperature. Loss of the DGK1 -encoded diacylglycerol kinase, which converts diacylglycerol to phosphatidate, partially suppressed the pah1 Δ-mediated induction of Cho1 and PSS activity. Collectively, these data showed that PAP activity controls the expression of PSS for membrane phospholipid synthesis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Characterization of Nrf2 activation and heme oxygenase-1 expression in NIH3T3 cells exposed to aqueous extracts of cigarette smoke.

PubMed

Knörr-Wittmann, Constanze; Hengstermann, Arnd; Gebel, Stephan; Alam, Jawed; Müller, Thomas

2005-12-01

Cigarette smoke (CS) is a complex chemical mixture estimated to be composed of up to 5000 different chemicals, many of which are prooxidant. Here we show that, at least in vitro, the cellular response designed to combat oxidative stress resulting from CS exposure is primarily controlled by the transcription factor Nrf2, a principal inducer of antioxidant and phase II-related genes. The prominent role of Nrf2 in the cellular response to CS is substantiated by the following observations: In NIH3T3 cells exposed to aqueous extracts of CS (i) Nrf2 is strongly stabilized and becomes detectable in nuclear extracts. (ii) Nuclear localization of Nrf2 coincides with increased DNA binding of a putative Nrf2/MafK heterodimer to its cognate cis-regulatory site, i.e., the antioxidant-responsive element (ARE). (iii) Studies on the regulatory elements of the oxidative stress-inducible gene heme oxygenase-1 (hmox1) using various hmox1 promoter/luciferase reporter constructs revealed that the strong CS-dependent expression of this gene is primarily governed by the distal enhancers 1 ("E1") and 2 ("E2"), which both contain three canonical ARE-like stress-responsive elements (StREs). Notably, depletion of Nrf2 levels caused by RNA interference significantly compromised CS-induced hmox1 promoter activation, based on the distinct Nrf2 sensitivity exhibited by E1 and E2. Finally, (iv) siRNA-dependent knock-down of Nrf2 completely abrogated CS-induced expression of phase II-related genes. Taken together, these results confirm the outstanding role of Nrf2 both in sensing (oxidant) stress and in orchestrating an efficient transcriptional response aimed at resolving the stressing conditions.

Heart grafts tolerized through third-party multipotent adult progenitor cells can be retransplanted to secondary hosts with no immunosuppression.

PubMed

Eggenhofer, Elke; Popp, Felix C; Mendicino, Michael; Silber, Paula; Van't Hof, Wouter; Renner, Philipp; Hoogduijn, Martin J; Pinxteren, Jef; van Rooijen, Nico; Geissler, Edward K; Deans, Robert; Schlitt, Hans J; Dahlke, Marc H

2013-08-01

Multipotent adult progenitor cells (MAPCs) are an adherent stem cell population that belongs to the mesenchymal-type progenitor cell family. Although MAPCs are emerging as candidate agents for immunomodulation after solid organ transplantation, their value requires further validation in a clinically relevant cell therapy model using an organ donor- and organ recipient-independent, third-party cell product. We report that stable allograft survival can be achieved following third-party MAPC infusion in a rat model of fully allogeneic, heterotopic heart transplantation. Furthermore, long-term accepted heart grafts recovered from MAPC-treated animals can be successfully retransplanted to naïve animals without additional immunosuppression. This prolongation of MAPC-mediated allograft acceptance depends upon a myeloid cell population since depletion of macrophages by clodronate abrogates the tolerogenic MAPC effect. We also show that MAPC-mediated allograft acceptance differs mechanistically from drug-induced tolerance regarding marker gene expression, T regulatory cell induction, retransplantability, and macrophage dependence. MAPC-based immunomodulation represents a promising pathway for clinical immunotherapy that has led us to initiate a phase I clinical trial for testing safety and feasibility of third-party MAPC therapy after liver transplantation.

Hyperforin is a novel type of 5-lipoxygenase inhibitor with high efficacy in vivo.

PubMed

Feisst, Christian; Pergola, Carlo; Rakonjac, Marija; Rossi, Antonietta; Koeberle, Andreas; Dodt, Gabriele; Hoffmann, Marika; Hoernig, Christina; Fischer, Lutz; Steinhilber, Dieter; Franke, Lutz; Schneider, Gisbert; Rådmark, Olof; Sautebin, Lidia; Werz, Oliver

2009-08-01

We previously showed that, in vitro, hyperforin from St. John's wort (Hypericum perforatum) inhibits 5-lipoxygenase (5-LO), the key enzyme in leukotriene biosynthesis. Here, we demonstrate that hyperforin possesses a novel and unique molecular pharmacological profile as a 5-LO inhibitor with remarkable efficacy in vivo. Hyperforin (4 mg/kg, i.p.) significantly suppressed leukotriene B(4) formation in pleural exudates of carrageenan-treated rats associated with potent anti-inflammatory effectiveness. Inhibition of 5-LO by hyperforin, but not by the iron-ligand type 5-LO inhibitor BWA4C or the nonredox-type inhibitor ZM230487, was abolished in the presence of phosphatidylcholine and strongly reduced by mutation (W13A-W75A-W102A) of the 5-LO C2-like domain. Moreover, hyperforin impaired the interaction of 5-LO with coactosin-like protein and abrogated 5-LO nuclear membrane translocation in ionomycin-stimulated neutrophils, processes that are typically mediated via the regulatory 5-LO C2-like domain. Together, hyperforin is a novel type of 5-LO inhibitor apparently acting by interference with the C2-like domain, with high effectiveness in vivo.

Cbl-phosphatidylinositol 3 kinase interaction differentially regulates macrophage colony-stimulating factor-mediated osteoclast survival and cytoskeletal reorganization.

PubMed

Adapala, Naga Suresh; Barbe, Mary F; Langdon, Wallace Y; Tsygankov, Alexander Y; Sanjay, Archana

2010-03-01

The Cbl protein is a key player in macrophage colony-stimulating factor (M-CSF)-induced signaling. To examine the role of Cbl in M-CSF-mediated cellular events, we used Cbl(YF/YF) knockin mice in which the regulatory tyrosine 737, which when phosphorylated binds to the p85 subunit of phosphatidylinositol 3 kinase (PI3K), is substituted to phenylalanine. In ex vivo cultures, M-CSF and receptor activator of nuclear factor-kappaB ligand-mediated differentiation of bone marrow precursors from Cbl(YF/YF) mice generated increased number of osteoclasts; however, osteoclast numbers in Cbl(YF/YF) cultures were unchanged with increasing doses of M-CSF. We found that Cbl(YF/YF) osteoclasts have enhanced intrinsic ability to survive, and this response was further augmented upon exposure to M-CSF. Treatment of osteoclasts with M-CSF-induced actin reorganization and lamellipodia formation in wild-type osteoclasts; however, in Cbl(YF/YF) osteoclasts lamellipodia formation was compromised. Collectively, these results indicate that abrogation of the Cbl-PI3K interaction, although not affecting M-CSF-induced proliferation and differentiation of precursors, is required for regulation of survival and actin cytoskeletal reorganization of mature osteoclasts.

The Arf-GDP-regulated recruitment of GBF1 to Golgi membranes requires domains HDS1 and HDS2 and a Golgi-localized protein receptor.

PubMed

Quilty, Douglas; Chan, Calvin J; Yurkiw, Katherine; Bain, Alexandra; Babolmorad, Ghazal; Melançon, Paul

2018-04-19

We previously proposed a novel mechanism by which the enzyme Golgi-specific Brefeldin A resistance factor 1 (GBF1) is recruited to the membranes of the cis -Golgi, based on in vivo experiments. Here, we extended our in vivo analysis on the production of regulatory Arf-GDP and observed that ArfGAP2 and ArfGAP3 do not play a role in GBF1 recruitment. We confirm that Arf-GDP localization is critical, as a TGN-localized Arf-GDP mutant protein fails to promote GBF1 recruitment. We also reported the establishment of an in vitro GBF1 recruitment assay that supports the regulation of GBF1 recruitment by Arf-GDP. This in vitro assay yielded further evidence for the requirement of a Golgi-localized protein because heat denaturation or protease treatment of Golgi membranes abrogated GBF1 recruitment. Finally, combined in vivo and in vitro measurements indicated that the recruitment to Golgi membranes via a putative receptor requires only the HDS1 and HDS2 domains in the C-terminal half of GBF1. © 2018. Published by The Company of Biologists Ltd.

rSNPBase 3.0: an updated database of SNP-related regulatory elements, element-gene pairs and SNP-based gene regulatory networks.

PubMed

Guo, Liyuan; Wang, Jing

2018-01-04

Here, we present the updated rSNPBase 3.0 database (http://rsnp3.psych.ac.cn), which provides human SNP-related regulatory elements, element-gene pairs and SNP-based regulatory networks. This database is the updated version of the SNP regulatory annotation database rSNPBase and rVarBase. In comparison to the last two versions, there are both structural and data adjustments in rSNPBase 3.0: (i) The most significant new feature is the expansion of analysis scope from SNP-related regulatory elements to include regulatory element-target gene pairs (E-G pairs), therefore it can provide SNP-based gene regulatory networks. (ii) Web function was modified according to data content and a new network search module is provided in the rSNPBase 3.0 in addition to the previous regulatory SNP (rSNP) search module. The two search modules support data query for detailed information (related-elements, element-gene pairs, and other extended annotations) on specific SNPs and SNP-related graphic networks constructed by interacting transcription factors (TFs), miRNAs and genes. (3) The type of regulatory elements was modified and enriched. To our best knowledge, the updated rSNPBase 3.0 is the first data tool supports SNP functional analysis from a regulatory network prospective, it will provide both a comprehensive understanding and concrete guidance for SNP-related regulatory studies. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

rSNPBase 3.0: an updated database of SNP-related regulatory elements, element-gene pairs and SNP-based gene regulatory networks

PubMed Central

2018-01-01

Abstract Here, we present the updated rSNPBase 3.0 database (http://rsnp3.psych.ac.cn), which provides human SNP-related regulatory elements, element-gene pairs and SNP-based regulatory networks. This database is the updated version of the SNP regulatory annotation database rSNPBase and rVarBase. In comparison to the last two versions, there are both structural and data adjustments in rSNPBase 3.0: (i) The most significant new feature is the expansion of analysis scope from SNP-related regulatory elements to include regulatory element–target gene pairs (E–G pairs), therefore it can provide SNP-based gene regulatory networks. (ii) Web function was modified according to data content and a new network search module is provided in the rSNPBase 3.0 in addition to the previous regulatory SNP (rSNP) search module. The two search modules support data query for detailed information (related-elements, element-gene pairs, and other extended annotations) on specific SNPs and SNP-related graphic networks constructed by interacting transcription factors (TFs), miRNAs and genes. (3) The type of regulatory elements was modified and enriched. To our best knowledge, the updated rSNPBase 3.0 is the first data tool supports SNP functional analysis from a regulatory network prospective, it will provide both a comprehensive understanding and concrete guidance for SNP-related regulatory studies. PMID:29140525

Th17 cell cytokine secretion profile in host defense and autoimmunity.

PubMed

Graeber, Kristen E; Olsen, Nancy J

2012-02-01

The goal of this review is to examine the effector functions of Th17 cells in host defense and autoimmunity. Published literature on Th17 cells was reviewed with a focus on the secreted products that mediate effector activities of these cells. Th17 cells secrete an array of cytokines that contribute to host defense and that bridge the innate and adaptive arms of the immune response. When this subset of T cells is dysregulated, autoimmune phenomena develop that contribute to the manifestations of many autoimmune diseases. Th17 cells are positioned at a crossroads between innate and adaptive immunity and provide mediators that are essential for host defense. Current interest in harnessing this system for treatment of autoimmune disease will be challenged by the need to avoid abrogating these many protective functions.

Functional conservation of the human EXT1 tumor suppressor gene and its Drosophila homolog tout velu.

PubMed

Dasgupta, Ujjaini; Dixit, Bharat L; Rusch, Melissa; Selleck, Scott; The, Inge

2007-08-01

Heparan sulfate proteoglycans play a vital role in signaling of various growth factors in both Drosophila and vertebrates. In Drosophila, mutations in the tout velu (ttv) gene, a homolog of the mammalian EXT1 tumor suppressor gene, leads to abrogation of glycosaminoglycan (GAG) biosynthesis. This impairs distribution and signaling activities of various morphogens such as Hedgehog (Hh), Wingless (Wg), and Decapentaplegic (Dpp). Mutations in members of the exostosin (EXT) gene family lead to hereditary multiple exostosis in humans leading to bone outgrowths and tumors. In this study, we provide genetic and biochemical evidence that the human EXT1 (hEXT1) gene is conserved through species and can functionally complement the ttv mutation in Drosophila. The hEXT1 gene was able to rescue a ttv null mutant to adulthood and restore GAG biosynthesis.

IFN-α and CD46 stimulation are associated with active lupus and skew natural T regulatory cell differentiation to type 1 regulatory T (Tr1) cells

PubMed Central

Le Buanec, Hélène; Gougeon, Marie-Lise; Mathian, Alexis; Lebon, Pierre; Dupont, Jean-Michel; Peltre, Gabriel; Hemon, Patrice; Schmid, Michel; Bizzini, Bernard; Künding, Thomas; Burny, Arsène; Bensussan, Armand; Amoura, Zahir; Gallo, Robert C.; Zagury, Daniel

2011-01-01

Immune suppressive activities exerted by regulatory T-cell subsets have several specific functions, including self-tolerance and regulation of adaptive immune reactions, and their dysfunction can lead to autoimmune diseases and contribute to AIDS and cancer. Two functionally distinct regulatory T-cell subsets are currently identified in peripheral tissues: thymus-developed natural T regulatory cells (nTregs) controlling self-tolerance and antiinflammatory IL-10–secreting type 1 regulatory T cells (Tr1) derived from Ag-stimulated T cells, which regulate inflammation-dependent adaptive immunity and minimize immunopathology. We establish herein that cell contact-mediated nTreg regulatory function is inhibited by inflammation, especially in the presence of the complement C3b receptor (CD46). Instead, as with other T-cell subsets, the latter inflammatory conditions of stimulation skew nTreg differentiation to Tr1 cells secreting IL-10, an effect potentiated by IFN-α. The clinical relevance of these findings was verified in a study of 152 lupus patients, in which we showed that lupus nTreg dysfunction is not due to intrinsic defects but is rather induced by C3b stimulation of CD46 and IFN-α and that these immune components of inflammation are directly associated with active lupus. These results provide a rationale for using anti–IFN-α Ab immunotherapy in lupus patients. PMID:22065791

ATM-NFκB axis-driven TIGAR regulates sensitivity of glioma cells to radiomimetics in the presence of TNFα.

PubMed

Sinha, S; Ghildiyal, R; Mehta, V S; Sen, E

2013-05-02

Gliomas are resistant to radiation therapy, as well as to TNFα induced killing. Radiation-induced TNFα triggers Nuclear factor κB (NFκB)-mediated radioresistance. As inhibition of NFκB activation sensitizes glioma cells to TNFα-induced apoptosis, we investigated whether TNFα modulates the responsiveness of glioma cells to ionizing radiation-mimetic Neocarzinostatin (NCS). TNFα enhanced the ability of NCS to induce glioma cell apoptosis. NCS-mediated death involved caspase-9 activation, reduction of mitochondrial copy number and lactate production. Death was concurrent with NFκB, Akt and Erk activation. Abrogation of Akt and NFκB activation further potentiated the death inducing ability of NCS in TNFα cotreated cells. NCS-induced p53 expression was accompanied by increase in TP53-induced glycolysis and apoptosis regulator (TIGAR) levels and ATM phosphorylation. siRNA-mediated knockdown of TIGAR abrogated NCS-induced apoptosis. While DN-IκB abrogated NCS-induced TIGAR both in the presence and absence of TNFα, TIGAR had no effect on NFκB activation. Transfection with TIGAR mutant (i) decreased apoptosis and γH2AX foci formation (ii) decreased p53 (iii) elevated ROS and (iv) increased Akt/Erk activation in cells cotreated with NCS and TNFα. Heightened TIGAR expression was observed in GBM tumors. While NCS induced ATM phosphorylation in a NFκB independent manner, ATM inhibition abrogated TIGAR and NFκB activation. Metabolic gene profiling indicated that TNFα affects NCS-mediated regulation of several genes associated with glycolysis. The existence of ATM-NFκB axis that regulate metabolic modeler TIGAR to overcome prosurvival response in NCS and TNFα cotreated cells, suggests mechanisms through which inflammation could affect resistance and adaptation to radiomimetics despite concurrent induction of death.

ATM-NFκB axis-driven TIGAR regulates sensitivity of glioma cells to radiomimetics in the presence of TNFα

PubMed Central

Sinha, S; Ghildiyal, R; Mehta, V S; Sen, E

2013-01-01

Gliomas are resistant to radiation therapy, as well as to TNFα induced killing. Radiation-induced TNFα triggers Nuclear factor κB (NFκB)-mediated radioresistance. As inhibition of NFκB activation sensitizes glioma cells to TNFα-induced apoptosis, we investigated whether TNFα modulates the responsiveness of glioma cells to ionizing radiation-mimetic Neocarzinostatin (NCS). TNFα enhanced the ability of NCS to induce glioma cell apoptosis. NCS-mediated death involved caspase-9 activation, reduction of mitochondrial copy number and lactate production. Death was concurrent with NFκB, Akt and Erk activation. Abrogation of Akt and NFκB activation further potentiated the death inducing ability of NCS in TNFα cotreated cells. NCS-induced p53 expression was accompanied by increase in TP53-induced glycolysis and apoptosis regulator (TIGAR) levels and ATM phosphorylation. siRNA-mediated knockdown of TIGAR abrogated NCS-induced apoptosis. While DN-IκB abrogated NCS-induced TIGAR both in the presence and absence of TNFα, TIGAR had no effect on NFκB activation. Transfection with TIGAR mutant (i) decreased apoptosis and γH2AX foci formation (ii) decreased p53 (iii) elevated ROS and (iv) increased Akt/Erk activation in cells cotreated with NCS and TNFα. Heightened TIGAR expression was observed in GBM tumors. While NCS induced ATM phosphorylation in a NFκB independent manner, ATM inhibition abrogated TIGAR and NFκB activation. Metabolic gene profiling indicated that TNFα affects NCS-mediated regulation of several genes associated with glycolysis. The existence of ATM-NFκB axis that regulate metabolic modeler TIGAR to overcome prosurvival response in NCS and TNFα cotreated cells, suggests mechanisms through which inflammation could affect resistance and adaptation to radiomimetics despite concurrent induction of death. PMID:23640457

A 3D bioprinting exemplar of the consequences of the regulatory requirements on customized processes.

PubMed

Hourd, Paul; Medcalf, Nicholas; Segal, Joel; Williams, David J

2015-01-01

Computer-aided 3D printing approaches to the industrial production of customized 3D functional living constructs for restoration of tissue and organ function face significant regulatory challenges. Using the manufacture of a customized, 3D-bioprinted nasal implant as a well-informed but hypothetical exemplar, we examine how these products might be regulated. Existing EU and USA regulatory frameworks do not account for the differences between 3D printing and conventional manufacturing methods or the ability to create individual customized products using mechanized rather than craft approaches. Already subject to extensive regulatory control, issues related to control of the computer-aided design to manufacture process and the associated software system chain present additional scientific and regulatory challenges for manufacturers of these complex 3D-bioprinted advanced combination products.

Enhancer scanning to locate regulatory regions in genomic loci

PubMed Central

Buckley, Melissa; Gjyshi, Anxhela; Mendoza-Fandiño, Gustavo; Baskin, Rebekah; Carvalho, Renato S.; Carvalho, Marcelo A.; Woods, Nicholas T.; Monteiro, Alvaro N.A.

2016-01-01

The present protocol provides a rapid, streamlined and scalable strategy to systematically scan genomic regions for the presence of transcriptional regulatory regions active in a specific cell type. It creates genomic tiles spanning a region of interest that are subsequently cloned by recombination into a luciferase reporter vector containing the Simian Virus 40 promoter. Tiling clones are transfected into specific cell types to test for the presence of transcriptional regulatory regions. The protocol includes testing of different SNP (single nucleotide polymorphism) alleles to determine their effect on regulatory activity. This procedure provides a systematic framework to identify candidate functional SNPs within a locus during functional analysis of genome-wide association studies. This protocol adapts and combines previous well-established molecular biology methods to provide a streamlined strategy, based on automated primer design and recombinational cloning to rapidly go from a genomic locus to a set of candidate functional SNPs in eight weeks. PMID:26658467

On the Concept of Cis-regulatory Information: From Sequence Motifs to Logic Functions

NASA Astrophysics Data System (ADS)

Tarpine, Ryan; Istrail, Sorin

The regulatory genome is about the “system level organization of the core genomic regulatory apparatus, and how this is the locus of causality underlying the twin phenomena of animal development and animal evolution” (E.H. Davidson. The Regulatory Genome: Gene Regulatory Networks in Development and Evolution, Academic Press, 2006). Information processing in the regulatory genome is done through regulatory states, defined as sets of transcription factors (sequence-specific DNA binding proteins which determine gene expression) that are expressed and active at the same time. The core information processing machinery consists of modular DNA sequence elements, called cis-modules, that interact with transcription factors. The cis-modules “read” the information contained in the regulatory state of the cell through transcription factor binding, “process” it, and directly or indirectly communicate with the basal transcription apparatus to determine gene expression. This endowment of each gene with the information-receiving capacity through their cis-regulatory modules is essential for the response to every possible regulatory state to which it might be exposed during all phases of the life cycle and in all cell types. We present here a set of challenges addressed by our CYRENE research project aimed at studying the cis-regulatory code of the regulatory genome. The CYRENE Project is devoted to (1) the construction of a database, the cis-Lexicon, containing comprehensive information across species about experimentally validated cis-regulatory modules; and (2) the software development of a next-generation genome browser, the cis-Browser, specialized for the regulatory genome. The presentation is anchored on three main computational challenges: the Gene Naming Problem, the Consensus Sequence Bottleneck Problem, and the Logic Function Inference Problem.

CD4+CD25+ T-Cells Control Autoimmunity in the Absence of B-Cells

PubMed Central

Mariño, Eliana; Villanueva, Jeanette; Walters, Stacey; Liuwantara, David; Mackay, Fabienne; Grey, Shane T.

2009-01-01

OBJECTIVE Tumor necrosis factor ligand family members B-cell–activating factor (BAFF) and a proliferation-inducing ligand (APRIL) can exert powerful effects on B-cell activation and development, type 1 T-helper cell (Th1) immune responses, and autoimmunity. We examined the effect of blocking BAFF and APRIL on the development of autoimmune diabetes. RESEARCH DESIGN AND METHODS Female NOD mice were administered B-cell maturation antigen (BCMA)-Fc from 9 to 15 weeks of age. Diabetes incidence, islet pathology, and T- and B-cell populations were examined. RESULTS BCMA-Fc treatment reduced the severity of insulitis and prevented diabetes development in NOD mice. BCMA-Fc–treated mice showed reduced follicular, marginal-zone, and T2MZ B-cells. B-cell reduction was accompanied by decreased frequencies of pathogenic CD4+CD40+ T-cells and reduced Th1 cytokines IL-7, IL-15, and IL-17. Thus, T-cell activation was blunted with reduced B-cells. However, BCMA-Fc–treated mice still harbored detectable diabetogenic T-cells, suggesting that regulatory mechanisms contributed to diabetes prevention. Indeed, BCMA-Fc–treated mice accumulated increased CD4+CD25+ regulatory T-cells (Tregs) with age. CD4+CD25+ cells were essential for maintaining euglycemia because their depletion abrogated BCMA-Fc–mediated protection. BCMA-Fc did not directly affect Treg homeostasis given that CD4+CD25+Foxp3+ T-cells did not express TACI or BR3 receptors and that CD4+CD25+Foxp3+ T-cell frequencies were equivalent in wild-type, BAFF−/−, TACI−/−, BCMA−/−, and BR3−/− mice. Rather, B-cell depletion resulted in CD4+CD25+ T-cell–mediated protection from diabetes because anti-CD25 monoclonal antibody treatment precipitated diabetes in both diabetes-resistant NOD.μMT−/− and BCMA-Fc–treated mice. CONCLUSIONS BAFF/APRIL blockade prevents diabetes. BCMA-Fc reduces B-cells, subsequently blunting autoimmune activity and allowing endogenous regulatory mechanisms to preserve a prehyperglycemic state. PMID:19336675

Tissue-Specific and Genetic Regulation of Insulin Sensitivity-Associated Transcripts in African Americans

PubMed Central

Sharma, Neeraj K.; Sajuthi, Satria P.; Chou, Jeff W.; Calles-Escandon, Jorge; Demons, Jamehl; Rogers, Samantha; Ma, Lijun; Palmer, Nicholette D.; McWilliams, David R.; Beal, John; Comeau, Mary E.; Cherry, Kristina; Hawkins, Gregory A.; Menon, Lata; Kouba, Ethel; Davis, Donna; Burris, Marcie; Byerly, Sara J.; Easter, Linda; Bowden, Donald W.; Freedman, Barry I.; Langefeld, Carl D.

2016-01-01

Context: Compared with European Americans, African Americans (AAs) are more insulin resistant, have a higher insulin secretion response to glucose, and develop type 2 diabetes more often. Molecular processes and/or genetic variations contributing to altered glucose homeostasis in high-risk AAs remain uncharacterized. Objective: Adipose and muscle transcript expression profiling and genotyping were performed in 260 AAs to identify genetic regulatory mechanisms associated with insulin sensitivity (SI). We hypothesized that: 1) transcription profiles would reveal tissue-specific modulation of physiologic pathways with SI, and 2) a subset of SI-associated transcripts would be controlled by DNA sequence variants as expression quantitative traits, and these variants in turn would be associated with SI. Design and Settings: The cross-sectional research study was performed in a clinical research unit. Participants: Unrelated nondiabetic AAs were recruited for the study. Main Outcome Measures: SI was measured by frequently sampled iv glucose tolerance test. Results: The expression levels of 2212 transcripts in adipose and 145 transcripts in muscle were associated with SI. Genes involved in eIF2, eIF4-p70S6K, and mTOR signaling were modulated with SI in both tissues. Genes involved in leukocyte extravasation signaling showed adipose-specific regulation, and genes involved in oxidative phosphorylation had discordant regulation between tissues. Intersecting cis-expression quantitative trait loci results with data from transcript-SI association analysis identified cis-regulatory single nucleotide polymorphisms for 363 and 42 SI-associated transcripts in adipose and muscle, respectively. Cis-eSNPs for three SI-associated adipose transcripts, NINJ1, AGA, and CLEC10A were associated with SI. Abrogation of NINJ1 induction in THP1 macrophages modulated expression of genes in chemokine signaling, cell adhesion, and angiogenesis pathways. Conclusion: This study identified multiple pathways associated with SI; particularly discordant tissue-specific regulation of the oxidative phosphorylation pathway, and adipose-specific regulation of transcripts in the leukocyte extravasation signaling pathway that seem to be important in insulin resistance. Identification of single nucleotide polymorphisms associated with SI and with modulation of expression of SI-associated transcripts, including NINJ1, reveals novel genetic regulatory mechanisms of insulin resistance in AAs. PMID:26789776

Identification and role of regulatory non-coding RNAs in Listeria monocytogenes.

PubMed

Izar, Benjamin; Mraheil, Mobarak Abu; Hain, Torsten

2011-01-01

Bacterial regulatory non-coding RNAs control numerous mRNA targets that direct a plethora of biological processes, such as the adaption to environmental changes, growth and virulence. Recently developed high-throughput techniques, such as genomic tiling arrays and RNA-Seq have allowed investigating prokaryotic cis- and trans-acting regulatory RNAs, including sRNAs, asRNAs, untranslated regions (UTR) and riboswitches. As a result, we obtained a more comprehensive view on the complexity and plasticity of the prokaryotic genome biology. Listeria monocytogenes was utilized as a model system for intracellular pathogenic bacteria in several studies, which revealed the presence of about 180 regulatory RNAs in the listerial genome. A regulatory role of non-coding RNAs in survival, virulence and adaptation mechanisms of L. monocytogenes was confirmed in subsequent experiments, thus, providing insight into a multifaceted modulatory function of RNA/mRNA interference. In this review, we discuss the identification of regulatory RNAs by high-throughput techniques and in their functional role in L. monocytogenes.

Genomic dissection of conserved transcriptional regulation in intestinal epithelial cells

PubMed Central

Camp, J. Gray; Weiser, Matthew; Cocchiaro, Jordan L.; Kingsley, David M.; Furey, Terrence S.; Sheikh, Shehzad Z.; Rawls, John F.

2017-01-01

The intestinal epithelium serves critical physiologic functions that are shared among all vertebrates. However, it is unknown how the transcriptional regulatory mechanisms underlying these functions have changed over the course of vertebrate evolution. We generated genome-wide mRNA and accessible chromatin data from adult intestinal epithelial cells (IECs) in zebrafish, stickleback, mouse, and human species to determine if conserved IEC functions are achieved through common transcriptional regulation. We found evidence for substantial common regulation and conservation of gene expression regionally along the length of the intestine from fish to mammals and identified a core set of genes comprising a vertebrate IEC signature. We also identified transcriptional start sites and other putative regulatory regions that are differentially accessible in IECs in all 4 species. Although these sites rarely showed sequence conservation from fish to mammals, surprisingly, they drove highly conserved IEC expression in a zebrafish reporter assay. Common putative transcription factor binding sites (TFBS) found at these sites in multiple species indicate that sequence conservation alone is insufficient to identify much of the functionally conserved IEC regulatory information. Among the rare, highly sequence-conserved, IEC-specific regulatory regions, we discovered an ancient enhancer upstream from her6/HES1 that is active in a distinct population of Notch-positive cells in the intestinal epithelium. Together, these results show how combining accessible chromatin and mRNA datasets with TFBS prediction and in vivo reporter assays can reveal tissue-specific regulatory information conserved across 420 million years of vertebrate evolution. We define an IEC transcriptional regulatory network that is shared between fish and mammals and establish an experimental platform for studying how evolutionarily distilled regulatory information commonly controls IEC development and physiology. PMID:28850571

The development of regulatory functions from birth to 5 years: insights from premature infants.

PubMed

Feldman, Ruth

2009-01-01

This study examined physiological, emotional, and attentional regulatory functions as predictors of self-regulation in 125 infants followed 7 times from birth to 5 years. Physiological regulation was assessed by neonatal vagal tone and sleep-wake cyclicity; emotion regulation by response to stress at 3, 6, and 12 months; and attention regulation by focused attention and delayed response in the 2nd year. Executive functions, behavior adaptation, and self-restraint were measured at 5 years. Regulatory functions showed stability across time, measures, and levels. Structural modeling demonstrated both mediated paths from physiological to self-regulation through emotional and attentional processes and direct continuity between vagal tone and each level of regulation. Results support the coherence of the regulation construct and are consistent with neurobiological models on self and consciousness.

