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Sample records for positively regulates euchromatic

  1. Juxtaposition of heterochromatic and euchromatic regions by chromosomal translocation mediates a heterochromatic long-range position effect associated with a severe neurological phenotype

    PubMed Central

    2012-01-01

    Background The term "position effect" is used when the expression of a gene is deleteriously affected by an alteration in its chromosomal environment even though the integrity of the protein coding sequences is maintained. We describe a patient affected by epilepsy and severe neurodevelopment delay carrying a balanced translocation t(15;16)(p11.2;q12.1)dn that we assume caused a position effect as a result of the accidental juxtaposition of heterochromatin in the euchromatic region. Results FISH mapped the translocation breakpoints (bkps) to 15p11.2 within satellite III and the 16q12.1 euchromatic band within the ITFG1 gene. The expression of the genes located on both sides of the translocation were tested by means of real-time PCR and three, all located on der(16), were found to be variously perturbed: the euchromatic gene NETO2/BTCL2 was silenced, whereas VPS35 and SHCBP1, located within the major heterochromatic block of chromosome 16q11.2, were over-expressed. Pyrosequencing and chromatin immunoprecipitation of NETO2/BTCL2 and VPS35 confirmed the expression findings. Interphase FISH analysis showed that der(16) localised to regions occupied by the beta satellite heterochromatic blocks more frequently than der(15). Conclusions To the best of our knowledge, this is the first report of a heterochromatic position effect in humans caused by the juxtaposition of euchromatin/heterochromatin as a result of chromosomal rearrangement. The overall results are fully in keeping with the observations in Drosophila and suggest the occurrence of a human heterochromatin position effect associated with the nuclear repositioning of the der(16) and its causative role in the patient's syndromic phenotype. PMID:22475481

  2. PHD Finger Recognition of Unmodified Histone H3R2 Links UHRF1 to Regulation of Euchromatic Gene Expression

    SciTech Connect

    E Rajakumara; Z Wang; H Ma; L Hu; H Chen; Y Lin; R Guo; F Wu; H Li; et al.

    2011-12-31

    Histone methylation occurs on both lysine and arginine residues, and its dynamic regulation plays a critical role in chromatin biology. Here we identify the UHRF1 PHD finger (PHD{sub UHRF1}), an important regulator of DNA CpG methylation, as a histone H3 unmodified arginine 2 (H3R2) recognition modality. This conclusion is based on binding studies and cocrystal structures of PHD{sub UHRF1} bound to histone H3 peptides, where the guanidinium group of unmodified R2 forms an extensive intermolecular hydrogen bond network, with methylation of H3R2, but not H3K4 or H3K9, disrupting complex formation. We have identified direct target genes of UHRF1 from microarray and ChIP studies. Importantly, we show that UHRF1's ability to repress its direct target gene expression is dependent on PHD{sub UHRF1} binding to unmodified H3R2, thereby demonstrating the functional importance of this recognition event and supporting the potential for crosstalk between histone arginine methylation and UHRF1 function.

  3. PHD Finger Recognition of Unmodified Histone H3R2 Links UHRF1 to Regulation of Euchromatic Gene Expression

    SciTech Connect

    Rajakumara, Eerappa; Wang, Zhentian; Ma, Honghui; Hu, Lulu; Chen, Hao; Lin, Yan; Guo, Rui; Wu, Feizhen; Li, Haitao; Lan, Fei; Shi, Yujiang Geno; Xu, Yanhui; Patel, Dinshaw J.; Shi, Yang

    2011-08-29

    Histone methylation occurs on both lysine and arginine residues, and its dynamic regulation plays a critical role in chromatin biology. Here we identify the UHRF1 PHD finger (PHD{sub UHRF1}), an important regulator of DNA CpG methylation, as a histone H3 unmodified arginine 2 (H3R2) recognition modality. This conclusion is based on binding studies and cocrystal structures of PHD{sub UHRF1} bound to histone H3 peptides, where the guanidinium group of unmodified R2 forms an extensive intermolecular hydrogen bond network, with methylation of H3R2, but not H3K4 or H3K9, disrupting complex formation. We have identified direct target genes of UHRF1 from microarray and ChIP studies. Importantly, we show that UHRF1's ability to repress its direct target gene expression is dependent on PHD{sub UHRF1} binding to unmodified H3R2, thereby demonstrating the functional importance of this recognition event and supporting the potential for crosstalk between histone arginine methylation and UHRF1 function.

  4. Finishing The Euchromatic Sequence Of The Human Genome

    SciTech Connect

    Rubin, Edward M.; Lucas, Susan; Richardson, Paul; Rokhsar, Daniel; Pennacchio, Len

    2004-09-07

    The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process.The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers {approx}99% of the euchromatic genome and is accurate to an error rate of {approx}1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number,birth and death. Notably, the human genome seems to encode only20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead.

  5. Positively regulated bacterial expression systems

    PubMed Central

    Brautaset, Trygve; Lale, Rahmi; Valla, Svein

    2009-01-01

    Summary Regulated promoters are useful tools for many aspects related to recombinant gene expression in bacteria, including for high‐level expression of heterologous proteins and for expression at physiological levels in metabolic engineering applications. In general, it is common to express the genes of interest from an inducible promoter controlled either by a positive regulator or by a repressor protein. In this review, we discuss established and potentially useful positively regulated bacterial promoter systems, with a particular emphasis on those that are controlled by the AraC‐XylS family of transcriptional activators. The systems function in a wide range of microorganisms, including enterobacteria, soil bacteria, lactic bacteria and streptomycetes. The available systems that have been applied to express heterologous genes are regulated either by sugars (l‐arabinose, l‐rhamnose, xylose and sucrose), substituted benzenes, cyclohexanone‐related compounds, ε‐caprolactam, propionate, thiostrepton, alkanes or peptides. It is of applied interest that some of the inducers require the presence of transport systems, some are more prone than others to become metabolized by the host and some have been applied mainly in one or a limited number of species. Based on bioinformatics analyses, the AraC‐XylS family of regulators contains a large number of different members (currently over 300), but only a small fraction of these, the XylS/Pm, AraC/PBAD, RhaR‐RhaS/rhaBAD, NitR/PnitA and ChnR/Pb regulator/promoter systems, have so far been explored for biotechnological applications. PMID:21261879

  6. Directly transmitted unbalanced chromosome abnormalities and euchromatic variants

    PubMed Central

    Barber, J

    2005-01-01

    In total, 200 families were reviewed with directly transmitted, cytogenetically visible unbalanced chromosome abnormalities (UBCAs) or euchromatic variants (EVs). Both the 130 UBCA and 70 EV families were divided into three groups depending on the presence or absence of an abnormal phenotype in parents and offspring. No detectable phenotypic effect was evident in 23/130 (18%) UBCA families ascertained mostly through prenatal diagnosis (group 1). In 30/130 (23%) families, the affected proband had the same UBCA as other phenotypically normal family members (group 2). In the remaining 77/130 (59%) families, UBCAs had consistently mild consequences (group 3). In the 70 families with established EVs of 8p23.1, 9p12, 9q12, 15q11.2, and 16p11.2, no phenotypic effect was apparent in 38/70 (54%). The same EV was found in affected probands and phenotypically normal family members in 30/70 families (43%) (group 2), and an EV co-segregated with mild phenotypic anomalies in only 2/70 (3%) families (group 3). Recent evidence indicates that EVs involve copy number variation of common paralogous gene and pseudogene sequences that are polymorphic in the normal population and only become visible at the cytogenetic level when copy number is high. The average size of the deletions and duplications in all three groups of UBCAs was close to 10 Mb, and these UBCAs and EVs form the "Chromosome Anomaly Collection" at http://www.ngrl.org.uk/Wessex/collection. The continuum of severity associated with UBCAs and the variability of the genome at the sub-cytogenetic level make further close collaboration between medical and laboratory staff essential to distinguish clinically silent variation from pathogenic rearrangement. PMID:16061560

  7. Drosophila Sir2 is required for heterochromatic silencing and by euchromatic Hairy/E(Spl) bHLH repressors in segmentation and sex determination.

    PubMed

    Rosenberg, Miriam I; Parkhurst, Susan M

    2002-05-17

    Yeast SIR2 is a NAD+-dependent histone deacetylase required for heterochromatic silencing at telomeres, rDNA, and mating-type loci. We find that the Drosophila homolog of Sir2 (dSir2) also encodes deacetylase activity and is required for heterochromatic silencing, but unlike ySir2, is not required for silencing at telomeres. We show that dSir2 interacts genetically and physically with members of the Hairy/Deadpan/E(Spl) family of bHLH euchromatic repressors, key regulators of Drosophila development. dSir2 is an essential gene whose loss of function results in both segmentation defects and skewed sex ratios, associated with reduced activities of the Hairy and Deadpan bHLH repressors. These results indicate that Sir2 in higher organisms plays an essential role in both euchromatic repression and heterochromatic silencing.

  8. Stomagen positively regulates stomatal density in Arabidopsis.

    PubMed

    Sugano, Shigeo S; Shimada, Tomoo; Imai, Yu; Okawa, Katsuya; Tamai, Atsushi; Mori, Masashi; Hara-Nishimura, Ikuko

    2010-01-14

    Stomata in the epidermal tissues of leaves are valves through which passes CO(2), and as such they influence the global carbon cycle. The two-dimensional pattern and density of stomata in the leaf epidermis are genetically and environmentally regulated to optimize gas exchange. Two putative intercellular signalling factors, EPF1 and EPF2, function as negative regulators of stomatal development in Arabidopsis, possibly by interacting with the receptor-like protein TMM. One or more positive intercellular signalling factors are assumed to be involved in stomatal development, but their identities are unknown. Here we show that a novel secretory peptide, which we designate as stomagen, is a positive intercellular signalling factor that is conserved among vascular plants. Stomagen is a 45-amino-rich peptide that is generated from a 102-amino-acid precursor protein designated as STOMAGEN. Both an in planta analysis and a semi-in-vitro analysis with recombinant and chemically synthesized stomagen peptides showed that stomagen has stomata-inducing activity in a dose-dependent manner. A genetic analysis showed that TMM is epistatic to STOMAGEN (At4g12970), suggesting that stomatal development is finely regulated by competitive binding of positive and negative regulators to the same receptor. Notably, STOMAGEN is expressed in inner tissues (the mesophyll) of immature leaves but not in the epidermal tissues where stomata develop. This study provides evidence of a mesophyll-derived positive regulator of stomatal density. Our findings provide a conceptual advancement in understanding stomatal development: inner photosynthetic tissues optimize their function by regulating stomatal density in the epidermis for efficient uptake of CO(2).