Phylum-Level Conservation of Regulatory Information in Nematodes despite Extensive Non-coding Sequence Divergence

PubMed Central

Gordon, Kacy L.; Arthur, Robert K.; Ruvinsky, Ilya

2015-01-01

Gene regulatory information guides development and shapes the course of evolution. To test conservation of gene regulation within the phylum Nematoda, we compared the functions of putative cis-regulatory sequences of four sets of orthologs (unc-47, unc-25, mec-3 and elt-2) from distantly-related nematode species. These species, Caenorhabditis elegans, its congeneric C. briggsae, and three parasitic species Meloidogyne hapla, Brugia malayi, and Trichinella spiralis, represent four of the five major clades in the phylum Nematoda. Despite the great phylogenetic distances sampled and the extensive sequence divergence of nematode genomes, all but one of the regulatory elements we tested are able to drive at least a subset of the expected gene expression patterns. We show that functionally conserved cis-regulatory elements have no more extended sequence similarity to their C. elegans orthologs than would be expected by chance, but they do harbor motifs that are important for proper expression of the C. elegans genes. These motifs are too short to be distinguished from the background level of sequence similarity, and while identical in sequence they are not conserved in orientation or position. Functional tests reveal that some of these motifs contribute to proper expression. Our results suggest that conserved regulatory circuitry can persist despite considerable turnover within cis elements. PMID:26020930

Can microbiota transplantation abrogate murine colonization resistance against Campylobacter jejuni?

PubMed

Heimesaat, M M; Plickert, R; Fischer, A; Göbel, U B; Bereswill, S

2013-03-01

Enterocolitis caused by Campylobacter jejuni represents an important socioeconomic burden worldwide. The host-specific intestinal microbiota is essential for maintaining colonization resistance (CR) against C. jejuni in conventional mice. Notably, CR is abrogated by shifts of the intestinal microbiota towards overgrowth with commensal E. coli during acute ileitis. Thus, we investigated whether oral transplantation (TX) of ileal microbiota derived from C. jejuni susceptible mice with acute ileitis overcomes CR of healthy conventional animals. Four days following ileitis microbiota TX or ileitis induction and right before C. jejuni infection, mice displayed comparable loads of main intestinal bacterial groups as shown by culture. Eight days following ileitis induction, but not ileal microbiota TX, however, C. jejuni could readily colonize the gastrointestinal tract of conventional mice and also translocate to extra-intestinal tissue sites such as mesenteric lymph nodes, spleen, liver, and blood within 4 days following oral infection. Of note, C. jejuni did not further deteriorate histopathology following ileitis induction. Lack of C. jejuni colonization in TX mice was accompanied by a decrease of commensal E. coli loads in the feces 4 days following C. jejuni infection. In summary, oral ileal microbiota TX from susceptible donors is not sufficient to abrogate murine CR against C. jejuni.

The P2X7 receptor antagonist, oxidized adenosine triphosphate, ameliorates renal ischemia-reperfusion injury by expansion of regulatory T cells.

PubMed

Koo, Tai Yeon; Lee, Jae-Ghi; Yan, Ji-Jing; Jang, Joon Young; Ju, Kyung Don; Han, Miyeun; Oh, Kook-Hwan; Ahn, Curie; Yang, Jaeseok

2017-08-01

Extracellular adenosine triphosphate (ATP) binds to purinergic receptors and, as a danger molecule, promotes inflammatory responses. Here we tested whether periodate-oxidized ATP (oATP), a P2X7 receptor (P2X7R) antagonist can attenuate renal ischemia-reperfusion injury and clarify the related cellular mechanisms. Treatment with oATP prior to ischemia-reperfusion injury decreased blood urea nitrogen, serum creatinine, the tubular injury score, and tubular epithelial cell apoptosis after injury. The infiltration of dendritic cells, neutrophils, macrophages, CD69 + CD4 + , and CD44 + CD4 + T cells was attenuated, but renal Foxp3 + CD4 + Treg infiltration was increased by oATP. The levels of IL-6 and CCL2 were reduced in the oATP group. Additionally, oATP treatment following injury improved renal function, decreased the infiltration of innate and adaptive effector cells, and increased the renal infiltration of Foxp3 + CD4 + Tregs. Post-ischemia-reperfusion injury oATP treatment increased tubular cell proliferation and reduced renal fibrosis. oATP treatment attenuated renal functional deterioration after ischemia-reperfusion injury in RAG-1 knockout mice; however, Treg depletion using PC61 abrogated the beneficial effects of oATP in wild-type mice. Furthermore, oATP treatment after transfer of Tregs from wild-type mice improved the beneficial effects of Tregs on ischemia-reperfusion injury, but treatment after transfer of Tregs from P2X7R knockout mice did not. Renal ischemia-reperfusion injury was also attenuated in P2X7R knockout mice. Experiments using bone marrow chimeras established that P2X7R expression on hematopoietic cells rather than non-hematopoietic cells, such as tubular epithelial cells, plays a major role in ischemia-reperfusion injury. Thus, oATP attenuated acute renal damage and facilitated renal recovery in ischemia-reperfusion injury by expansion of Tregs. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Bone marrow-resident NK cells prime monocytes for regulatory function during infection

PubMed Central

Askenase, Michael H.; Han, Seong-Ji; Byrd, Allyson L.; da Fonseca, Denise Morais; Bouladoux, Nicolas; Wilhelm, Christoph; Konkel, Joanne E.; Hand, Timothy W.; Lacerda-Queiroz, Norinne; Su, Xin-Zhuan; Trinchieri, Giorgio; Grainger, John R.; Belkaid, Yasmine

2015-01-01

SUMMARY Tissue-infiltrating Ly6Chi monocytes play diverse roles in immunity, ranging from pathogen killing to immune regulation. How and where this diversity of function is imposed remains poorly understood. Here we show that during acute gastrointestinal infection, priming of monocytes for regulatory function preceded systemic inflammation and was initiated prior to bone marrow egress. Notably, natural killer (NK) cell-derived IFN-γ promoted a regulatory program in monocyte progenitors during development. Early bone marrow NK cell activation was controlled by systemic interleukin-12 (IL-12) produced by Batf3-dependent dendritic cells (DC) in the mucosal-associated lymphoid tissue (MALT). This work challenges the paradigm that monocyte function is dominantly imposed by local signals following tissue recruitment, and instead proposes a sequential model of differentiation in which monocytes are pre-emptively educated during development in the bone marrow to promote their tissue-specific function. PMID:26070484

Biosynthesis and function of chondroitin sulfate.

PubMed

Mikami, Tadahisa; Kitagawa, Hiroshi

2013-10-01

Chondroitin sulfate proteoglycans (CSPGs) are principal pericellular and extracellular components that form regulatory milieu involving numerous biological and pathophysiological phenomena. Diverse functions of CSPGs can be mainly attributed to structural variability of their polysaccharide moieties, chondroitin sulfate glycosaminoglycans (CS-GAG). Comprehensive understanding of the regulatory mechanisms for CS biosynthesis and its catabolic processes is required in order to understand those functions. Here, we focus on recent advances in the study of enzymatic regulatory pathways for CS biosynthesis including successive modification/degradation, distinct CS functions, and disease phenotypes that have been revealed by perturbation of the respective enzymes in vitro and in vivo. Fine-tuned machineries for CS production/degradation are crucial for the functional expression of CS chains in developmental and pathophysiological processes. Control of enzymes responsible for CS biosynthesis/catabolism is a potential target for therapeutic intervention for the CS-associated disorders. Copyright © 2013 Elsevier B.V. All rights reserved.

T Cell Receptor Signaling in the Control of Regulatory T Cell Differentiation and Function

PubMed Central

Li, Ming O.; Rudensky, Alexander Y.

2016-01-01

Regulatory T cells (TReg cells), a specialized T cell lineage, have a pivotal function in the control of self-tolerance and inflammatory responses. Recent studies have revealed a discrete mode of TCR signaling that regulates Treg cell differentiation, maintenance and function and that impacts on gene expression, metabolism, cell adhesion and migration of these cells. Here, we discuss the emerging understanding of TCR-guided differentiation of Treg cells in the context of their function in health and disease. PMID:27026074

Dynactin functions as both a dynamic tether and brake during dynein-driven motility

NASA Astrophysics Data System (ADS)

Ayloo, Swathi; Lazarus, Jacob E.; Dodda, Aditya; Tokito, Mariko; Ostap, E. Michael; Holzbaur, Erika L. F.

2014-09-01

Dynactin is an essential cofactor for most cellular functions of the microtubule motor cytoplasmic dynein, but the mechanism by which dynactin activates dynein remains unclear. Here we use single molecule approaches to investigate dynein regulation by the dynactin subunit p150Glued. We investigate the formation and motility of a dynein-p150Glued co-complex using dual-colour total internal reflection fluorescence microscopy. p150Glued recruits and tethers dynein to the microtubule in a concentration-dependent manner. Single molecule imaging of motility in cell extracts demonstrates that the CAP-Gly domain of p150Glued decreases the detachment rate of the dynein-dynactin complex from the microtubule and also acts as a brake to slow the dynein motor. Consistent with this important role, two neurodegenerative disease-causing mutations in the CAP-Gly domain abrogate these functions in our assays. Together, these observations support a model in which dynactin enhances the initial recruitment of dynein onto microtubules and promotes the sustained engagement of dynein with its cytoskeletal track.

Functional expression of IL-12 receptor by human eosinophils: IL-12 promotes eosinophil apoptosis.

PubMed

Nutku, E; Zhuang, Q; Soussi-Gounni, A; Aris, F; Mazer, B D; Hamid, Q

2001-07-15

In murine models of allergic inflammation, IL-12 has been shown to decrease tissue eosinophilia, but the underlying mechanisms are not known. We evaluated the expression of IL-12R and the effect of IL-12 on eosinophil survival. In situ hybridization demonstrated the presence of mRNA and immunoreactivity for IL-12Rbeta1 and -beta2 subunits in human peripheral blood eosinophils. Surface expression of IL-12Rbeta1 and -beta2 subunits on freshly isolated human eosinophils was optimally expressed after incubation with PMA. To determine the functional significance of IL-12R studies, we studied cell viability and apoptosis. Morphological analysis and propidium iodide staining for cell cycle demonstrated that recombinant human IL-12 increased in vitro human eosinophil apoptosis in a dose-dependent manner. Addition of IL-5 together with IL-12 abrogated eosinophil apoptosis, suggesting that IL-12 and IL-5 have antagonistic effects. Our findings provide evidence for a novel role for IL-12 in regulating eosinophil function by increasing eosinophil apoptosis.

LMP1-Induced Sumoylation Influences the Maintenance of Epstein-Barr Virus Latency through KAP1

PubMed Central

Moss, Charles Randall; Whitehurst, Christopher B.; Moody, Cary A.

2015-01-01

ABSTRACT As a herpesvirus, Epstein-Barr virus (EBV) establishes a latent infection that can periodically undergo reactivation, resulting in lytic replication and the production of new infectious virus. Latent membrane protein-1 (LMP1), the principal viral oncoprotein, is a latency-associated protein implicated in regulating viral reactivation and the maintenance of latency. We recently found that LMP1 hijacks the SUMO-conjugating enzyme Ubc9 via its C-terminal activating region-3 (CTAR3) and induces the sumoylation of cellular proteins. Because protein sumoylation can promote transcriptional repression, we hypothesized that LMP1-induced protein sumoylation induces the repression of EBV lytic promoters and helps maintain the viral genome in its latent state. We now show that with inhibition of LMP1-induced protein sumoylation, the latent state becomes less stable or leakier in EBV-transformed lymphoblastoid cell lines. The cells are also more sensitive to viral reactivation induced by irradiation, which results in the increased production and release of infectious virus, as well as increased susceptibility to ganciclovir treatment. We have identified a target of LMP1-mediated sumoylation that contributes to the maintenance of latency in this context: KRAB-associated protein-1 (KAP1). LMP1 CTAR3-mediated sumoylation regulates the function of KAP1. KAP1 also binds to EBV OriLyt and immediate early promoters in a CTAR3-dependent manner, and inhibition of sumoylation processes abrogates the binding of KAP1 to these promoters. These data provide an additional line of evidence that supports our findings that CTAR3 is a distinct functioning regulatory region of LMP1 and confirm that LMP1-induced sumoylation may help stabilize the maintenance of EBV latency. IMPORTANCE Epstein-Barr virus (EBV) latent membrane protein-1 (LMP1) plays an important role in the maintenance of viral latency. Previously, we documented that LMP1 targets cellular proteins to be modified by a ubiquitin-like protein (SUMO). We have now identified a function for this LMP1-induced modification of cellular proteins in the maintenance of EBV latency. Because latently infected cells have to undergo viral reactivation in order to be vulnerable to antiviral drugs, these findings identify a new way to increase the rate of EBV reactivation, which increases cell susceptibility to antiviral therapies. PMID:25948750

Radiation and the regulatory landscape of neo2-Darwinism.

PubMed

Rollo, C David

2006-05-11

Several recently revealed features of eukaryotic genomes were not predicted by earlier evolutionary paradigms, including the relatively small number of genes, the very large amounts of non-functional code and its quarantine in heterochromatin, the remarkable conservation of many functionally important genes across relatively enormous phylogenetic distances, and the prevalence of extra-genomic information associated with chromatin structure and histone proteins. All of these emphasize a paramount role for regulatory evolution, which is further reinforced by recent perspectives highlighting even higher-order regulation governing epigenetics and development (EVO-DEVO). Modern neo2-Darwinism, with its emphasis on regulatory mechanisms and regulatory evolution provides new vision for understanding radiation biology, particularly because free radicals and redox states are central to many regulatory mechanisms and free radicals generated by radiation mimic and amplify endogenous signalling. This paper explores some of these aspects and their implications for low-dose radiation biology.

Disentangling the many layers of eukaryotic transcriptional regulation.

PubMed

Lelli, Katherine M; Slattery, Matthew; Mann, Richard S

2012-01-01

Regulation of gene expression in eukaryotes is an extremely complex process. In this review, we break down several critical steps, emphasizing new data and techniques that have expanded current gene regulatory models. We begin at the level of DNA sequence where cis-regulatory modules (CRMs) provide important regulatory information in the form of transcription factor (TF) binding sites. In this respect, CRMs function as instructional platforms for the assembly of gene regulatory complexes. We discuss multiple mechanisms controlling complex assembly, including cooperative DNA binding, combinatorial codes, and CRM architecture. The second section of this review places CRM assembly in the context of nucleosomes and condensed chromatin. We discuss how DNA accessibility and histone modifications contribute to TF function. Lastly, new advances in chromosomal mapping techniques have provided increased understanding of intra- and interchromosomal interactions. We discuss how these topological maps influence gene regulatory models.

Abundant raw material for cis-regulatory evolution in humans

NASA Technical Reports Server (NTRS)

Rockman, Matthew V.; Wray, Gregory A.

2002-01-01

Changes in gene expression and regulation--due in particular to the evolution of cis-regulatory DNA sequences--may underlie many evolutionary changes in phenotypes, yet little is known about the distribution of such variation in populations. We present in this study the first survey of experimentally validated functional cis-regulatory polymorphism. These data are derived from more than 140 polymorphisms involved in the regulation of 107 genes in Homo sapiens, the eukaryote species with the most available data. We find that functional cis-regulatory variation is widespread in the human genome and that the consequent variation in gene expression is twofold or greater for 63% of the genes surveyed. Transcription factor-DNA interactions are highly polymorphic, and regulatory interactions have been gained and lost within human populations. On average, humans are heterozygous at more functional cis-regulatory sites (>16,000) than at amino acid positions (<13,000), in part because of an overrepresentation among the former in multiallelic tandem repeat variation, especially (AC)(n) dinucleotide microsatellites. The role of microsatellites in gene expression variation may provide a larger store of heritable phenotypic variation, and a more rapid mutational input of such variation, than has been realized. Finally, we outline the distinctive consequences of cis-regulatory variation for the genotype-phenotype relationship, including ubiquitous epistasis and genotype-by-environment interactions, as well as underappreciated modes of pleiotropy and overdominance. Ordinary small-scale mutations contribute to pervasive variation in transcription rates and consequently to patterns of human phenotypic variation.

Abasy Atlas: a comprehensive inventory of systems, global network properties and systems-level elements across bacteria.

PubMed

Ibarra-Arellano, Miguel A; Campos-González, Adrián I; Treviño-Quintanilla, Luis G; Tauch, Andreas; Freyre-González, Julio A

2016-01-01

The availability of databases electronically encoding curated regulatory networks and of high-throughput technologies and methods to discover regulatory interactions provides an invaluable source of data to understand the principles underpinning the organization and evolution of these networks responsible for cellular regulation. Nevertheless, data on these sources never goes beyond the regulon level despite the fact that regulatory networks are complex hierarchical-modular structures still challenging our understanding. This brings the necessity for an inventory of systems across a large range of organisms, a key step to rendering feasible comparative systems biology approaches. In this work, we take the first step towards a global understanding of the regulatory networks organization by making a cartography of the functional architectures of diverse bacteria. Abasy ( A: cross- BA: cteria SY: stems) Atlas provides a comprehensive inventory of annotated functional systems, global network properties and systems-level elements (global regulators, modular genes shaping functional systems, basal machinery genes and intermodular genes) predicted by the natural decomposition approach for reconstructed and meta-curated regulatory networks across a large range of bacteria, including pathogenically and biotechnologically relevant organisms. The meta-curation of regulatory datasets provides the most complete and reliable set of regulatory interactions currently available, which can even be projected into subsets by considering the force or weight of evidence supporting them or the systems that they belong to. Besides, Abasy Atlas provides data enabling large-scale comparative systems biology studies aimed at understanding the common principles and particular lifestyle adaptions of systems across bacteria. Abasy Atlas contains systems and system-level elements for 50 regulatory networks comprising 78 649 regulatory interactions covering 42 bacteria in nine taxa, containing 3708 regulons and 1776 systems. All this brings together a large corpus of data that will surely inspire studies to generate hypothesis regarding the principles governing the evolution and organization of systems and the functional architectures controlling them.Database URL: http://abasy.ccg.unam.mx. © The Author(s) 2016. Published by Oxford University Press.

Form and function in gene regulatory networks: the structure of network motifs determines fundamental properties of their dynamical state space.

PubMed

Ahnert, S E; Fink, T M A

2016-07-01

Network motifs have been studied extensively over the past decade, and certain motifs, such as the feed-forward loop, play an important role in regulatory networks. Recent studies have used Boolean network motifs to explore the link between form and function in gene regulatory networks and have found that the structure of a motif does not strongly determine its function, if this is defined in terms of the gene expression patterns the motif can produce. Here, we offer a different, higher-level definition of the 'function' of a motif, in terms of two fundamental properties of its dynamical state space as a Boolean network. One is the basin entropy, which is a complexity measure of the dynamics of Boolean networks. The other is the diversity of cyclic attractor lengths that a given motif can produce. Using these two measures, we examine all 104 topologically distinct three-node motifs and show that the structural properties of a motif, such as the presence of feedback loops and feed-forward loops, predict fundamental characteristics of its dynamical state space, which in turn determine aspects of its functional versatility. We also show that these higher-level properties have a direct bearing on real regulatory networks, as both basin entropy and cycle length diversity show a close correspondence with the prevalence, in neural and genetic regulatory networks, of the 13 connected motifs without self-interactions that have been studied extensively in the literature. © 2016 The Authors.

[Regulatory T cells].

PubMed

Marinić, Igor; Gagro, Alenka; Rabatić, Sabina

2006-12-01

Regulatory T-cells are a subset of T cells that have beene extensively studied in modern immunology. They are important for the maintenance of peripheral tolerance, and have an important role in various clinical conditions such as allergy, autoimmune disorders, tumors, infections, and in transplant medicine. Basically, this population has a suppressive effect on the neighboring immune cells, thus contributing to the local modulation and control of immune response. There are two main populations of regulatory T cells - natural regulatory T cells, which form a distinct cellular lineage, develop in thymus and perform their modulatory action through direct intercellular contact, along with the secreted cytokines; and inducible regulatory T cells, which develop in the periphery after contact with the antigen that is presented on the antigen presenting cell, and their primary mode of action is through the interleukin 10 (IL-10) and transforming growth factor beta (TGF-alpha) cytokines. Natural regulatory T cells are activated through T cell receptor after contact with specific antigen and inhibit proliferation of other T cells in an antigen independent manner. One of the major difficulties in the research of regulatory T cells is the lack of specific molecular markers that would identify these cells. Natural regulatory T cells constitutively express surface molecule CD25, but many other surface and intracellular molecules (HLA-DR, CD122, CD45RO, CD62, CTLA-4, GITR, PD-1, Notch, FOXP3, etc.) are being investigated for further phenotypic characterization of these cells. Because regulatory T cells have an important role in establishing peripheral tolerance, their importance is manifested in a number of clinical conditions. In the IPEX syndrome (immunodysregulation, polyendocrinopathy and enteropathy, X-linked), which is caused by mutation in Foxp3 gene that influences the development and function of regulatory T cells, patients develop severe autoimmune reactions that involve autoimmune endocrine disorders (type 1 diabetes, thyroiditis), respiratory and nutritive allergy, eczema and severe infections. In different types of allergy (pollen allergy, dust mite, nutritive allergens, contact hypersensitivity, etc.) and autoimmune diseases (such as rheumatoid arthritis, multiple sclerosis and type 1 diabetes) a lower number or decreased functional capability of regulatory T cells have been described. In inflammatory conditions and infections, this cell population has an important task in restricting immune response and protecting the host from excessive damage. This ability of regulatory T cells can be used by some pathogens (Epstein Barr virus, Mycobacterium tuberculosis, Leishmania major, etc.) and tumor cells to avoid host response and therefore contribute to the development of some pathological conditions. The knowledge gained on the phenotype and function of regulatory T cells could be useful in many medical conditions. In allergy, autoimmune diseases and in transplant procedures in medicine it would be desirable to increase their function, thus to partially suppress the immune system activity. On the other hand, in some infections and tumors, it would be preferable to decrease the activity of regulatory T cells and boost the function of effector T cells. Regulatory T cells comprise a very active field of immunology, therefore monitoring and modulating of their activity is of great potential significance in a broad spectrum of clinical conditions. By developing and standardizing methods for their monitoring, it would be possible to follow additional parameters of certain clinical conditions and possibly utilize them in therapy.

A hairpin within YAP mRNA 3′UTR functions in regulation at post-transcription level

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Yuen; Wang, Yuan; Feng, Jinyan

2015-04-03

The central dogma of gene expression is that DNA is transcribed into messenger RNAs, which in turn serve as the template for protein synthesis. Recently, it has been reported that mRNAs display regulatory roles that rely on their ability to compete for microRNA binding, independent of their protein-coding function. However, the regulatory mechanism of mRNAs remains poorly understood. Here, we report that a hairpin within YAP mRNA 3′untranslated region (3′UTR) functions in regulation at post-transcription level through generating endogenous siRNAs (esiRNAs). Bioinformatics analysis for secondary structure showed that YAP mRNA displayed a hairpin structure (termed standard hairpin, S-hairpin) within itsmore » 3′UTR. Surprisingly, we observed that the overexpression of S-hairpin derived from YAP 3′UTR (YAP-sh) increased the luciferase reporter activities of transcriptional factor NF-κB and AP-1 in 293T cells. Moreover, we identified that a fragment from YAP-sh, an esiRNA, was able to target mRNA 3′UTR of NF2 (a member of Hippo-signaling pathway) and YAP mRNA 3′UTR itself in hepatoma cells. Thus, we conclude that the YAP-sh within YAP mRNA 3′UTR may serve as a novel regulatory element, which functions in regulation at post-transcription level. Our finding provides new insights into the mechanism of mRNAs in regulatory function. - Highlights: • An S-hairpin within YAP mRNA 3′UTR possesses regulatory function. • YAP-sh acts as a regulatory element for YAP at post-transcription level. • YAP-sh-3p20, an esiRNA derived from YAP-sh, targets mRNAs of YAP and NF2. • YAP-sh-3p20 depresses the proliferation of HepG2 cells in vitro.« less

Hypomethylation associated enhanced transcription of trefoil factor-3 mediates tamoxifen-stimulated oncogenicity of ER+ endometrial carcinoma cells.

PubMed

Pandey, Vijay; Zhang, Min; Chong, Qing-Yun; You, Mingliang; Raquib, Ainiah Rushdiana; Pandey, Amit K; Liu, Dong-Xu; Liu, Liang; Ma, Lan; Jha, Sudhakar; Wu, Zheng-Sheng; Zhu, Tao; Lobie, Peter E

2017-09-29

Tamoxifen (TAM) is widely used as an adjuvant therapy for women with breast cancer (BC). However, TAM possesses partial oestrogenic activity in the uterus and its use has been associated with an increased incidence of endometrial carcinoma (EC). The molecular mechanism for these observations is not well understood. Herein, we demonstrated that forced expression of Trefoil factor 3 ( TFF3) , in oestrogen receptor-positive (ER+) EC cells significantly increased cell cycle progression, cell survival, anchorage-independent growth, invasiveness and tumour growth in xenograft models. Clinically, elevated TFF3 protein expression was observed in EC compared with normal endometrial tissue, and its increased expression in EC was significantly associated with myometrial invasion. TAM exposure increased expression of TFF3 in ER+ EC cells and its elevated expression resulted in increased oncogenicity and invasiveness. TAM-stimulated expression of TFF3 in EC cells was associated with hypomethylation of the TFF3 promoter sequence and c-JUN/SP1-dependent transcriptional activation. In addition, small interfering ( si) RNA -mediated depletion or polyclonal antibody inhibition of TFF3 significantly abrogated oncogenicity and invasiveness in EC cells consequent to TAM induction or forced expression of TFF3. Hence, TAM-stimulated upregulation of TFF3 in EC cells was critical in promoting EC progression associated with TAM treatment. Importantly, inhibition of TFF3 function might be an attractive molecular modality to abrogate the stimulatory effects of TAM on endometrial tissue and to limit the progression of EC.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Changhong; Li, Ming; Chen, Pan

Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of L-glutamate and, in animals, is extensively regulated by a number of metabolites. Gain of function mutations in GDH that abrogate GTP inhibition cause the hyperinsulinism/hyperammonemia syndrome (HHS), resulting in increased pancreatic {beta}-cell responsiveness to leucine and susceptibility to hypoglycemia following high protein meals. We have previously shown that two of the polyphenols from green tea (epigallocatechin gallate (EGCG) and epicatechin gallate (ECG)) inhibit GDH in vitro and that EGCG blocks GDH-mediated insulin secretion in wild type rat islets. Using structural and site-directed mutagenesis studies, we demonstrate that ECG binds to the samemore » site as the allosteric regulator, ADP. Perifusion assays using pancreatic islets from transgenic mice expressing a human HHS form of GDH demonstrate that the hyperresponse to glutamine caused by dysregulated GDH is blocked by the addition of EGCG. As observed in HHS patients, these transgenic mice are hypersensitive to amino acid feeding, and this is abrogated by oral administration of EGCG prior to challenge. Finally, the low basal blood glucose level in the HHS mouse model is improved upon chronic administration of EGCG. These results suggest that this common natural product or some derivative thereof may prove useful in controlling this genetic disorder. Of broader clinical implication is that other groups have shown that restriction of glutamine catabolism via these GDH inhibitors can be useful in treating various tumors. This HHS transgenic mouse model offers a highly useful means to test these agents in vivo.« less

A compact, in vivo screen of all 6-mers reveals drivers of tissue-specific expression and guides synthetic regulatory element design.

PubMed

Smith, Robin P; Riesenfeld, Samantha J; Holloway, Alisha K; Li, Qiang; Murphy, Karl K; Feliciano, Natalie M; Orecchia, Lorenzo; Oksenberg, Nir; Pollard, Katherine S; Ahituv, Nadav

2013-07-18

Large-scale annotation efforts have improved our ability to coarsely predict regulatory elements throughout vertebrate genomes. However, it is unclear how complex spatiotemporal patterns of gene expression driven by these elements emerge from the activity of short, transcription factor binding sequences. We describe a comprehensive promoter extension assay in which the regulatory potential of all 6 base-pair (bp) sequences was tested in the context of a minimal promoter. To enable this large-scale screen, we developed algorithms that use a reverse-complement aware decomposition of the de Bruijn graph to design a library of DNA oligomers incorporating every 6-bp sequence exactly once. Our library multiplexes all 4,096 unique 6-mers into 184 double-stranded 15-bp oligomers, which is sufficiently compact for in vivo testing. We injected each multiplexed construct into zebrafish embryos and scored GFP expression in 15 tissues at two developmental time points. Twenty-seven constructs produced consistent expression patterns, with the majority doing so in only one tissue. Functional sequences are enriched near biologically relevant genes, match motifs for developmental transcription factors, and are required for enhancer activity. By concatenating tissue-specific functional sequences, we generated completely synthetic enhancers for the notochord, epidermis, spinal cord, forebrain and otic lateral line, and show that short regulatory sequences do not always function modularly. This work introduces a unique in vivo catalog of short, functional regulatory sequences and demonstrates several important principles of regulatory element organization. Furthermore, we provide resources for designing compact, reverse-complement aware k-mer libraries.

Synthesis, structural characterization and effect on human granulocyte intracellular cAMP levels of abscisic acid analogs.

PubMed

Bellotti, Marta; Salis, Annalisa; Grozio, Alessia; Damonte, Gianluca; Vigliarolo, Tiziana; Galatini, Andrea; Zocchi, Elena; Benatti, Umberto; Millo, Enrico

2015-01-01

The phytohormone abscisic acid (ABA), in addition to regulating physiological functions in plants, is also produced and released by several mammalian cell types, including human granulocytes, where it stimulates innate immune functions via an increase of the intracellular cAMP concentration ([cAMP]i). We synthesized several ABA analogs and evaluated the structure-activity relationship, by the systematical modification of selected regions of these analogs. The resulting molecules were tested for their ability to inhibit the ABA-induced increase of [cAMP]i in human granulocytes. The analogs with modified configurations at C-2' and C-3' abrogated the ABA-induced increase of the [cAMP]i and also inhibited several pro-inflammatory effects induced by exogenous ABA on granulocytes and monocytes. Accordingly, these analogs could be suitable as novel putative anti-inflammatory compounds. Copyright © 2014 Elsevier Ltd. All rights reserved.

Effects of Sex, Strain, and Energy Intake on Hallmarks of Aging in Mice.

PubMed

Mitchell, Sarah J; Madrigal-Matute, Julio; Scheibye-Knudsen, Morten; Fang, Evandro; Aon, Miguel; González-Reyes, José A; Cortassa, Sonia; Kaushik, Susmita; Gonzalez-Freire, Marta; Patel, Bindi; Wahl, Devin; Ali, Ahmed; Calvo-Rubio, Miguel; Burón, María I; Guiterrez, Vincent; Ward, Theresa M; Palacios, Hector H; Cai, Huan; Frederick, David W; Hine, Christopher; Broeskamp, Filomena; Habering, Lukas; Dawson, John; Beasley, T Mark; Wan, Junxiang; Ikeno, Yuji; Hubbard, Gene; Becker, Kevin G; Zhang, Yongqing; Bohr, Vilhelm A; Longo, Dan L; Navas, Placido; Ferrucci, Luigi; Sinclair, David A; Cohen, Pinchas; Egan, Josephine M; Mitchell, James R; Baur, Joseph A; Allison, David B; Anson, R Michael; Villalba, José M; Madeo, Frank; Cuervo, Ana Maria; Pearson, Kevin J; Ingram, Donald K; Bernier, Michel; de Cabo, Rafael

2016-06-14

Calorie restriction (CR) is the most robust non-genetic intervention to delay aging. However, there are a number of emerging experimental variables that alter CR responses. We investigated the role of sex, strain, and level of CR on health and survival in mice. CR did not always correlate with lifespan extension, although it consistently improved health across strains and sexes. Transcriptional and metabolomics changes driven by CR in liver indicated anaplerotic filling of the Krebs cycle together with fatty acid fueling of mitochondria. CR prevented age-associated decline in the liver proteostasis network while increasing mitochondrial number, preserving mitochondrial ultrastructure and function with age. Abrogation of mitochondrial function negated life-prolonging effects of CR in yeast and worms. Our data illustrate the complexity of CR in the context of aging, with a clear separation of outcomes related to health and survival, highlighting complexities of translation of CR into human interventions. Published by Elsevier Inc.

Structure of the Pds5-Scc1 Complex and Implications for Cohesin Function.

PubMed

Muir, Kyle W; Kschonsak, Marc; Li, Yan; Metz, Jutta; Haering, Christian H; Panne, Daniel

2016-03-08

Sister chromatid cohesion is a fundamental prerequisite to faithful genome segregation. Cohesion is precisely regulated by accessory factors that modulate the stability with which the cohesin complex embraces chromosomes. One of these factors, Pds5, engages cohesin through Scc1 and is both a facilitator of cohesion, and, conversely also mediates the release of cohesin from chromatin. We present here the crystal structure of a complex between budding yeast Pds5 and Scc1, thus elucidating the molecular basis of Pds5 function. Pds5 forms an elongated HEAT repeat that binds to Scc1 via a conserved surface patch. We demonstrate that the integrity of the Pds5-Scc1 interface is indispensable for the recruitment of Pds5 to cohesin, and that its abrogation results in loss of sister chromatid cohesion and cell viability. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Immune privilege as an intrinsic CNS property: astrocytes protect the CNS against T-cell-mediated neuroinflammation.

PubMed

Gimsa, Ulrike; Mitchison, N Avrion; Brunner-Weinzierl, Monika C

2013-01-01

Astrocytes have many functions in the central nervous system (CNS). They support differentiation and homeostasis of neurons and influence synaptic activity. They are responsible for formation of the blood-brain barrier (BBB) and make up the glia limitans. Here, we review their contribution to neuroimmune interactions and in particular to those induced by the invasion of activated T cells. We discuss the mechanisms by which astrocytes regulate pro- and anti-inflammatory aspects of T-cell responses within the CNS. Depending on the microenvironment, they may become potent antigen-presenting cells for T cells and they may contribute to inflammatory processes. They are also able to abrogate or reprogram T-cell responses by inducing apoptosis or secreting inhibitory mediators. We consider apparently contradictory functions of astrocytes in health and disease, particularly in their interaction with lymphocytes, which may either aggravate or suppress neuroinflammation.

Small Molecule Bcl2 BH4 Antagonist for Lung Cancer Therapy

PubMed Central

Han, Bingshe; Park, Dongkyoo; Li, Rui; Xie, Maohua; Owonikoko, Taofeek K.; Zhang, Guojing; Sica, Gabriel L.; Ding, Chunyong; Zhou, Jia; Magis, Andrew T.; Chen, Zhuo G.; Shin, Dong M.; Ramalingam, Suresh S.; Khuri, Fadlo R.; Curran, Walter J.; Deng, Xingming

2015-01-01

SUMMARY The BH4 domain of Bcl2 is required for its antiapoptotic function, thus constituting a promising anticancer target. We identified a small molecule Bcl2-BH4 domain-antagonist (BDA-366) that binds BH4 with high affinity and selectivity. BDA-366-Bcl2 binding induces conformational change in Bcl2 that abrogates its antiapoptotic function, converting it from a survival to a cell death inducer. BDA-366 suppresses growth of lung cancer xenografts derived from cell lines and patient without significant normal tissue toxicity at effective doses. mTOR inhibition up-regulates Bcl2 in lung cancer cells and tumor tissues from clinical trial patients. Combined BDA-366 and RAD001 treatment exhibits strong synergy against lung cancer in vivo. Development of this Bcl2-BH4 antagonist may provide a strategy to improve lung cancer outcome. PMID:26004684

Dysfunctional tubular endoplasmic reticulum constitutes a pathological feature of Alzheimer's disease.

PubMed

Sharoar, M G; Shi, Q; Ge, Y; He, W; Hu, X; Perry, G; Zhu, X; Yan, R

2016-09-01

Pathological features in Alzheimer's brains include mitochondrial dysfunction and dystrophic neurites (DNs) in areas surrounding amyloid plaques. Using a mouse model that overexpresses reticulon 3 (RTN3) and spontaneously develops age-dependent hippocampal DNs, here we report that DNs contain both RTN3 and REEPs, topologically similar proteins that can shape tubular endoplasmic reticulum (ER). Importantly, ultrastructural examinations of such DNs revealed gradual accumulation of tubular ER in axonal termini, and such abnormal tubular ER inclusion is found in areas surrounding amyloid plaques in biopsy samples from Alzheimer's disease (AD) brains. Functionally, abnormally clustered tubular ER induces enhanced mitochondrial fission in the early stages of DN formation and eventual mitochondrial degeneration at later stages. Furthermore, such DNs are abrogated when RTN3 is ablated in aging and AD mouse models. Hence, abnormally clustered tubular ER can be pathogenic in brain regions: disrupting mitochondrial integrity, inducing DNs formation and impairing cognitive function in AD and aging brains.

The sigma-1 receptor: a regulator of cancer cell electrical plasticity?

PubMed Central

Crottès, David; Guizouarn, Hélène; Martin, Patrick; Borgese, Franck; Soriani, Olivier

2013-01-01

Originally mistaken as an opioid receptor, the sigma-1 receptor (Sig1R) is a ubiquitous membrane protein that has been involved in many cellular processes. While the precise function of Sig1R has long remained mysterious, recent studies have shed light on its role and the molecular mechanisms triggered. Sig1R is in fact a stress-activated chaperone mainly associated with the ER-mitochondria interface that can regulate cell survival through the control of calcium homeostasis. Sig1R functionally regulates ion channels belonging to various molecular families and it has thus been involved in neuronal plasticity and central nervous system diseases. Interestingly, Sig1R is frequently expressed in tumors but its function in cancer has not been yet clarified. In this review, we discuss the current understanding of Sig1R. We suggest herein that Sig1R shapes cancer cell electrical signature upon environmental conditions. Thus, Sig1R may be used as a novel therapeutic target to specifically abrogate pro-invasive functions of ion channels in cancer tissue. PMID:23882221

The structure of human SFPQ reveals a coiled-coil mediated polymer essential for functional aggregation in gene regulation

PubMed Central

Lee, Mihwa; Sadowska, Agata; Bekere, Indra; Ho, Diwei; Gully, Benjamin S.; Lu, Yanling; Iyer, K. Swaminathan; Trewhella, Jill; Fox, Archa H.; Bond, Charles S.

2015-01-01

SFPQ, (a.k.a. PSF), is a human tumor suppressor protein that regulates many important functions in the cell nucleus including coordination of long non-coding RNA molecules into nuclear bodies. Here we describe the first crystal structures of Splicing Factor Proline and Glutamine Rich (SFPQ), revealing structural similarity to the related PSPC1/NONO heterodimer and a strikingly extended structure (over 265 Å long) formed by an unusual anti-parallel coiled-coil that results in an infinite linear polymer of SFPQ dimers within the crystals. Small-angle X-ray scattering and transmission electron microscopy experiments show that polymerization is reversible in solution and can be templated by DNA. We demonstrate that the ability to polymerize is essential for the cellular functions of SFPQ: disruptive mutation of the coiled-coil interaction motif results in SFPQ mislocalization, reduced formation of nuclear bodies, abrogated molecular interactions and deficient transcriptional regulation. The coiled-coil interaction motif thus provides a molecular explanation for the functional aggregation of SFPQ that directs its role in regulating many aspects of cellular nucleic acid metabolism. PMID:25765647

PlantTFDB 4.0: toward a central hub for transcription factors and regulatory interactions in plants.

PubMed

Jin, Jinpu; Tian, Feng; Yang, De-Chang; Meng, Yu-Qi; Kong, Lei; Luo, Jingchu; Gao, Ge

2017-01-04

With the goal of providing a comprehensive, high-quality resource for both plant transcription factors (TFs) and their regulatory interactions with target genes, we upgraded plant TF database PlantTFDB to version 4.0 (http://planttfdb.cbi.pku.edu.cn/). In the new version, we identified 320 370 TFs from 165 species, presenting a more comprehensive genomic TF repertoires of green plants. Besides updating the pre-existing abundant functional and evolutionary annotation for identified TFs, we generated three new types of annotation which provide more directly clues to investigate functional mechanisms underlying: (i) a set of high-quality, non-redundant TF binding motifs derived from experiments; (ii) multiple types of regulatory elements identified from high-throughput sequencing data; (iii) regulatory interactions curated from literature and inferred by combining TF binding motifs and regulatory elements. In addition, we upgraded previous TF prediction server, and set up four novel tools for regulation prediction and functional enrichment analyses. Finally, we set up a novel companion portal PlantRegMap (http://plantregmap.cbi.pku.edu.cn) for users to access the regulation resource and analysis tools conveniently. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Role of WDHD1 in Human Papillomavirus-Mediated Oncogenesis Identified by Transcriptional Profiling of E7-Expressing Cells

PubMed Central

Zhou, Yunying; Zhang, Qishu; Gao, Ge; Zhang, Xiaoli; Liu, Yafei; Yuan, Shoudao

2016-01-01

ABSTRACT The E7 oncoprotein of the high-risk human papillomavirus (HPV) plays a major role in HPV-induced carcinogenesis. E7 abrogates the G1 cell cycle checkpoint and induces genomic instability, but the mechanism is not fully understood. In this study, we performed RNA sequencing (RNA-seq) to characterize the transcriptional profile of keratinocytes expressing HPV 16 (HPV-16) E7. At the transcriptome level, 236 genes were differentially expressed between E7 and vector control cells. A subset of the differentially expressed genes, most of them novel to E7-expressing cells, was further confirmed by real-time PCR. Of interest, the activities of multiple transcription factors were altered in E7-expressing cells. Through bioinformatics analysis, pathways altered in E7-expressing cells were investigated. The upregulated genes were enriched in cell cycle and DNA replication, as well as in the DNA metabolic process, transcription, DNA damage, DNA repair, and nucleotide metabolism. Specifically, we focused our studies on the gene encoding WDHD1 (WD repeat and high mobility group [HMG]-box DNA-binding protein), one of the genes that was upregulated in E7-expressing cells. WDHD1 is a component of the replisome that regulates DNA replication. Recent studies suggest that WDHD1 may also function as a DNA replication initiation factor as well as a G1 checkpoint regulator. We found that in E7-expressing cells, the steady-state level of WDHD1 protein was increased along with the half-life. Moreover, downregulation of WDHD1 reduced E7-induced G1 checkpoint abrogation and rereplication, demonstrating a novel function for WDHD1. These studies shed light on mechanisms by which HPV induces genomic instability and have therapeutic implications. IMPORTANCE The high-risk HPV types induce cervical cancer and encode an E7 oncoprotein that plays a major role in HPV-induced carcinogenesis. However, the mechanism by which E7 induces carcinogenesis is not fully understood; specific anti-HPV agents are not available. In this study, we performed RNA-seq to characterize transcriptional profiling of keratinocytes expressing HPV-16 E7 and identified more than 200 genes that were differentially expressed between E7 and vector control cells. Through bioinformatics analysis, pathways altered in E7-expressing cells were identified. Significantly, the WDHD1 gene, one of the genes that is upregulated in E7-expressing cells, was found to play an important role in E7-induced G1 checkpoint abrogation and rereplication. These studies shed light on mechanisms by which HPV induces genomic instability and have therapeutic implications. PMID:27099318

Regulatory RNAs derived from transfer RNA?

PubMed

Pederson, Thoru

2010-10-01

Four recent studies suggest that cleavages of transfer RNAs generate products with microRNA-like features, with some evidence of function. If their regulatory functions were to be confirmed, these newly revealed RNAs would add to the expanding repertoire of small noncoding RNAs and would also provide new perspectives on the coevolution of transfer RNA and messenger RNA.

The Development of Regulatory Functions from Birth to 5 Years: Insights from Premature Infants

ERIC Educational Resources Information Center

Feldman, Ruth

2009-01-01

This study examined physiological, emotional, and attentional regulatory functions as predictors of self-regulation in 125 infants followed 7 times from birth to 5 years. Physiological regulation was assessed by neonatal vagal tone and sleep-wake cyclicity; emotion regulation by response to stress at 3, 6, and 12 months; and attention regulation…

75 FR 43584 - Self-Regulatory Organizations; EDGX Exchange, Inc; Order Approving a Proposed Rule Change...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-07-26

... SECURITIES AND EXCHANGE COMMISSION [Release No. 34-62515; File No. SR-EDGX-2010-02] Self-Regulatory Organizations; EDGX Exchange, Inc; Order Approving a Proposed Rule Change Relating to Direct Edge... DE Holdings, and DE Holdings will be the sole stockholder of DEI. The self-regulatory functions of...

75 FR 43582 - Self-Regulatory Organizations; EDGA Exchange, Inc.; Order Approving a Proposed Rule Change...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-07-26

... SECURITIES AND EXCHANGE COMMISSION [Release No. 34-62514; File No. SR-EDGA-2010-02] Self-Regulatory Organizations; EDGA Exchange, Inc.; Order Approving a Proposed Rule Change Relating to Direct Edge... Holdings will be the sole stockholder of DEI. The self-regulatory functions of the Exchange will remain...

Regulatory Mechanisms Controlling Maturation of Serotonin Neuron Identity and Function

PubMed Central

Spencer, William C.; Deneris, Evan S.

2017-01-01

The brain serotonin (5-hydroxytryptamine; 5-HT) system has been extensively studied for its role in normal physiology and behavior, as well as, neuropsychiatric disorders. The broad influence of 5-HT on brain function, is in part due to the vast connectivity pattern of 5-HT-producing neurons throughout the CNS. 5-HT neurons are born and terminally specified midway through embryogenesis, then enter a protracted period of maturation, where they functionally integrate into CNS circuitry and then are maintained throughout life. The transcriptional regulatory networks controlling progenitor cell generation and terminal specification of 5-HT neurons are relatively well-understood, yet the factors controlling 5-HT neuron maturation are only recently coming to light. In this review, we first provide an update on the regulatory network controlling 5-HT neuron development, then delve deeper into the properties and regulatory strategies governing 5-HT neuron maturation. In particular, we discuss the role of the 5-HT neuron terminal selector transcription factor (TF) Pet-1 as a key regulator of 5-HT neuron maturation. Pet-1 was originally shown to positively regulate genes needed for 5-HT synthesis, reuptake and vesicular transport, hence 5-HT neuron-type transmitter identity. It has now been shown to regulate, both positively and negatively, many other categories of genes in 5-HT neurons including ion channels, GPCRs, transporters, neuropeptides, and other transcription factors. Its function as a terminal selector results in the maturation of 5-HT neuron excitability, firing characteristics, and synaptic modulation by several neurotransmitters. Furthermore, there is a temporal requirement for Pet-1 in the control of postmitotic gene expression trajectories thus indicating a direct role in 5-HT neuron maturation. Proper regulation of the maturation of cellular identity is critical for normal neuronal functioning and perturbations in the gene regulatory networks controlling these processes may result in long-lasting changes in brain function in adulthood. Further study of 5-HT neuron gene regulatory networks is likely to provide additional insight into how neurons acquire their mature identities and how terminal selector-type TFs function in postmitotic vertebrate neurons. PMID:28769770

Regulatory Mechanisms Controlling Maturation of Serotonin Neuron Identity and Function.

PubMed

Spencer, William C; Deneris, Evan S

2017-01-01

The brain serotonin (5-hydroxytryptamine; 5-HT) system has been extensively studied for its role in normal physiology and behavior, as well as, neuropsychiatric disorders. The broad influence of 5-HT on brain function, is in part due to the vast connectivity pattern of 5-HT-producing neurons throughout the CNS. 5-HT neurons are born and terminally specified midway through embryogenesis, then enter a protracted period of maturation, where they functionally integrate into CNS circuitry and then are maintained throughout life. The transcriptional regulatory networks controlling progenitor cell generation and terminal specification of 5-HT neurons are relatively well-understood, yet the factors controlling 5-HT neuron maturation are only recently coming to light. In this review, we first provide an update on the regulatory network controlling 5-HT neuron development, then delve deeper into the properties and regulatory strategies governing 5-HT neuron maturation. In particular, we discuss the role of the 5-HT neuron terminal selector transcription factor (TF) Pet-1 as a key regulator of 5-HT neuron maturation. Pet-1 was originally shown to positively regulate genes needed for 5-HT synthesis, reuptake and vesicular transport, hence 5-HT neuron-type transmitter identity. It has now been shown to regulate, both positively and negatively, many other categories of genes in 5-HT neurons including ion channels, GPCRs, transporters, neuropeptides, and other transcription factors. Its function as a terminal selector results in the maturation of 5-HT neuron excitability, firing characteristics, and synaptic modulation by several neurotransmitters. Furthermore, there is a temporal requirement for Pet-1 in the control of postmitotic gene expression trajectories thus indicating a direct role in 5-HT neuron maturation. Proper regulation of the maturation of cellular identity is critical for normal neuronal functioning and perturbations in the gene regulatory networks controlling these processes may result in long-lasting changes in brain function in adulthood. Further study of 5-HT neuron gene regulatory networks is likely to provide additional insight into how neurons acquire their mature identities and how terminal selector-type TFs function in postmitotic vertebrate neurons.

Robustness, evolvability, and the logic of genetic regulation.

PubMed

Payne, Joshua L; Moore, Jason H; Wagner, Andreas

2014-01-01

In gene regulatory circuits, the expression of individual genes is commonly modulated by a set of regulating gene products, which bind to a gene's cis-regulatory region. This region encodes an input-output function, referred to as signal-integration logic, that maps a specific combination of regulatory signals (inputs) to a particular expression state (output) of a gene. The space of all possible signal-integration functions is vast and the mapping from input to output is many-to-one: For the same set of inputs, many functions (genotypes) yield the same expression output (phenotype). Here, we exhaustively enumerate the set of signal-integration functions that yield identical gene expression patterns within a computational model of gene regulatory circuits. Our goal is to characterize the relationship between robustness and evolvability in the signal-integration space of regulatory circuits, and to understand how these properties vary between the genotypic and phenotypic scales. Among other results, we find that the distributions of genotypic robustness are skewed, so that the majority of signal-integration functions are robust to perturbation. We show that the connected set of genotypes that make up a given phenotype are constrained to specific regions of the space of all possible signal-integration functions, but that as the distance between genotypes increases, so does their capacity for unique innovations. In addition, we find that robust phenotypes are (i) evolvable, (ii) easily identified by random mutation, and (iii) mutationally biased toward other robust phenotypes. We explore the implications of these latter observations for mutation-based evolution by conducting random walks between randomly chosen source and target phenotypes. We demonstrate that the time required to identify the target phenotype is independent of the properties of the source phenotype.

Robustness, Evolvability, and the Logic of Genetic Regulation

PubMed Central

Moore, Jason H.; Wagner, Andreas

2014-01-01

In gene regulatory circuits, the expression of individual genes is commonly modulated by a set of regulating gene products, which bind to a gene’s cis-regulatory region. This region encodes an input-output function, referred to as signal-integration logic, that maps a specific combination of regulatory signals (inputs) to a particular expression state (output) of a gene. The space of all possible signal-integration functions is vast and the mapping from input to output is many-to-one: for the same set of inputs, many functions (genotypes) yield the same expression output (phenotype). Here, we exhaustively enumerate the set of signal-integration functions that yield idential gene expression patterns within a computational model of gene regulatory circuits. Our goal is to characterize the relationship between robustness and evolvability in the signal-integration space of regulatory circuits, and to understand how these properties vary between the genotypic and phenotypic scales. Among other results, we find that the distributions of genotypic robustness are skewed, such that the majority of signal-integration functions are robust to perturbation. We show that the connected set of genotypes that make up a given phenotype are constrained to specific regions of the space of all possible signal-integration functions, but that as the distance between genotypes increases, so does their capacity for unique innovations. In addition, we find that robust phenotypes are (i) evolvable, (ii) easily identified by random mutation, and (iii) mutationally biased toward other robust phenotypes. We explore the implications of these latter observations for mutation-based evolution by conducting random walks between randomly chosen source and target phenotypes. We demonstrate that the time required to identify the target phenotype is independent of the properties of the source phenotype. PMID:23373974

Regulatory protein BBD18 of the lyme disease spirochete: essential role during tick acquisition?

PubMed

Hayes, Beth M; Dulebohn, Daniel P; Sarkar, Amit; Tilly, Kit; Bestor, Aaron; Ambroggio, Xavier; Rosa, Patricia A

2014-04-01

The Lyme disease spirochete Borrelia burgdorferi senses and responds to environmental cues as it transits between the tick vector and vertebrate host. Failure to properly adapt can block transmission of the spirochete and persistence in either vector or host. We previously identified BBD18, a novel plasmid-encoded protein of B. burgdorferi, as a putative repressor of the host-essential factor OspC. In this study, we investigate the in vivo role of BBD18 as a regulatory protein, using an experimental mouse-tick model system that closely resembles the natural infectious cycle of B. burgdorferi. We show that spirochetes that have been engineered to constitutively produce BBD18 can colonize and persist in ticks but do not infect mice when introduced by either tick bite or needle inoculation. Conversely, spirochetes lacking BBD18 can persistently infect mice but are not acquired by feeding ticks. Through site-directed mutagenesis, we have demonstrated that abrogation of spirochete infection in mice by overexpression of BBD18 occurs only with bbd18 alleles that can suppress OspC synthesis. Finally, we demonstrate that BBD18-mediated regulation does not utilize a previously described ospC operator sequence required by B. burgdorferi for persistence in immunocompetent mice. These data lead us to conclude that BBD18 does not represent the putative repressor utilized by B. burgdorferi for the specific downregulation of OspC in the mammalian host. Rather, we suggest that BBD18 exhibits features more consistent with those of a global regulatory protein whose critical role occurs during spirochete acquisition by feeding ticks. IMPORTANCE Lyme disease, caused by Borrelia burgdorferi, is the most common arthropod-borne disease in North America. B. burgdorferi is transmitted to humans and other vertebrate hosts by ticks as they take a blood meal. Transmission between vectors and hosts requires the bacterium to sense changes in the environment and adapt. However, the mechanisms involved in this process are not well understood. By determining how B. burgdorferi cycles between two very different environments, we can potentially establish novel ways to interfere with transmission and limit infection of this vector-borne pathogen. We are studying a regulatory protein called BBD18 that we recently described. We found that too much BBD18 interferes with the spirochete's ability to establish infection in mice, whereas too little BBD18 appears to prevent colonization in ticks. Our study provides new insight into key elements of the infectious cycle of the Lyme disease spirochete.

EWS and FUS bind a subset of transcribed genes encoding proteins enriched in RNA regulatory functions.

PubMed

Luo, Yonglun; Blechingberg, Jenny; Fernandes, Ana Miguel; Li, Shengting; Fryland, Tue; Børglum, Anders D; Bolund, Lars; Nielsen, Anders Lade

2015-11-14

FUS (TLS) and EWS (EWSR1) belong to the FET-protein family of RNA and DNA binding proteins. FUS and EWS are structurally and functionally related and participate in transcriptional regulation and RNA processing. FUS and EWS are identified in translocation generated cancer fusion proteins and involved in the human neurological diseases amyotrophic lateral sclerosis and fronto-temporal lobar degeneration. To determine the gene regulatory functions of FUS and EWS at the level of chromatin, we have performed chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq). Our results show that FUS and EWS bind to a subset of actively transcribed genes, that binding often is downstream the poly(A)-signal, and that binding overlaps with RNA polymerase II. Functional examinations of selected target genes identified that FUS and EWS can regulate gene expression at different levels. Gene Ontology analyses showed that FUS and EWS target genes preferentially encode proteins involved in regulatory processes at the RNA level. The presented results yield new insights into gene interactions of EWS and FUS and have identified a set of FUS and EWS target genes involved in pathways at the RNA regulatory level with potential to mediate normal and disease-associated functions of the FUS and EWS proteins.

DiRE: identifying distant regulatory elements of co-expressed genes

PubMed Central

Gotea, Valer; Ovcharenko, Ivan

2008-01-01

Regulation of gene expression in eukaryotic genomes is established through a complex cooperative activity of proximal promoters and distant regulatory elements (REs) such as enhancers, repressors and silencers. We have developed a web server named DiRE, based on the Enhancer Identification (EI) method, for predicting distant regulatory elements in higher eukaryotic genomes, namely for determining their chromosomal location and functional characteristics. The server uses gene co-expression data, comparative genomics and profiles of transcription factor binding sites (TFBSs) to determine TFBS-association signatures that can be used for discriminating specific regulatory functions. DiRE's unique feature is its ability to detect REs outside of proximal promoter regions, as it takes advantage of the full gene locus to conduct the search. DiRE can predict common REs for any set of input genes for which the user has prior knowledge of co-expression, co-function or other biologically meaningful grouping. The server predicts function-specific REs consisting of clusters of specifically-associated TFBSs and it also scores the association of individual transcription factors (TFs) with the biological function shared by the group of input genes. Its integration with the Array2BIO server allows users to start their analysis with raw microarray expression data. The DiRE web server is freely available at http://dire.dcode.org. PMID:18487623

Repression of enhancer II activity by a negative regulatory element in the hepatitis B virus genome.

PubMed Central

Lo, W Y; Ting, L P

1994-01-01

Enhancer II of human hepatitis B virus has dual functions in vivo. Located at nucleotides (nt) 1646 to 1741, it can stimulate the surface and X promoters from a downstream position. Moreover, the same sequence can also function as upstream regulatory element that activates the core promoter in a position- and orientation-dependent manner. In this study, we report the identification and characterization of a negative regulatory element (NRE) upstream of enhancer II (nt 1613 to 1636) which can repress both the enhancer and upstream stimulatory function of the enhancer II sequence in differentiated liver cells. This NRE has marginal inhibitory effect by itself but a strong repressive function in the presence of a functional enhancer II. Mutational analysis reveals that sequence from nt 1616 to 1621 is required for repression of enhancer activity by the NRE. Gel shift analysis reveals that this negative regulatory region can be recognized by a specific protein factor(s) present at the 0.4 M NaCl fraction of HepG2 nuclear extracts. The discovery of the NRE indicates that HBV gene transcription is controlled by combined effects of both positive and negative regulation. It also provides a unique system with which to study the mechanism of negative regulation of gene expression. Images PMID:8107237

[Neuronal and hormonal regulatory mechanisms of tears production and secretion].

PubMed

Mrugacz, Małgorzata; Zywalewska, Nella; Bakunowicz-Lazarczyk, Alina

2005-01-01

The ocular surface, tear film, lacrimal glands act as a functional unit to preserve the quality of the refractive surface of the eye, and to resist injury and protect the eye against bodily and environmental conditions. Homeostasis of this functional unit involves neuronal and hormonal regulatory mechanisms. The eye appears to be a target organ for sex hormones particulary the androgen, as they modulate the immune system and trophic functions of the lacrimal and Meibomian glands.

Genome-wide inference of regulatory networks in Streptomyces coelicolor.

PubMed

Castro-Melchor, Marlene; Charaniya, Salim; Karypis, George; Takano, Eriko; Hu, Wei-Shou

2010-10-18

The onset of antibiotics production in Streptomyces species is co-ordinated with differentiation events. An understanding of the genetic circuits that regulate these coupled biological phenomena is essential to discover and engineer the pharmacologically important natural products made by these species. The availability of genomic tools and access to a large warehouse of transcriptome data for the model organism, Streptomyces coelicolor, provides incentive to decipher the intricacies of the regulatory cascades and develop biologically meaningful hypotheses. In this study, more than 500 samples of genome-wide temporal transcriptome data, comprising wild-type and more than 25 regulatory gene mutants of Streptomyces coelicolor probed across multiple stress and medium conditions, were investigated. Information based on transcript and functional similarity was used to update a previously-predicted whole-genome operon map and further applied to predict transcriptional networks constituting modules enriched in diverse functions such as secondary metabolism, and sigma factor. The predicted network displays a scale-free architecture with a small-world property observed in many biological networks. The networks were further investigated to identify functionally-relevant modules that exhibit functional coherence and a consensus motif in the promoter elements indicative of DNA-binding elements. Despite the enormous experimental as well as computational challenges, a systems approach for integrating diverse genome-scale datasets to elucidate complex regulatory networks is beginning to emerge. We present an integrated analysis of transcriptome data and genomic features to refine a whole-genome operon map and to construct regulatory networks at the cistron level in Streptomyces coelicolor. The functionally-relevant modules identified in this study pose as potential targets for further studies and verification.

Transcription Factor Networks Directing the Development, Function, and Evolution of Innate Lymphoid Effectors

PubMed Central

Kang, Joonsoo; Malhotra, Nidhi

2015-01-01

Mammalian lymphoid immunity is mediated by fast and slow responders to pathogens. Fast innate lymphocytes are active within hours after infections in mucosal tissues. Slow adaptive lymphocytes are conventional T and B cells with clonal antigen receptors that function days after pathogen exposure. A transcription factor (TF) regulatory network guiding early T cell development is at the core of effector function diversification in all innate lymphocytes, and the kinetics of immune responses is set by developmental programming. Operational units within the innate lymphoid system are not classified by the types of pathogen-sensing machineries but rather by discrete effector functions programmed by regulatory TF networks. Based on the evolutionary history of TFs of the regulatory networks, fast effectors likely arose earlier in the evolution of animals to fortify body barriers, and in mammals they often develop in fetal ontogeny prior to the establishment of fully competent adaptive immunity. PMID:25650177

Zinc in Cellular Regulation: The Nature and Significance of "Zinc Signals".

PubMed

Maret, Wolfgang

2017-10-31

In the last decade, we witnessed discoveries that established Zn 2+ as a second major signalling metal ion in the transmission of information within cells and in communication between cells. Together with Ca 2+ and Mg 2+ , Zn 2+ covers biological regulation with redox-inert metal ions over many orders of magnitude in concentrations. The regulatory functions of zinc ions, together with their functions as a cofactor in about three thousand zinc metalloproteins, impact virtually all aspects of cell biology. This article attempts to define the regulatory functions of zinc ions, and focuses on the nature of zinc signals and zinc signalling in pathways where zinc ions are either extracellular stimuli or intracellular messengers. These pathways interact with Ca 2+ , redox, and phosphorylation signalling. The regulatory functions of zinc require a complex system of precise homeostatic control for transients, subcellular distribution and traffic, organellar homeostasis, and vesicular storage and exocytosis of zinc ions.

78 FR 13132 - Self-Regulatory Organizations; NASDAQ OMX PHLX LLC; Notice of Filing of Proposed Rule Change To...

Federal Register 2010, 2011, 2012, 2013, 2014

2013-02-26

... SECURITIES AND EXCHANGE COMMISSION [Release No. 34-68960; File No. SR-Phlx-2013-09] Self-Regulatory Organizations; NASDAQ OMX PHLX LLC; Notice of Filing of Proposed Rule Change To Enhance the Functionality Offered on Its Options Floor Broker Management System (``FBMS'') by, Among Other Things, Automating Functions Currently Performed by Floor...

The Role of Executive Functions Skills and Self-Regulation Behaviors in School Readiness and Adjustment

ERIC Educational Resources Information Center

Sasser, Tyler R.; Bierman, Karen L.