  9. Euchromatic Subdomains in Rice Centromeres Are Associated with Genes and Transcription[W

    PubMed Central

    Wu, Yufeng; Kikuchi, Shinji; Yan, Huihuang; Zhang, Wenli; Rosenbaum, Heidi; Iniguez, A. Leonardo; Jiang, Jiming

    2011-01-01

    The presence of the centromere-specific histone H3 variant, CENH3, defines centromeric (CEN) chromatin, but poorly understood epigenetic mechanisms determine its establishment and maintenance. CEN chromatin is embedded within pericentromeric heterochromatin in most higher eukaryotes, but, interestingly, it can show euchromatic characteristics; for example, the euchromatic histone modification mark dimethylated H3 Lys 4 (H3K4me2) is uniquely associated with animal centromeres. To examine the histone marks and chromatin properties of plant centromeres, we developed a genomic tiling array for four fully sequenced rice (Oryza sativa) centromeres and used chromatin immunoprecipitation–chip to study the patterns of four euchromatic histone modification marks: H3K4me2, trimethylated H3 Lys 4, trimethylated H3 Lys 36, and acetylated H3 Lys 4, 9. The vast majority of the four histone marks were associated with genes located in the H3 subdomains within the centromere cores. We demonstrate that H3K4me2 is not a ubiquitous component of rice CEN chromatin, and the euchromatic characteristics of rice CEN chromatin are hallmarks of the transcribed sequences embedded in the centromeric H3 subdomains. We propose that the transcribed sequences located in rice centromeres may provide a barrier preventing loading of CENH3 into the H3 subdomains. The separation of CENH3 and H3 subdomains in the centromere core may be favorable for the formation of three-dimensional centromere structure and for rice centromere function. PMID:22080597

  10. Euchromatic histone methyltransferase 2 inhibitor, BIX-01294, sensitizes human promyelocytic leukemia HL-60 and NB4 cells to growth inhibition and differentiation.

    PubMed

    Savickiene, Jurate; Treigyte, Grazina; Stirblyte, Ieva; Valiuliene, Giedre; Navakauskiene, Ruta

    2014-07-01

    The involvement of histone lysine methyltransferases (HMT) in carcinogenesis is not well understood. Here, we describe a dose-dependent growth and survival inhibitory effects of BIX-01294, a specific inhibitor of euchromatic HMT2, in promyelocytic leukemia HL-60 and NB4 cells. BIX-01294 combined with all-trans retinoic acid or together with histone deacetylase and DNA methyltransferase inhibitors enhanced cell differentiation to granulocytes and induced cell line-specific changes in the expression of cell cycle-, survival- and differentiation regulating genes and proteins in association with histone modification state. Our results suggest that targeting EHMT2 may be of therapeutical benefits in myeloid leukemia.

  11. G9a histone methyltransferase plays a dominant role in euchromatic histone H3 lysine 9 methylation and is essential for early embryogenesis.

    PubMed

    Tachibana, Makoto; Sugimoto, Kenji; Nozaki, Masami; Ueda, Jun; Ohta, Tsutomu; Ohki, Misao; Fukuda, Mikiko; Takeda, Naoki; Niida, Hiroyuki; Kato, Hiroyuki; Shinkai, Yoichi

    2002-07-15

    Covalent modification of histone tails is crucial for transcriptional regulation, mitotic chromosomal condensation, and heterochromatin formation. Histone H3 lysine 9 (H3-K9) methylation catalyzed by the Suv39h family proteins is essential for establishing the architecture of pericentric heterochromatin. We recently identified a mammalian histone methyltransferase (HMTase), G9a, which has strong HMTase activity towards H3-K9 in vitro. To investigate the in vivo functions of G9a, we generated G9a-deficient mice and embryonic stem (ES) cells. We found that H3-K9 methylation was drastically decreased in G9a-deficient embryos, which displayed severe growth retardation and early lethality. G9a-deficient ES cells also exhibited reduced H3-K9 methylation compared to wild-type cells, indicating that G9a is a dominant H3-K9 HMTase in vivo. Importantly, the loss of G9a abolished methylated H3-K9 mostly in euchromatic regions. Finally, G9a exerted a transcriptionally suppressive function that depended on its HMTase activity. Our results indicate that euchromatic H3-K9 methylation regulated by G9a is essential for early embryogenesis and is involved in the transcriptional repression of developmental genes.

  12. The fourth chromosome of Drosophila melanogaster: Interspersed euchromatic and heterochromatic domains

    PubMed Central

    Sun, Fang-Lin; Cuaycong, Matthew H.; Craig, Carolyn A.; Wallrath, Lori L.; Locke, John; Elgin, Sarah C. R.

    2000-01-01

    The small fourth chromosome of Drosophila melanogaster (3.5% of the genome) presents a puzzle. Cytological analysis suggests that the bulk of the fourth, including the portion that appears banded in the polytene chromosomes, is heterochromatic; the banded region includes blocks of middle repetitious DNA associated with heterochromatin protein 1 (HP1). However, genetic screens indicate 50–75 genes in this region, a density similar to that in other euchromatic portions of the genome. Using a P element containing an hsp70-white gene and a copy of hsp26 (marked with a fragment of plant DNA designated pt), we have identified domains that allow for full expression of the white marker (R domains), and others that induce a variegating phenotype (V domains). In the former case, the hsp26-pt gene shows an accessibility and heat-shock-inducible activity similar to that seen in euchromatin, whereas in the latter case, accessibility and inducible expression are reduced to levels typical of heterochromatin. Mapping by in situ hybridization and by hybridization of flanking DNA sequences to a collection of cosmid and bacterial artificial chromosome clones shows that the R domains (euchromatin-like) and V domains (heterochromatin-like) are interspersed. Examination of the effect of genetic modifiers on the variegating transgenes shows some differences among these domains. The results suggest that heterochromatic and euchromatic domains are interspersed and closely associated within this 1.2-megabase region of the genome. PMID:10779561

  13. Molecular topography of the secondary constriction region (qh) of human chromosome 9 with an unusual euchromatic band

    SciTech Connect

    Verma, R.S.; Luk, S.; Brennan, J.P.; Mathews, T.; Conte, R.A.; Macera, M.J. )

    1993-05-01

    Heterochromatin confined to pericentromeric (c) and secondary constriction (qh) regions plays a major role in morphological variation of chromosome 9, because of its size and affinity for pericentric inversion. Consequently, pairing at pachytene may lead to some disturbances between homologous chromosomes having such extreme variations and may result in abnormalities involving bands adjacent to the qh region. The authors encountered such a case, where a G-positive band has originated de nova, suggesting a maternal origin from the chromosome 9 that has had a complete pericentric inversion. In previously reported cases, the presence of an extra G-positive band within the 9qh region has been familial, and in the majority of those cases it was not associated with any clinical consequences. Therefore, this anomaly has been referred to as a [open quotes]rare[close quotes] variant. The qh region consists of a mixture of various tandemly repeated DNA sequences, and routine banding techniques have failed to characterize the origin of this extra genetic material. By the chromosome in situ suppression hybridization technique using whole chromosome paint, the probe annealed with the extra G-band, suggesting a euchromatic origin from chromosome 9, presumably band p12. By the fluorescence in situ hybridization technique using alpha- and beta-satellite probes, the dicentric nature was further revealed, supporting the concept of unequal crossing-over during maternal meiosis I, which could account for a duplication of the h region. The G-positive band most likely became genetically inert when it was sandwiched between two blocks of heterochromatin, resulting in a phenotypically normal child. Therefore, an earlier hypothesis, suggesting its origin from heterochromatin through so-called euchromatinization, is refuted here. If the proband's progeny inherit this chromosome, it shall be envisaged as a rare familial variant whose clinical consequences remain obscure. 52 refs., 3 figs.

  14. Regulating positive and negative emotions in daily life.

    PubMed

    Nezlek, John B; Kuppens, Peter

    2008-06-01

    The present study examined how people regulate their emotions in daily life and how such regulation is related to their daily affective experience and psychological adjustment. Each day for an average of 3 weeks, participants described how they had regulated their emotions in terms of the reappraisal and suppression (inhibiting the expression) of positive and negative emotions, and they described their emotional experience, self-esteem, and psychological adjustment in terms of Beck's triadic model of depression. Reappraisal was used more often than suppression, and suppressing positive emotions was used less than the other three strategies. In general, regulation through reappraisal was found to be beneficial, whereas regulation by suppression was not. Reappraisal of positive emotions was associated with increases in positive affect, self-esteem, and psychological adjustment, whereas suppressing positive emotions was associated with decreased positive emotion, self-esteem, and psychological adjustment, and increased negative emotions. Moreover, relationships between reappraisal and psychological adjustment and self-esteem were mediated by experienced positive affect, whereas relationships between suppression of positive emotions and self-esteem adjustment were mediated by negative affect.

  15. 102. Giullotine type gate (inclosed position to regulate furnace exhaust ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    102. Giullotine type gate (inclosed position to regulate furnace exhaust gases to stoves during heating cycle. - Sloss-Sheffield Steel & Iron, First Avenue North Viaduct at Thirty-second Street, Birmingham, Jefferson County, AL

  16. Impact of multiple insertions of two retroelements, ZAM and Idefix at an euchromatic locus.

    PubMed

    Conte, C; Calco, V; Desset, S; Leblanc, P; Dastugue, B; Vaury, C

    2000-01-01

    Transposable elements represent a large fraction of eukaryotic genomes and they are thought to play an important role in chromatin structure. ZAM and Idefix are two LTR-retrotransposons from Drosophila melanogaster very similar in structure to vertebrate retroviruses. In all the strains where their distribution has been studied, ZAM appears to be present exclusively in the intercalary heterochromatin while Idefix copies are mainly found in the centromeric heterochromatin with very few copies in euchromatin. Their distribution varies in a specific strain called RevI in which the mobilization of ZAM and Idefix is highly induced. In this strain, 15 copies of ZAM and 30 copies of Idefix are found on the chromosomal arms in addition to their usual distribution. Amongst the loci where new copies are detected, a hotspot for their insertion has been detected at the white locus where up to four elements occurred within a 3-kb fragment at the 5' end of this gene. This property of ZAM and Idefix to accumulate at a defined site provides an interesting paradigm to bring insight into the effect exerted by multiple insertions of transposable elements at an euchromatic locus.

  17. An Integrative Theory-Driven Positive Emotion Regulation Intervention

    PubMed Central

    Weytens, Fanny; Luminet, Olivier; Verhofstadt, Lesley L.; Mikolajczak, Moïra

    2014-01-01

    Over the past fifteen years, positive psychology research has validated a set of happiness enhancing techniques. These techniques are relatively simple exercises that allow happiness seekers to mimic thoughts and behavior of naturally happy people, in order to increase their level of well-being. Because research has shown that the joint use of these exercises increases their effects, practitioners who want to help happiness seekers need validated interventions that combine several of these techniques. To meet this need, we have developed and tested an integrative intervention (Positive Emotion Regulation program – PER program) incorporating a number of validated techniques structured around a theoretical model: the Process Model of Positive Emotion Regulation. To test the effectiveness of this program and to identify its added value relative to existing interventions, 113 undergraduate students were randomly assigned to a 6-week positive emotion regulation pilot program, a loving-kindness meditation training program, or a wait-list control group. Results indicate that fewer participants dropped out from the PER program than from the Loving-Kindness Meditation training. Furthermore, subjects in the PER group showed a significant increase in subjective well-being and life satisfaction and a significant decrease in depression and physical symptoms when compared to controls. Our results suggest that the Process Model of Positive Emotion Regulation can be an effective option to organize and deliver positive integrative interventions. PMID:24759870

  18. An integrative theory-driven positive emotion regulation intervention.

    PubMed

    Weytens, Fanny; Luminet, Olivier; Verhofstadt, Lesley L; Mikolajczak, Moïra

    2014-01-01

    Over the past fifteen years, positive psychology research has validated a set of happiness enhancing techniques. These techniques are relatively simple exercises that allow happiness seekers to mimic thoughts and behavior of naturally happy people, in order to increase their level of well-being. Because research has shown that the joint use of these exercises increases their effects, practitioners who want to help happiness seekers need validated interventions that combine several of these techniques. To meet this need, we have developed and tested an integrative intervention (Positive Emotion Regulation program - PER program) incorporating a number of validated techniques structured around a theoretical model: the Process Model of Positive Emotion Regulation. To test the effectiveness of this program and to identify its added value relative to existing interventions, 113 undergraduate students were randomly assigned to a 6-week positive emotion regulation pilot program, a loving-kindness meditation training program, or a wait-list control group. Results indicate that fewer participants dropped out from the PER program than from the Loving-Kindness Meditation training. Furthermore, subjects in the PER group showed a significant increase in subjective well-being and life satisfaction and a significant decrease in depression and physical symptoms when compared to controls. Our results suggest that the Process Model of Positive Emotion Regulation can be an effective option to organize and deliver positive integrative interventions.