2012-01-01

The aim of this study was to examine co-variation in the development of self-regulatory skills evident in pre-kindergarten and evaluate the implications of that variation for school adjustment in kindergarten and first grade. Measures of self-regulatory skill development included: direct assessments of EF (executive function) (e.g., Peg Tapping,…

Understanding regulatory networks requires more than computing a multitude of graph statistics. Comment on "Drivers of structural features in gene regulatory networks: From biophysical constraints to biological function" by O.C. Martin et al.

NASA Astrophysics Data System (ADS)

Tkačik, Gašper

2016-07-01

The article by O. Martin and colleagues provides a much needed systematic review of a body of work that relates the topological structure of genetic regulatory networks to evolutionary selection for function. This connection is very important. Using the current wealth of genomic data, statistical features of regulatory networks (e.g., degree distributions, motif composition, etc.) can be quantified rather easily; it is, however, often unclear how to interpret the results. On a graph theoretic level the statistical significance of the results can be evaluated by comparing observed graphs to ;randomized; ones (bravely ignoring the issue of how precisely to randomize!) and comparing the frequency of appearance of a particular network structure relative to a randomized null expectation. While this is a convenient operational test for statistical significance, its biological meaning is questionable. In contrast, an in-silico genotype-to-phenotype model makes explicit the assumptions about the network function, and thus clearly defines the expected network structures that can be compared to the case of no selection for function and, ultimately, to data.

Interfacing cellular networks of S. cerevisiae and E. coli: Connecting dynamic and genetic information

PubMed Central

2013-01-01

Background In recent years, various types of cellular networks have penetrated biology and are nowadays used omnipresently for studying eukaryote and prokaryote organisms. Still, the relation and the biological overlap among phenomenological and inferential gene networks, e.g., between the protein interaction network and the gene regulatory network inferred from large-scale transcriptomic data, is largely unexplored. Results We provide in this study an in-depth analysis of the structural, functional and chromosomal relationship between a protein-protein network, a transcriptional regulatory network and an inferred gene regulatory network, for S. cerevisiae and E. coli. Further, we study global and local aspects of these networks and their biological information overlap by comparing, e.g., the functional co-occurrence of Gene Ontology terms by exploiting the available interaction structure among the genes. Conclusions Although the individual networks represent different levels of cellular interactions with global structural and functional dissimilarities, we observe crucial functions of their network interfaces for the assembly of protein complexes, proteolysis, transcription, translation, metabolic and regulatory interactions. Overall, our results shed light on the integrability of these networks and their interfacing biological processes. PMID:23663484

Fungal Genes in Context: Genome Architecture Reflects Regulatory Complexity and Function

PubMed Central

Noble, Luke M.; Andrianopoulos, Alex

2013-01-01

Gene context determines gene expression, with local chromosomal environment most influential. Comparative genomic analysis is often limited in scope to conserved or divergent gene and protein families, and fungi are well suited to this approach with low functional redundancy and relatively streamlined genomes. We show here that one aspect of gene context, the amount of potential upstream regulatory sequence maintained through evolution, is highly predictive of both molecular function and biological process in diverse fungi. Orthologs with large upstream intergenic regions (UIRs) are strongly enriched in information processing functions, such as signal transduction and sequence-specific DNA binding, and, in the genus Aspergillus, include the majority of experimentally studied, high-level developmental and metabolic transcriptional regulators. Many uncharacterized genes are also present in this class and, by implication, may be of similar importance. Large intergenic regions also share two novel sequence characteristics, currently of unknown significance: they are enriched for plus-strand polypyrimidine tracts and an information-rich, putative regulatory motif that was present in the last common ancestor of the Pezizomycotina. Systematic consideration of gene UIR in comparative genomics, particularly for poorly characterized species, could help reveal organisms’ regulatory priorities. PMID:23699226

Functional analysis of two sterol regulatory element binding proteins in Penicillium digitatum

PubMed Central

Ruan, Ruoxin; Wang, Mingshuang; Liu, Xin; Sun, Xuepeng; Chung, Kuang-Ren

2017-01-01

The sterol regulatory element binding proteins (SREBPs) are key regulators for sterol homeostasis in most fungi. In the citrus postharvest pathogen Penicillium digitatum, the SREBP homolog is required for fungicide resistance and regulation of CYP51 expression. In this study, we identified another SREBP transcription factor PdSreB in P. digitatum, and the biological functions of both SREBPs were characterized and compared. Inactivation of PdsreA, PdsreB or both genes in P. digitatum reduced ergosterol contents and increased sensitivities to sterol 14-α-demethylation inhibitors (DMIs) and cobalt chloride. Fungal strains impaired at PdsreA but not PdsreB increased sensitivity to tridemorph and an iron chelator 2,2’-dipyridyl. Virulence assays on citrus fruit revealed that fungal strains impaired at PdsreA, PdsreB or both induce maceration lesions similar to those induced by wild-type. However, ΔPdsreA, ΔPdsreB or the double mutant strain rarely produce aerial mycelia on infected citrus fruit peels. RNA-Seq analysis showed the broad regulatory functions of both SREBPs in biosynthesis, transmembrane transportation and stress responses. Our results provide new insights into the conserved and differentiated regulatory functions of SREBP homologs in plant pathogenic fungi. PMID:28467453

Validation of Skeletal Muscle cis-Regulatory Module Predictions Reveals Nucleotide Composition Bias in Functional Enhancers

PubMed Central

Kwon, Andrew T.; Chou, Alice Yi; Arenillas, David J.; Wasserman, Wyeth W.

2011-01-01

We performed a genome-wide scan for muscle-specific cis-regulatory modules (CRMs) using three computational prediction programs. Based on the predictions, 339 candidate CRMs were tested in cell culture with NIH3T3 fibroblasts and C2C12 myoblasts for capacity to direct selective reporter gene expression to differentiated C2C12 myotubes. A subset of 19 CRMs validated as functional in the assay. The rate of predictive success reveals striking limitations of computational regulatory sequence analysis methods for CRM discovery. Motif-based methods performed no better than predictions based only on sequence conservation. Analysis of the properties of the functional sequences relative to inactive sequences identifies nucleotide sequence composition can be an important characteristic to incorporate in future methods for improved predictive specificity. Muscle-related TFBSs predicted within the functional sequences display greater sequence conservation than non-TFBS flanking regions. Comparison with recent MyoD and histone modification ChIP-Seq data supports the validity of the functional regions. PMID:22144875

Alternative end-joining repair pathways are the ultimate backup for abrogated classical non-homologous end-joining and homologous recombination repair: Implications for the formation of chromosome translocations.

PubMed

Iliakis, George; Murmann, Tamara; Soni, Aashish

2015-11-01

DNA double strand breaks (DSB) are the most deleterious lesions for the integrity of the genome, as their misrepair can lead to the formation of chromosome translocations. Cells have evolved two main repair pathways to suppress the formation of these genotoxic lesions: homology-dependent, error-free homologous recombination repair (HRR), and potentially error-prone, classical, DNA-PK-dependent non-homologous end-joining (c-NHEJ). The most salient feature of c-NHEJ, speed, will largely suppress chromosome translocation formation, while sequence alterations at the junction remain possible. It is now widely accepted that when c-NHEJ is inactivated, globally or locally, an alternative form of end-joining (alt-EJ) removes DSBs. Alt-EJ operates with speed and fidelity markedly lower than c-NHEJ, causing thus with higher probability chromosome translocations, and generating more extensive sequence alterations at the junction. Our working hypothesis is that alt-EJ operates as a backup to c-NHEJ. Recent results show that alt-EJ can also backup abrogated HRR in G2 phase cells, again at the cost of elevated formation of chromosome translocations. These observations raise alt-EJ to a global rescuing mechanism operating on ends that have lost their chromatin context in ways that compromise processing by HRR or c-NHEJ. While responsible for eliminating from the genome highly cytotoxic DNA ends, alt-EJ provides this function at the price of increased translocation formation. Here, we analyze recent literature on the mechanisms of chromosome translocation formation and propose a functional hierarchy among DSB processing pathways that makes alt-EJ the global backup pathway. We discuss possible ramifications of this model in cellular DSB management and pathway choice, and analyze its implications in radiation carcinogenesis and the design of novel therapeutic approaches. Copyright © 2015 Elsevier B.V. All rights reserved.

Dabigatran abrogates brain endothelial cell permeability in response to thrombin

PubMed Central

Hawkins, Brian Thomas; Gu, Yu-Huan; Izawa, Yoshikane; del Zoppo, Gregory John

2015-01-01

Atrial fibrillation (AF) increases the risk and severity of thromboembolic stroke. Generally, antithrombotic agents increase the hemorrhagic risk of thromboembolic stroke. However, significant reductions in thromboembolism and intracerebral hemorrhage have been shown with the antithrombin dabigatran compared with warfarin. As thrombin has been implicated in microvessel injury during cerebral ischemia, we hypothesized that dabigatran decreases the risk of intracerebral hemorrhage by direct inhibition of the thrombin-mediated increase in cerebral endothelial cell permeability. Primary murine brain endothelial cells (mBECs) were exposed to murine thrombin before measuring permeability to 4-kDa fluorescein isothiocyanate-dextran. Thrombin increased mBEC permeability in a concentration-dependent manner, without significant endothelial cell death. Pretreatment of mBECs with dabigatran completely abrogated the effect of thrombin on permeability. Neither the expressions of the endothelial cell β1-integrins nor the tight junction protein claudin-5 were affected by thrombin exposure. Oxygen-glucose deprivation (OGD) also increased permeability; this effect was abrogated by treatment with dabigatran, as was the additive effect of thrombin and OGD on permeability. Taken together, these results indicate that dabigatran could contribute to a lower risk of intracerebral hemorrhage during embolism-associated ischemia from AF by protection of the microvessel permeability barrier from local thrombin challenge. PMID:25669912

A Novel Clinically Relevant Strategy to Abrogate Autoimmunity and Regulate Alloimmunity in NOD Mice

PubMed Central

Vergani, Andrea; D'Addio, Francesca; Jurewicz, Mollie; Petrelli, Alessandra; Watanabe, Toshihiko; Liu, Kaifeng; Law, Kenneth; Schuetz, Christian; Carvello, Michele; Orsenigo, Elena; Deng, Shaoping; Rodig, Scott J.; Ansari, Javeed M.; Staudacher, Carlo; Abdi, Reza; Williams, John; Markmann, James; Atkinson, Mark; Sayegh, Mohamed H.; Fiorina, Paolo

2010-01-01

OBJECTIVE To investigate a new clinically relevant immunoregulatory strategy based on treatment with murine Thymoglobulin mATG Genzyme and CTLA4-Ig in NOD mice to prevent allo- and autoimmune activation using a stringent model of islet transplantation and diabetes reversal. RESEARCH DESIGN AND METHODS Using allogeneic islet transplantation models as well as NOD mice with recent onset type 1 diabetes, we addressed the therapeutic efficacy and immunomodulatory mechanisms associated with a new immunoregulatory protocol based on prolonged low-dose mATG plus CTLA4-Ig. RESULTS BALB/c islets transplanted into hyperglycemic NOD mice under prolonged mATG+CTLA4-Ig treatment showed a pronounced delay in allograft rejection compared with untreated mice (mean survival time: 54 vs. 8 days, P < 0.0001). Immunologic analysis of mice receiving transplants revealed a complete abrogation of autoimmune responses and severe downregulation of alloimmunity in response to treatment. The striking effect on autoimmunity was confirmed by 100% diabetes reversal in newly hyperglycemic NOD mice and 100% indefinite survival of syngeneic islet transplantation (NOD.SCID into NOD mice). CONCLUSIONS The capacity to regulate alloimmunity and to abrogate the autoimmune response in NOD mice in different settings confirmed that prolonged mATG+CTLA4-Ig treatment is a clinically relevant strategy to translate to humans with type 1 diabetes. PMID:20805386

Genome-wide comparative analysis reveals human-mouse regulatory landscape and evolution.

PubMed

Denas, Olgert; Sandstrom, Richard; Cheng, Yong; Beal, Kathryn; Herrero, Javier; Hardison, Ross C; Taylor, James

2015-02-14

Because species-specific gene expression is driven by species-specific regulation, understanding the relationship between sequence and function of the regulatory regions in different species will help elucidate how differences among species arise. Despite active experimental and computational research, relationships among sequence, conservation, and function are still poorly understood. We compared transcription factor occupied segments (TFos) for 116 human and 35 mouse TFs in 546 human and 125 mouse cell types and tissues from the Human and the Mouse ENCODE projects. We based the map between human and mouse TFos on a one-to-one nucleotide cross-species mapper, bnMapper, that utilizes whole genome alignments (WGA). Our analysis shows that TFos are under evolutionary constraint, but a substantial portion (25.1% of mouse and 25.85% of human on average) of the TFos does not have a homologous sequence on the other species; this portion varies among cell types and TFs. Furthermore, 47.67% and 57.01% of the homologous TFos sequence shows binding activity on the other species for human and mouse respectively. However, 79.87% and 69.22% is repurposed such that it binds the same TF in different cells or different TFs in the same cells. Remarkably, within the set of repurposed TFos, the corresponding genome regions in the other species are preferred locations of novel TFos. These events suggest exaptation of some functional regulatory sequences into new function. Despite TFos repurposing, we did not find substantial changes in their predicted target genes, suggesting that CRMs buffer evolutionary events allowing little or no change in the TFos - target gene associations. Thus, the small portion of TFos with strictly conserved occupancy underestimates the degree of conservation of regulatory interactions. We mapped regulatory sequences from an extensive number of TFs and cell types between human and mouse using WGA. A comparative analysis of this correspondence unveiled the extent of the shared regulatory sequence across TFs and cell types under study. Importantly, a large part of the shared regulatory sequence is repurposed on the other species. This sequence, fueled by turnover events, provides a strong case for exaptation in regulatory elements.

Self-Regulation and Executive Functioning as Related to Survival in Motor Neuron Disease: Preliminary Findings.

PubMed

Garcia-Willingham, Natasha E; Roach, Abbey R; Kasarskis, Edward J; Segerstrom, Suzanne C

2018-05-16

Disease progression varies widely among patients with motor neuron disease (MND). Patients with MND and coexisting dementia have shorter survival. However, implications of mild cognitive and behavioral difficulties are unclear. The present study examined the relative contribution of executive functioning and self-regulation difficulties on survival over a 6-year period among patients with MND, who scored largely within normal limits on cognitive and behavioral indices. Patients with MND (N=37, age=59.97±11.57, 46% female) completed the Wisconsin Card Sorting Task (WCST) as an executive functioning perseveration index. The Behavior Rating Inventory of Executive Functions (BRIEF-A) was used as a behavioral measure of self-regulation in two subdomains self-regulatory behavior (Behavioral Regulation) and self-regulatory problem-solving (Metacognition). Cox proportional hazard regression analyses were used. In total, 23 patients died during follow-up. In Cox proportional hazard regressions adjusted for a priori covariates, each 10-point T-score increment in patient-reported BRIEF-A self-regulatory behavior and problem-solving difficulties increased mortality risk by 94% and103%, respectively (adjusted HR=1.94, 95% CI [1.07, 3.52]; adjusted HR=2.03, 95% CI [1.19, 3.48]). In sensitivity analyses, patient-reported self-regulatory problem-solving remained significant independent of disease severity and a priori covariates (adjusted HR=1.68, 95% CI [1.01, 2.78], though the predictive value of self-regulatory behavior was attenuated in adjusted models (HR=1.67, 95% CI [0.85, 3.27). Caregiver-reported BRIEF-A ratings of patients and WCST perseverative errors did not significantly predict survival. Preliminary evidence suggests patient-reported self-regulatory problem-solving difficulties indicate poorer prognosis in MND. Further research is needed to uncover mechanisms that negatively affect patient survival.

Update: the role of FoxP3 in allergic disease.

PubMed

Paik, Young; Dahl, Matthew; Fang, Deyu; Calhoun, Karen

2008-06-01

T-regulatory cells play a key role in allergic and asthmatic inflammatory airway diseases. This review discusses the importance of a critical gene associated with T-regulatory cells. Forkhead box P3 is a forkhead-winged helix transcription factor gene involved in immune function in allergy and asthma. Recently, many functions of forkhead box P3 and its influence on the immune system have been elucidated. T-regulatory cells that are CD4+CD25+ and express forkhead box P3, influence the development and expression of atopy and allergic response. The exact mechanisms are not yet delineated, but multiple recent studies provide greater understanding of the mechanism of forkhead box P3 and its influence on these T-regulatory cells. Greater understanding of the molecular and immunological mechanisms underlying the T-regulatory cells and forkhead box P3 will permit the development of targeted treatment modalities to influence disease processes such as allergic rhinitis and bronchial asthma.

Functional and topological characteristics of mammalian regulatory domains

PubMed Central

Symmons, Orsolya; Uslu, Veli Vural; Tsujimura, Taro; Ruf, Sandra; Nassari, Sonya; Schwarzer, Wibke; Ettwiller, Laurence; Spitz, François

2014-01-01

Long-range regulatory interactions play an important role in shaping gene-expression programs. However, the genomic features that organize these activities are still poorly characterized. We conducted a large operational analysis to chart the distribution of gene regulatory activities along the mouse genome, using hundreds of insertions of a regulatory sensor. We found that enhancers distribute their activities along broad regions and not in a gene-centric manner, defining large regulatory domains. Remarkably, these domains correlate strongly with the recently described TADs, which partition the genome into distinct self-interacting blocks. Different features, including specific repeats and CTCF-binding sites, correlate with the transition zones separating regulatory domains, and may help to further organize promiscuously distributed regulatory influences within large domains. These findings support a model of genomic organization where TADs confine regulatory activities to specific but large regulatory domains, contributing to the establishment of specific gene expression profiles. PMID:24398455

Design Principles of Regulatory Networks: Searching for the Molecular Algorithms of the Cell

PubMed Central

Lim, Wendell A.; Lee, Connie M.; Tang, Chao

2013-01-01

A challenge in biology is to understand how complex molecular networks in the cell execute sophisticated regulatory functions. Here we explore the idea that there are common and general principles that link network structures to biological functions, principles that constrain the design solutions that evolution can converge upon for accomplishing a given cellular task. We describe approaches for classifying networks based on abstract architectures and functions, rather than on the specific molecular components of the networks. For any common regulatory task, can we define the space of all possible molecular solutions? Such inverse approaches might ultimately allow the assembly of a design table of core molecular algorithms that could serve as a guide for building synthetic networks and modulating disease networks. PMID:23352241

CTCF and Cohesin in Genome Folding and Transcriptional Gene Regulation.

PubMed

Merkenschlager, Matthias; Nora, Elphège P

2016-08-31

Genome function, replication, integrity, and propagation rely on the dynamic structural organization of chromosomes during the cell cycle. Genome folding in interphase provides regulatory segmentation for appropriate transcriptional control, facilitates ordered genome replication, and contributes to genome integrity by limiting illegitimate recombination. Here, we review recent high-resolution chromosome conformation capture and functional studies that have informed models of the spatial and regulatory compartmentalization of mammalian genomes, and discuss mechanistic models for how CTCF and cohesin control the functional architecture of mammalian chromosomes.

Genetic moderation of the association between regulatory focus and reward responsiveness: a proof-of-concept study.

PubMed

Goetz, Elena L; Hariri, Ahmad R; Pizzagalli, Diego A; Strauman, Timothy J

2013-02-01

Recent studies implicate individual differences in regulatory focus as contributing to self-regulatory dysfunction, particularly not responding to positive outcomes. How such individual differences emerge, however, is unclear. We conducted a proof-of-concept study to examine the moderating effects of genetically driven variation in dopamine signaling, a key modulator of neural reward circuits, on the association between regulatory focus and reward cue responsiveness. Healthy Caucasians (N=59) completed a measure of chronic regulatory focus and a probabilistic reward task. A common functional genetic polymorphism impacting prefrontal dopamine signaling (COMT rs4680) was evaluated. Response bias, the participants' propensity to modulate behavior as a function of reward, was predicted by an interaction of regulatory focus and COMT genotype. Specifically, self-perceived success at achieving promotion goals predicted total response bias, but only for individuals with the COMT genotype (Val/Val) associated with relatively increased phasic dopamine signaling and cognitive flexibility. The combination of success in promotion goal pursuit and Val/Val genotype appears to facilitate responding to reward opportunities in the environment. This study is among the first to integrate an assessment of self-regulatory style with an examination of genetic variability that underlies responsiveness to positive outcomes in goal pursuit.

Cis-regulatory Evolution of Chalcone-Synthase Expression in the Genus Arabidopsis

PubMed Central

de Meaux, Juliette; Pop, A.; Mitchell-Olds, T.

2006-01-01

The contribution of cis-regulation to adaptive evolutionary change is believed to be essential, yet little is known about the evolutionary rules that govern regulatory sequences. Here, we characterize the short-term evolutionary dynamics of a cis-regulatory region within and among two closely related species, A. lyrata and A. halleri, and compare our findings to A. thaliana. We focused on the cis-regulatory region of chalcone synthase (CHS), a key enzyme involved in the synthesis of plant secondary metabolites. We observed patterns of nucleotide diversity that differ among species but do not depart from neutral expectations. Using intra- and interspecific F1 progeny, we have evaluated functional cis-regulatory variation in response to light and herbivory, environmental cues, which are known to induce CHS expression. We find that substantial cis-regulatory variation segregates within and among populations as well as between species, some of which results from interspecific genetic introgression. We further demonstrate that, in A. thaliana, CHS cis-regulation in response to herbivory is greater than in A. lyrata or A. halleri. Our work indicates that the evolutionary dynamics of a cis-regulatory region is characterized by pervasive functional variation, achieved mostly by modification of response modules to one but not all environmental cues. Our study did not detect the footprint of selection on this variation. PMID:17028316

T regulatory cells in contact hypersensitivity.

PubMed

Cavani, Andrea

2008-08-01

The review summarizes the recent investigations focused on T regulatory cells in hapten diseases. Multiple mechanisms ensure tolerance to small chemicals penetrating the skin. Among these, specific T regulatory cells play a major role in controlling harmful immune responses to environmental antigens. Most of the T regulatory cells involved in this process belongs to the CD4 subset and suppress hapten-specific immune response through the release of IL-10 and through direct interaction with effector T cells, blocking their function. Methods for in-vitro and in-vivo expansion of specific T regulatory cells may represent an innovative approach for the cure of contact hypersensitivity.

Immune Tolerance in Multiple Sclerosis

PubMed Central

Goverman, Joan M.

2011-01-01

Summary Multiple sclerosis is believed to be mediated by T cells specific for myelin antigens that circulate harmlessly in the periphery of healthy individuals until they are erroneously by an environmental stimulus. Upon activation, the T cells enter the central nervous system and orchestrate an immune response against myelin. To understand the initial steps in the pathogenesis of multiple sclerosis, it is important to identify the mechanisms that maintain T-cell tolerance to myelin antigens and to understand how some myelin-specific T cells escape tolerance and what conditions lead to their activation. Central tolerance strongly shapes the peripheral repertoire of myelin-specific T cells, as most myelin-specific T cells are eliminated by clonal deletion in the thymus. Self-reactive T cells that escape central tolerance are generally capable only of low-avidity interactions with antigen-presenting cells. Despite the low avidity of these interactions, peripheral tolerance mechanisms are required to prevent spontaneous autoimmunity. Multiple peripheral tolerance mechanisms for myelin-specific T cells have been indentified, the most important of which appears to be regulatory T cells. While most studies have focused on CD4+ myelin-specific T cells, interesting differences in tolerance mechanisms and the conditions that abrogate these mechanisms have recently been described for CD8+ myelin-specific T cells. PMID:21488900

Chronic Trichuris muris infection causes neoplastic change in the intestine and exacerbates tumour formation in APC min/+ mice

PubMed Central

Cliffe, Laura J.; Bancroft, Alison J.; Forman, Simon P.; Thompson, Seona; Booth, Cath

2017-01-01

Incidences of infection-related cancers are on the rise in developing countries where the prevalence of intestinal nematode worm infections are also high. Trichuris muris (T. muris) is a murine gut-dwelling nematode that is the direct model for human T. trichiura, one of the major soil-transmitted helminth infections of humans. In order to assess whether chronic infection with T. muris does indeed influence the development of cancer hallmarks, both wild type mice and colon cancer model (APC min/+) mice were infected with this parasite. Parasite infection in wild type mice led to the development of neoplastic change similar to that seen in mice that had been treated with the carcinogen azoxymethane. Additionally, both chronic and acute infection in the APCmin/+ mice led to an enhanced tumour development that was distinct to the site of infection suggesting systemic control. By blocking the parasite induced T regulatory response in these mice, the increase in the number of tumours following infection was abrogated. Thus T. muris infection alone causes an increase in gut pathologies that are known to be markers of cancer but also increases the incidence of tumour formation in a colon cancer model. The influence of parasitic worm infection on the development of cancer may therefore be significant. PMID:28650985

Chronic Trichuris muris infection causes neoplastic change in the intestine and exacerbates tumour formation in APC min/+ mice.

PubMed

Hayes, Kelly S; Cliffe, Laura J; Bancroft, Alison J; Forman, Simon P; Thompson, Seona; Booth, Cath; Grencis, Richard K

2017-06-01

Incidences of infection-related cancers are on the rise in developing countries where the prevalence of intestinal nematode worm infections are also high. Trichuris muris (T. muris) is a murine gut-dwelling nematode that is the direct model for human T. trichiura, one of the major soil-transmitted helminth infections of humans. In order to assess whether chronic infection with T. muris does indeed influence the development of cancer hallmarks, both wild type mice and colon cancer model (APC min/+) mice were infected with this parasite. Parasite infection in wild type mice led to the development of neoplastic change similar to that seen in mice that had been treated with the carcinogen azoxymethane. Additionally, both chronic and acute infection in the APCmin/+ mice led to an enhanced tumour development that was distinct to the site of infection suggesting systemic control. By blocking the parasite induced T regulatory response in these mice, the increase in the number of tumours following infection was abrogated. Thus T. muris infection alone causes an increase in gut pathologies that are known to be markers of cancer but also increases the incidence of tumour formation in a colon cancer model. The influence of parasitic worm infection on the development of cancer may therefore be significant.

Live fate-mapping of joint-associated fibroblasts visualizes expansion of cell contributions during zebrafish fin regeneration.

PubMed

Tornini, Valerie A; Thompson, John D; Allen, Raymond L; Poss, Kenneth D

2017-08-15

The blastema is a mass of progenitor cells responsible for regeneration of amputated salamander limbs and fish fins. Previous studies have indicated that resident cell sources producing the blastema contribute lineage-restricted progeny to regenerating tissue. However, these studies have labeled general cell types rather than granular cell subpopulations, and they do not explain the developmental transitions that must occur for distal structures to arise from cells with proximal identities in the appendage stump. Here, we find that regulatory sequences of tph1b , which encodes an enzyme that synthesizes serotonin, mark a subpopulation of fibroblast-like cells restricted to the joints of uninjured adult zebrafish fins. Amputation stimulates serotonin production in regenerating fin fibroblasts, yet targeted tph1b mutations abrogating this response do not disrupt fin regeneration. In uninjured animals, tph1b -expressing cells contribute fibroblast progeny that remain restricted to joints throughout life. By contrast, upon amputation, tph1b + joint cells give rise to fibroblasts that distribute across the entire lengths of regenerating fin rays. Our experiments visualize and quantify how incorporation into an appendage blastema broadens the progeny contributions of a cellular subpopulation that normally has proximodistal restrictions. © 2017. Published by The Company of Biologists Ltd.

COT phosphorylates prolyl-isomerase Pin1 to promote tumorigenesis in breast cancer.

PubMed

Kim, Garam; Khanal, Prem; Kim, Jin Young; Yun, Hyo-Jeong; Lim, Sung-Chul; Shim, Jung-Hyun; Choi, Hong Seok

2015-06-01

Pin1, a conserved eukaryotic Peptidyl-prolyl cis/trans isomerase, has profound effects on numerous key-signaling molecules, and its deregulation contributes to disease, particularly cancer. Although Pin1-mediated prolyl isomerization is an essential and novel regulatory mechanism for protein phosphorylation, little is known about the upstream signaling pathway(s) that regulates Pin1 activity. Here, we identify MAP3K-related serine-threonine kinase (the gene encoding COT/Tpl2) as a kinase responsible for phosphorylation of Pin1 Ser16. COT interacts with and phosphorylates Pin1 on Ser16. Consequently, Pin1 Ser16 phosphorylation by COT increases cyclin D1 abundance and enhances tumorigenecity of MCF7 cells. In contrast, depletion of COT in MCF7 cells leads to downregulation of Pin1 Ser16 phosphorylation, which subsequently decrease cyclin D1 levels, inhibiting tumorigenecity of MCF7 cells. In a xenograft model, treatment of TKI, a COT inhibitor, and Juglone, a Pin1 inhibitor, abrogates tumor growth. In human breast cancer patients, immunohistochemical staining shows that Pin1 pSer16 levels are positively correlated with COT levels, providing strong evidence for an essential role of the COT/Pin1 axis in conveying oncogenic signals to promote aggressiveness in human breast cancer. © 2013 Wiley Periodicals, Inc.

Epithelial-derived IL-33 promotes intestinal tumorigenesis in Apc Min/+ mice.

PubMed

He, Zhengxiang; Chen, Lili; Souto, Fabricio O; Canasto-Chibuque, Claudia; Bongers, Gerold; Deshpande, Madhura; Harpaz, Noam; Ko, Huaibin M; Kelley, Kevin; Furtado, Glaucia C; Lira, Sergio A

2017-07-14

Increased expression of Interleukin (IL)-33 has been detected in intestinal samples of patients with ulcerative colitis, a condition associated with increased risk for colon cancer, but its role in the development of colorectal cancer has yet to be fully examined. Here, we investigated the role of epithelial expressed IL-33 during development of intestinal tumors. IL-33 expression was detected in epithelial cells in colorectal cancer specimens and in the Apc Min/+ mice. To better understand the role of epithelial-derived IL-33 in the intestinal tumorigenesis, we generated transgenic mice expressing IL-33 in intestinal epithelial cells (V33 mice). V33 Apc Min/+ mice, resulting from the cross of V33 with Apc Min/+ mice, had increased intestinal tumor burden compared with littermate Apc Min/+ mice. Consistently, Apc Min/+ mice deficient for IL-33 receptor (ST2), had reduced polyp burden. Mechanistically, overexpression of IL-33 promoted expansion of ST2 + regulatory T cells, increased Th2 cytokine milieu, and induced alternatively activated macrophages in the gut. IL-33 promoted marked changes in the expression of antimicrobial peptides, and antibiotic treatment of V33 Apc Min/+ mice abrogated the tumor promoting-effects of IL-33 in the colon. In conclusion, elevated IL-33 signaling increases tumor development in the Apc Min/+ mice.

HOXA9 Methylation by PRMT5 Is Essential for Endothelial Cell Expression of Leukocyte Adhesion Molecules

PubMed Central

Bandyopadhyay, Smarajit; Harris, Daniel P.; Adams, Gregory N.; Lause, Gregory E.; McHugh, Anne; Tillmaand, Emily G.; Money, Angela; Willard, Belinda; Fox, Paul L.

2012-01-01

The induction of proinflammatory proteins in stimulated endothelial cells (EC) requires activation of multiple transcription programs. The homeobox transcription factor HOXA9 has an important regulatory role in cytokine induction of the EC-leukocyte adhesion molecules (ELAM) E-selectin and vascular cell adhesion molecule 1 (VCAM-1). However, the mechanism underlying stimulus-dependent activation of HOXA9 is completely unknown. Here, we elucidate the molecular mechanism of HOXA9 activation by tumor necrosis factor alpha (TNF-α) and show an unexpected requirement for arginine methylation by protein arginine methyltransferase 5 (PRMT5). PRMT5 was identified as a TNF-α-dependent binding partner of HOXA9 by mass spectrometry. Small interfering RNA (siRNA)-mediated depletion of PRMT5 abrogated stimulus-dependent HOXA9 methylation with concomitant loss in E-selectin or VCAM-1 induction. Chromatin immunoprecipitation analysis revealed that PRMT5 is recruited to the E-selectin promoter following transient HOXA9 binding to its cognate recognition sequence. PRMT5 induces symmetric dimethylation of Arg140 on HOXA9, an event essential for E-selectin induction. In summary, PRMT5 is a critical coactivator component in a newly defined, HOXA9-containing transcription complex. Moreover, stimulus-dependent methylation of HOXA9 is essential for ELAM expression during the EC inflammatory response. PMID:22269951

Management of fear of radiation exposure in carers of outpatients treated with iodine-131.

PubMed

Calais, Phillipe J; Page, Andrew C; Turner, J Harvey

2012-07-01

To characterise potential fear of radiation exposure in a normal population of individuals who have volunteered to care for a radioactive family member or friend after outpatient radioimmunotherapy (RIT) treatment for cancer, and obtain their knowing and willing acceptance of the risk. Over 750 carers of 300 patients confined to their homes for 1 week following outpatient iodine-131 rituximab RIT of lymphoma were interviewed by a nuclear medicine physicist according to a multi-visit integrated protocol designed to minimise radiation exposure, define risk and gain informed consent. Median radiation exposure of carers was 0.49 mSv (range 0.01-3.7 mSv) which is below the Western Australian regulatory limit of 5 mSv for consenting adult carers of radioactive patients. After signing a declaration of consent, only 2 carers of 750 abrogated their responsibility and none of those who carried out their duties expressed residual concerns at the end of the exit interview with respect to their radiation exposure. Fear of radiation exposure in a normal population may be characterised as a normal emotional response. In the special case of carers of radioactive patients, this fear may be successfully managed by rational, authoritative and empathic explanation to define the risk and gain willing acceptance within the context of domiciliary patient care.

WAVE2, N-WASP, and Mena facilitate cell invasion via phosphatidylinositol 3-kinase-dependent local accumulation of actin filaments.

PubMed

Takahashi, Kazuhide; Suzuki, Katsuo

2011-11-01

Cell migration is accomplished by the formation of cellular protrusions such as lamellipodia and filopodia. These protrusions result from actin filament (F-actin) rearrangement at the cell cortex by WASP/WAVE family proteins and Drosophila enabled (Ena)/vasodilator-stimulated factor proteins. However, the role of each of these actin cytoskeletal regulatory proteins in the regulation of three-dimensional cell invasion remains to be clarified. We found that platelet-derived growth factor (PDGF) induces invasion of MDA-MB-231 human breast cancer cells through invasion chamber membrane pores. This invasion was accompanied by intensive F-actin accumulation at the sites of cell infiltration. After PDGF stimulation, WAVE2, N-WASP, and a mammalian Ena (Mena) colocalized with F-actin at the sites of cell infiltration in a phosphatidylinositol 3-kinase (PI3K)-dependent manner. Depletion of WAVE2, N-WASP, or Mena by RNA interference (RNAi) abrogated both cell invasion and intensive F-actin accumulation at the invasion site. These results indicate that by mediating intensive F-actin accumulation at the sites of cell infiltration, WAVE2, N-WASP, and Mena are crucial for PI3K-dependent cell invasion induced by PDGF. Copyright © 2011 Wiley Periodicals, Inc.

Regulatory role of XynR (YagI) in catabolism of xylonate in Escherichia coli K-12.

PubMed

Shimada, Tomohiro; Momiyama, Eri; Yamanaka, Yuki; Watanabe, Hiroki; Yamamoto, Kaneyoshi; Ishihama, Akira

2017-12-01

The genome of Escherichia coli K-12 contains ten cryptic phages, altogether constituting about 3.6% of the genome in sequence. Among more than 200 predicted genes in these cryptic phages, 14 putative transcription factor (TF) genes exist, but their regulatory functions remain unidentified. As an initial attempt to make a breakthrough for understanding the regulatory roles of cryptic phage-encoded TFs, we tried to identify the regulatory function of CP4-6 cryptic prophage-encoded YagI with unknown function. After SELEX screening, YagI was found to bind mainly at a single site within the spacer of bidirectional transcription units, yagA (encoding another uncharacterized TF) and yagEF (encoding 2-keto-3-deoxy gluconate aldolase, and dehydratase, respectively) within this prophage region. YagEF enzymes are involved in the catabolism of xylose downstream from xylonate. We then designated YagI as XynR (regulator of xylonate catabolism), one of the rare single-target TFs. In agreement with this predicted regulatory function, the activity of XynR was suggested to be controlled by xylonate. Even though low-affinity binding sites of XynR were identified in the E. coli K-12 genome, they all were inside open reading frames, implying that the regulation network of XynR is still fixed within the CR4-6 prophage without significant influence over the host E. coli K-12. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Conserved Noncoding Elements in the Most Distant Genera of Cephalochordates: The Goldilocks Principle

PubMed Central

Yue, Jia-Xing; Kozmikova, Iryna; Ono, Hiroki; Nossa, Carlos W.; Kozmik, Zbynek; Putnam, Nicholas H.; Yu, Jr-Kai; Holland, Linda Z.

2016-01-01

Cephalochordates, the sister group of vertebrates + tunicates, are evolving particularly slowly. Therefore, genome comparisons between two congeners of Branchiostoma revealed so many conserved noncoding elements (CNEs), that it was not clear how many are functional regulatory elements. To more effectively identify CNEs with potential regulatory functions, we compared noncoding sequences of genomes of the most phylogenetically distant cephalochordate genera, Asymmetron and Branchiostoma, which diverged approximately 120–160 million years ago. We found 113,070 noncoding elements conserved between the two species, amounting to 3.3% of the genome. The genomic distribution, target gene ontology, and enriched motifs of these CNEs all suggest that many of them are probably cis-regulatory elements. More than 90% of previously verified amphioxus regulatory elements were re-captured in this study. A search of the cephalochordate CNEs around 50 developmental genes in several vertebrate genomes revealed eight CNEs conserved between cephalochordates and vertebrates, indicating sequence conservation over >500 million years of divergence. The function of five CNEs was tested in reporter assays in zebrafish, and one was also tested in amphioxus. All five CNEs proved to be tissue-specific enhancers. Taken together, these findings indicate that even though Branchiostoma and Asymmetron are distantly related, as they are evolving slowly, comparisons between them are likely optimal for identifying most of their tissue-specific cis-regulatory elements laying the foundation for functional characterizations and a better understanding of the evolution of developmental regulation in cephalochordates. PMID:27412606

GWAS4D: multidimensional analysis of context-specific regulatory variant for human complex diseases and traits.

PubMed

Huang, Dandan; Yi, Xianfu; Zhang, Shijie; Zheng, Zhanye; Wang, Panwen; Xuan, Chenghao; Sham, Pak Chung; Wang, Junwen; Li, Mulin Jun

2018-05-16

Genome-wide association studies have generated over thousands of susceptibility loci for many human complex traits, and yet for most of these associations the true causal variants remain unknown. Tissue/cell type-specific prediction and prioritization of non-coding regulatory variants will facilitate the identification of causal variants and underlying pathogenic mechanisms for particular complex diseases and traits. By leveraging recent large-scale functional genomics/epigenomics data, we develop an intuitive web server, GWAS4D (http://mulinlab.tmu.edu.cn/gwas4d or http://mulinlab.org/gwas4d), that systematically evaluates GWAS signals and identifies context-specific regulatory variants. The updated web server includes six major features: (i) updates the regulatory variant prioritization method with our new algorithm; (ii) incorporates 127 tissue/cell type-specific epigenomes data; (iii) integrates motifs of 1480 transcriptional regulators from 13 public resources; (iv) uniformly processes Hi-C data and generates significant interactions at 5 kb resolution across 60 tissues/cell types; (v) adds comprehensive non-coding variant functional annotations; (vi) equips a highly interactive visualization function for SNP-target interaction. Using a GWAS fine-mapped set for 161 coronary artery disease risk loci, we demonstrate that GWAS4D is able to efficiently prioritize disease-causal regulatory variants.

Evolutionary rewiring of bacterial regulatory networks

PubMed Central

Taylor, Tiffany B.; Mulley, Geraldine; McGuffin, Liam J.; Johnson, Louise J.; Brockhurst, Michael A.; Arseneault, Tanya; Silby, Mark W.; Jackson, Robert W.

2015-01-01

Bacteria have evolved complex regulatory networks that enable integration of multiple intracellular and extracellular signals to coordinate responses to environmental changes. However, our knowledge of how regulatory systems function and evolve is still relatively limited. There is often extensive homology between components of different networks, due to past cycles of gene duplication, divergence, and horizontal gene transfer, raising the possibility of cross-talk or redundancy. Consequently, evolutionary resilience is built into gene networks - homology between regulators can potentially allow rapid rescue of lost regulatory function across distant regions of the genome. In our recent study [Taylor, et al. Science (2015), 347(6225)] we find that mutations that facilitate cross-talk between pathways can contribute to gene network evolution, but that such mutations come with severe pleiotropic costs. Arising from this work are a number of questions surrounding how this phenomenon occurs. PMID:28357301

cis-Regulatory control of the initial neurogenic pattern of onecut gene expression in the sea urchin embryo.

PubMed

Barsi, Julius C; Davidson, Eric H

2016-01-01

Specification of the ciliated band (CB) of echinoid embryos executes three spatial functions essential for postgastrular organization. These are establishment of a band about 5 cells wide which delimits and bounds other embryonic territories; definition of a neurogenic domain within this band; and generation within it of arrays of ciliary cells that bear the special long cilia from which the structure derives its name. In Strongylocentrotus purpuratus the spatial coordinates of the future ciliated band are initially and exactly determined by the disposition of a ring of cells that transcriptionally activate the onecut homeodomain regulatory gene, beginning in blastula stage, long before the appearance of the CB per se. Thus the cis-regulatory apparatus that governs onecut expression in the blastula directly reveals the genomic sequence code by which these aspects of the spatial organization of the embryo are initially determined. We screened the entire onecut locus and its flanking region for transcriptionally active cis-regulatory elements, and by means of BAC recombineered deletions identified three separated and required cis-regulatory modules that execute different functions. The operating logic of the crucial spatial control module accounting for the spectacularly precise and beautiful early onecut expression domain depends on spatial repression. Previously predicted oral ectoderm and aboral ectoderm repressors were identified by cis-regulatory mutation as the products of goosecoid and irxa genes respectively, while the pan-ectodermal activator SoxB1 supplies a transcriptional driver function. Copyright © 2015. Published by Elsevier Inc.

Parallel evolution of chordate cis-regulatory code for development.

PubMed

Doglio, Laura; Goode, Debbie K; Pelleri, Maria C; Pauls, Stefan; Frabetti, Flavia; Shimeld, Sebastian M; Vavouri, Tanya; Elgar, Greg

2013-11-01

Urochordates are the closest relatives of vertebrates and at the larval stage, possess a characteristic bilateral chordate body plan. In vertebrates, the genes that orchestrate embryonic patterning are in part regulated by highly conserved non-coding elements (CNEs), yet these elements have not been identified in urochordate genomes. Consequently the evolution of the cis-regulatory code for urochordate development remains largely uncharacterised. Here, we use genome-wide comparisons between C. intestinalis and C. savignyi to identify putative urochordate cis-regulatory sequences. Ciona conserved non-coding elements (ciCNEs) are associated with largely the same key regulatory genes as vertebrate CNEs. Furthermore, some of the tested ciCNEs are able to activate reporter gene expression in both zebrafish and Ciona embryos, in a pattern that at least partially overlaps that of the gene they associate with, despite the absence of sequence identity. We also show that the ability of a ciCNE to up-regulate gene expression in vertebrate embryos can in some cases be localised to short sub-sequences, suggesting that functional cross-talk may be defined by small regions of ancestral regulatory logic, although functional sub-sequences may also be dispersed across the whole element. We conclude that the structure and organisation of cis-regulatory modules is very different between vertebrates and urochordates, reflecting their separate evolutionary histories. However, functional cross-talk still exists because the same repertoire of transcription factors has likely guided their parallel evolution, exploiting similar sets of binding sites but in different combinations.

Finding functional features in Saccharomyces genomes by phylogenetic footprinting.

PubMed

Cliften, Paul; Sudarsanam, Priya; Desikan, Ashwin; Fulton, Lucinda; Fulton, Bob; Majors, John; Waterston, Robert; Cohen, Barak A; Johnston, Mark

2003-07-04

The sifting and winnowing of DNA sequence that occur during evolution cause nonfunctional sequences to diverge, leaving phylogenetic footprints of functional sequence elements in comparisons of genome sequences. We searched for such footprints among the genome sequences of six Saccharomyces species and identified potentially functional sequences. Comparison of these sequences allowed us to revise the catalog of yeast genes and identify sequence motifs that may be targets of transcriptional regulatory proteins. Some of these conserved sequence motifs reside upstream of genes with similar functional annotations or similar expression patterns or those bound by the same transcription factor and are thus good candidates for functional regulatory sequences.

Activation of p21-Dependent G1/G2 Arrest in the Absence of DNA Damage as an Antiapoptotic Response to Metabolic Stress

PubMed Central

Hoeferlin, L. Alexis; Oleinik, Natalia V.; Krupenko, Natalia I.

2011-01-01

The folate enzyme, FDH (10-formyltetrahydrofolate dehydrogenase, ALDH1L1), a metabolic regulator of proliferation, activates p53-dependent G1 arrest and apoptosis in A549 cells. In the present study, we have demonstrated that FDH-induced apoptosis is abrogated upon siRNA knockdown of the p53 downstream target PUMA. Conversely, siRNA knockdown of p21 eliminated FDH-dependent G1 arrest and resulted in an early apoptosis onset. The acceleration of FDH-dependent apoptosis was even more profound in another cell line, HCT116, in which the p21 gene was silenced through homologous recombination (p21−/− cells). In contrast to A549 cells, FDH caused G2 instead of G1 arrest in HCT116 p21+/+ cells; such an arrest was not seen in p21-deficient (HCT116 p21−/−) cells. In agreement with the cell cycle regulatory function of p21, its strong accumulation in nuclei was seen upon FDH expression. Interestingly, our study did not reveal DNA damage upon FDH elevation in either cell line, as judged by comet assay and the evaluation of histone H2AX phosphorylation. In both A549 and HCT116 cell lines, FDH induced a strong decrease in the intracellular ATP pool (2-fold and 30-fold, respectively), an indication of a decrease in de novo purine biosynthesis as we previously reported. The underlying mechanism for the drop in ATP was the strong decrease in intracellular 10-formyltetrahydrofolate, a substrate in two reactions of the de novo purine pathway. Overall, we have demonstrated that p21 can activate G1 or G2 arrest in the absence of DNA damage as a response to metabolite deprivation. In the case of FDH-related metabolic alterations, this response delays apoptosis but is not sufficient to prevent cell death. PMID:22593801

Regulation of PSMB5 Protein and β Subunits of Mammalian Proteasome by Constitutively Activated Signal Transducer and Activator of Transcription 3 (STAT3)

PubMed Central

Vangala, Janakiram Reddy; Dudem, Srikanth; Jain, Nishant; Kalivendi, Shasi V.

2014-01-01

The ubiquitin-proteasome system facilitates the degradation of ubiquitin-tagged proteins and performs a regulatory role in cells. Elevated proteasome activity and subunit expression are found in several cancers. However, the inherent molecular mechanisms responsible for increased proteasome function in cancers remain unclear despite the well investigated and defined role of the mammalian proteasome. This study was initiated to elucidate the mechanisms involved in the regulation of β subunits of the mammalian proteasome. Suppression of STAT3 tyrosine phosphorylation coordinately decreased the mRNA and protein levels of the β subunits of the 20 S core complex in DU145 cells. Notably, PSMB5, a molecular target of bortezomib, was shown to be a target of STAT3. Knockdown of STAT3 decreased PSMB5 protein. Inhibition of phospho-STAT3 substantially reduced PSMB5 protein levels in cells expressing constitutively active-STAT3. Accumulation of activated STAT3 resulted in the induction of PSMB5 promoter and protein levels. In addition, a direct correlation was observed between the endogenous levels of PSMB5 and constitutively active STAT3. PSMB5 and STAT3 protein levels remained unaltered following the inhibition of proteasome activity. The EGF-induced concerted increase of β subunits was blocked by inhibition of the EGF receptor or STAT3 but not by the PI3K/AKT or MEK/ERK pathways. Decreased proteasome activities were due to reduced protein levels of catalytic subunits of the proteasome in STAT3-inhibited cells. Combined treatments with bortezomib and inhibitor of STAT3 abrogated proteasome activity and enhanced cellular apoptosis. Overall, we demonstrate that aberrant activation of STAT3 regulates the expression of β subunits, in particular PSMB5, and the catalytic activity of the proteasome. PMID:24627483

Dengue Virus Subverts Host Innate Immunity by Targeting Adaptor Protein MAVS

PubMed Central

He, Zhenjian; Zhu, Xun; Wen, Weitao; Yuan, Jie; Hu, Yiwen; Chen, Jiahui; An, Shu; Dong, Xinhuai; Lin, Cuiji; Yu, Jianchen; Wu, Jueheng; Yang, Yi; Cai, Junchao; Li, Jun

2016-01-01

ABSTRACT Dengue virus (DENV) is the most common mosquito-borne virus infecting humans and is currently a serious global health challenge. To establish infection in its host cells, DENV must subvert the production and/or antiviral effects of interferon (IFN). The aim of this study was to understand the mechanisms by which DENV suppresses IFN production. We determined that DENV NS4A interacts with mitochondrial antiviral signaling protein (MAVS), which was previously found to activate NF-κB and IFN regulatory factor 3 (IRF3), thus inducing type I IFN in the mitochondrion-associated endoplasmic reticulum membranes (MAMs). We further demonstrated that NS4A is associated with the N-terminal CARD-like (CL) domain and the C-terminal transmembrane (TM) domain of MAVS. This association prevented the binding of MAVS to RIG-I, resulting in the repression of RIG-I-induced IRF3 activation and, consequently, the abrogation of IFN production. Collectively, our findings illustrate a new molecular mechanism by which DENV evades the host immune system and suggest new targets for anti-DENV strategies. IMPORTANCE Type I interferon (IFN) constitutes the first line of host defense against invading viruses. To successfully establish infection, dengue virus (DENV) must counteract either the production or the function of IFN. The mechanism by which DENV suppresses IFN production is poorly understood and characterized. In this study, we demonstrate that the DENV NS4A protein plays an important role in suppressing interferon production through binding MAVS and disrupting the RIG-I–MAVS interaction in mitochondrion-associated endoplasmic reticulum membranes (MAMs). Our study reveals that MAVS is a novel host target of NS4A and provides a molecular mechanism for DENV evasion of the host innate immune response. These findings have important implications for understanding the pathogenesis of DENV and may provide new insights into using NS4A as a therapeutic and/or prevention target. PMID:27252539

Epidermal Growth Factor and Interleukin-1β Utilize Divergent Signaling Pathways to Synergistically Upregulate Cyclooxygenase-2 Gene Expression in Human Amnion-Derived WISH Cells1

PubMed Central

Ackerman, William E.; Rovin, Brad H.; Kniss, Douglas A.

2006-01-01

In human parturition, uterotonic prostaglandins (PGs) arise predominantly via increased expression of cyclooxygenase-2 (COX-2 [also known as prostaglandin synthase 2]) within intra-uterine tissues. Interleukin-1 (IL-1) and epidermal growth factor (EGF), both inducers of COX-2 transcription, are among numerous factors that accumulate within amniotic fluid with advancing gestation. It was previously demonstrated that EGF could potentiate IL-1β-driven PGE2 production in amnion and amnion-derived (WISH) cells. To define the mechanism for this observation, we hypothesized that EGF and IL-1β might exhibit synergism in regulating COX-2 gene expression. In WISH cells, combined treatment with EGF and IL-1β resulted in a greater-than-additive increase in COX-2 mRNA relative to challenge with either agent independently. Augmentation of IL-1β-induced transactivation by EGF was not observed in cells harboring reporter plasmids bearing nuclear factor-kappa B (NFκB) regulatory elements alone, but was evident when a fragment (−891/+9) of the COX-2 gene 5′-promoter was present. Both agents transiently activated intermediates of multiple signaling pathways potentially involved in the regulation of COX-2 gene expression. The 26 S proteasome inhibitor, MG-132, selectively abrogated IL-1β-driven NFκB activation and COX-2 mRNA expression. Only pharmacologic blockade of the p38 mitogen-activated protein kinase eliminated COX-2 expression following EGF stimulation. We conclude that EGF and IL-1β appear to signal through different signaling cascades leading to COX-2 gene expression. IL-1β employs the NFκB pathway predominantly, while the spectrum of EGF signaling is broader and includes p38 kinase. The synergism observed between IL-1β and EGF does not rely on augmented NFκB function, but rather, occurs through differential use of independent response elements within the COX-2 promoter. PMID:15329330

Arctigenin exerts anti-colitis efficacy through inhibiting the differentiation of Th1 and Th17 cells via an mTORC1-dependent pathway.

PubMed

Wu, Xin; Dou, Yannong; Yang, Yan; Bian, Difei; Luo, Jinque; Tong, Bei; Xia, Yufeng; Dai, Yue

2015-08-15

Arctigenin, the main effective constituent of Arctium lappa L. fruit, has previously been proven to dramatically attenuate dextran sulfate sodium (DSS)-induced colitis in mice, a frequently used animal model of inflammatory bowel disease (IBD). As Th1 and Th17 cells play a crucial role in the pathogenesis of IBD, the present study addressed whether and how arctigenin exerted anti-colitis efficacy by interfering with the differentiation and activation of Th1/Th17 cells. In vitro, arctigenin was shown to markedly inhibit the differentiation of Th17 cells from naïve T cells, and moderately inhibit the differentiation of Th1 cells, which was accompanied by lowered phosphorylation of STAT3 and STAT4, respectively. In contrast, arctigenin was lack of marked effect on the differentiation of either Th2 or regulatory T cells. Furthermore, arctigenin was shown to suppress the mammalian target of rapamycin complex 1 (mTORC1) pathway in T cells as demonstrated by down-regulated phosphorylation of the downstream target genes p70S6K and RPS6, and it functioned independent of two well-known upstream kinases PI3K/AKT and ERK. Arctigenin was also able to inhibit the activity of mTORC1 by dissociating raptor from mTOR. Interestingly, the inhibitory effect of arctigenin on T cell differentiation disappeared under a status of mTORC1 overactivation via knockdown of tuberous sclerosis complex 2 (TSC2, a negative regulator of mTORC1) or pretreatment of leucine (an agonist of mTOR). In DSS-induced mice, the inhibition of Th1/Th17 responses and anti-colitis effect of arctigenin were abrogated by leucine treatment. In conclusion, arctigenin ameliorates colitis through down-regulating the differentiation of Th1 and Th17 cells via mTORC1 pathway. Copyright © 2015 Elsevier Inc. All rights reserved.

E3Net: a system for exploring E3-mediated regulatory networks of cellular functions.

PubMed

Han, Youngwoong; Lee, Hodong; Park, Jong C; Yi, Gwan-Su

2012-04-01

Ubiquitin-protein ligase (E3) is a key enzyme targeting specific substrates in diverse cellular processes for ubiquitination and degradation. The existing findings of substrate specificity of E3 are, however, scattered over a number of resources, making it difficult to study them together with an integrative view. Here we present E3Net, a web-based system that provides a comprehensive collection of available E3-substrate specificities and a systematic framework for the analysis of E3-mediated regulatory networks of diverse cellular functions. Currently, E3Net contains 2201 E3s and 4896 substrates in 427 organisms and 1671 E3-substrate specific relations between 493 E3s and 1277 substrates in 42 organisms, extracted mainly from MEDLINE abstracts and UniProt comments with an automatic text mining method and additional manual inspection and partly from high throughput experiment data and public ubiquitination databases. The significant functions and pathways of the extracted E3-specific substrate groups were identified from a functional enrichment analysis with 12 functional category resources for molecular functions, protein families, protein complexes, pathways, cellular processes, cellular localization, and diseases. E3Net includes interactive analysis and navigation tools that make it possible to build an integrative view of E3-substrate networks and their correlated functions with graphical illustrations and summarized descriptions. As a result, E3Net provides a comprehensive resource of E3s, substrates, and their functional implications summarized from the regulatory network structures of E3-specific substrate groups and their correlated functions. This resource will facilitate further in-depth investigation of ubiquitination-dependent regulatory mechanisms. E3Net is freely available online at http://pnet.kaist.ac.kr/e3net.

Long Noncoding RNAs: a New Regulatory Code in Metabolic Control

PubMed Central

Zhao, Xu-Yun; Lin, Jiandie D.

2015-01-01

Long noncoding RNAs (lncRNAs) are emerging as an integral part of the regulatory information encoded in the genome. LncRNAs possess the unique capability to interact with nucleic acids and proteins and exert discrete effects on numerous biological processes. Recent studies have delineated multiple lncRNA pathways that control metabolic tissue development and function. The expansion of the regulatory code that links nutrient and hormonal signals to tissue metabolism gives new insights into the genetic and pathogenic mechanisms underlying metabolic disease. This review discusses lncRNA biology with a focus on its role in the development, signaling, and function of key metabolic tissues. PMID:26410599

78 FR 35650 - Self-Regulatory Organizations; Chicago Board Options Exchange, Incorporated; Notice of Filing and...

Federal Register 2010, 2011, 2012, 2013, 2014

2013-06-13

... this new AIM feature will function as described in rule filing SR-CBOE-2013- 048. B. Self-Regulatory... the necessary system work and perform sufficient testing to ensure that this new AIM feature functions... Effectiveness of a Proposed Rule Change Relating to Rule 6.74A June 7, 2013. Pursuant to Section 19(b)(1) of the...

Role of the glucocorticoid-induced TNFR-related protein (GITR)-GITR ligand pathway in innate and adaptive immunity.

PubMed

Azuma, Miyuki

2010-01-01

Glucocorticoid-induced TNF receptor-related protein (GITR) is expressed in regulatory T cells at high levels, but is also inducible in conventional effector T cells after activation. Initial studies using an agonistic anti- GITR mAb mislead this line of research with respect to the contribution of GITR stimulation on the function of regulatory T cells. In fact, GITR acts as a costimulatory receptor for both effector and regulatory T cells by enhancing effector and regulatory functions, respectively. Unlike other costimulatory ligands, GITR ligand (GITRL) expression on mature myeloid dendritic cells (DCs) is extremely limited and the GITR-GITRL pathway does not contribute markedly to direct interactions with T cells and antigen-presenting cells in the secondary lymphoid tissues. Rather, GITRL is constitutively expressed on parenchymal tissue cells and interacts with GITR expressed on tissue-infiltrating macrophages and DCs, or effector and regulatory T cells. Interactions with GITR and GITRL at local inflammatory sites induce site-specific production of cytokines and chemokines, resulting in control activation of tissue-infiltrating effector or regulatory cells and their migration. This review summarizes recent reports on the GITR-GITRL pathway, which controls both innate and adaptive immune responses.

Isolation of (S)-(+)-naproxene from Musa acuminata. Inhibitory effect of naproxene and its 7-methoxy isomer on constitutive COX-1 and inducible COX-2.

PubMed

Abad, T; McNaughton-Smith, G; Fletcher, W Q; Echeverri, F; Diaz-Peñate, R; Tabraue, C; Ruiz de Galarreta, C M; López-Blanco, F; Luis, J G

2000-06-01

The isolation and characterisation of (S)-(+)-6-methoxy-alpha-methyl-2-naphthaleneacetic acid, a well known synthetic non-steroidal anti-inflammatory drug (naproxene), from a natural source is described for the first time. We evaluated the ability of naproxene and its 7-methoxy isomer to abrogate constitutive COX-1 and inducible COX-2 activity in human A549 cells. Naproxene inhibited COX-1 (IC50 = 3.42 microM) and COX-2 (IC50 = 1.53 microM), whereas the 7-methoxy isomer had no appreciable effect on COX-1 (IC50 > 100 microM) but also abrogated the activity of COX-2 enzyme (IC50 = 14.42 microM).

Regulatory Mutants at the his1 Locus of Yeast

PubMed Central

Lax, Carol; Fogel, Seymour; Cramer, Carole

1979-01-01

The his1 gene in Saccharomyces cerevisiae codes for phosphoribosyl transferase, an allosteric enzyme that catalyzes the initial step in histidine biosynthesis. Mutants that specifically alter the feedback regulatory function were isolated by selecting his1 prototrophic revertants that overproduce and excrete histidine. The prototrophs were obtained from diploids homoallelic for his1–7 and heterozygous for the flanking markers thr3 and arg6. Among six independently derived mutant isolates, three distinct levels of histidine excretion were detected. The mutants were shown to be second-site alterations mapping at the his1 locus by recovery of the original auoxtrophic parental alleles. The double mutants, HIS1–7e, are dominant with respect to catalytic function but recessive in regulatory function. When removed from this his1–7 background, the mutant regulatory site (HIS1–e) still confers prototrophy but not histidine excretion. To yield the excretion phenotype, the primary and altered secondary sites are required in cis array. Differences in histidine excretion levels correlate with resistance to the histidine analogue, triazoalanine. PMID:385447

Shared Enhancer Activity in the Limbs and Phallus and Functional Divergence of a Limb-Genital cis-Regulatory Element in Snakes.

PubMed

Infante, Carlos R; Mihala, Alexandra G; Park, Sungdae; Wang, Jialiang S; Johnson, Kenji K; Lauderdale, James D; Menke, Douglas B

2015-10-12

The amniote phallus and limbs differ dramatically in their morphologies but share patterns of signaling and gene expression in early development. Thus far, the extent to which genital and limb transcriptional networks also share cis-regulatory elements has remained unexplored. We show that many limb enhancers are retained in snake genomes, suggesting that these elements may function in non-limb tissues. Consistent with this, our analysis of cis-regulatory activity in mice and Anolis lizards reveals that patterns of enhancer activity in embryonic limbs and genitalia overlap heavily. In mice, deletion of HLEB, an enhancer of Tbx4, produces defects in hindlimbs and genitalia, establishing the importance of this limb-genital enhancer for development of these different appendages. Further analyses demonstrate that the HLEB of snakes has lost hindlimb enhancer function while retaining genital activity. Our findings identify roles for Tbx4 in genital development and highlight deep similarities in cis-regulatory activity between limbs and genitalia. Copyright © 2015 Elsevier Inc. All rights reserved.

Regulatory Snapshots: integrative mining of regulatory modules from expression time series and regulatory networks.