  19. CAPRICE positively regulates stomatal formation in the Arabidopsis hypocotyl.

    PubMed

    Serna, Laura

    2008-12-01

    In the Arabidopsis hypocotyl, stomata develop only from a set of epidermal cell files. Previous studies have identified several negative regulators of stomata formation. Such regulators also trigger non-hair cell fate in the root. Here, it is shown that TOO MANY MOUTHS (TMM) positively regulates CAPRICE (CPC) expression in differentiating stomaless-forming cell files, and that the CPC protein might move to the nucleus of neighbouring stoma-forming cells, where it promotes stomata formation in a redundant manner with TRIPTYCHON (TRY). Unexpectedly, the CPC protein was also localized in the nucleus and peripheral cytoplasm of hypocotyl fully differentiated epidermal cells, suggesting that CPC plays an additional role to those related to stomata formation. These results identify CPC and TRY as positive regulators of stomata formation in the embryonic stem, which increases the similarity between the genetic control of root hair and stoma cell fate determination.

  20. Studies of Normal and Position-Affected Expression of ROSY Region Genes in DROSOPHILA MELANOGASTER

    PubMed Central

    Clark, Stephen H.; Chovnick, Arthur

    1986-01-01

    Transformant complementation, intragenic deletions and Northern blot analyses provide unambiguous localization of the l(3) S12 gene immediately proximal to the 5' end of the rosy locus. We have characterized an array of transformants with respect to l( 3)S12 and rosy expression. The l(3) S12 gene is exceedingly sensitive to euchromatic site-specific position effects. Unlike the rosy locus, l(3)S12 is insensitive to heterochromatic position effect in rearrangements, as well as in a transformant located in heterochromatin. Cotransformants for both l(3)S12 and rosy elicit no apparent pattern of concordance with respect to euchromatic site-specific position effects. Heterochromatic-euchromatic rearrangements are examined with respect to position effects on expression of the rosy region genes l(3) 12, rosy, snake and piccolo, as well as suppressor effects. Clear distinction is seen between euchromatic and heterochromatic effects. PMID:3098623

  1. Regulation of positive and negative emotion: effects of sociocultural context

    PubMed Central

    Snyder, Sara A.; Heller, S. Megan; Lumian, Daniel S.; McRae, Kateri

    2013-01-01

    Previous research has demonstrated that the use of emotion regulation strategies can vary by sociocultural context. In a previous study, we reported changes in the use of two different emotion regulation strategies at an annual alternative cultural event, Burning Man (McRae et al., 2011). In this sociocultural context, as compared to typically at home, participants reported less use of expressive suppression (a strategy generally associated with maladaptive outcomes), and greater use of cognitive reappraisal (a strategy generally associated with adaptive outcomes). What remained unclear was whether these changes in self-reported emotion regulation strategy use were characterized by changes in the regulation of positive emotion, negative emotion, or both. We addressed this issue in the current study by asking Burning Man participants separate questions about positive and negative emotion. Using multiple datasets, we replicated our previous findings, and found that the decreased use of suppression is primarily driven by reports of decreased suppression of positive emotion at Burning Man. By contrast, the increased use of reappraisal is not characterized by differential reappraisal of positive and negative emotion at Burning Man. Moreover, we observed novel individual differences in the magnitude of these effects. The contextual changes in self-reported suppression that we observe are strongest for men and younger participants. For those who had previously attended Burning Man, we observed lower levels of self-reported suppression in both sociocultural contexts: Burning Man and typically at home. These findings have implications for understanding the ways in which certain sociocultural contexts may decrease suppression, and possibly minimize its associated maladaptive effects. PMID:23840191

  2. Positive cooperative regulation of double binding sites for human acetylcholinesterase.

    PubMed

    Liu, Hao; Ye, Wei; Chen, Hai-Feng

    2016-10-25

    Acetylcholinesterase is a potent enzyme that regulates neurotransmission by rapidly hydrolyzing the neurotransmitter acetylcholine in synapses of the nervous system. As drug target of anti-AD, it has catalytic and peripheral anionic sites. However, the regulation relation between these two sites is unclear. Therefore, we constructed dynamics fluctuation network based on all-atom molecular dynamics simulations to reveal the regulation mechanism. The results suggest that the correlation network in double-site system (hAChE/TZ5) is distinctly different from that in the free state and single-site systems (hAChE/huprine and hAChE/1YL). The community network analysis indicates that the information freely transfers from the peripheral anionic site to the catalytic active site in hAChE/TZ5. Furthermore, the binding free energy between the inhibitor and hAChE for hAChE/TZ5 is significantly lower than of either hAChE/huprine or hAChE/1YL. Thus, a hypothesis of 'positive cooperative regulation' is proposed for the regulation of double binding sites and further confirmed by the weakening and mutation community analyses. Finally, one possible cooperative regulation pathway of W86-TZ5-W286 was identified based on the shortest path algorithm and was confirmed by the network perturbation analysis. Interestingly, the regulation pathway for single-site systems is significantly different from that of dual-site system. The process targeting on the shortest pathway can better regulate the hydrolyzing the neurotransmitter acetylcholine and significantly inhibit the aggregation of Aβ amyloid.

  3. Compassion-based emotion regulation up-regulates experienced positive affect and associated neural networks

    PubMed Central

    Singer, Tania

    2015-01-01

    Emotion regulation research has primarily focused on techniques that attenuate or modulate the impact of emotional stimuli. Recent evidence suggests that this mode regulation can be problematic in the context of regulation of emotion elicited by the suffering of others, resulting in reduced emotional connectedness. Here, we investigated the effects of an alternative emotion regulation technique based on the up-regulation of positive affect via Compassion-meditation on experiential and neural affective responses to depictions of individuals in distress, and compared these with the established emotion regulation strategy of Reappraisal. Using fMRI, we scanned 15 expert practitioners of Compassion-meditation either passively viewing, or using Compassion-meditation or Reappraisal to modulate their emotional reactions to film clips depicting people in distress. Both strategies effectively, but differentially regulated experienced affect, with Compassion primarily increasing positive and Reappraisal primarily decreasing negative affect. Imaging results showed that Compassion, relative to both passive-viewing and Reappraisal increased activation in regions involved in affiliation, positive affect and reward processing including ventral striatum and medial orbitfrontal cortex. This network was shown to be active prior to stimulus presentation, suggesting that the regulatory mechanism of Compassion is the stimulus-independent endogenous generation of positive affect. PMID:25698699

  4. Compassion-based emotion regulation up-regulates experienced positive affect and associated neural networks.

    PubMed

    Engen, Haakon G; Singer, Tania

    2015-09-01

    Emotion regulation research has primarily focused on techniques that attenuate or modulate the impact of emotional stimuli. Recent evidence suggests that this mode regulation can be problematic in the context of regulation of emotion elicited by the suffering of others, resulting in reduced emotional connectedness. Here, we investigated the effects of an alternative emotion regulation technique based on the up-regulation of positive affect via Compassion-meditation on experiential and neural affective responses to depictions of individuals in distress, and compared these with the established emotion regulation strategy of Reappraisal. Using fMRI, we scanned 15 expert practitioners of Compassion-meditation either passively viewing, or using Compassion-meditation or Reappraisal to modulate their emotional reactions to film clips depicting people in distress. Both strategies effectively, but differentially regulated experienced affect, with Compassion primarily increasing positive and Reappraisal primarily decreasing negative affect. Imaging results showed that Compassion, relative to both passive-viewing and Reappraisal increased activation in regions involved in affiliation, positive affect and reward processing including ventral striatum and medial orbitfrontal cortex. This network was shown to be active prior to stimulus presentation, suggesting that the regulatory mechanism of Compassion is the stimulus-independent endogenous generation of positive affect.

  5. The MYB107 Transcription Factor Positively Regulates Suberin Biosynthesis

    SciTech Connect

    Gou, Mingyue; Hou, Guichuan; Yang, Huijun; Zhang, Xuebin; Cai, Yuanheng; Kai, Guoyin; Liu, Chang-Jun

    2016-12-13

    Suberin, a lipophilic polymer deposited in the outer integument of the Arabidopsis (Arabidopsis thaliana) seed coat, represents an essential sealing component controlling water and solute movement and protecting seed from pathogenic infection. Although many genes responsible for suberin synthesis are identified, the regulatory components controlling its biosynthesis have not been definitively determined. Here, we show that the Arabidopsis MYB107 transcription factor acts as a positive regulator controlling suberin biosynthetic gene expression in the seed coat. MYB107 coexpresses with suberin biosynthetic genes in a temporal manner during seed development. Disrupting MYB107 particularly suppresses the expression of genes involved in suberin but not cutin biosynthesis, lowers seed coat suberin accumulation, alters suberin lamellar structure, and consequently renders higher seed coat permeability and susceptibility to abiotic stresses. Furthermore, MYB107 directly binds to the promoters of suberin biosynthetic genes, verifying its primary role in regulating their expression. Identifying MYB107 as a positive regulator for seed coat suberin synthesis offers a basis for discovering the potential transcriptional network behind one of the most abundant lipid-based polymers in nature.

  6. Interferon gamma Signaling Positively Regulates Hematopoietic Stem Cell Emergence

    PubMed Central

    Sawamiphak, Suphansa; Kontarakis, Zacharias; Stainier, Didier Y.R.

    2014-01-01

    Summary Vertebrate hematopoietic stem cells (HSCs) emerge in the aorta-gonad-mesonephros (AGM) region from “hemogenic” endothelium. Here we show that the pro-inflammatory cytokine Ifn-γ and its receptor Crfb17 positively regulate HSC development in zebrafish. This regulation does not appear to modulate the proliferation or survival of HSCs or endothelial cells, but rather the endothelial to HSC transition. Notch signaling and blood flow positively regulate the expression of ifng and crfb17 in the AGM. Notably, Ifn-γ overexpression partially rescues the HSC loss observed in the absence of blood flow or Notch signaling. Importantly, Ifn-γ signaling acts cell-autonomously to control the endothelial to HSC transition. Ifn-γ activates Stat3, an atypical transducer of Ifn-γ signaling, in the AGM, and Stat3 inhibition decreases HSC formation. Together, our findings uncover a developmental role for an inflammatory cytokine and place its action downstream of Notch signaling and blood flow to control Stat3 activation and HSC emergence. PMID:25490269

  7. PLAP-1/Asporin Positively Regulates FGF-2 Activity.

    PubMed

    Awata, T; Yamada, S; Tsushima, K; Sakashita, H; Yamaba, S; Kajikawa, T; Yamashita, M; Takedachi, M; Yanagita, M; Kitamura, M; Murakami, S

    2015-10-01

    PLAP-1 is an extracellular matrix protein that is predominantly expressed in the periodontal ligament within periodontal tissue. It was previously revealed that PLAP-1 negatively regulates bone morphogenetic protein 2 and transforming growth factor β activity through direct interactions. However, the interaction between PLAP-1 and other growth factors has not been defined. Here, we revealed that PLAP-1 positively regulates the activity of fibroblast growth factor 2 (FGF-2), a critical growth factor in tissue homeostasis and repair. In this study, we isolated mouse embryonic fibroblasts (MEFs) from Plap-1(-/-) mice generated in our laboratory. Interestingly, Plap-1(-/-) MEFs exhibited enhanced responses to bone morphogenetic protein 2 but defective responses to FGF-2, and Plap-1 transfection into Plap-1(-/-) MEFs rescued these defective responses. In addition, binding assays revealed that PLAP-1 promotes FGF-2-FGF receptor 1 (FGFR1) complex formation by direct binding to FGF-2. Immunocytochemistry analyses revealed colocalization of PLAP-1 and FGF-2 in wild-type MEFs and reduced colocalization of FGF-2 and FGFR1 in Plap-1(-/-) MEFs compared with wild-type MEFs. Taken together, PLAP-1 positively regulates FGF-2 activity through a direct interaction. Extracellular matrix-growth factor interactions have considerable effects; thus, this approach may be useful in several regenerative medicine applications.

  8. The MYB107 Transcription Factor Positively Regulates Suberin Biosynthesis

    DOE PAGES

    Gou, Mingyue; Hou, Guichuan; Yang, Huijun; ...

    2016-12-13

    Suberin, a lipophilic polymer deposited in the outer integument of the Arabidopsis (Arabidopsis thaliana) seed coat, represents an essential sealing component controlling water and solute movement and protecting seed from pathogenic infection. Although many genes responsible for suberin synthesis are identified, the regulatory components controlling its biosynthesis have not been definitively determined. Here, we show that the Arabidopsis MYB107 transcription factor acts as a positive regulator controlling suberin biosynthetic gene expression in the seed coat. MYB107 coexpresses with suberin biosynthetic genes in a temporal manner during seed development. Disrupting MYB107 particularly suppresses the expression of genes involved in suberin butmore » not cutin biosynthesis, lowers seed coat suberin accumulation, alters suberin lamellar structure, and consequently renders higher seed coat permeability and susceptibility to abiotic stresses. Furthermore, MYB107 directly binds to the promoters of suberin biosynthetic genes, verifying its primary role in regulating their expression. Identifying MYB107 as a positive regulator for seed coat suberin synthesis offers a basis for discovering the potential transcriptional network behind one of the most abundant lipid-based polymers in nature.« less

  9. Quorum sensing positively regulates flagellar motility in pathogenic Vibrio harveyi.

    PubMed

    Yang, Qian; Defoirdt, Tom

    2015-04-01

    Vibrios belonging to the Harveyi clade are among the major pathogens of aquatic organisms. Quorum sensing (QS) is essential for virulence of V. harveyi towards different hosts. However, most virulence factors reported to be controlled by QS to date are negatively regulated by QS, therefore suggesting that their impact on virulence is limited. In this study, we report that QS positively regulates flagellar motility. We found that autoinducer synthase mutants showed significantly lower swimming motility than the wild type, and the swimming motility could be restored by adding synthetic signal molecules. Further, motility of a luxO mutant with inactive QS (LuxO D47E) was significantly lower than that of the wild type and of a luxO mutant with constitutively maximal QS activity (LuxO D47A). Furthermore, we found that the expression of flagellar genes (both early, middle and late genes) was significantly lower in the luxO mutant with inactive QS when compared with wild type and the luxO mutant with maximal QS activity. Motility assays and gene expression also revealed the involvement of the quorum-sensing master regulator LuxR in the QS regulation of motility. Finally, the motility inhibitor phenamil significantly decreased the virulence of V. harveyi towards gnotobiotic brine shrimp larvae.

  10. HMGCR positively regulated the growth and migration of glioblastoma cells.

    PubMed

    Qiu, Zhihua; Yuan, Wen; Chen, Tao; Zhou, Chenzhi; Liu, Chao; Huang, Yongkai; Han, Deqing; Huang, Qinghui

    2016-01-15

    The metabolic program of cancer cells is significant different from the normal cells, which makes it possible to develop novel strategies targeting cancer cells. Mevalonate pathway and its rate-limiting enzyme HMG-CoA reductase (HMGCR) have shown important roles in the progression of several cancer types. However, their roles in glioblastoma cells remain unknown. In this study, up-regulation of HMGCR in the clinical glioblastoma samples was observed. Forced expression of HMGCR promoted the growth and migration of U251 and U373 cells, while knocking down the expression of HMGCR inhibited the growth, migration and metastasis of glioblastoma cells. Molecular mechanism studies revealed that HMGCR positively regulated the expression of TAZ, an important mediator of Hippo pathway, and the downstream target gene connective tissue growth factor (CTGF), suggesting HMGCR might activate Hippo pathway in glioblastoma cells. Taken together, our study demonstrated the oncogenic roles of HMGCR in glioblastoma cells and HMGCR might be a promising therapeutic target.

  11. Autophagy positively regulates DNA damage recognition by nucleotide excision repair.

    PubMed

    Qiang, Lei; Zhao, Baozhong; Shah, Palak; Sample, Ashley; Yang, Seungwon; He, Yu-Ying

    2016-01-01

    Macroautophagy (hereafter autophagy) is a cellular catabolic process that is essential for maintaining tissue homeostasis and regulating various normal and pathologic processes in human diseases including cancer. One cancer-driving process is accumulation of genetic mutations due to impaired DNA damage repair, including nucleotide excision repair. Here we show that autophagy positively regulates nucleotide excision repair through enhancing DNA damage recognition by the DNA damage sensor proteins XPC and DDB2 via 2 pathways. First, autophagy deficiency downregulates the transcription of XPC through TWIST1-dependent activation of the transcription repressor complex E2F4-RBL2. Second, autophagy deficiency impairs the recruitment of DDB2 to ultraviolet radiation (UV)-induced DNA damage sites through TWIST1-mediated inhibition of EP300. In mice, the pharmacological autophagy inhibitor Spautin-1 promotes UVB-induced tumorigenesis, whereas the autophagy inducer rapamycin reduces UVB-induced tumorigenesis. These findings demonstrate the crucial role of autophagy in maintaining proper nucleotide excision repair in mammalian cells and suggest a previously unrecognized tumor-suppressive mechanism of autophagy in cancer.

  12. LATS2 Positively Regulates Polycomb Repressive Complex 2

    PubMed Central

    Torigata, Kosuke; Daisuke, Okuzaki; Mukai, Satomi; Hatanaka, Akira; Ohka, Fumiharu; Motooka, Daisuke; Nakamura, Shota; Ohkawa, Yasuyuki; Yabuta, Norikazu; Kondo, Yutaka; Nojima, Hiroshi

    2016-01-01

    LATS2, a pivotal Ser/Thr kinase of the Hippo pathway, plays important roles in many biological processes. LATS2 also function in Hippo-independent pathway, including mitosis, DNA damage response and epithelial to mesenchymal transition. However, the physiological relevance and molecular basis of these LATS2 functions remain obscure. To understand novel functions of LATS2, we constructed a LATS2 knockout HeLa-S3 cell line using TAL-effector nuclease (TALEN). Integrated omics profiling of this cell line revealed that LATS2 knockout caused genome-wide downregulation of Polycomb repressive complex 2 (PRC2) and H3K27me3. Cell-cycle analysis revealed that downregulation of PRC2 was not due to cell cycle aberrations caused by LATS2 knockout. Not LATS1, a homolog of LATS2, but LATS2 bound PRC2 on chromatin and phosphorylated it. LATS2 positively regulates histone methyltransferase activity of PRC2 and their expression at both the mRNA and protein levels. Our findings reveal a novel signal upstream of PRC2, and provide insight into the crucial role of LATS2 in coordinating the epigenome through regulation of PRC2. PMID:27434182

  13. HUA ENHANCER1 is involved in posttranscriptional regulation of positive and negative regulators in Arabidopsis photomorphogenesis.

    PubMed

    Tsai, Huang-Lung; Li, Yi-Hang; Hsieh, Wen-Ping; Lin, Meng-Chun; Ahn, Ji Hoon; Wu, Shu-Hsing

    2014-07-01

    Light regulates growth and developmental processes in plants via global transcriptome adjustment, translational control, and multilayered posttranslational modification of proteins. The transcriptional activation and repression of light-responsive genes has been well documented; however, the impact of posttranscriptional regulation on conveying light signals has been less addressed. Here, we examined whether optimal photomorphogenesis in Arabidopsis thaliana requires the proper biogenesis of small regulatory RNAs that play pivotal roles in the posttranscriptional regulation of gene expression. Arabidopsis carrying a mutation in HUA ENHANCER1 (HEN1), required for stabilization of small regulatory RNAs, showed defects in multiple aspects of photomorphogenic and skotomorphogenic development. HEN1 negatively regulated Arabidopsis photomorphogenesis. Light-activated HEN1 expression depended on the photoreceptors phytochrome A (phyA), phyB, cryptochrome 1 (cry1), and cry2 and key transcriptional regulators ELONGATED HYPOCOTYL5 (HY5) and HY5-HOMOLOG. We also demonstrate the involvement of the small regulatory RNAs miR157d and miR319 in modulating the expression of a positive regulator, HY5, and negative regulators TEOSINTE BRANCHED1, CYCLOIDEA AND PCF family proteins, respectively, for optimal photomorphogenic development in Arabidopsis.

  14. LMTK1 regulates dendritic formation by regulating movement of Rab11A-positive endosomes

    PubMed Central

    Takano, Tetsuya; Urushibara, Tomoki; Yoshioka, Nozomu; Saito, Taro; Fukuda, Mitsunori; Tomomura, Mineko; Hisanaga, Shin-ichi

    2014-01-01

    Neurons extend two types of neurites—axons and dendrites—that differ in structure and function. Although it is well understood that the cytoskeleton plays a pivotal role in neurite differentiation and extension, the mechanisms by which membrane components are supplied to growing axons or dendrites is largely unknown. We previously reported that the membrane supply to axons is regulated by lemur kinase 1 (LMTK1) through Rab11A-positive endosomes. Here we investigate the role of LMTK1 in dendrite formation. Down-regulation of LMTK1 increases dendrite growth and branching of cerebral cortical neurons in vitro and in vivo. LMTK1 knockout significantly enhances the prevalence, velocity, and run length of anterograde movement of Rab11A-positive endosomes to levels similar to those expressing constitutively active Rab11A-Q70L. Rab11A-positive endosome dynamics also increases in the cell body and growth cone of LMTK1-deficient neurons. Moreover, a nonphosphorylatable LMTK1 mutant (Ser34Ala, a Cdk5 phosphorylation site) dramatically promotes dendrite growth. Thus LMTK1 negatively controls dendritic formation by regulating Rab11A-positive endosomal trafficking in a Cdk5-dependent manner, indicating the Cdk5-LMTK1-Rab11A pathway as a regulatory mechanism of dendrite development as well as axon outgrowth. PMID:24672056