PubMed

Gonçalves, Joana P; Aires, Ricardo S; Francisco, Alexandre P; Madeira, Sara C

2012-01-01

Explaining regulatory mechanisms is crucial to understand complex cellular responses leading to system perturbations. Some strategies reverse engineer regulatory interactions from experimental data, while others identify functional regulatory units (modules) under the assumption that biological systems yield a modular organization. Most modular studies focus on network structure and static properties, ignoring that gene regulation is largely driven by stimulus-response behavior. Expression time series are key to gain insight into dynamics, but have been insufficiently explored by current methods, which often (1) apply generic algorithms unsuited for expression analysis over time, due to inability to maintain the chronology of events or incorporate time dependency; (2) ignore local patterns, abundant in most interesting cases of transcriptional activity; (3) neglect physical binding or lack automatic association of regulators, focusing mainly on expression patterns; or (4) limit the discovery to a predefined number of modules. We propose Regulatory Snapshots, an integrative mining approach to identify regulatory modules over time by combining transcriptional control with response, while overcoming the above challenges. Temporal biclustering is first used to reveal transcriptional modules composed of genes showing coherent expression profiles over time. Personalized ranking is then applied to prioritize prominent regulators targeting the modules at each time point using a network of documented regulatory associations and the expression data. Custom graphics are finally depicted to expose the regulatory activity in a module at consecutive time points (snapshots). Regulatory Snapshots successfully unraveled modules underlying yeast response to heat shock and human epithelial-to-mesenchymal transition, based on regulations documented in the YEASTRACT and JASPAR databases, respectively, and available expression data. Regulatory players involved in functionally enriched processes related to these biological events were identified. Ranking scores further suggested ability to discern the primary role of a gene (target or regulator). Prototype is available at: http://kdbio.inesc-id.pt/software/regulatorysnapshots.

Regulatory Snapshots: Integrative Mining of Regulatory Modules from Expression Time Series and Regulatory Networks

PubMed Central

Gonçalves, Joana P.; Aires, Ricardo S.; Francisco, Alexandre P.; Madeira, Sara C.

2012-01-01

Explaining regulatory mechanisms is crucial to understand complex cellular responses leading to system perturbations. Some strategies reverse engineer regulatory interactions from experimental data, while others identify functional regulatory units (modules) under the assumption that biological systems yield a modular organization. Most modular studies focus on network structure and static properties, ignoring that gene regulation is largely driven by stimulus-response behavior. Expression time series are key to gain insight into dynamics, but have been insufficiently explored by current methods, which often (1) apply generic algorithms unsuited for expression analysis over time, due to inability to maintain the chronology of events or incorporate time dependency; (2) ignore local patterns, abundant in most interesting cases of transcriptional activity; (3) neglect physical binding or lack automatic association of regulators, focusing mainly on expression patterns; or (4) limit the discovery to a predefined number of modules. We propose Regulatory Snapshots, an integrative mining approach to identify regulatory modules over time by combining transcriptional control with response, while overcoming the above challenges. Temporal biclustering is first used to reveal transcriptional modules composed of genes showing coherent expression profiles over time. Personalized ranking is then applied to prioritize prominent regulators targeting the modules at each time point using a network of documented regulatory associations and the expression data. Custom graphics are finally depicted to expose the regulatory activity in a module at consecutive time points (snapshots). Regulatory Snapshots successfully unraveled modules underlying yeast response to heat shock and human epithelial-to-mesenchymal transition, based on regulations documented in the YEASTRACT and JASPAR databases, respectively, and available expression data. Regulatory players involved in functionally enriched processes related to these biological events were identified. Ranking scores further suggested ability to discern the primary role of a gene (target or regulator). Prototype is available at: http://kdbio.inesc-id.pt/software/regulatorysnapshots. PMID:22563474

Sphingosine kinase-1 mediates androgen-induced osteoblast cell growth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Martin, Claire; Universite de Toulouse, UPS, IPBS, Toulouse F-31000; Lafosse, Jean-Michel

Herein we report that the lipid kinase sphingosine kinase-1 (SphK1) is instrumental in mediating androgen-induced cell proliferation in osteoblasts. Dihydrotestosterone (DHT) triggered cell growth in steroid-deprived MC3T3 cells, which was associated with a rapid stimulation of SphK1 and activation of both Akt and ERK signaling pathways. This mechanism relied on functional androgen receptor/PI3K/Akt nongenotropic signaling as pharmacological antagonists could block SphK1 stimulation by DHT and its consequences. Finally, SphK1 inhibition not only abrogated DHT-induced ERK activation but also blocked cell proliferation, while ERK inhibition had no impact, suggesting that SphK1 was critical for DHT signaling yet independently of the ERK.

Genetic evidence for conserved non-coding element function across species–the ears have it

PubMed Central

Turner, Eric E.; Cox, Timothy C.

2014-01-01

Comparison of genomic sequences from diverse vertebrate species has revealed numerous highly conserved regions that do not appear to encode proteins or functional RNAs. Often these “conserved non-coding elements,” or CNEs, can direct gene expression to specific tissues in transgenic models, demonstrating they have regulatory function. CNEs are frequently found near “developmental” genes, particularly transcription factors, implying that these elements have essential regulatory roles in development. However, actual examples demonstrating CNE regulatory functions across species have been few, and recent loss-of-function studies of several CNEs in mice have shown relatively minor effects. In this Perspectives article, we discuss new findings in “fancy” rats and Highland cattle demonstrating that function of a CNE near the Hmx1 gene is crucial for normal external ear development and when disrupted can mimic loss-of function Hmx1 coding mutations in mice and humans. These findings provide important support for conserved developmental roles of CNEs in divergent species, and reinforce the concept that CNEs should be examined systematically in the ongoing search for genetic causes of human developmental disorders in the era of genome-scale sequencing. PMID:24478720

Regulatory B-cell induction by helminths: implications for allergic disease.

PubMed

Hussaarts, Leonie; van der Vlugt, Luciën E P M; Yazdanbakhsh, Maria; Smits, Hermelijn H

2011-10-01

Chronic helminth infections are often associated with a reduced prevalence of inflammatory disorders, including allergic diseases. Helminths influence the host immune system by downregulating T-cell responses; the cytokine IL-10 appears to play a central role in this process. Over the last decade, evidence has emerged toward a new regulatory cell type: IL-10-producing B cells, capable of regulating immunity and therefore termed regulatory B cells. Initially, regulatory B cells have been described in autoimmunity models where they dampen inflammation, but recently they were also found in several helminth infection models. Importantly, regulatory B cells have recently been identified in humans, and it has been suggested that patients suffering from autoimmunity have an impaired regulatory B-cell function. As such, it is of therapeutic interest to study the conditions in which IL-10-producing B cells can be induced. Chronic helminth infections appear to hold promise in this context as emerging evidence suggests that helminth-induced regulatory B cells strongly suppress allergic inflammation. In this review, we will discuss the conditions under which regulatory B cells are present, leading to a state of tolerance, as well as the conditions where their absence or functional impairment leads to exacerbated disease. We will summarize their phenotypic characteristics and their mechanisms of action and elaborate on possible mechanisms whereby regulatory B cells can be induced or expanded, as this may open novel avenues for the treatment of inflammatory diseases, such as allergic asthma. Copyright © 2011 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

10 CFR 51.68 - Environmental report-rulemaking.

Code of Federal Regulations, 2010 CFR

2010-01-01

... Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing... special nuclear material shall submit with the petition a separate document entitled “Petitioner's...

Subnuclear organization and trafficking of regulatory proteins: implications for biological control and cancer.

PubMed

Stein, G S; van Wijnen, A J; Stein, J L; Lian, J B; Montecino, M; Zaidi, K; Javed, A

2000-01-01

The regulated and regulatory components that interrelate nuclear structure and function must be experimentally established. A formidable challenge is to define further the control of transcription factor targeting to acceptor sites associated with the nuclear matrix. It will be important to determine whether acceptor proteins are associated with a pre-existing core-filament structural lattice or whether a compositely organized scaffold of regulatory factors is dynamically assembled. An inclusive model for all steps in the targeting of proteins to subnuclear sites cannot yet be proposed. However, this model must account for the apparent diversity of intranuclear targeting signals. It is also important to assess the extent to which regulatory discrimination is mediated by subnuclear domain-specific trafficking signals. Furthermore, the checkpoints that monitor subnuclear distribution of regulatory factors and the sorting steps that ensure both structural and functional fidelity of nuclear domains in which replication and expression of genes occur must be biochemically and mechanistically defined. There is emerging recognition that placement of regulatory components of gene expression must be temporally and spatially coordinated to facilitate biological control. The consequences of breaches in nuclear structure-function relationships are observed in an expanding series of diseases that include cancer [Weis et al., 1994; Rogaia et al., 1997; Yano et al., 1997; Rowley, 1998; Zeng et al., 1998; McNeil et al., 1999; Tao and Levine, 1999a] and neurological disorders [Skinner et al., 1997]. As the repertoire of architecture-associated regulatory factors and cofactors expands, workers in the field are becoming increasingly confident that nuclear organization contributes significantly to control of transcription. To gain increased appreciation for the complexities of subnuclear organization and gene regulation, we must continue to characterize mechanisms that direct regulatory proteins to specific transcription sites within the nucleus so that these proteins are in the right place at the right time. J. Cell. Biochem. Suppl. 35:84-92, 2000. Copyright 2001 Wiley-Liss, Inc.

Functions of MicroRNAs in Cardiovascular Biology and Disease

PubMed Central

Hata, Akiko

2015-01-01

In 1993, lin-4 was discovered as a critical modulator of temporal development in Caenorhabditis elegans and, most notably, as the first in the class of small, single-stranded noncoding RNAs now defined as microRNAs (miRNAs). Another eight years elapsed before miRNA expression was detected in mammalian cells. Since then, explosive advancements in the field of miRNA biology have elucidated the basic mechanism of miRNA biogenesis, regulation, and gene-regulatory function. The discovery of this new class of small RNAs has augmented the complexity of gene-regulatory programs as well as the understanding of developmental and pathological processes in the cardiovascular system. Indeed, the contributions of miRNAs in cardiovascular development and function have been widely explored, revealing the extensive role of these small regulatory RNAs in cardiovascular physiology. PMID:23157557

The impact of transposable elements on mammalian development

PubMed Central

Garcia-Perez, Jose L.; Widmann, Thomas J.; Adams, Ian R.

2018-01-01

Summary Despite often being classified as selfish or junk DNA, transposable elements (TEs) are a group of abundant genetic sequences that significantly impact on mammalian development and genome regulation. In recent years, our understanding of how pre-existing TEs affect genome architecture, gene regulatory networks and protein function during mammalian embryogenesis has dramatically expanded. In addition, the mobilization of active TEs in selected cell types has been shown to generate genetic variation during development and in fully differentiated tissues. Importantly, the ongoing domestication and evolution of TEs appears to provide a rich source of regulatory elements, functional modules and genetic variation that fuels the evolution of mammalian developmental processes. Here, we review the functional impact that TEs exert on mammalian developmental processes and how the somatic activity of TEs can influence gene regulatory networks. PMID:27875251

A Sphingosine-1 Phosphate agonist (FTY720) limits trauma/hemorrhagic shock induced multiple organ dysfunction syndrome

PubMed Central

Bonitz, Joyce A.; Son, Julie Y.; Chandler, Benjamin; Tomaio, Jacquelyn N.; Qin, Yong; Prescott, Lauriston M.; Feketeova, Eleonora; Deitch, Edwin A.

2014-01-01

BACKGROUND Trauma/hemorrhagic shock is one of the major consequences of battlefield injury as well as civilian trauma. FTY720 (sphingosine-1 phosphate agonist) has the capability to decrease the activity of the innate and adaptive immune systems and, at the same time, maintain endothelial cell barrier function and vascular homeostasis during stress. For this reason, we hypothesize that FTY720, as part of resuscitation therapy, would limit T/HS induced multiple organ dysfunction syndrome (MODS) in a rodent trauma-hemorrhagic shock (T/HS) model. METHODS Rats subjected to trauma/sham-shock (T/SS) or T/HS (30 mm Hg × 90 min), were administered FTY720 (1 mg/kg) post-T/HS during volume resuscitation. Lung injury (permeability to Evans Blue dye), PMN priming (respiratory burst activity), and RBC rigidity were measured. In addition, lymph duct cannulated rats were used to quantify the effect of FTY720 on gut injury (permeability and morphology) and the biologic activity of T/HS vs. T/SS lymph on PMN-RB and RBC deformability. RESULTS T/HS-induced increased lung permeability, PMN priming and RBC rigidity were all abrogated by FTY720. The systemic protective effect of FTY720 was only partially at the gut level, since FTY720 did not prevent T/HS-induced gut injury (morphology or permeability,) however, it did abrogate T/HS lymph-induced increased respiratory burst and RBC rigidity. CONCLUSION FTY720 limited T/HS-induced MODS (lung injury, red cell injury, and neutrophil priming) as well as T/HS lymph bioactivity, although it did not limit gut injury. PMID:25004059

Amino Acid Substitutions of Coiled-Coil Protein Tpr Abrogate Anchorage to the Nuclear Pore Complex but Not Parallel, In-Register Homodimerization

PubMed Central

Hase, Manuela E.; Kuznetsov, Nikolai V.; Cordes, Volker C.

2001-01-01

Tpr is a protein component of nuclear pore complex (NPC)-attached intranuclear filaments. Secondary structure predictions suggest a bipartite structure, with a large N-terminal domain dominated by heptad repeats (HRs) typical for coiled-coil–forming proteins. Proposed functions for Tpr have included roles as a homo- or heteropolymeric architectural element of the nuclear interior. To gain insight into Tpr's ultrastructural properties, we have studied recombinant Tpr segments by circular dichroism spectroscopy, chemical cross-linking, and rotary shadowing electron microscopy. We show that polypeptides of the N-terminal domain homodimerize in vitro and represent α-helical molecules of extended rod-like shape. With the use of a yeast two-hybrid approach, arrangement of the coiled-coil is found to be in parallel and in register. To clarify whether Tpr can self-assemble further into homopolymeric filaments, the full-length protein and deletion mutants were overexpressed in human cells and then analyzed by confocal immunofluorescence microscopy, cell fractionation, and immuno-electron microscopy. Surplus Tpr, which does not bind to the NPC, remains in a soluble state of ∼7.5 S and occasionally forms aggregates of entangled molecules but neither self-assembles into extended linear filaments nor stably binds to other intranuclear structures. Binding to the NPC is shown to depend on the integrity of individual HRs; amino acid substitutions within these HRs abrogate NPC binding and render the protein soluble but do not abolish Tpr's general ability to homodimerize. Possible contributions of Tpr to the structural organization of the nuclear periphery in somatic cells are discussed. PMID:11514627

Smad4 Deficiency in Smooth Muscle Cells Initiates the Formation of Aortic Aneurysm.

PubMed

Zhang, Peng; Hou, Siyuan; Chen, Jicheng; Zhang, Jishuai; Lin, Fuyu; Ju, Renjie; Cheng, Xuan; Ma, Xiaowei; Song, Yao; Zhang, Youyi; Zhu, Minsheng; Du, Jie; Lan, Yu; Yang, Xiao

2016-02-05

Aortic aneurysm is a life-threatening cardiovascular disorder caused by the predisposition for dissection and rupture. Genetic studies have proved the involvement of the transforming growth factor-β (TGF-β) pathway in aortic aneurysm. Smad4 is the central mediator of the canonical TGF-β signaling pathway. However, the exact role of Smad4 in smooth muscle cells (SMCs) leading to the pathogenesis of aortic aneurysms is largely unknown. To determine the role of smooth muscle Smad4 in the pathogenesis of aortic aneurysms. Conditional gene knockout strategy combined with histology and expression analysis showed that Smad4 or TGF-β receptor type II deficiency in SMCs led to the occurrence of aortic aneurysms along with an upregulation of cathepsin S and matrix metallopeptidase-12, which are proteases essential for elastin degradation. We further demonstrated a previously unknown downregulation of matrix metallopeptidase-12 by TGF-β in the aortic SMCs, which is largely abrogated in the absence of Smad4. Chemotactic assay and pharmacologic treatment demonstrated that Smad4-deficient SMCs directly triggered aortic wall inflammation via the excessive production of chemokines to recruit macrophages. Monocyte/macrophage depletion or blocking selective chemokine axis largely abrogated the progression of aortic aneurysm caused by Smad4 deficiency in SMCs. The findings reveal that Smad4-dependent TGF-β signaling in SMCs protects against aortic aneurysm formation and dissection. The data also suggest important implications for novel therapeutic strategies to limit the progression of the aneurysm resulting from TGF-β signaling loss-of-function mutations. © 2015 American Heart Association, Inc.

JPRS Report, Science & Technology, USSR: Life Sciences

DTIC Science & Technology

1989-02-10

29 Effects of Regulatory Peptides on Recovery of Visual Functions in Retinitis Pigmentosa [N. B. Kostelyanets, O. B. Ilyinskiy, et al; FIZIOLOGIYA...Western. UDC 612.812 Effects of Regulatory Peptides on Recovery of Visual Functions in Retinitis Pigmentosa 18400030a Moscow FIZIOLOGIYA CHELOVEKA in...et al.; BIOORGANICHESKAYA KHIMIYA, Vol 14 No 3, Mar 88] .. 3 Synthesis and Properties of C13-Dependent Retinals [S. V. Yeremin, B. I. Mitsner, et

Identification of herpesvirus proteins that contribute to G1/S arrest.

PubMed

Paladino, Patrick; Marcon, Edyta; Greenblatt, Jack; Frappier, Lori

2014-04-01

Lytic infection by herpesviruses induces cell cycle arrest at the G1/S transition. This appears to be a function of multiple herpesvirus proteins, but only a minority of herpesvirus proteins have been examined for cell cycle effects. To gain a more comprehensive understanding of the viral proteins that contribute to G1/S arrest, we screened a library of over 200 proteins from herpes simplex virus type 1, human cytomegalovirus, and Epstein-Barr virus (EBV) for effects on the G1/S interface, using HeLa fluorescent, ubiquitination-based cell cycle indicator (Fucci) cells in which G1/S can be detected colorimetrically. Proteins from each virus were identified that induce accumulation of G1/S cells, predominantly tegument, early, and capsid proteins. The identification of several capsid proteins in this screen suggests that incoming viral capsids may function to modulate cellular processes. The cell cycle effects of selected EBV proteins were further verified and examined for effects on p53 and p21 as regulators of the G1/S transition. Two EBV replication proteins (BORF2 and BMRF1) were found to induce p53 but not p21, while a previously uncharacterized tegument protein (BGLF2) was found to induce p21 protein levels in a p53-independent manner. Proteomic analyses of BGLF2-interacting proteins identified interactions with the NIMA-related protein kinase (NEK9) and GEM-interacting protein (GMIP). Silencing of either NEK9 or GMIP induced p21 without affecting p53 and abrogated the ability of BGLF2 to further induce p21. Collectively, these results suggest multiple viral proteins contribute to G1/S arrest, including BGLF2, which induces p21 levels likely by interfering with the functions of NEK9 and GMIP. Most people are infected with multiple herpesviruses, whose proteins alter the infected cells in several ways. During lytic infection, the viral proteins block cell proliferation just before the cellular DNA replicates. We used a novel screening method to identify proteins from three different herpesviruses that contribute to this block. Several of the proteins we identified had previously unknown functions or were structural components of the virion. Subsets of these proteins from Epstein-Barr virus were studied for their effects on the cell cycle regulatory proteins p53 and p21, thereby identifying two proteins that induce p53 and one that induces p21 (BGLF2). We identified interactions of BGLF2 with two human proteins, both of which regulate p21, suggesting that BGLF2 induces p21 by interfering with the functions of these two host proteins. Our study indicates that multiple herpesvirus proteins contribute to the cell proliferation block, including components of the incoming virions.

Modularity and design principles in the sea urchin embryo gene regulatory network

PubMed Central

Peter, Isabelle S.; Davidson, Eric H.

2010-01-01

The gene regulatory network (GRN) established experimentally for the pre-gastrular sea urchin embryo provides causal explanations of the biological functions required for spatial specification of embryonic regulatory states. Here we focus on the structure of the GRN which controls the progressive increase in complexity of territorial regulatory states during embryogenesis; and on the types of modular subcircuits of which the GRN is composed. Each of these subcircuit topologies executes a particular operation of spatial information processing. The GRN architecture reflects the particular mode of embryogenesis represented by sea urchin development. Network structure not only specifies the linkages constituting the genomic regulatory code for development, but also indicates the various regulatory requirements of regional developmental processes. PMID:19932099

Individual differences in regulatory focus predict neural response to reward.

PubMed

Scult, Matthew A; Knodt, Annchen R; Hanson, Jamie L; Ryoo, Minyoung; Adcock, R Alison; Hariri, Ahmad R; Strauman, Timothy J

2017-08-01

Although goal pursuit is related to both functioning of the brain's reward circuits and psychological factors, the literatures surrounding these concepts have often been separate. Here, we use the psychological construct of regulatory focus to investigate individual differences in neural response to reward. Regulatory focus theory proposes two motivational orientations for personal goal pursuit: (1) promotion, associated with sensitivity to potential gain, and (2) prevention, associated with sensitivity to potential loss. The monetary incentive delay task was used to manipulate reward circuit function, along with instructional framing corresponding to promotion and prevention in a within-subject design. We observed that the more promotion oriented an individual was, the lower their ventral striatum response to gain cues. Follow-up analyses revealed that greater promotion orientation was associated with decreased ventral striatum response even to no-value cues, suggesting that promotion orientation may be associated with relatively hypoactive reward system function. The findings are also likely to represent an interaction between the cognitive and motivational characteristics of the promotion system with the task demands. Prevention orientation did not correlate with ventral striatum response to gain cues, supporting the discriminant validity of regulatory focus theory. The results highlight a dynamic association between individual differences in self-regulation and reward system function.

Coordination of meristem and boundary functions by transcription factors in the SHOOT MERISTEMLESS regulatory network.

PubMed

Scofield, Simon; Murison, Alexander; Jones, Angharad; Fozard, John; Aida, Mitsuhiro; Band, Leah R; Bennett, Malcolm; Murray, James A H

2018-04-30

The Arabidopsis homeodomain transcription factor SHOOT MERISTEMLESS (STM) is crucial for shoot apical meristem (SAM) function, yet the components and structure of the STM gene regulatory network (GRN) are largely unknown. Here, we show that transcriptional regulators are overrepresented among STM-regulated genes and, using these as GRN components in Bayesian network analysis, we infer STM GRN associations and reveal regulatory relationships between STM and factors involved in multiple aspects of SAM function. These include hormone regulation, TCP-mediated control of cell differentiation, AIL/PLT-mediated regulation of pluripotency and phyllotaxis, and specification of meristem-organ boundary zones via CUC1. We demonstrate a direct positive transcriptional feedback loop between STM and CUC1, despite their distinct expression patterns in the meristem and organ boundary, respectively. Our further finding that STM activates expression of the CUC1-targeting microRNA miR164c combined with mathematical modelling provides a potential solution for this apparent contradiction, demonstrating that these proposed regulatory interactions coupled with STM mobility could be sufficient to provide a mechanism for CUC1 localisation at the meristem-organ boundary. Our findings highlight the central role for the STM GRN in coordinating SAM functions. © 2018. Published by The Company of Biologists Ltd.

Directed evolution of a synthetic phylogeny of programmable Trp repressors.

PubMed

Ellefson, Jared W; Ledbetter, Michael P; Ellington, Andrew D

2018-04-01

As synthetic regulatory programs expand in sophistication, an ever increasing number of biological components with predictable phenotypes is required. Regulators are often 'part mined' from a diverse, but uncharacterized, array of genomic sequences, often leading to idiosyncratic behavior. Here, we generate an entire synthetic phylogeny from the canonical allosteric transcription factor TrpR. Iterative rounds of positive and negative compartmentalized partnered replication (CPR) led to the exponential amplification of variants that responded with high affinity and specificity to halogenated tryptophan analogs and novel operator sites. Fourteen repressor variants were evolved with unique regulatory profiles across five operators and three ligands. The logic of individual repressors can be modularly programmed by creating heterodimeric fusions, resulting in single proteins that display logic functions, such as 'NAND'. Despite the evolutionarily limited regulatory role of TrpR, vast functional spaces exist around this highly conserved protein scaffold and can be harnessed to create synthetic regulatory programs.

Innate immune activity conditions the effect of regulatory variants upon monocyte gene expression.

PubMed

Fairfax, Benjamin P; Humburg, Peter; Makino, Seiko; Naranbhai, Vivek; Wong, Daniel; Lau, Evelyn; Jostins, Luke; Plant, Katharine; Andrews, Robert; McGee, Chris; Knight, Julian C

2014-03-07

To systematically investigate the impact of immune stimulation upon regulatory variant activity, we exposed primary monocytes from 432 healthy Europeans to interferon-γ (IFN-γ) or differing durations of lipopolysaccharide and mapped expression quantitative trait loci (eQTLs). More than half of cis-eQTLs identified, involving hundreds of genes and associated pathways, are detected specifically in stimulated monocytes. Induced innate immune activity reveals multiple master regulatory trans-eQTLs including the major histocompatibility complex (MHC), coding variants altering enzyme and receptor function, an IFN-β cytokine network showing temporal specificity, and an interferon regulatory factor 2 (IRF2) transcription factor-modulated network. Induced eQTL are significantly enriched for genome-wide association study loci, identifying context-specific associations to putative causal genes including CARD9, ATM, and IRF8. Thus, applying pathophysiologically relevant immune stimuli assists resolution of functional genetic variants.

Regulatory T cells: present facts and future hopes.

PubMed

Becker, Christian; Stoll, Sabine; Bopp, Tobias; Schmitt, Edgar; Jonuleit, Helmut

2006-09-01

Naturally occurring CD4(+)CD25(+)Foxp3(+) regulatory T cells and several subsets of induced suppressor T cells are key players of the immune tolerance network and control the induction and effector phase of our immunological defense system. These T cell populations actively control the properties of other immune cells by suppressing their functional activity to prevent autoimmunity and transplant rejection but also influence the immune response to allergens as well as against tumor cells and pathogens. Even though we are far from completely understanding the molecular and cellular mechanisms that manage the different regulatory T cell populations, increasing evidence exists about their functional importance. The knowledge on their induction and activation opens the possibility for their selective manipulation in vivo as an attractive approach for an immunotherapy of unwanted immune responses. This review summarizes this knowledge and discusses the potential of regulatory T cells for novel immunointervention strategies in the future.

A transcription factor collective defines the HSN serotonergic neuron regulatory landscape

PubMed Central

Artacho, Alejandro; Jimeno-Martín, Ángela; Chirivella, Laura; Weinberg, Peter

2018-01-01

Cell differentiation is controlled by individual transcription factors (TFs) that together activate a selection of enhancers in specific cell types. How these combinations of TFs identify and activate their target sequences remains poorly understood. Here, we identify the cis-regulatory transcriptional code that controls the differentiation of serotonergic HSN neurons in Caenorhabditis elegans. Activation of the HSN transcriptome is directly orchestrated by a collective of six TFs. Binding site clusters for this TF collective form a regulatory signature that is sufficient for de novo identification of HSN neuron functional enhancers. Among C. elegans neurons, the HSN transcriptome most closely resembles that of mouse serotonergic neurons. Mouse orthologs of the HSN TF collective also regulate serotonergic differentiation and can functionally substitute for their worm counterparts which suggests deep homology. Our results identify rules governing the regulatory landscape of a critically important neuronal type in two species separated by over 700 million years. PMID:29553368

Sheep genome functional annotation reveals proximal regulatory elements contributed to the evolution of modern breeds.

PubMed

Naval-Sanchez, Marina; Nguyen, Quan; McWilliam, Sean; Porto-Neto, Laercio R; Tellam, Ross; Vuocolo, Tony; Reverter, Antonio; Perez-Enciso, Miguel; Brauning, Rudiger; Clarke, Shannon; McCulloch, Alan; Zamani, Wahid; Naderi, Saeid; Rezaei, Hamid Reza; Pompanon, Francois; Taberlet, Pierre; Worley, Kim C; Gibbs, Richard A; Muzny, Donna M; Jhangiani, Shalini N; Cockett, Noelle; Daetwyler, Hans; Kijas, James

2018-02-28

Domestication fundamentally reshaped animal morphology, physiology and behaviour, offering the opportunity to investigate the molecular processes driving evolutionary change. Here we assess sheep domestication and artificial selection by comparing genome sequence from 43 modern breeds (Ovis aries) and their Asian mouflon ancestor (O. orientalis) to identify selection sweeps. Next, we provide a comparative functional annotation of the sheep genome, validated using experimental ChIP-Seq of sheep tissue. Using these annotations, we evaluate the impact of selection and domestication on regulatory sequences and find that sweeps are significantly enriched for protein coding genes, proximal regulatory elements of genes and genome features associated with active transcription. Finally, we find individual sites displaying strong allele frequency divergence are enriched for the same regulatory features. Our data demonstrate that remodelling of gene expression is likely to have been one of the evolutionary forces that drove phenotypic diversification of this common livestock species.

I-motif DNA structures are formed in the nuclei of human cells

NASA Astrophysics Data System (ADS)

Zeraati, Mahdi; Langley, David B.; Schofield, Peter; Moye, Aaron L.; Rouet, Romain; Hughes, William E.; Bryan, Tracy M.; Dinger, Marcel E.; Christ, Daniel

2018-06-01

Human genome function is underpinned by the primary storage of genetic information in canonical B-form DNA, with a second layer of DNA structure providing regulatory control. I-motif structures are thought to form in cytosine-rich regions of the genome and to have regulatory functions; however, in vivo evidence for the existence of such structures has so far remained elusive. Here we report the generation and characterization of an antibody fragment (iMab) that recognizes i-motif structures with high selectivity and affinity, enabling the detection of i-motifs in the nuclei of human cells. We demonstrate that the in vivo formation of such structures is cell-cycle and pH dependent. Furthermore, we provide evidence that i-motif structures are formed in regulatory regions of the human genome, including promoters and telomeric regions. Our results support the notion that i-motif structures provide key regulatory roles in the genome.

Investigation of the multifunctional gene AOP3 expands the regulatory network fine-tuning glucosinolate production in Arabidopsis

PubMed Central

Jensen, Lea M.; Kliebenstein, Daniel J.; Burow, Meike

2015-01-01

Quantitative trait loci (QTL) mapping studies enable identification of loci that are part of regulatory networks controlling various phenotypes. Detailed investigations of genes within these loci are required to ultimately understand the function of individual genes and how they interact with other players in the network. In this study, we use transgenic plants in combination with natural variation to investigate the regulatory role of the AOP3 gene found in GS-AOP locus previously suggested to contribute to the regulation of glucosinolate defense compounds. Phenotypic analysis and QTL mapping in F2 populations with different AOP3 transgenes support that the enzymatic function and the AOP3 RNA both play a significant role in controlling glucosinolate accumulation. Furthermore, we find different loci interacting with either the enzymatic activity or the RNA of AOP3 and thereby extend the regulatory network controlling glucosinolate accumulation. PMID:26442075

The secreted form of the p40 subunit of interleukin (IL)-12 inhibits IL-23 functions and abrogates IL-23-mediated antitumour effects

PubMed Central

Shimozato, Osamu; Ugai, Shin-ichi; Chiyo, Masako; Takenobu, Hisanori; Nagakawa, Hiroyasu; Wada, Akihiko; Kawamura, Kiyoko; Yamamoto, Hiroshi; Tagawa, Masatoshi

2006-01-01

Interleukin (IL)-23 is a heterodimeric cytokine consisting of a novel p19 molecule and the p40 subunit of IL-12. Since secreted p40 can act as an antagonist for IL-12, we investigated whether p40 also inhibited IL-23-mediated immunological functions. p40 did not induce interferon (IFN)-γ or IL-17 production from splenocytes but impaired IL-23-induced cytokine production by competitive binding to the IL-23 receptors. Furthermore, a mixed population of murine colon carcinoma Colon 26 cells transduced with the p40 gene and those transduced with the IL-23 gene developed tumours in syngenic mice, whereas the IL-23-expressing Colon 26 cells were completely rejected. p40 also suppressed IFN-γ production of antigen-stimulated splenocytes and IL-23-mediated cytotoxic T-lymphocyte activities in the mice that rejected Colon 26 cells expressing IL-23. p40 can thereby antagonize IL-23 and is a possible therapeutic agent for suppression of IL-23 functions. PMID:16423037

PinX1 is recruited to the mitotic chromosome periphery by Nucleolin and facilitates chromosome congression.