  15. Positive and negative elements regulate a melanocyte-specific promoter.

    PubMed Central

    Lowings, P; Yavuzer, U; Goding, C R

    1992-01-01

    Melanocytes are specialized cells residing in the hair follicles, the eye, and the basal layer of the human epidermis whose primary function is the production of the pigment melanin, giving rise to skin, hair, and eye color. Melanogenesis, a process unique to melanocytes that involves the processing of tyrosine by a number of melanocyte-specific enzymes, including tyrosinase and tyrosinase-related protein 1 (TRP-1), occurs only after differentiation from the melanocyte precursor, the melanoblast. In humans, melanogenesis is inducible by UV irradiation, with melanin being transferred from the melanocyte in the epidermis to the surrounding keratinocytes as protection from UV-induced damage. Excessive exposure to UV, however, is the primary cause of malignant melanoma, an increasingly common and highly aggressive disease. As an initial approach to understanding the regulation of melanocyte differentiation and melanocyte-specific transcription, we have isolated the gene encoding TRP-1 and examined the cis- and trans-acting factors required for cell-type-specific expression. We find that the TRP-1 promoter comprises both positive and negative regulatory elements which confer efficient expression in a TRP-1-expressing, pigmented melanoma cell line but not in NIH 3T3 or JEG3 cells and that a minimal promoter extending between -44 and +107 is sufficient for cell-type-specific expression. Assays for DNA-protein interactions coupled with extensive mutagenesis identified three factors, whose binding correlated with the function of two positive and one negative regulatory element. One of these factors, termed M-box-binding factor 1, binds to an 11-bp motif, the M box, which acts as a positive regulatory element both in TRP-1-expressing and -nonexpressing cell lines, despite being entirely conserved between the melanocyte-specific tyrosinase and TRP-1 promoters. The possible mechanisms underlying melanocyte-specific gene expression are discussed. Images PMID:1321344

  16. Interleukin-32 Positively Regulates Radiation-Induced Vascular Inflammation

    SciTech Connect

    Kobayashi, Hanako; Yazlovitskaya, Eugenia M.; Lin, P. Charles

    2009-08-01

    Purpose: To study the role of interleukin-32 (IL-32), a novel protein only detected in human tissues, in ionizing radiation (IR)-induced vascular inflammation. Methods and Materials: Irradiated (0-6 Gy) human umbilical vein endothelial cells treated with or without various agents-a cytosolic phospholipase A2 (cPLA2) inhibitor, a cyclooxygenase-2 (Cox-2) inhibitor, or lysophosphatidylcholines (LPCs)-were used to assess IL-32 expression by Northern blot analysis and quantitative reverse transcriptase-polymerase chain reaction. Expression of cell adhesion molecules and leukocyte adhesion to endothelial cells using human acute monocytic leukemia cell line (THP-1) cells was also analyzed. Results: Ionizing radiation dramatically increased IL-32 expression in vascular endothelial cells through multiple pathways. Ionizing radiation induced IL-32 expression through nuclear factor {kappa}B activation, through induction of cPLA2 and LPC, as well as induction of Cox-2 and subsequent conversion of arachidonic acid to prostacyclin. Conversely, blocking nuclear factor {kappa}B, cPLA2, and Cox-2 activity impaired IR-induced IL-32 expression. Importantly, IL-32 significantly enhanced IR-induced expression of vascular cell adhesion molecules and leukocyte adhesion on endothelial cells. Conclusion: This study identifies IL-32 as a positive regulator in IR-induced vascular inflammation, and neutralization of IL-32 may be beneficial in protecting from IR-induced inflammation.

  17. Regulation of Apoptosis by Gram-Positive Bacteria

    PubMed Central

    Ulett, Glen C.; Adderson, Elisabeth E.

    2008-01-01

    Apoptosis, or programmed cell death (PCD), is an important physiological mechanism, through which the human immune system regulates homeostasis and responds to diverse forms of cellular damage. PCD may also be involved in immune counteraction to microbial infection. Over the past decade, the amount of research on bacteria-induced PCD has grown tremendously, and the implications of this mechanism on immunity are being elucidated. Some pathogenic bacteria actively trigger the suicide response in critical lineages of leukocytes that orchestrate both the innate and adaptive immune responses; other bacteria proactively prevent PCD to benefit their own survival and persistence. Currently, the microbial virulence factors, which represent the keys to unlocking the suicide response in host cells, are a primary focus of this field. In this review, we discuss these bacterial “apoptosis regulatory molecules” and the apoptotic events they either trigger or prevent, the host target cells of this regulatory activity, and the possible ramifications for immunity to infection. Gram-positive pathogens including Staphylococcus, Streptococcus, Bacillus, Listeria, and Clostridia species are discussed as important agents of human infection that modulate PCD pathways in eukaryotic cells. PMID:19081777

  18. Phosphatidylserine directly and positively regulates fusion of myoblasts into myotubes

    SciTech Connect

    Jeong, Jaemin; Conboy, Irina M.

    2011-10-14

    Highlights: {yields} PS broadly and persistently trans-locates to the outer leaflet of plasma membrane during myoblast fusion into myotubes. {yields} Robust myotubes are formed when PS liposomes are added exogenously. {yields} PS increases the width of de novo myotubes and the numbers of myonuclei, but not the myotube length. {yields} Annexin V or PS antibody inhibits myotube formation by masking exposed PS. -- Abstract: Cell membrane consists of various lipids such as phosphatidylserine (PS), phosphatidylcholine (PC), and phosphatidylethanolamine (PE). Among them, PS is a molecular marker of apoptosis, because it is located to the inner leaflet of plasma membrane generally but it is moved to the outer leaflet during programmed cell death. The process of apoptosis has been implicated in the fusion of muscle progenitor cells, myoblasts, into myotubes. However, it remained unclear whether PS regulates muscle cell differentiation directly. In this paper, localization of PS to the outer leaflet of plasma membrane in proliferating primary myoblasts and during fusion of these myoblasts into myotubes is validated using Annexin V. Moreover, we show the presence of PS clusters at the cell-cell contact points, suggesting the importance of membrane ruffling and PS exposure for the myogenic cell fusion. Confirming this conclusion, experimentally constructed PS, but not PC liposomes dramatically enhance the formation of myotubes from myoblasts, thus demonstrating a direct positive effect of PS on the muscle cell fusion. In contrast, myoblasts exposed to PC liposomes produce long myotubes with low numbers of myonuclei. Moreover, pharmacological masking of PS on the myoblast surface inhibits fusion of these cells into myotubes in a dose-dependent manner.

  19. Spontaneous Emotion Regulation to Positive and Negative Stimuli

    ERIC Educational Resources Information Center

    Volokhov, Rachael N.; Demaree, Heath A.

    2010-01-01

    The ability to regulate one's emotions is an integral part of human social behavior. One antecedent emotion regulation strategy, known as reappraisal, is characterized by cognitively evaluating an emotional stimulus to alter its emotional impact and one response-focused strategy, suppression, is aimed at reducing behavioral output. People are…

  20. Patient with three euchromatic supernumerary marker chromosomes derived from chromosomes 1, 12, and 18: characterization and evaluation of the aberrations.

    PubMed

    Schwanitz, Gesa; Hagh, Javad Karim Zad; Rad, Isa Abdi; Omrani, Mir Davood; Gamerdinger, Ulrike; Schubert, Regine; Elbracht, Miriam; Eggermann, Thomas; Eggermann, Katja; Spengler, Sabrina; Schüler, Herdit; Gogiel, Magdalena

    2014-03-01

    The genetic relevance of small supernumerary marker chromosomes (sSMCs) depends on their content of euchromatin. In case of mosaicism, the phenotype of the carrier furthermore is influenced by the distribution of the marker in the body. In the majority of reported cases no correlation of the degree of mosaicism in the tissue(s) analyzed and the phenotype could be detected. In particular, non-acrocentric derived sSMCs show a strong tendency to appear in mosaic state irrespective of the clinical picture. We present a patient with cognitive disability and mild craniofacial dysmorphisms with mosaicism of three different autosomal marker chromosomes. The extra chromosomes were analyzed by a combination of SNP array and a variety of fluorescence in situ hybridization (FISH) probes. All three markers were identified as ring chromosomes containing different amounts of euchromatic material derived from chromosome 1 (1p12 → q21), 12 (12p13.1 → q13.11) and 18 (18p11.21 → q11.2). The size and the frequency of the sSMCs were strikingly different, besides, we observed an unequal combination of the three derivates.

  1. Genome-wide analysis of tandem repeats in Tribolium castaneum genome reveals abundant and highly dynamic tandem repeat families with satellite DNA features in euchromatic chromosomal arms.

    PubMed

    Pavlek, Martina; Gelfand, Yevgeniy; Plohl, Miroslav; Meštrović, Nevenka

    2015-12-01

    Although satellite DNAs are well-explored components of heterochromatin and centromeres, little is known about emergence, dispersal and possible impact of comparably structured tandem repeats (TRs) on the genome-wide scale. Our bioinformatics analysis of assembled Tribolium castaneum genome disclosed significant contribution of TRs in euchromatic chromosomal arms and clear predominance of satellite DNA-typical 170 bp monomers in arrays of ≥5 repeats. By applying different experimental approaches, we revealed that the nine most prominent TR families Cast1-Cast9 extracted from the assembly comprise ∼4.3% of the entire genome and reside almost exclusively in euchromatic regions. Among them, seven families that build ∼3.9% of the genome are based on ∼170 and ∼340 bp long monomers. Results of phylogenetic analyses of 2500 monomers originating from these families show high-sequence dynamics, evident by extensive exchanges between arrays on non-homologous chromosomes. In addition, our analysis shows that concerted evolution acts more efficiently on longer than on shorter arrays. Efficient genome-wide distribution of nine TR families implies the role of transposition only in expansion of the most dispersed family, and involvement of other mechanisms is anticipated. Despite similarities in sequence features, FISH experiments indicate high-level compartmentalization of centromeric and euchromatic tandem repeats.

  2. A gene expression map for the euchromatic genome of Drosophila melanogaster.

    PubMed

    Stolc, Viktor; Gauhar, Zareen; Mason, Christopher; Halasz, Gabor; van Batenburg, Marinus F; Rifkin, Scott A; Hua, Sujun; Herreman, Tine; Tongprasit, Waraporn; Barbano, Paolo Emilio; Bussemaker, Harmen J; White, Kevin P

    2004-10-22

    We used a maskless photolithography method to produce DNA oligonucleotide microarrays with unique probe sequences tiled throughout the genome of Drosophila melanogaster and across predicted splice junctions. RNA expression of protein coding and nonprotein coding sequences was determined for each major stage of the life cycle, including adult males and females. We detected transcriptional activity for 93% of annotated genes and RNA expression for 41% of the probes in intronic and intergenic sequences. Comparison to genome-wide RNA interference data and to gene annotations revealed distinguishable levels of expression for different classes of genes and higher levels of expression for genes with essential cellular functions. Differential splicing was observed in about 40% of predicted genes, and 5440 previously unknown splice forms were detected. Genes within conserved regions of synteny with D. pseudoobscura had highly correlated expression; these regions ranged in length from 10 to 900 kilobase pairs. The expressed intergenic and intronic sequences are more likely to be evolutionarily conserved than nonexpressed ones, and about 15% of them appear to be developmentally regulated. Our results provide a draft expression map for the entire nonrepetitive genome, which reveals a much more extensive and diverse set of expressed sequences than was previously predicted.

  3. Positive regulation of the Escherichia coli glycine cleavage enzyme system.

    PubMed Central

    Wilson, R L; Steiert, P S; Stauffer, G V

    1993-01-01

    A new mutation in Escherichia coli, designated gcvA1, that results in noninducible expression of both gcv and a gcvT-lacZ gene fusion was isolated. A plasmid carrying the wild-type gcvA gene complemented the mutation and restored glycine-inducible gcv and gcvT-lacZ gene expression. These results suggest that gcvA encodes a positive-acting regulatory protein that acts in trans to increase expression of gcv. PMID:8423160

  4. YY1 positively regulates human UBIAD1 expression

    SciTech Connect

    Funahashi, Nobuaki; Hirota, Yoshihisa; Nakagawa, Kimie; Sawada, Natumi; Watanabe, Masato; Suhara, Yoshitomo; Okano, Toshio

    2015-05-01

    Vitamin K is involved in bone formation and blood coagulation. Natural vitamin K compounds are composed of the plant form phylloquinone (vitamin K{sub 1}) and a series of bacterial menaquionones (MK-n; vitamin K{sub 2}). Menadione (vitamin K{sub 3}) is an artificial vitamin K compound. MK-4 contains 4-isoprenyl as a side group in the 2-methyl-1,4-naphthoquinone common structure and has various bioactivities. UbiA prenyltransferase domain containing 1 (UBIAD1 or TERE1) is the menaquinone-4 biosynthetic enzyme. UBIAD1 transcript expression significantly decreases in patients with prostate carcinoma and overexpressing UBIAD1 inhibits proliferation of a tumour cell line. UBIAD1 mRNA expression is ubiquitous in mouse tissues, and higher UBIAD1 mRNA expression levels are detected in the brain, heart, kidneys and pancreas. Several functions of UBIAD1 have been reported; however, regulation of the human UBIAD1 gene has not been elucidated. Here we report cloning and characterisation of the human UBIAD1 promoter. A 5′ rapid amplification of cDNA ends analysis revealed that the main transcriptional start site was 306 nucleotides upstream of the translation initiation codon. Deletion and mutation analyses revealed the functional importance of the YY1 consensus motif. Electrophoretic gel mobility shift and chromatin immunoprecipitation assays demonstrated that YY1 binds the UBIAD1 promoter in vitro and in vivo. In addition, YY1 small interfering RNA decreased endogenous UBIAD1 mRNA expression and UBIAD1 conversion activity. These results suggest that YY1 up-regulates UBIAD1 expression and UBIAD1 conversion activity through the UBIAD1 promoter. - Highlights: • We cloned the human UBIAD1 promoter. • The functional importance of the YY1 motif was identified in the UBIAD1 promoter. • YY1 binds the UBIAD1 promoter in vitro and in vivo. • Knockdown of YY1 significantly decreased UBIAD1 expression. • YY1 up-regulates UBIAD1 conversion activity through the UBIAD1

  5. Regulation of meiotic chromatin loop size by chromosomal position.

    PubMed Central

    Heng, H H; Chamberlain, J W; Shi, X M; Spyropoulos, B; Tsui, L C; Moens, P B

    1996-01-01

    At meiotic prophase, chromatin loops around a proteinaceous core, with the sizes of these loops varying between species. Comparison of the morphology of sequence-related inserts at different sites in transgenic mice demonstrates that loop size also varies with chromosomal geography. Similarly, chromatin loop lengths differ dramatically for interstitially and terminally located hamster telomeric sequences. Sequences, telomeric or otherwise, located at chromosome termini, closely associate with the meiotic proteinaceous core, forming shorter loops than identical interstitial sequences. Thus, we present evidence that different chromatin packaging mechanisms exist for interstitial versus terminal chromosomal regions, which act separately from those operating at the level of the DNA sequence. Chromosomal position plays the dominant role in chromatin packaging. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8610120

  6. Dampening or savoring positive emotions: a dialectical cultural script guides emotion regulation.

    PubMed

    Miyamoto, Yuri; Ma, Xiaoming

    2011-12-01

    Four studies examined the hypothesis that, although people may generally want to savor, rather than to dampen, their positive emotions (i.e., hedonic emotion regulation), such a hedonic emotion regulation tendency should be less pronounced for Easterners than for Westerners. Using retrospective memory procedures, Study 1 found that Easterners recalled engaging in hedonic emotion regulation less than Westerners did, even after controlling for their initial emotional reactions. Studies 2-3 showed that cultural differences in emotion regulation were mediated by dialectical beliefs about positive emotions. Study 4 replicated the findings by examining online reports of emotion regulation strategies on the day students received a good grade. Furthermore, there were cultural differences in actual emotion change over time, which was partly explained by dialectical beliefs about positive emotions. These findings highlight the active role cultural scripts play in shaping emotion regulation and emotional experiences.

  7. [The regulation of negative and positive emotions during picture viewing: an ERP study].

    PubMed

    Reva, N V; Pavlov, S V; Korenek, V V; Loktev, K V; Tumialis, A V; Brak, I V; Aftanas, L I

    2015-01-01

    The study examines the effects of cognitive reappraisal on the event-related potentials (ERPs) to affective stimuli. Participants (n = 53) were asked either to attend affective images, or to down-regulate negative affect, or to up-regulate positive affect. Reappraisal of negative images was associated with attenuation of the P300 and late positive potential (LPP) over parietal regions, whereas reappraisal of positive images had no significant effect on ERP components. The weak P300 reduction correlated with high personality scores of negative affectivity. We assume that only down-regulation of negative emotions is associated with the changes in primary appraisals, and so far reflected in ERP modulation.

  8. The Late Positive Potential: A Neurophysiological Marker for Emotion Regulation in Children

    ERIC Educational Resources Information Center

    Dennis, Tracy A.; Hajcak, Greg

    2009-01-01

    Background: The ability to modulate emotional responses, or emotion regulation, is a key mechanism in the development of mood disruptions. Detection of a neural marker for emotion regulation thus has the potential to inform early detection and intervention for mood problems. One such neural marker may be the late positive potential (LPP), which is…

  9. The Older Adult Positivity Effect in Evaluations of Trustworthiness: Emotion Regulation or Cognitive Capacity?

    PubMed Central

    Zebrowitz, Leslie A.; Boshyan, Jasmine; Ward, Noreen; Gutchess, Angela; Hadjikhani, Nouchine

    2017-01-01

    An older adult positivity effect, i.e., the tendency for older adults to favor positive over negative stimulus information more than do younger adults, has been previously shown in attention, memory, and evaluations. This effect has been attributed to greater emotion regulation in older adults. In the case of attention and memory, this explanation has been supported by some evidence that the older adult positivity effect is most pronounced for negative stimuli, which would motivate emotion regulation, and that it is reduced by cognitive load, which would impede emotion regulation. We investigated whether greater older adult positivity in the case of evaluative responses to faces is also enhanced for negative stimuli and attenuated by cognitive load, as an emotion regulation explanation would predict. In two studies, younger and older adults rated trustworthiness of faces that varied in valence both under low and high cognitive load, with the latter manipulated by a distracting backwards counting task. In Study 1, face valence was manipulated by attractiveness (low /disfigured faces, medium, high/fashion models’ faces). In Study 2, face valence was manipulated by trustworthiness (low, medium, high). Both studies revealed a significant older adult positivity effect. However, contrary to an emotion regulation account, this effect was not stronger for more negative faces, and cognitive load increased rather than decreased the rated trustworthiness of negatively valenced faces. Although inconsistent with emotion regulation, the latter effect is consistent with theory and research arguing that more cognitive resources are required to process negative stimuli, because they are more cognitively elaborated than positive ones. The finding that increased age and increased cognitive load both enhanced the positivity of trustworthy ratings suggests that the older adult positivity effect in evaluative ratings of faces may reflect age-related declines in cognitive capacity

  10. Positive allosteric feedback regulation of the stringent response enzyme RelA by its product

    PubMed Central

    Shyp, Viktoriya; Tankov, Stoyan; Ermakov, Andrey; Kudrin, Pavel; English, Brian P; Ehrenberg, Måns; Tenson, Tanel; Elf, Johan; Hauryliuk, Vasili

    2012-01-01

    During the stringent response, Escherichia coli enzyme RelA produces the ppGpp alarmone, which in turn regulates transcription, translation and replication. We show that ppGpp dramatically increases the turnover rate of its own ribosome-dependent synthesis by RelA, resulting in direct positive regulation of an enzyme by its product. Positive allosteric regulation therefore constitutes a new mechanism of enzyme activation. By integrating the output of individual RelA molecules and ppGpp degradation pathways, this regulatory circuit contributes to a fast and coordinated transition to stringency. PMID:22814757

  11. 75 FR 50950 - Federal Speculative Position Limits for Referenced Energy Contracts and Associated Regulations

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-18

    ... COMMISSION 17 CFR Parts 1, 20, and 151 RIN 3038-AC85 Federal Speculative Position Limits for Referenced...'' or ``Commission'') proposed to implement position limits for futures and option contracts based on a... Limits for Referenced Energy Contracts and Associated Regulations,'' for ease of reference,...

  12. A positive role for polycomb in transcriptional regulation via H4K20me1.

    PubMed

    Lv, Xiangdong; Han, Zhijun; Chen, Hao; Yang, Bo; Yang, Xiaofeng; Xia, Yuanxin; Pan, Chenyu; Fu, Lin; Zhang, Shuo; Han, Hui; Wu, Min; Zhou, Zhaocai; Zhang, Lei; Li, Lin; Wei, Gang; Zhao, Yun

    2016-05-01

    The highly conserved polycomb group (PcG) proteins maintain heritable transcription repression of the genes essential for development from fly to mammals. However, sporadic reports imply a potential role of PcGs in positive regulation of gene transcription, although systematic investigation of such function and the underlying mechanism has rarely been reported. Here, we report a Pc-mediated, H3K27me3-dependent positive transcriptional regulation of Senseless (Sens), a key transcription factor required for development. Mechanistic studies show that Pc regulates Sens expression by promoting H4K20me1 at the Sens locus. Further bioinformatic analysis at genome-wide level indicates that the existence of H4K20me1 acts as a selective mark for positive transcriptional regulation by Pc/H3K27me3. Both the intensities and specific patterns of Pc and H3K27me3 are important for the fates of target gene transcription. Moreover, binding of transcription factor Broad (Br), which physically interacts with Pc and positively regulates the transcription of Sens, is observed in Pc(+)H3K27me3(+)H4K20me1(+) genes, but not in Pc(+)H3K27me3(+)H4K20me1(-) genes. Taken together, our study reveals that, coupling with the transcription factor Br, Pc positively regulates transcription of Pc(+)H3K27me3(+)H4K20me1(+) genes in developing Drosophila wing disc.