PubMed

Li, Na; Yuan, Kai; Yan, Feng; Huo, Yuda; Zhu, Tongge; Liu, Xing; Guo, Zhen; Yao, Xuebiao

2009-06-19

Mitotic chromosome movements are orchestrated by interactions between spindle microtubules and chromosomes. It is well known that kinetochore is the major site where microtubule-chromosome attachment occurs. However, the functions of other domains of chromosome such as chromosome periphery have remained elusive. Our previous studies show that PinX1 distributes to chromosome periphery and kinetochore during mitosis, and harbors the microtubule binding activity. Here we report that PinX1 interacts with Nucleolin, a chromosome periphery protein, through its C-termini. Deconvolution microscopic analyses show PinX1 mainly co-localizes with Nucleolin at chromosome periphery in prometaphase. Moreover, depletion of Nucleolin abolishes chromosome periphery localizations of PinX1, suggesting a functional interrelationship between PinX1 and Nucleolin. Importantly, repression of PinX1 and Nucleolin abrogates chromosome segregation in real-time mitosis, validating the functional importance of PinX1-Nucleolin interaction. We propose PinX1 is recruited to chromosome periphery by Nucleolin and a complex of PinX1 and Nucleolin is essential for faithful chromosome congression.

Natural killer cell biology illuminated by primary immunodeficiency syndromes in humans.

PubMed

Voss, Matthias; Bryceson, Yenan T

2017-04-01

Natural killer (NK) cells are innate immune cytotoxic effector cells well known for their role in antiviral immunity and tumor immunosurveillance. In parts, this knowledge stems from rare inherited immunodeficiency disorders in humans that abrogate NK cell function leading to immune impairments, most notably associated with a high susceptibility to viral infections. Phenotypically, these disorders range from deficiencies selectively affecting NK cells to complex general immune defects that affect NK cells but also other immune cell subsets. Moreover, deficiencies may be associated with reduced NK cell numbers or rather impair specific NK cell effector functions. In recent years, genetic defects underlying the various NK cell deficiencies have been uncovered and have triggered investigative efforts to decipher the molecular mechanisms underlying these disorders. Here we review the associations between inherited human diseases and NK cell development as well as function, with a particular focus on defects in NK cell exocytosis and cytotoxicity. Furthermore we outline how reports of diverse genetic defects have shaped our understanding of NK cell biology. Copyright © 2015. Published by Elsevier Inc.

Macrophage PPARγ inhibits Gpr132 to mediate the anti-tumor effects of rosiglitazone

PubMed Central

Cheng, Wing Yin; Huynh, HoangDinh; Chen, Peiwen; Peña-Llopis, Samuel; Wan, Yihong

2016-01-01

Tumor-associated macrophage (TAM) significantly contributes to cancer progression. Human cancer is enhanced by PPARγ loss-of-function mutations, but inhibited by PPARγ agonists such as TZD diabetes drugs including rosiglitazone. However, it remains enigmatic whether and how macrophage contributes to PPARγ tumor-suppressive functions. Here we report that macrophage PPARγ deletion in mice not only exacerbates mammary tumor development but also impairs the anti-tumor effects of rosiglitazone. Mechanistically, we identify Gpr132 as a novel direct PPARγ target in macrophage whose expression is enhanced by PPARγ loss but repressed by PPARγ activation. Functionally, macrophage Gpr132 is pro-inflammatory and pro-tumor. Genetic Gpr132 deletion not only retards inflammation and cancer growth but also abrogates the anti-tumor effects of PPARγ and rosiglitazone. Pharmacological Gpr132 inhibition significantly impedes mammary tumor malignancy. These findings uncover macrophage PPARγ and Gpr132 as critical TAM modulators, new cancer therapeutic targets, and essential mediators of TZD anti-cancer effects. DOI: http://dx.doi.org/10.7554/eLife.18501.001 PMID:27692066

Membrane raft association is a determinant of plasma membrane localization.

PubMed

Diaz-Rohrer, Blanca B; Levental, Kandice R; Simons, Kai; Levental, Ilya

2014-06-10

The lipid raft hypothesis proposes lateral domains driven by preferential interactions between sterols, sphingolipids, and specific proteins as a central mechanism for the regulation of membrane structure and function; however, experimental limitations in defining raft composition and properties have prevented unequivocal demonstration of their functional relevance. Here, we establish a quantitative, functional relationship between raft association and subcellular protein sorting. By systematic mutation of the transmembrane and juxtamembrane domains of a model transmembrane protein, linker for activation of T-cells (LAT), we generated a panel of variants possessing a range of raft affinities. These mutations revealed palmitoylation, transmembrane domain length, and transmembrane sequence to be critical determinants of membrane raft association. Moreover, plasma membrane (PM) localization was strictly dependent on raft partitioning across the entire panel of unrelated mutants, suggesting that raft association is necessary and sufficient for PM sorting of LAT. Abrogation of raft partitioning led to mistargeting to late endosomes/lysosomes because of a failure to recycle from early endosomes. These findings identify structural determinants of raft association and validate lipid-driven domain formation as a mechanism for endosomal protein sorting.

Membrane raft association is a determinant of plasma membrane localization

PubMed Central

Diaz-Rohrer, Blanca B.; Levental, Kandice R.; Simons, Kai; Levental, Ilya

2014-01-01

The lipid raft hypothesis proposes lateral domains driven by preferential interactions between sterols, sphingolipids, and specific proteins as a central mechanism for the regulation of membrane structure and function; however, experimental limitations in defining raft composition and properties have prevented unequivocal demonstration of their functional relevance. Here, we establish a quantitative, functional relationship between raft association and subcellular protein sorting. By systematic mutation of the transmembrane and juxtamembrane domains of a model transmembrane protein, linker for activation of T-cells (LAT), we generated a panel of variants possessing a range of raft affinities. These mutations revealed palmitoylation, transmembrane domain length, and transmembrane sequence to be critical determinants of membrane raft association. Moreover, plasma membrane (PM) localization was strictly dependent on raft partitioning across the entire panel of unrelated mutants, suggesting that raft association is necessary and sufficient for PM sorting of LAT. Abrogation of raft partitioning led to mistargeting to late endosomes/lysosomes because of a failure to recycle from early endosomes. These findings identify structural determinants of raft association and validate lipid-driven domain formation as a mechanism for endosomal protein sorting. PMID:24912166

Investigating Maternal Touch and Infants' Self-Regulatory Behaviours during a Modified Face-to-Face Still-Face with Touch Procedure

ERIC Educational Resources Information Center

Jean, Amélie D. L.; Stack, Dale M.; Arnold, Sharon

2014-01-01

Maternal touch and infants' self-regulatory behaviours were examined during a modified Still-Face with Touch (SF?+?T) procedure. Mothers and their 5½-month-old infants participated in one period of Normal interaction followed by three SF?+?T periods. Maternal functions of touch, and infants' self-regulatory behaviour, affect, and…

Metabolic functions of AMPK: aspects of structure and of natural mutations in the regulatory gamma subunits.

PubMed

Moffat, Cynthia; Harper, Mary Ellen

2010-10-01

AMP-activated protein kinase, AMPK, is widely accepted as the master regulator of energy levels within the cell. Responding quickly to changing energy demands, AMPK works to restore levels of ATP during times of cellular stress by promoting ATP producing catabolic pathways and inhibiting ATP consuming anabolic ones. As a heterotrimeric protein complex, AMPK's subunits each act in unique and crucial ways to control AMPK function and its localization within the cell. Research in the last decade has identified and begun to characterize the impact of naturally occurring mutations in the gamma regulatory subunits. Mutations in the γ2 subunit have implications for cardiac function and disease, while the R225W mutation in the γ3 subunit have implications for skeletal muscle fuel metabolism and resistance to fatigue. Research focused on structure-function aspects of AMPK regulatory subunits will lead to a better understanding of the roles of AMPK in health and disease.

Pain acceptance, psychological functioning, and self-regulatory fatigue in temporomandibular disorder.

PubMed

Eisenlohr-Moul, Tory A; Burris, Jessica L; Evans, Daniel R

2013-12-01

A growing body of evidence suggests that chronic pain patients suffer from chronic self-regulatory fatigue: difficulty controlling thoughts, emotions, and behavior. Pain acceptance, which involves responding to pain and related experiences without attempts to control or avoid them (pain willingness), and pursuit of valued life activities regardless of pain (activity engagement) has been associated with various favorable outcomes in chronic pain patients, including better psychological functioning. The study presented here tested the hypotheses that pain acceptance is associated with less psychological distress, higher psychological well-being, and reduced self-regulatory fatigue in temporomandibular disorder (TMD) patients, particularly for those with longer pain duration. Cross-sectional data were provided by 135 TMD patients during an initial evaluation at a university-based tertiary orofacial pain clinic. Results of hierarchical linear regression models indicated that, controlling for pain severity, pain willingness is associated with less psychological distress and lower self-regulatory fatigue, and activity engagement is associated with greater psychological well-being. Furthermore, the effect of pain willingness on psychological distress was moderated by pain duration such that pain willingness was more strongly associated with less psychological distress in patients with longer pain duration; this moderating effect was fully mediated by self-regulatory fatigue. These findings suggest pain willingness may buffer against self-regulatory fatigue in those with longer pain duration, and such conservation of self-regulatory resources may protect against psychological symptoms.

Regulatory T cells in the control of host-microorganism interactions (*).

PubMed

Belkaid, Yasmine; Tarbell, Kristin

2009-01-01

Each microenvironment requires a specific set of regulatory elements that are finely and constantly tuned to maintain local homeostasis. Various populations of regulatory T cells contribute to the maintenance of this equilibrium and establishment of controlled immune responses. In particular, regulatory T cells limit the magnitude of effector responses, which may result in failure to adequately control infection. However, regulatory T cells also help limit collateral tissue damage caused by vigorous antimicrobial immune responses against pathogenic microbes as well as commensals. In this review, we describe various situations in which the balance between regulatory T cells and effector immune functions influence the outcome of host-microorganism coexistence and discuss current hypotheses and points of polemic associated with the origin, target, and antigen specificity of both endogenous and induced regulatory T cells during these interactions.

Deciphering the transcriptional cis-regulatory code.

PubMed

Yáñez-Cuna, J Omar; Kvon, Evgeny Z; Stark, Alexander

2013-01-01

Information about developmental gene expression resides in defined regulatory elements, called enhancers, in the non-coding part of the genome. Although cells reliably utilize enhancers to orchestrate gene expression, a cis-regulatory code that would allow their interpretation has remained one of the greatest challenges of modern biology. In this review, we summarize studies from the past three decades that describe progress towards revealing the properties of enhancers and discuss how recent approaches are providing unprecedented insights into regulatory elements in animal genomes. Over the next years, we believe that the functional characterization of regulatory sequences in entire genomes, combined with recent computational methods, will provide a comprehensive view of genomic regulatory elements and their building blocks and will enable researchers to begin to understand the sequence basis of the cis-regulatory code. Copyright © 2012 Elsevier Ltd. All rights reserved.

Identification of a regulatory T cell specific cell surface molecule that mediates suppressive signals and induces Foxp3 expression.

PubMed

Wang, Rui; Wan, Qi; Kozhaya, Lina; Fujii, Hodaka; Unutmaz, Derya

2008-07-16

Regulatory T (T(reg)) cells control immune activation and maintain tolerance. How T(regs) mediate their suppressive function is unclear. Here we identified a cell surface molecule, called GARP, (or LRRC32), which within T cells is specifically expressed in T(regs) activated through the T cell receptor (TCR). Ectopic expression of GARP in human naïve T (T(N)) cells inhibited their proliferation and cytokine secretion upon TCR activation. Remarkably, GARP over-expression in T(N) cells induced expression of T(reg) master transcription factor Foxp3 and endowed them with a partial suppressive function. The extracellular but not the cytoplasmic region of GARP, was necessary for these functions. Silencing Foxp3 in human T(reg) cells reduced expression of GARP and attenuated their suppressive function. However, GARP function was not affected when Foxp3 was downregulated in GARP-overexpressing cells, while silencing GARP in Foxp3-overexpressing cells reduced their suppressive activity. These findings reveal a novel cell surface molecule-mediated regulatory mechanism, with implications for modulating aberrant immune responses.

Exploring associations between self-regulatory mechanisms and neuropsychological functioning and driver behaviour after brain injury.

PubMed

Rike, Per-Ola; Johansen, Hans J; Ulleberg, Pål; Lundqvist, Anna; Schanke, Anne-Kristine

2018-04-01

The objective of this prospective one-year follow-up study was to explore the associations between self-regulatory mechanisms and neuropsychological tests as well as baseline and follow-up ratings of driver behaviour. The participants were a cohort of subjects with stroke and traumatic brain injury (TBI) who were found fit to drive after a multi-disciplinary driver assessment (baseline). Baseline measures included neuropsychological tests and ratings of self-regulatory mechanisms, i.e., executive functions (Behavior Rating Inventory of Executive Function-Adult Version; BRIEF-A) and impulsive personality traits (UPPS Impulsive Behavior Scale). The participants rated pre-injury driving behaviour on the Driver Behaviour Qestionnaire (DBQ) retrospectively at baseline and after one year of post-injury driving (follow-up). Better performance on neuropsychological tests was significantly associated with more post-injury DBQ Violations. The BRIEF-A main indexes were significantly associated with baseline and follow-up ratings of DBQ Mistakes and follow-up DBQ Inattention. UPPS (lack of) Perseverance was significantly associated with baseline DBQ Inattention, whereas UPPS Urgency was significantly associated with baseline DBQ Inexperience and post-injury DBQ Mistakes. There were no significant changes in DBQ ratings from baseline (pre-injury) to follow-up (post-injury). It was concluded that neuropsychological functioning and self-regulatory mechanisms are related to driver behaviour. Some aspects of driver behaviour do not necessarily change after brain injury, reflecting the influence of premorbid driving behaviour or impaired awareness of deficits on post-injury driving behaviour. Further evidence is required to predict the role of self-regulatory mechanisms on driver behaviour and crashes or near misses.

Analysis of functional importance of binding sites in the Drosophila gap gene network model.

PubMed

Kozlov, Konstantin; Gursky, Vitaly V; Kulakovskiy, Ivan V; Dymova, Arina; Samsonova, Maria

2015-01-01

The statistical thermodynamics based approach provides a promising framework for construction of the genotype-phenotype map in many biological systems. Among important aspects of a good model connecting the DNA sequence information with that of a molecular phenotype (gene expression) is the selection of regulatory interactions and relevant transcription factor bindings sites. As the model may predict different levels of the functional importance of specific binding sites in different genomic and regulatory contexts, it is essential to formulate and study such models under different modeling assumptions. We elaborate a two-layer model for the Drosophila gap gene network and include in the model a combined set of transcription factor binding sites and concentration dependent regulatory interaction between gap genes hunchback and Kruppel. We show that the new variants of the model are more consistent in terms of gene expression predictions for various genetic constructs in comparison to previous work. We quantify the functional importance of binding sites by calculating their impact on gene expression in the model and calculate how these impacts correlate across all sites under different modeling assumptions. The assumption about the dual interaction between hb and Kr leads to the most consistent modeling results, but, on the other hand, may obscure existence of indirect interactions between binding sites in regulatory regions of distinct genes. The analysis confirms the previously formulated regulation concept of many weak binding sites working in concert. The model predicts a more or less uniform distribution of functionally important binding sites over the sets of experimentally characterized regulatory modules and other open chromatin domains.

Regulatory T cells: mechanisms of differentiation and function.

PubMed

Josefowicz, Steven Z; Lu, Li-Fan; Rudensky, Alexander Y

2012-01-01

The immune system has evolved to mount an effective defense against pathogens and to minimize deleterious immune-mediated inflammation caused by commensal microorganisms, immune responses against self and environmental antigens, and metabolic inflammatory disorders. Regulatory T (Treg) cell-mediated suppression serves as a vital mechanism of negative regulation of immune-mediated inflammation and features prominently in autoimmune and autoinflammatory disorders, allergy, acute and chronic infections, cancer, and metabolic inflammation. The discovery that Foxp3 is the transcription factor that specifies the Treg cell lineage facilitated recent progress in understanding the biology of regulatory T cells. In this review, we discuss cellular and molecular mechanisms in the differentiation and function of these cells.

The regulatory role of B cells in autoimmunity, infections and cancer: Perspectives beyond IL10 production.

PubMed

Gorosito Serrán, Melisa; Fiocca Vernengo, Facundo; Beccaria, Cristian G; Acosta Rodriguez, Eva V; Montes, Carolina L; Gruppi, Adriana

2015-11-14

The term regulatory B cells (B regs) is ascribed to a heterogeneous population of B cells with the function of suppressing inflammatory responses. They have been described mainly during the last decade in the context of different immune-mediated diseases. Most of the work on B regs has been focused on IL-10-producing B cells. However, B cells can exert regulatory functions independently of IL-10 production. Here we discuss the phenotypes, development and effector mechanisms of B regs and advances in their role in autoimmunity, infections and cancer. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Unraveling transcriptional control and cis-regulatory codes using the software suite GeneACT

PubMed Central

Cheung, Tom Hiu; Kwan, Yin Lam; Hamady, Micah; Liu, Xuedong

2006-01-01

Deciphering gene regulatory networks requires the systematic identification of functional cis-acting regulatory elements. We present a suite of web-based bioinformatics tools, called GeneACT , that can rapidly detect evolutionarily conserved transcription factor binding sites or microRNA target sites that are either unique or over-represented in differentially expressed genes from DNA microarray data. GeneACT provides graphic visualization and extraction of common regulatory sequence elements in the promoters and 3'-untranslated regions that are conserved across multiple mammalian species. PMID:17064417

Effects of natalizumab treatment on Foxp3+ T regulatory cells.

PubMed

Stenner, Max-Philipp; Waschbisch, Anne; Buck, Dorothea; Doerck, Sebastian; Einsele, Hermann; Toyka, Klaus V; Wiendl, Heinz

2008-10-06

Natalizumab, a monoclonal humanized antibody targeting the alpha-4 chain of very late activation antigen 4 (VLA-4) exerts impressive therapeutic effects in patients with relapsing-remitting multiple sclerosis. Our objective was to study impacts of Natalizumab therapy on Foxp3+ T regulatory cells (Tregs) in multiple sclerosis (MS) patients. A combined approach of in vitro and ex vivo experiments using T cells isolated from the peripheral blood of healthy donors and Natalizumab treated MS patients was chosen. We determined binding of Natalizumab and its effects on the frequency, transmigratory behaviour and suppressive function of Tregs. Binding of Natalizumab and expression of CD49d (alpha-4 chain of VLA-4) differed between non-regulatory and regulatory cells. Albeit Foxp3+ Tregs had lower levels of CD49d, Natalizumab blocked the transmigration of Foxp3+ Tregs similar to non-regulatory T cells. The frequency of peripheral blood Tregs was unaffected by Natalizumab treatment. Natalizumab does not alter the suppressive capacity of CD4+CD25(high)CD127(low)Foxp3+ Tregs under in vitro conditions. Furthermore, the impaired function of Tregs in MS patients is not restored by Natalizumab treatment. We provide a first detailed analysis of Natalizumab effects on the regulatory T cell population. Our prospective study shows that Foxp3+ Tregs express lower levels of VLA-4 and bind less Natalizumab. We further the understanding of the mechanisms of action of Natalizumab by demonstrating that unlike other immunomodulatory drugs the beneficial therapeutic effects of the monoclonal antibody are largely independent of alterations in Treg frequency or function.

Effects of Natalizumab Treatment on Foxp3+ T Regulatory Cells

PubMed Central

Buck, Dorothea; Doerck, Sebastian; Einsele, Hermann; Toyka, Klaus V.; Wiendl, Heinz

2008-01-01

Background Natalizumab, a monoclonal humanized antibody targeting the alpha-4 chain of very late activation antigen 4 (VLA-4) exerts impressive therapeutic effects in patients with relapsing-remitting multiple sclerosis. Our objective was to study impacts of Natalizumab therapy on Foxp3+ T regulatory cells (Tregs) in multiple sclerosis (MS) patients. Methodology A combined approach of in vitro and ex vivo experiments using T cells isolated from the peripheral blood of healthy donors and Natalizumab treated MS patients was chosen. We determined binding of Natalizumab and its effects on the frequency, transmigratory behaviour and suppressive function of Tregs. Principal Findings Binding of Natalizumab and expression of CD49d (alpha-4 chain of VLA-4) differed between non-regulatory and regulatory cells. Albeit Foxp3+ Tregs had lower levels of CD49d, Natalizumab blocked the transmigration of Foxp3+ Tregs similar to non-regulatory T cells. The frequency of peripheral blood Tregs was unaffected by Natalizumab treatment. Natalizumab does not alter the suppressive capacity of CD4+CD25highCD127lowFoxp3+ Tregs under in vitro conditions. Furthermore, the impaired function of Tregs in MS patients is not restored by Natalizumab treatment. Conclusions We provide a first detailed analysis of Natalizumab effects on the regulatory T cell population. Our prospective study shows that Foxp3+ Tregs express lower levels of VLA-4 and bind less Natalizumab. We further the understanding of the mechanisms of action of Natalizumab by demonstrating that unlike other immunomodulatory drugs the beneficial therapeutic effects of the monoclonal antibody are largely independent of alterations in Treg frequency or function. PMID:18836525

A gene regulatory network armature for T-lymphocyte specification

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fung, Elizabeth-sharon

Choice of a T-lymphoid fate by hematopoietic progenitor cells depends on sustained Notch-Delta signaling combined with tightly-regulated activities of multiple transcription factors. To dissect the regulatory network connections that mediate this process, we have used high-resolution analysis of regulatory gene expression trajectories from the beginning to the end of specification; tests of the short-term Notchdependence of these gene expression changes; and perturbation analyses of the effects of overexpression of two essential transcription factors, namely PU.l and GATA-3. Quantitative expression measurements of >50 transcription factor and marker genes have been used to derive the principal components of regulatory change through whichmore » T-cell precursors progress from primitive multipotency to T-lineage commitment. Distinct parts of the path reveal separate contributions of Notch signaling, GATA-3 activity, and downregulation of PU.l. Using BioTapestry, the results have been assembled into a draft gene regulatory network for the specification of T-cell precursors and the choice of T as opposed to myeloid dendritic or mast-cell fates. This network also accommodates effects of E proteins and mutual repression circuits of Gfil against Egr-2 and of TCF-l against PU.l as proposed elsewhere, but requires additional functions that remain unidentified. Distinctive features of this network structure include the intense dose-dependence of GATA-3 effects; the gene-specific modulation of PU.l activity based on Notch activity; the lack of direct opposition between PU.l and GATA-3; and the need for a distinct, late-acting repressive function or functions to extinguish stem and progenitor-derived regulatory gene expression.« less

Paeoniflorin inhibits cell growth and induces cell cycle arrest through inhibition of FoxM1 in colorectal cancer cells.

PubMed

Yue, Meng; Li, Shiquan; Yan, Guoqiang; Li, Chenyao; Kang, Zhenhua

2018-01-01

Paeoniflorin (PF) exhibits tumor suppressive functions in a variety of human cancers. However, the function of PF and molecular mechanism in colorectal cancer are elusive. In the present study, we investigated whether PF could exert its antiproliferative activity, anti-migration, and anti-invasive function in colorectal cancer cells. We found that PF inhibited cell growth and induced apoptosis and blocked cell cycle progression in the G0/G1 phase in colorectal cancer cells. Moreover, we found that PF suppressed cell migration and invasion in colorectal cancer cells. FoxM1 has been reported to play an important oncogenic role in human cancers. We also determine whether PF inhibited the expression of FoxM1, leading to its anti-cancer activity. We found that PF treatment in colorectal cancer cells resulted in down-regulation of FoxM1. The rescue experiments showed that overexpression of FoxM1 abrogated the tumor suppressive function induced by PF treatment. Notably, depletion of FoxM1 promoted the anti-tumor activity of PF in colorectal cancer cells. Therefore, inhibition of FoxM1 could participate in the anti-tumor activity of PF in colorectal cancer cells.

10 CFR 51.30 - Environmental assessment.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Environmental assessment. 51.30 Section 51.30 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2...

10 CFR 51.99 - [Reserved

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false [Reserved] 51.99 Section 51.99 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2) Final Environmental...

10 CFR 51.99 - [Reserved

Code of Federal Regulations, 2014 CFR

2014-01-01

... 10 Energy 2 2014-01-01 2014-01-01 false [Reserved] 51.99 Section 51.99 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2) Final Environmental...

10 CFR 51.16 - Proprietary information.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Proprietary information. 51.16 Section 51.16 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2...

10 CFR 51.16 - Proprietary information.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Proprietary information. 51.16 Section 51.16 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2...

A conserved RNA structural element within the hepatitis B virus post-transcriptional regulatory element enhance nuclear export of intronless transcripts and repress the splicing mechanism.

PubMed

Visootsat, Akasit; Payungporn, Sunchai; T-Thienprasert, Nattanan P

2015-12-01

Hepatitis B virus (HBV) infection is a primary cause of hepatocellular carcinoma and liver cirrhosis worldwide. To develop novel antiviral drugs, a better understanding of HBV gene expression regulation is vital. One important aspect is to understand how HBV hijacks the cellular machinery to export unspliced RNA from the nucleus. The HBV post-transcriptional regulatory element (HBV PRE) has been proposed to be the HBV RNA nuclear export element. However, the function remains controversial, and the core element is unclear. This study, therefore, aimed to identify functional regulatory elements within the HBV PRE and investigate their functions. Using bioinformatics programs based on sequence conservation and conserved RNA secondary structures, three regulatory elements were predicted, namely PRE 1151-1410, PRE 1520-1620 and PRE 1650-1684. PRE 1151-1410 significantly increased intronless and unspliced luciferase activity in both HepG2 and COS-7 cells. Likewise, PRE 1151-1410 significantly elevated intronless and unspliced HBV surface transcripts in liver cancer cells. Moreover, motif analysis predicted that PRE 1151-1410 contains several regulatory motifs. This study reported the roles of PRE 1151-1410 in intronless transcript nuclear export and the splicing mechanism. Additionally, these results provide knowledge in the field of HBV RNA regulation. Moreover, PRE 1151-1410 may be used to enhance the expression of other mRNAs in intronless reporter plasmids.

Emotion: The Self-regulatory Sense

PubMed Central

2014-01-01

While emotion is a central component of human health and well-being, traditional approaches to understanding its biological function have been wanting. A dynamic systems model, however, broadly redefines and recasts emotion as a primary sensory system—perhaps the first sensory system to have emerged, serving the ancient autopoietic function of “self-regulation.” Drawing upon molecular biology and revelations from the field of epigenetics, the model suggests that human emotional perceptions provide an ongoing stream of “self-relevant” sensory information concerning optimally adaptive states between the organism and its immediate environment, along with coupled behavioral corrections that honor a universal self-regulatory logic, one still encoded within cellular signaling and immune functions. Exemplified by the fundamental molecular circuitry of sensorimotor control in the E coli bacterium, the model suggests that the hedonic (affective) categories emerge directly from positive and negative feedback processes, their good/bad binary appraisals relating to dual self-regulatory behavioral regimes—evolutionary purposes, through which organisms actively participate in natural selection, and through which humans can interpret optimal or deficit states of balanced being and becoming. The self-regulatory sensory paradigm transcends anthropomorphism, unites divergent theoretical perspectives and isolated bodies of literature, while challenging time-honored assumptions. While suppressive regulatory strategies abound, it suggests that emotions are better understood as regulating us, providing a service crucial to all semantic language, learning systems, evaluative decision-making, and fundamental to optimal physical, mental, and social health. PMID:24808986

Cardiac Expression of ms1/STARS, a Novel Gene Involved in Cardiac Development and Disease, Is Regulated by GATA4

PubMed Central

Kobayashi, Satoru; Peterson, Richard E.; He, Aibin; Motterle, Anna; Samani, Nilesh J.; Menick, Donald R.; Pu, William T.; Liang, Qiangrong

2012-01-01

Ms1/STARS is a novel muscle-specific actin-binding protein that specifically modulates the myocardin-related transcription factor (MRTF)-serum response factor (SRF) regulatory axis within striated muscle. This ms1/STARS-dependent regulatory axis is of central importance within the cardiac gene regulatory network and has been implicated in cardiac development and postnatal cardiac function/homeostasis. The dysregulation of ms1/STARS is associated with and causative of pathological cardiac phenotypes, including cardiac hypertrophy and cardiomyopathy. In order to gain an understanding of the mechanisms governing ms1/STARS expression in the heart, we have coupled a comparative genomic in silico analysis with reporter, gain-of-function, and loss-of-function approaches. Through this integrated analysis, we have identified three evolutionarily conserved regions (ECRs), α, SINA, and DINA, that act as cis-regulatory modules and confer differential cardiac cell-specific activity. Two of these ECRs, α and DINA, displayed distinct regulatory sensitivity to the core cardiac transcription factor GATA4. Overall, our results demonstrate that within embryonic, neonatal, and adult hearts, GATA4 represses ms1/STARS expression with the pathologically associated depletion of GATA4 (type 1/type 2 diabetic models), resulting in ms1/STARS upregulation. This GATA4-dependent repression of ms1/STARS expression has major implications for MRTF-SRF signaling in the context of cardiac development and disease. PMID:22431517

Function does not follow form in gene regulatory circuits.

PubMed

Payne, Joshua L; Wagner, Andreas

2015-08-20

Gene regulatory circuits are to the cell what arithmetic logic units are to the chip: fundamental components of information processing that map an input onto an output. Gene regulatory circuits come in many different forms, distinct structural configurations that determine who regulates whom. Studies that have focused on the gene expression patterns (functions) of circuits with a given structure (form) have examined just a few structures or gene expression patterns. Here, we use a computational model to exhaustively characterize the gene expression patterns of nearly 17 million three-gene circuits in order to systematically explore the relationship between circuit form and function. Three main conclusions emerge. First, function does not follow form. A circuit of any one structure can have between twelve and nearly thirty thousand distinct gene expression patterns. Second, and conversely, form does not follow function. Most gene expression patterns can be realized by more than one circuit structure. And third, multifunctionality severely constrains circuit form. The number of circuit structures able to drive multiple gene expression patterns decreases rapidly with the number of these patterns. These results indicate that it is generally not possible to infer circuit function from circuit form, or vice versa.

Stretch-activated ion channel blockade attenuates adaptations to eccentric exercise.