  13. Miro1 Regulates Activity-Driven Positioning of Mitochondria within Astrocytic Processes Apposed to Synapses to Regulate Intracellular Calcium Signaling

    PubMed Central

    Stephen, Terri-Leigh; Higgs, Nathalie F.; Sheehan, David F.; Al Awabdh, Sana; López-Doménech, Guillermo; Arancibia-Carcamo, I. Lorena

    2015-01-01

    It is fast emerging that maintaining mitochondrial function is important for regulating astrocyte function, although the specific mechanisms that govern astrocyte mitochondrial trafficking and positioning remain poorly understood. The mitochondrial Rho-GTPase 1 protein (Miro1) regulates mitochondrial trafficking and detachment from the microtubule transport network to control activity-dependent mitochondrial positioning in neurons. However, whether Miro proteins are important for regulating signaling-dependent mitochondrial dynamics in astrocytic processes remains unclear. Using live-cell confocal microscopy of rat organotypic hippocampal slices, we find that enhancing neuronal activity induces transient mitochondrial remodeling in astrocytes, with a concomitant, transient reduction in mitochondrial trafficking, mediated by elevations in intracellular Ca2+. Stimulating neuronal activity also induced mitochondrial confinement within astrocytic processes in close proximity to synapses. Furthermore, we show that the Ca2+-sensing EF-hand domains of Miro1 are important for regulating mitochondrial trafficking in astrocytes and required for activity-driven mitochondrial confinement near synapses. Additionally, activity-dependent mitochondrial positioning by Miro1 reciprocally regulates the levels of intracellular Ca2+ in astrocytic processes. Thus, the regulation of intracellular Ca2+ signaling, dependent on Miro1-mediated mitochondrial positioning, could have important consequences for astrocyte Ca2+ wave propagation, gliotransmission, and ultimately neuronal function. SIGNIFICANCE STATEMENT Mitochondria are key cellular organelles that play important roles in providing cellular energy and buffering intracellular calcium ions. The mechanisms that control mitochondrial distribution within the processes of glial cells called astrocytes and the impact this may have on calcium signaling remains unclear. We show that activation of glutamate receptors or increased neuronal

  14. Rethinking emotion: cognitive reappraisal is an effective positive and negative emotion regulation strategy in bipolar disorder.

    PubMed

    Gruber, June; Hay, Aleena C; Gross, James J

    2014-04-01

    Bipolar disorder involves difficulties with emotion regulation, yet the precise nature of these emotion regulatory difficulties is unclear. The current study examined whether individuals with remitted bipolar I disorder (n = 23) and healthy controls (n = 23) differ in their ability to use one effective and common form of emotion regulation, cognitive reappraisal. Positive, negative, and neutral films were used to elicit emotion, and participants were cued to watch the film carefully (i.e., uninstructed condition) or reappraise while measures of affect, behavior, and psychophysiology were obtained. Results showed that reappraisal was associated with reductions in emotion reactivity across subjective (i.e., positive and negative affect), behavioral (i.e., positive facial displays), and physiological (i.e., skin conductance) response domains across all participants. Results suggest that reappraisal may be an effective regulation strategy for both negative and positive emotion across both healthy adults and individuals with bipolar disorder. Discussion focuses on clinical and treatment implications for bipolar disorder.

  15. Regulating the Intersection of Metabolism and Pathogenesis in Gram-positive Bacteria

    PubMed Central

    RICHARDSON, ANTHONY R.; SOMERVILLE, GREG A.; SONENSHEIN, ABRAHAM L.

    2015-01-01

    Pathogenic bacteria must contend with immune systems that actively restrict the availability of nutrients and cofactors, and create a hostile growth environment. To deal with these hostile environments, pathogenic bacteria have evolved or acquired virulence determinants that aid in the acquisition of nutrients. This connection between pathogenesis and nutrition may explain why regulators of metabolism in nonpathogenic bacteria are used by pathogenic bacteria to regulate both metabolism and virulence. Such coordinated regulation is presumably advantageous because it conserves carbon and energy by aligning synthesis of virulence determinants with the nutritional environment. In Gram-positive bacterial pathogens, at least three metabolite-responsive global regulators, CcpA, CodY, and Rex, have been shown to coordinate the expression of metabolism and virulence genes. In this chapter, we discuss how environmental challenges alter metabolism, the regulators that respond to this altered metabolism, and how these regulators influence the host-pathogen interaction. PMID:26185086

  16. p21-activated kinase 4 regulates mitotic spindle positioning and orientation.

    PubMed

    Bompard, Guillaume; Morin, Nathalie

    2012-01-01

    During mitosis, microtubules (MTs) are massively rearranged into three sets of highly dynamic MTs that are nucleated from the centrosomes to form the mitotic spindle. Tight regulation of spindle positioning in the dividing cell and chromosome alignment at the center of the metaphase spindle are required to ensure perfect chromosome segregation and to position the cytokinetic furrow that will specify the two daughter cells. Spindle positioning requires regulation of MT dynamics, involving depolymerase activities together with cortical and kinetochore-mediated pushing and pulling forces acting on astral MTs and kinetochore fibres. These forces rely on MT motor activities. Cortical pulling forces exerted on astral MTs depend upon dynein/dynactin complexes and are essential in both symmetric and asymmetric cell division. A well-established spindle positioning pathway regulating the cortical targeting of dynein/dynactin involves the conserved LGN (Leu-Gly-Asn repeat-enriched-protein) and NuMA (microtubule binding nuclear mitotic apparatus protein) complex. Spindle orientation is also regulated by integrin-mediated cell adhesion and actin retraction fibres that respond to mechanical stress and are influenced by the microenvironment of the dividing cell. Altering the capture of astral MTs or modulating pulling forces affects spindle position, which can impair cell division, differentiation and embryogenesis. In this general scheme, the activity of mitotic kinases such as Auroras and Plk1 (Polo-like kinase 1) is crucial. Recently, the p21-activated kinases (PAKs) emerged as novel important players in mitotic progression. In our recent article, we demonstrated that PAK4 regulates spindle positioning in symmetric cell division. In this commentary, and in light of recent published studies, we discuss how PAK4 could participate in the regulation of mechanisms involved in spindle positioning and orientation.

  17. miRNA863-3p sequentially targets negative immune regulator ARLPKs and positive regulator SERRATE upon bacterial infection

    PubMed Central

    Niu, Dongdong; Lii, Yifan E.; Chellappan, Padmanabhan; Lei, Lei; Peralta, Karl; Jiang, Chunhao; Guo, Jianhua; Coaker, Gitta; Jin, Hailing

    2016-01-01

    Plant small RNAs play important roles in gene regulation during pathogen infection. Here we show that miR863-3p is induced by the bacterial pathogen Pseudomonas syringae carrying various effectors. Early during infection, miR863-3p silences two negative regulators of plant defence, atypical receptor-like pseudokinase1 (ARLPK1) and ARLPK2, both lacking extracellular domains and kinase activity, through mRNA degradation to promote immunity. ARLPK1 associates with, and may function through another negative immune regulator ARLPK1-interacting receptor-like kinase 1 (AKIK1), an active kinase with an extracellular domain. Later during infection, miR863-3p silences SERRATE, which is essential for miRNA accumulation and positively regulates defence, through translational inhibition. This results in decreased miR863-3p levels, thus forming a negative feedback loop to attenuate immune responses after successful defence. This is an example of a miRNA that sequentially targets both negative and positive regulators of immunity through two modes of action to fine-tune the timing and amplitude of defence responses. PMID:27108563

  18. Positive regulation of botulinum neurotoxin gene expression by CodY in Clostridium botulinum ATCC 3502.

    PubMed

    Zhang, Zhen; Dahlsten, Elias; Korkeala, Hannu; Lindström, Miia

    2014-12-01

    Botulinum neurotoxin, produced mainly by the spore-forming bacterium Clostridium botulinum, is the most poisonous biological substance known. Here, we show that CodY, a global regulator conserved in low-G+C Gram-positive bacteria, positively regulates the botulinum neurotoxin gene expression. Inactivation of codY resulted in decreased expression of botA, encoding the neurotoxin, as well as in reduced neurotoxin synthesis. Complementation of the codY mutation in trans rescued neurotoxin synthesis, and overexpression of codY in trans caused elevated neurotoxin production. Recombinant CodY was found to bind to a 30-bp region containing the botA transcription start site, suggesting regulation of the neurotoxin gene transcription through direct interaction. GTP enhanced the binding affinity of CodY to the botA promoter, suggesting that CodY-dependent neurotoxin regulation is associated with nutritional status.

  19. Positive Regulation of Botulinum Neurotoxin Gene Expression by CodY in Clostridium botulinum ATCC 3502

    PubMed Central

    Zhang, Zhen; Dahlsten, Elias; Korkeala, Hannu

    2014-01-01

    Botulinum neurotoxin, produced mainly by the spore-forming bacterium Clostridium botulinum, is the most poisonous biological substance known. Here, we show that CodY, a global regulator conserved in low-G+C Gram-positive bacteria, positively regulates the botulinum neurotoxin gene expression. Inactivation of codY resulted in decreased expression of botA, encoding the neurotoxin, as well as in reduced neurotoxin synthesis. Complementation of the codY mutation in trans rescued neurotoxin synthesis, and overexpression of codY in trans caused elevated neurotoxin production. Recombinant CodY was found to bind to a 30-bp region containing the botA transcription start site, suggesting regulation of the neurotoxin gene transcription through direct interaction. GTP enhanced the binding affinity of CodY to the botA promoter, suggesting that CodY-dependent neurotoxin regulation is associated with nutritional status. PMID:25281376

  20. [Regulation of the Z ring positioning in bacterial cell division--a review].

    PubMed

    Sang, Yu; Tao, Jing; Yao, Yufeng

    2013-04-04

    The regulatory mechanism of bacterial cell division has long been a research focus. Forming a septum at the middle of the cell, the seemingly simple process is involved by multiple regulation factors. Zring (FtsZ ring) is the skeleton of the splitting complex. The locus where Z ring is formed is not only the position the septum formed but also determines the cell division site. Formation of Zring in the incorrect location results in inequality cell division. Several cell division regulation systems have been identified, including the Min system, nucleoid occlusion and the MipZ protein which effectively prevent Zring assembly by different mechanisms, ensuring formation of the fission complex at the correct position. Recent progresses about the formation process of Zring and regulation mechanism affecting the Z-ring positioning are summarized.

  1. Positioning.

    ERIC Educational Resources Information Center

    Conone, Ruth M.

    The key to positioning is the creation of a clear benefit image in the consumer's mind. One positioning strategy is creating in the prospect's mind a position that takes into consideration the company's or agency's strengths and weaknesses as well as those of its competitors. Another strategy is to gain entry into a position ladder owned by…

  2. The neural correlates of regulating positive and negative emotions in medication-free major depression.

    PubMed

    Greening, Steven G; Osuch, Elizabeth A; Williamson, Peter C; Mitchell, Derek G V

    2014-05-01

    Depressive cognitive schemas play an important role in the emergence and persistence of major depressive disorder (MDD). The current study adapted emotion regulation techniques to reflect elements of cognitive behavioural therapy (CBT) and related psychotherapies to delineate neurocognitive abnormalities associated with modulating the negative cognitive style in MDD. Nineteen non-medicated patients with MDD and 19 matched controls reduced negative or enhanced positive feelings elicited by emotional scenes while undergoing functional magnetic resonance imaging. Although both groups showed significant emotion regulation success as measured by subjective ratings of affect, the controls were significantly better at modulating both negative and positive emotion. Both groups recruited regions of dorsolateral prefrontal cortex and ventrolateral prefrontal cortex (VLPFC) when regulating negative emotions. Only in controls was this accompanied by reduced activity in sensory cortices and amygdala. Similarly, both groups showed enhanced activity in VLPFC and ventral striatum when enhancing positive affect; however, only in controls was ventral striatum activity correlated with regulation efficacy. The results suggest that depression is associated with both a reduced capacity to achieve relief from negative affect despite recruitment of ventral and dorsal prefrontal cortical regions implicated in emotion regulation, coupled with a disconnect between activity in reward-related regions and subjective positive affect.