PubMed

Butterfield, Timothy A; Best, Thomas M

2009-02-01

The purpose of this study was to test the hypothesis that stretch-activated ion channel (SAC) function is essential for the repeated bout effect (RBE) in skeletal muscle. Specifically, we investigated if daily injections of streptomycin (a known SAC blocker) would abrogate the muscle's adaptive resistance to the damaging effects of eccentric exercise over a 4-wk period. Furthermore, we hypothesized that the lack of an RBE would be due to the lack of functional adaptations that typically result from repeated bouts of eccentric exercise, including increased peak isometric torque, muscle hypertrophy, and rightward shift of the torque-angle relationship. Twelve New Zealand white rabbits were each subjected to 12 bouts of eccentric exercise over a 4-wk period while receiving either daily injections of streptomycin or sham injections. Although blocking the SAC function completely eliminated the expected adaptive response in biomechanical parameters during the exercise regimen, there remained evidence of an acquired RBE, albeit with an attenuated response when compared with the muscles with intact SAC function. Blocking sarcolemmal SAC eliminates functional adaptations of muscle after eccentric exercise. In the absence of SAC function, muscles subjected to chronic eccentric exercise still exhibit some degree of the RBE. As such, it appears that the signaling cascade that results in functional, biomechanical adaptations associated with the RBE during eccentric exercise is dependent upon intact SAC function.

Cis-regulatory signatures of orthologous stress-associated bZIP transcription factors from rice, sorghum and Arabidopsis based on phylogenetic footprints

PubMed Central

2012-01-01

Background The potential contribution of upstream sequence variation to the unique features of orthologous genes is just beginning to be unraveled. A core subset of stress-associated bZIP transcription factors from rice (Oryza sativa) formed ten clusters of orthologous groups (COG) with genes from the monocot sorghum (Sorghum bicolor) and dicot Arabidopsis (Arabidopsis thaliana). The total cis-regulatory information content of each stress-associated COG was examined by phylogenetic footprinting to reveal ortholog-specific, lineage-specific and species-specific conservation patterns. Results The most apparent pattern observed was the occurrence of spatially conserved ‘core modules’ among the COGs but not among paralogs. These core modules are comprised of various combinations of two to four putative transcription factor binding site (TFBS) classes associated with either developmental or stress-related functions. Outside the core modules are specific stress (ABA, oxidative, abiotic, biotic) or organ-associated signals, which may be functioning as ‘regulatory fine-tuners’ and further define lineage-specific and species-specific cis-regulatory signatures. Orthologous monocot and dicot promoters have distinct TFBS classes involved in disease and oxidative-regulated expression, while the orthologous rice and sorghum promoters have distinct combinations of root-specific signals, a pattern that is not particularly conserved in Arabidopsis. Conclusions Patterns of cis-regulatory conservation imply that each ortholog has distinct signatures, further suggesting that they are potentially unique in a regulatory context despite the presumed conservation of broad biological function during speciation. Based on the observed patterns of conservation, we postulate that core modules are likely primary determinants of basal developmental programming, which may be integrated with and further elaborated by additional intrinsic or extrinsic signals in conjunction with lineage-specific or species-specific regulatory fine-tuners. This synergy may be critical for finer-scale spatio-temporal regulation, hence unique expression profiles of homologous transcription factors from different species with distinct zones of ecological adaptation such as rice, sorghum and Arabidopsis. The patterns revealed from these comparisons set the stage for further empirical validation by functional genomics. PMID:22992304

Hepatocyte-induced CD4+ T cell alloresponse is associated with major histocompatibility complex class II up-regulation on hepatocytes and suppressible by regulatory T cells.

PubMed

DeTemple, Daphne E; Oldhafer, Felix; Falk, Christine S; Chen-Wacker, Chen; Figueiredo, Constanca; Kleine, Moritz; Ramackers, Wolf; Timrott, Kai; Lehner, Frank; Klempnauer, Juergen; Bock, Michael; Vondran, Florian W R

2018-03-01

Hepatocyte transplantation is a promising therapeutic approach for various liver diseases. Despite the liver's tolerogenic potential, early immune-mediated loss of transplanted cells is observed, and longterm acceptance has not been achieved yet. Patients deemed tolerant after liver transplantation presented an increased frequency of regulatory T cells (Tregs), which therefore also might enable reduction of posttransplant cell loss and enhance longterm allograft acceptance. We hence characterized hepatocyte-induced immune reactions and evaluated the immunomodulatory potential of Tregs applying mixed lymphocyte cultures and mixed lymphocyte hepatocyte cultures. These were set up using peripheral blood mononuclear cells and primary human hepatocytes, respectively. Polyclonally expanded CD4 + CD25 high CD127 low Tregs were added to cocultures in single-/trans-well setups with/without supplementation of anti-interferon γ (IFNγ) antibodies. Hepatocyte-induced alloresponses were then analyzed by multicolor flow cytometry. Measurements indicated that T cell response upon stimulation was associated with IFNγ-induced major histocompatibility complex (MHC) class II up-regulation on hepatocytes and mediated by CD4 + T cells. An indirect route of antigen presentation could be ruled out by use of fragmented hepatocytes and culture supernatants of hepatocytes. Allospecific proliferation was accompanied by inflammatory cytokine secretion. CD8 + T cells showed early up-regulation of CD69 despite lack of cell proliferation in the course of coculture. Supplementation of Tregs effectively abrogated hepatocyte-induced alloresponses and was primarily cell contact dependent. In conclusion, human hepatocytes induce a CD4 + T cell alloresponse in vitro, which is associated with MHC class II up-regulation on hepatocytes and is susceptible to suppression by Tregs. Liver Transplantation 24 407-419 2018 AASLD. © 2018 The Authors. Liver Transplantation published by Wiley Periodicals, Inc. on behalf of American Association for the Study of Liver Diseases.

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. © 2015 British Society for Immunology.

Activated glycoprotein A repetitions predominant (GARP)-expressing regulatory T cells inhibit allergen-induced intestinal inflammation in humanized mice.

PubMed

Eschborn, Melanie; Weigmann, Benno; Reissig, Sonja; Waisman, Ari; Saloga, Joachim; Bellinghausen, Iris

2015-07-01

Recently, we developed a humanized mouse model of allergen-induced IgE-dependent gut inflammation in PBMC-engrafted immunodeficient mice. In the present study, we wanted to investigate the role of regulatory T (Treg) cells and their activation status in this model. Nonobese diabetic-severe combined immunodeficiency-γc(-/-) mice were injected intraperitoneally with human PBMCs from allergic donors together with the respective allergen or NaCl as control in the presence or absence of different concentrations of CD4(+)CD25(+) Treg cells of the same donor. After an additional allergen boost 1 week later, mice were challenged with the allergen rectally on day 21 and gut inflammation was monitored by a high-resolution video mini-endoscopic system evaluating translucency, granularity, fibrin production, vascularity, and stool. Allergen-specific human IgE in mouse sera, which was detectable only in PBMC plus allergen-treated mice, was strongly inhibited by coinjection of Treg cells at a ratio of at least 1:10. Consequently, the presence of Treg cells significantly decreased IgE-dependent allergen-induced gut inflammation after rectal allergen challenge. In addition, Treg cells reduced allergen-specific proliferation and cytokine production of recovered human CD4(+) T cells in vitro. Activation of Treg cells before injection further increased all inhibitory effects. Prevention of gut inflammation also occurred by the administration of glycoprotein A repetitions predominant, a molecule expressed by activated Treg cells, whereas its blockade completely abrogated inhibition by Treg cells. These results demonstrate that allergen-specific gut inflammation in human PBMC-engrafted mice can be avoided by enhancing the numbers or activity of autologous Treg cells, which is of great interest for therapeutic intervention of allergic diseases of the intestine. Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Tumor evasion of the immune system by converting CD4+CD25- T cells into CD4+CD25+ T regulatory cells: role of tumor-derived TGF-beta.

PubMed

Liu, Victoria C; Wong, Larry Y; Jang, Thomas; Shah, Ali H; Park, Irwin; Yang, Ximing; Zhang, Qiang; Lonning, Scott; Teicher, Beverly A; Lee, Chung

2007-03-01

CD4+CD25+ T regulatory (T(reg)) cells were initially described for their ability to suppress autoimmune diseases in animal models. An emerging interest is the potential role of T(reg) cells in cancer development and progression because they have been shown to suppress antitumor immunity. In this study, CD4+CD25- T cells cultured in conditioned medium (CM) derived from tumor cells, RENCA or TRAMP-C2, possess similar characteristics as those of naturally occurring T(reg) cells, including expression of Foxp3, a crucial transcription factor of T(reg) cells, production of low levels of IL-2, high levels of IL-10 and TGF-beta, and the ability to suppress CD4+CD25- T cell proliferation. Further investigation revealed a critical role of tumor-derived TGF-beta in converting CD4+CD25- T cells into T(reg) cells because a neutralizing Ab against TGF-beta, 1D11, completely abrogated the induction of T(reg) cells. CM from a nontumorigenic cell line, NRP-152, or irradiated tumor cells did not convert CD4+CD25- T cells to T(reg) cells because they produce low levels of TGF-beta in CM. Finally, we observed a reduced tumor burden in animals receiving 1D11. The reduction in tumor burden correlated with a decrease in tumor-derived TGF-beta. Treatment of 1D11 also reduced the conversion of CD4+ T cells into T(reg) cells and subsequent T(reg) cell-mediated suppression of antitumor immunity. In summary, we have demonstrated that tumor cells directly convert CD4+CD25- T cells to T(reg) cells through production of high levels of TGF-beta, suggesting a possible mechanism through which tumor cells evade the immune system.

Escherichia coli heat-labile enterotoxin B subunit prevents autoimmune arthritis through induction of regulatory CD4+ T cells.

PubMed

Luross, Jeffrey A; Heaton, Tricia; Hirst, Timothy R; Day, Michael J; Williams, Neil A

2002-06-01

The receptor-binding B subunit of Escherichia coli heat-labile enterotoxin (EtxB) is a highly stable, nontoxic protein that is capable of modulating immune responses. This study was conducted to determine whether mucosal administration of EtxB can block collagen-induced arthritis (CIA) and to investigate the mechanisms involved. Clinical arthritis in DBA/1 mice was monitored following mucosal administration of EtxB on 4 occasions. The dependence of disease prevention on receptor binding by EtxB and the associated alterations to the immune response to type II collagen (CII) were assessed. Adoptive transfer experiments and lymph node cell cocultures were used to investigate the underlying mechanisms. Both intranasal and intragastric delivery of EtxB were effective in preventing CIA; a 1-microg dose of EtxB was protective after intranasal administration. A non-receptor-binding mutant of EtxB failed to prevent disease. Intranasal EtxB lowered both the incidence and severity of arthritis when given either at the time of disease induction or 25 days later. EtxB markedly reduced levels of anti-CII IgG2a antibodies and interferon-gamma (IFNgamma) production while not affecting levels of IgG1, interleukin-4 (IL-4), or IL-10. Disease protection could be transferred by CD4+ T cells from treated mice, an effect that was abrogated upon depletion of the CD25+ population. In addition, CD4+CD25+ T cells from treated mice were able to suppress anti-CII IFNgamma production by CII-primed lymph node cells. Mucosal administration of EtxB can be used to prevent or treat CIA. Modulation of the anti-CII immune response by EtxB is associated with a reduction in Th1 cell reactivity without a concomitant shift toward Th2. Instead, EtxB mediates its effects through enhancing the activity of a population of CD4+ regulatory T cells.

1alpha,25-dihydroxyvitamin D3 potentiates the beneficial effects of allergen immunotherapy in a mouse model of allergic asthma: role for IL-10 and TGF-beta.

PubMed

Taher, Yousef A; van Esch, Betty C A M; Hofman, Gerard A; Henricks, Paul A J; van Oosterhout, Antoon J M

2008-04-15

1alpha,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), a potent inhibitor of NF-kappaB expression, can prevent the maturation of dendritic cells in vitro leading to tolerogenic dendritic cells with increased potential to induce regulatory T cells. Herein, we investigated whether the combination of allergen immunotherapy with 1,25(OH)(2)D(3) potentiates the suppressive effects of immunotherapy and whether the immunoregulatory cytokines IL-10 and TGF-beta are involved in the effector phase. OVA-sensitized and challenged BALB/c mice displayed airway hyperresponsiveness (AHR) and increased serum OVA-specific IgE levels, bronchoalveolar lavage eosinophilia, and Th2 cytokine levels. In this model, the dose response of allergen immunotherapy 10 days before OVA inhalation challenge shows strong suppression of asthma manifestations at 1 mg of OVA, but partial suppression of bronchoalveolar lavage eosinophilia, IgE up-regulation, and no reduction of AHR at 100 microg. Interestingly, coadministration of 10 ng of 1,25(OH)(2)D(3) with 100 microg of OVA immunotherapy significantly inhibited AHR and potentiated the reduction of serum OVA-specific IgE levels, airway eosinophilia, and Th2-related cytokines concomitant with increased IL-10 levels in lung tissues and TGF-beta and OVA-specific IgA levels in serum. Similar effects on suboptimal immunotherapy were observed by inhibition of the NF-kappaB pathway using the selective IkappaB kinase 2 inhibitor PS-1145. The suppressive effects of this combined immunotherapy were partially reversed by treatment with mAb to either IL-10R or TGF-beta before OVA inhalation challenge but completely abrogated when both Abs were given. These data demonstrate that 1,25(OH)(2)D(3) potentiates the efficacy of immunotherapy and that the regulatory cytokines IL-10 and TGF-beta play a crucial role in the effector phase of this mouse model.

Thromboxane A2-induced bi-directional regulation of cerebral arterial tone.

PubMed

Neppl, Ronald L; Lubomirov, Lubomir T; Momotani, Ko; Pfitzer, Gabriele; Eto, Masumi; Somlyo, Avril V

2009-03-06

Myosin light chain phosphatase plays a critical role in modulating smooth muscle contraction in response to a variety of physiologic stimuli. A downstream target of the RhoA/Rho-kinase and nitric oxide (NO)/cGMP/cyclic GMP-dependent kinase (cGKI) pathways, myosin light chain phosphatase activity reflects the sum of both calcium sensitization and desensitization pathways through phosphorylation and dephosphorylation of the myosin phosphatase targeting subunit (MYPT1). As cerebral blood flow is highly spatio-temporally modulated under normal physiologic conditions, severe perturbations in normal cerebral blood flow, such as in cerebral vasospasm, can induce neurological deficits. In nonpermeabilized cerebral vessels stimulated with U-46619, a stable mimetic of endogenous thromboxane A2 implicated in the etiology of cerebral vasospasm, we observed significant increases in contractile force, RhoA activation, regulatory light chain phosphorylation, as well as phosphorylation of MYPT1 at Thr-696, Thr-853, and surprisingly Ser-695. Inhibition of nitric oxide signaling completely abrogated basal MYPT1 Ser-695 phosphorylation and significantly increased and potentiated U-46619-induced MYPT1 Thr-853 phosphorylation and contractile force, indicating that NO/cGMP/cGKI signaling maintains basal vascular tone through active inhibition of calcium sensitization. Surprisingly, a fall in Ser-695 phosphorylation did not result in an increase in phosphorylation of the Thr-696 site. Although activation of cGKI with exogenous cyclic nucleotides inhibited thromboxane A2-induced MYPT1 membrane association, RhoA activation, contractile force, and regulatory light chain phosphorylation, the anticipated decreases in MYPT1 phosphorylation at Thr-696/Thr-853 were not observed, indicating that the vasorelaxant effects of cGKI are not through dephosphorylation of MYPT1. Thus, thromboxane A2 signaling within the intact cerebral vasculature induces "buffered" vasoconstrictions, in which both the RhoA/Rho-kinase calcium-sensitizing and the NO/cGMP/cGKI calcium-desensitizing pathways are activated.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xiong, Shanshan; Pan, Xiujie; Xu, Long

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 onmore » 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.« less

Architectural and functional commonalities between enhancers and promoters

PubMed Central

Kim, Tae-Kyung; Shiekhattar, Ramin

2015-01-01

Summary With the explosion of genome-wide studies of regulated transcription, it has become clear that traditional definitions of enhancers and promoters need to be revisited. These control elements can now be characterized in terms of their local and regional architecture, their regulatory components including histone modifications and associated binding factors and their functional contribution to transcription. This review discusses unifying themes between promoters and enhancers in transcriptional regulatory mechanisms. PMID:26317464

The impact of transposable elements on mammalian development.

PubMed

Garcia-Perez, Jose L; Widmann, Thomas J; Adams, Ian R

2016-11-15

Despite often being classified as selfish or junk DNA, transposable elements (TEs) are a group of abundant genetic sequences that have a significant impact on mammalian development and genome regulation. In recent years, our understanding of how pre-existing TEs affect genome architecture, gene regulatory networks and protein function during mammalian embryogenesis has dramatically expanded. In addition, the mobilization of active TEs in selected cell types has been shown to generate genetic variation during development and in fully differentiated tissues. Importantly, the ongoing domestication and evolution of TEs appears to provide a rich source of regulatory elements, functional modules and genetic variation that fuels the evolution of mammalian developmental processes. Here, we review the functional impact that TEs exert on mammalian developmental processes and discuss how the somatic activity of TEs can influence gene regulatory networks. © 2016. Published by The Company of Biologists Ltd.

[T-lymphocytes--do they control rheumatic immune responses?].

PubMed

Wagner, U; Schulze-Koops, H

2005-09-01

T cells, in particular CD4(+) T cells, have been implicated in mediating many aspects of rheumatoid inflammation. In rheumatoid arthritis (RA), CD4(+) T cells display various functional abnormalities in the synovium as well as in the peripheral circulation. Current evidence suggests, however, that the role of CD4(+) T cells in the development of rheumatoid inflammation exceeds that of activated pro-inflammatory effector T cells that drive the chronic autoimmune response. Subsets of CD4(+) T cells with regulatory capacity, such as CD25(+) Tregs, have been identified in mice and man, and recent observations suggest that in RA, the function of these regulatory T cells is severely impaired. Thus, in RA, defective regulatory immune mechanisms might allow the breakdown of peripheral tolerance, following which the detrimental CD4(+) T-cell-driven immune response evolves and proceeds to chronic inflammation. Here, we review the functional abnormalities and the contribution of different T-cell subsets to rheumatoid inflammation.

Principles of dynamical modularity in biological regulatory networks

PubMed Central

Deritei, Dávid; Aird, William C.; Ercsey-Ravasz, Mária; Regan, Erzsébet Ravasz

2016-01-01

Intractable diseases such as cancer are associated with breakdown in multiple individual functions, which conspire to create unhealthy phenotype-combinations. An important challenge is to decipher how these functions are coordinated in health and disease. We approach this by drawing on dynamical systems theory. We posit that distinct phenotype-combinations are generated by interactions among robust regulatory switches, each in control of a discrete set of phenotypic outcomes. First, we demonstrate the advantage of characterizing multi-switch regulatory systems in terms of their constituent switches by building a multiswitch cell cycle model which points to novel, testable interactions critical for early G2/M commitment to division. Second, we define quantitative measures of dynamical modularity, namely that global cell states are discrete combinations of switch-level phenotypes. Finally, we formulate three general principles that govern the way coupled switches coordinate their function. PMID:26979940

Increase of CTGF mRNA expression by respiratory syncytial virus infection is abrogated by caffeine in lung epithelial cells.

PubMed

Kunzmann, Steffen; Krempl, Christine; Seidenspinner, Silvia; Glaser, Kirsten; Speer, Christian P; Fehrholz, Markus

2018-04-16

Respiratory syncytial virus (RSV) is a leading cause of severe lower respiratory tract infection in early childhood. Underlying pathomechanisms of elevated pulmonary morbidity in later infancy are largely unknown. We found that RSV-infected H441 cells showed increased mRNA expression of connective tissue growth factor (CTGF), a key factor in airway remodeling. Additional dexamethasone treatment led to further elevated mRNA levels, indicating additive effects. Caffeine treatment prevented RSV-mediated increase of CTGF mRNA. RSV may be involved in airway remodeling processes by increasing CTGF mRNA expression. Caffeine might abrogate these negative effects and thereby help to restore lung homeostasis. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Molecular switch-like regulation in motor proteins.

PubMed

Tafoya, Sara; Bustamante, Carlos

2018-06-19

Motor proteins are powered by nucleotide hydrolysis and exert mechanical work to carry out many fundamental biological tasks. To ensure their correct and efficient performance, the motors' activities are allosterically regulated by additional factors that enhance or suppress their NTPase activity. Here, we review two highly conserved mechanisms of ATP hydrolysis activation and repression operating in motor proteins-the glutamate switch and the arginine finger-and their associated regulatory factors. We examine the implications of these regulatory mechanisms in proteins that are formed by multiple ATPase subunits. We argue that the regulatory mechanisms employed by motor proteins display features similar to those described in small GTPases, which require external regulatory elements, such as dissociation inhibitors, exchange factors and activating proteins, to switch the protein's function 'on' and 'off'. Likewise, similar regulatory roles are taken on by the motor's substrate, additional binding factors, and even adjacent subunits in multimeric complexes. However, in motor proteins, more than one regulatory factor and the two mechanisms described here often underlie the machine's operation. Furthermore, ATPase regulation takes place throughout the motor's cycle, which enables a more complex function than the binary 'active' and 'inactive' states.This article is part of a discussion meeting issue 'Allostery and molecular machines'. © 2018 The Author(s).

Evolutionary conservation of regulatory elements in vertebrate HOX gene clusters

DOE Office of Scientific and Technical Information (OSTI.GOV)

Santini, Simona; Boore, Jeffrey L.; Meyer, Axel

2003-12-31

Due to their high degree of conservation, comparisons of DNA sequences among evolutionarily distantly-related genomes permit to identify functional regions in noncoding DNA. Hox genes are optimal candidate sequences for comparative genome analyses, because they are extremely conserved in vertebrates and occur in clusters. We aligned (Pipmaker) the nucleotide sequences of HoxA clusters of tilapia, pufferfish, striped bass, zebrafish, horn shark, human and mouse (over 500 million years of evolutionary distance). We identified several highly conserved intergenic sequences, likely to be important in gene regulation. Only a few of these putative regulatory elements have been previously described as being involvedmore » in the regulation of Hox genes, while several others are new elements that might have regulatory functions. The majority of these newly identified putative regulatory elements contain short fragments that are almost completely conserved and are identical to known binding sites for regulatory proteins (Transfac). The conserved intergenic regions located between the most rostrally expressed genes in the developing embryo are longer and better retained through evolution. We document that presumed regulatory sequences are retained differentially in either A or A clusters resulting from a genome duplication in the fish lineage. This observation supports both the hypothesis that the conserved elements are involved in gene regulation and the Duplication-Deletion-Complementation model.« less

Elements in the transcriptional regulatory region flanking herpes simplex virus type 1 oriS stimulate origin function.

PubMed

Wong, S W; Schaffer, P A

1991-05-01

Like other DNA-containing viruses, the three origins of herpes simplex virus type 1 (HSV-1) DNA replication are flanked by sequences containing transcriptional regulatory elements. In a transient plasmid replication assay, deletion of sequences comprising the transcriptional regulatory elements of ICP4 and ICP22/47, which flank oriS, resulted in a greater than 80-fold decrease in origin function compared with a plasmid, pOS-822, which retains these sequences. In an effort to identify specific cis-acting elements responsible for this effect, we conducted systematic deletion analysis of the flanking region with plasmid pOS-822 and tested the resulting mutant plasmids for origin function. Stimulation by cis-acting elements was shown to be both distance and orientation dependent, as changes in either parameter resulted in a decrease in oriS function. Additional evidence for the stimulatory effect of flanking sequences on origin function was demonstrated by replacement of these sequences with the cytomegalovirus immediate-early promoter, resulting in nearly wild-type levels of oriS function. In competition experiments, cotransfection of cells with the test plasmid, pOS-822, and increasing molar concentrations of a competitor plasmid which contained the ICP4 and ICP22/47 transcriptional regulatory regions but lacked core origin sequences resulted in a significant reduction in the replication efficiency of pOS-822, demonstrating that factors which bind specifically to the oriS-flanking sequences are likely involved as auxiliary proteins in oriS function. Together, these studies demonstrate that trans-acting factors and the sites to which they bind play a critical role in the efficiency of HSV-1 DNA replication from oriS in transient-replication assays.

Plasticity and innovation of regulatory mechanisms underlying seed oil content mediated by duplicated genes in the palaeopolyploid soybean.

PubMed

Zhang, Dajian; Zhao, Meixia; Li, Shuai; Sun, Lianjun; Wang, Weidong; Cai, Chunmei; Dierking, Emily C; Ma, Jianxin

2017-06-01

Many plants have undergone whole genome duplication (WGD). However, how regulatory networks underlying a particular trait are reshaped in polyploids has not been experimentally investigated. Here we show that the regulatory pathways modulating seed oil content, which involve WRINKLED1 (WRI1), LEAFY COTYLEDON1 (LEC1), and LEC2 in Arabidopsis, have been modified in the palaeopolyploid soybean. Such modifications include functional reduction of GmWRI1b of the GmWRI1a/GmWRI1b homoeologous pair relevant to WRI1, complementary non-allelic dosage effects of the GmLEC1a/GmLEC1b homoeologous pair relevant to LEC1, pseudogenization of the singleton GmLEC2 relevant to LEC2, and the rise of the LEC2-like function of GmABI3b, contrasting to its homoeolog GmABI3a, which maintains the ABSCISIC ACID INSENSITIVE 3 (ABI3)-like function in modulating seed maturation and dormancy. The function of GmABI3b in modulating seed oil biosynthesis was fulfilled by direct binding to a RY (CATGCA) cis-regulatory element in the GmWRI1a promoter, which was absent in the GmWRI1b promoter, resulting in reduction of the GmWRI1b expression. Nevertheless, the three regulators each exhibited similar intensities of purifying selection to their respective duplicates since these pairs were formed by a WGD event that is proposed to have occurred approximately 13 million years ago (mya), suggesting that the differentiation in spatiotemporal expression between the duplicated genes is more likely to be the outcome of neutral variation in regulatory sequences. This study thus exemplifies the plasticity, dynamics, and novelty of regulatory networks mediated by WGD. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

10 CFR 51.71 - Draft environmental impact statement-contents.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 10 Energy 2 2011-01-01 2011-01-01 false Draft environmental impact statement-contents. 51.71 Section 51.71 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.11 - Relationship to other subparts. [Reserved

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Relationship to other subparts. [Reserved] 51.11 Section 51.11 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.110 - [Reserved

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false [Reserved] 51.110 Section 51.110 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2) Rulemaking § 51.110...

10 CFR 51.13 - Emergencies.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Emergencies. 51.13 Section 51.13 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2) § 51.13 Emergencies...

10 CFR 51.41 - Requirement to submit environmental information.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Requirement to submit environmental information. 51.41 Section 51.41 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.31 - Determinations based on environmental assessment.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Determinations based on environmental assessment. 51.31 Section 51.31 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.88 - Proposals for legislation.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 10 Energy 2 2014-01-01 2014-01-01 false Proposals for legislation. 51.88 Section 51.88 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2...

10 CFR 51.31 - Determinations based on environmental assessment.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Determinations based on environmental assessment. 51.31 Section 51.31 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.5 - Interpretations.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Interpretations. 51.5 Section 51.5 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS § 51.5 Interpretations. Except as specifically authorized by the Commission in writing...

10 CFR 51.5 - Interpretations.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Interpretations. 51.5 Section 51.5 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS § 51.5 Interpretations. Except as specifically authorized by the Commission in writing...

10 CFR 51.125 - Responsible official.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 10 Energy 2 2014-01-01 2014-01-01 false Responsible official. 51.125 Section 51.125 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2) Responsible...

10 CFR 51.17 - Information collection requirements; OMB approval.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Information collection requirements; OMB approval. 51.17 Section 51.17 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.67 - Environmental information concerning geologic repositories.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Environmental information concerning geologic repositories. 51.67 Section 51.67 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act...

10 CFR 51.41 - Requirement to submit environmental information.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Requirement to submit environmental information. 51.41 Section 51.41 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.90 - Final environmental impact statement-general.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 10 Energy 2 2014-01-01 2014-01-01 false Final environmental impact statement-general. 51.90 Section 51.90 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.17 - Information collection requirements; OMB approval.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Information collection requirements; OMB approval. 51.17 Section 51.17 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.55 - Environmental report-standard design certification.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Environmental report-standard design certification. 51.55 Section 51.55 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.13 - Emergencies.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Emergencies. 51.13 Section 51.13 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2) § 51.13 Emergencies...

10 CFR 51.110 - [Reserved

Code of Federal Regulations, 2014 CFR

2014-01-01

... 10 Energy 2 2014-01-01 2014-01-01 false [Reserved] 51.110 Section 51.110 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2) Rulemaking § 51.110...

10 CFR 51.91 - Final environmental impact statement-contents.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 10 Energy 2 2014-01-01 2014-01-01 false Final environmental impact statement-contents. 51.91 Section 51.91 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.11 - Relationship to other subparts. [Reserved

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 2 2013-01-01 2013-01-01 false Relationship to other subparts. [Reserved] 51.11 Section 51.11 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations...

10 CFR 51.66 - Environmental report-number of copies; distribution.

Code of Federal Regulations, 2010 CFR

2010-01-01

... Section 51.66 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations... submit to the Director of Nuclear Material Safety and Safeguards an environmental report or any...

10 CFR 51.125 - Responsible official.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 2 2010-01-01 2010-01-01 false Responsible official. 51.125 Section 51.125 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) ENVIRONMENTAL PROTECTION REGULATIONS FOR DOMESTIC LICENSING AND RELATED REGULATORY FUNCTIONS National Environmental Policy Act-Regulations Implementing Section 102(2) Responsible...

Regulatory immune cells in regulation of intestinal inflammatory response to microbiota

PubMed Central

Cong, Y; Liu, Z

2015-01-01

The intestinal lumen harbors nearly 100 trillion commensal bacteria that exert crucial function for health. An elaborate balance between immune responses and tolerance to intestinal microbiota is required to maintain intestinal homeostasis. This process depends on diverse regulatory mechanisms, including both innate and adaptive immunity. Dysregulation of the homeostasis between intestinal immune systems and microbiota has been shown to be associated with the development of inflammatory bowel diseases (IBD) in genetically susceptible populations. In this review, we discuss the recent progress reported in studies of distinct types of regulatory immune cells in the gut, including intestinal intraepithelial lymphocytes, Foxp3+ regulatory T cells, regulatory B cells, alternatively activated macrophages, dendritic cells, and innate lymphoid cells, and how dysfunction of this immune regulatory system contributes to intestinal diseases such as IBD. Moreover, we discuss the manipulation of these regulatory immune cells as a potential therapeutic method for management of intestinal inflammatory disorders. PMID:26080708

Regulatory immune cells in regulation of intestinal inflammatory response to microbiota.

PubMed

Sun, M; He, C; Cong, Y; Liu, Z

2015-09-01

The intestinal lumen harbors nearly 100 trillion commensal bacteria that exert crucial function for health. An elaborate balance between immune responses and tolerance to intestinal microbiota is required to maintain intestinal homeostasis. This process depends on diverse regulatory mechanisms, including both innate and adaptive immunity. Dysregulation of the homeostasis between intestinal immune systems and microbiota has been shown to be associated with the development of inflammatory bowel diseases (IBD) in genetically susceptible populations. In this review, we discuss the recent progress reported in studies of distinct types of regulatory immune cells in the gut, including intestinal intraepithelial lymphocytes, Foxp3(+) regulatory T cells, regulatory B cells, alternatively activated macrophages, dendritic cells, and innate lymphoid cells, and how dysfunction of this immune regulatory system contributes to intestinal diseases such as IBD. Moreover, we discuss the manipulation of these regulatory immune cells as a potential therapeutic method for management of intestinal inflammatory disorders.