  3. Factor Structure and Initial Validation of a Multidimensional Measure of Difficulties in the Regulation of Positive Emotions: The DERS-Positive.

    PubMed

    Weiss, Nicole H; Gratz, Kim L; Lavender, Jason M

    2015-05-01

    Emotion regulation difficulties are a transdiagnostic construct relevant to numerous clinical difficulties. Although the Difficulties in Emotion Regulation Scale (DERS) is a multidimensional measure of maladaptive ways of responding to emotions, it focuses on difficulties with the regulation of negative emotions and does not assess emotion dysregulation in the form of problematic responding to positive emotions. The aim of this study was to develop and validate a measure of clinically relevant difficulties in the regulation of positive emotions (DERS-Positive). Findings revealed a three-factor structure and supported the internal consistency and construct validity of the total and subscale scores.

  4. A positive role for polycomb in transcriptional regulation via H4K20me1

    PubMed Central

    Lv, Xiangdong; Han, Zhijun; Chen, Hao; Yang, Bo; Yang, Xiaofeng; Xia, Yuanxin; Pan, Chenyu; Fu, Lin; Zhang, Shuo; Han, Hui; Wu, Min; Zhou, Zhaocai; Zhang, Lei; Li, Lin; Wei, Gang; Zhao, Yun

    2016-01-01

    The highly conserved polycomb group (PcG) proteins maintain heritable transcription repression of the genes essential for development from fly to mammals. However, sporadic reports imply a potential role of PcGs in positive regulation of gene transcription, although systematic investigation of such function and the underlying mechanism has rarely been reported. Here, we report a Pc-mediated, H3K27me3-dependent positive transcriptional regulation of Senseless (Sens), a key transcription factor required for development. Mechanistic studies show that Pc regulates Sens expression by promoting H4K20me1 at the Sens locus. Further bioinformatic analysis at genome-wide level indicates that the existence of H4K20me1 acts as a selective mark for positive transcriptional regulation by Pc/H3K27me3. Both the intensities and specific patterns of Pc and H3K27me3 are important for the fates of target gene transcription. Moreover, binding of transcription factor Broad (Br), which physically interacts with Pc and positively regulates the transcription of Sens, is observed in Pc+H3K27me3+H4K20me1+ genes, but not in Pc+H3K27me3+H4K20me1− genes. Taken together, our study reveals that, coupling with the transcription factor Br, Pc positively regulates transcription of Pc+H3K27me3+H4K20me1+ genes in developing Drosophila wing disc. PMID:27002220

  5. Positive regulation of the Vibrio cholerae porin OmpT by iron and fur.

    PubMed

    Craig, S A; Carpenter, C D; Mey, A R; Wyckoff, E E; Payne, S M

    2011-12-01

    The transcription factor Fur regulates the expression of a number of genes in Vibrio cholerae in response to changes in the level of available iron. Fur usually acts as a repressor, but here we show that Fur positively regulates the expression of ompT, which encodes a major outer membrane porin. OmpT levels increased when the bacteria were grown in medium containing relatively high levels of iron, and this effect required Fur. The level of ompT mRNA also is increased in the presence of iron and Fur. The effect of iron on OmpT levels was independent of the known ompT regulators ToxR and Crp, and it did not require RyhB, which has been shown to be responsible for positive regulation by iron of some V. cholerae genes. Electrophoretic mobility shift assays showed that Fur binds upstream of the ompT transcription start site in a region overlapping known binding sites for ToxR and Crp. These data suggest that Fur and iron positively regulate ompT expression through the direct binding of Fur to the ompT promoter.

  6. Exercise temperature regulation in man in the upright and supine positions.

    NASA Technical Reports Server (NTRS)

    Greenleaf, J. E.; Card, D. H.; Rapport, M.; Van Kessel, A. L.; Ruff, W.

    1971-01-01

    Rectal, auditory canal, and mean skin temperatures and various metabolic measurements were taken on subjects during 70-min exercise periods, in the upright and supine positions, on a bicycle ergometer. The results indicate nonlinear relationships between sweating and core skin temperatures and suggest the action of undefined thermal and/or nonthermal inputs in the control of exercise temperature regulation.

  7. Positive and negative roles for soybean MPK6 in regulating defense responses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    It has been well established that MPK6 is a positive regulator of defense responses in model plants such as Arabidopsis and tobacco. However, the functional importance of soybean MPK6 in disease resistance has not been investigated. Here, we showed that silencing of GmMPK6 in soybean using virus-ind...

  8. 77 FR 60343 - Federal Acquisition Regulation; Positive Law Codification of Title 41

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-03

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF DEFENSE GENERAL SERVICES ADMINISTRATION NATIONAL AERONAUTICS AND SPACE ADMINISTRATION 48 CFR Part 53 RIN 9000-AM30 Federal Acquisition Regulation; Positive Law Codification of Title 41 Correction In proposed rule...

  9. A long noncoding RNA positively regulates CD56 in human natural killer cells

    PubMed Central

    Fu, Binqing; Wu, Yang; Sun, Rui; Tian, Zhigang; Wei, Haiming

    2016-01-01

    Natural killer (NK) cells are innate immune lymphocytes that play critical roles in host defense against viral infection and surveillance against malignant transformation. Long noncoding RNAs (lncRNAs) are important immune system regulators. Here, we analyzed human primary lymphocyte lncRNA expression profiles to identify NK-lncRNA signatures. We detected numerous novel NK-specific lncRNAs with potential roles in regulating human NK cell differentiation and function. Expression of lnc-CD56, an NK-specific lncRNA, was positively correlated with that of CD56, a classical human NK cell surface marker. We showed that lnc-CD56 may function as a positive regulator of CD56 in primary human NK cells and differentiated NK cells from human CD34+ hematopoietic progenitor cells. Our data provide an annotated human NK cell lncRNA expression catalog and demonstrate a key role for lncRNAs in NK cell biology. PMID:27713137

  10. Individual differences in positive and negative emotion regulation: Which strategies explain variability in loneliness?

    PubMed

    Kearns, Sinead M; Creaven, Ann-Marie

    2017-02-01

    Loneliness is the distressing feeling accompanying the perception that one's social needs are not being met by one's social relationships. Conceptual models point to a role for cognitive factors in this experience. Because research on determinants of loneliness is sparse, this study investigates associations between individual differences in emotion regulation (ER) and loneliness. Participants (N = 116) completed measures of loneliness, and a vignette-based measure of adaptive and maladaptive ER in response to positive and negative scenarios. Regression analyses indicated that the regulation of positive and negative emotions explained comparable variance in loneliness, and associations were only partially reduced by the inclusion of social support. The specific strategies positive reappraisal, being present and negative mental time travel explained the most variance in loneliness. The findings are consistent with both the cognitive and the social needs models of loneliness and suggest that variability in ER strategies should be considered relevant to loneliness. Copyright © 2016 John Wiley & Sons, Ltd.

  11. Positive and Negative Associations between Adolescents’ Religiousness and Health Behaviors via Self-Regulation

    PubMed Central

    Holmes, Christopher J.; Kim-Spoon, Jungmeen

    2015-01-01

    It has been proposed that self-regulation may be the explanatory mechanism for the relation between religiousness and positive health behaviors. However, different religious motivations have differential effects on a variety of health related outcomes, which may explain the adverse effects of religiousness found in some studies. The current study hypothesized that higher identification as religious motivation would be linked to higher health-promoting behavior and lower health-risk behavior through higher self-regulation, whereas higher introjection would be linked to lower health-promoting behavior and higher health-risk behavior through lower self-regulation. The sample included 220 adolescents (mean age = 15 years, 55% male) and their primary caregivers. Structural equation modeling results supported the hypotheses and indicated that adolescent self-regulation mediated the relations between their religious motivation and health behavior. The findings suggest that different types of religious motivation may be promotive or hindering for adolescents’ health. PMID:27595048

  12. Transcriptional regulator LsrB of Sinorhizobium meliloti positively regulates the expression of genes involved in lipopolysaccharide biosynthesis.

    PubMed

    Tang, Guirong; Wang, Ying; Luo, Li

    2014-09-01

    Rhizobia induce nitrogen-fixing nodules on host legumes, which is important in agriculture and ecology. Lipopolysaccharide (LPS) produced by rhizobia is required for infection or bacteroid survival in host cells. Genes required for LPS biosynthesis have been identified in several Rhizobium species. However, the regulation of their expression is not well understood. Here, Sinorhizobium meliloti LsrB, a member of the LysR family of transcriptional regulators, was found to be involved in LPS biosynthesis by positively regulating the expression of the lrp3-lpsCDE operon. An lsrB in-frame deletion mutant displayed growth deficiency, sensitivity to the detergent sodium dodecyl sulfate, and acidic pH compared to the parent strain. This mutant produced slightly less LPS due to lower expression of the lrp3 operon. Analysis of the transcriptional start sites of the lrp3 and lpsCDE gene suggested that they constitute one operon. The expression of lsrB was positively autoregulated. The promoter region of lrp3 was specifically precipitated by anti-LsrB antibodies in vivo. The promoter DNA fragment containing TN11A motifs was bound by the purified LsrB protein in vitro. These new findings suggest that S. meliloti LsrB is associated with LPS biosynthesis, which is required for symbiotic nitrogen fixation on some ecotypes of alfalfa plants.

  13. A new Bacillus subtilis gene, med, encodes a positive regulator of comK.

    PubMed

    Ogura, M; Ohshiro, Y; Hirao, S; Tanaka, T

    1997-10-01

    Bacillus subtilis degR, a positive regulator of the production of degradative enzymes, is negatively regulated by the competence transcription factor ComK which is overproduced in mecA null mutants. We used transposon Tn10 to search for a mutation that reduced the repression level of degR caused by a mecA mutation. A new gene exerting positive regulation on comK was obtained and designated med (suppressor of mecA effect on degR). Sequence determination, Northern analysis, and primer extension analyses revealed that the med gene contained an open reading frame (ORF) composed of 317 codons and was transcribed into an approximately 1,250-nucleotide mRNA together with its short downstream gene. The expression of comK is positively regulated by factors such as ComK itself, ComS (SrfA)-MecA, DegU, SinR, and AbrB. Quantitative analyses using comK'-'lacZ, srfA-lacZ, degU'-'lacZ, and sinR'-'lacZ fusions showed that disruption of med caused a significant decrease in comK expression in both mecA+ and mecA strains, while expression of srfA, sinR, and degU was not affected by the mutation. An epistatic analysis revealed that overproduction of ComK resulted in alteration of med expression, suggesting a regulatory loop between comK and med. Several possible mechanisms for positive regulation of comK by Med are discussed.

  14. Neurokinin-1 receptor is a novel positive regulator of Wnt/β-catenin signaling in melanogenesis

    PubMed Central

    Song, Huizhu; Lv, Bei; Wang, Lei; Shang, Jing; Wang, Yong; Chang, Chunyan; Ping, Fengfeng; Qian, Jun

    2016-01-01

    Wnt/β-catenin signaling is essential for melanogenesis in melanocytes. Neurokinin-1 receptor (NK-1R) has recently been demonstrated to be involved in melanin production. However, the cross talk between NK-1R and Wnt/β-catenin is poorly understood. Here, [Sar9, Met(O2)11] substance P (SMSP) was used to activate NK-1R, while L-733060 was used to inhibit it. The effects of NK-1R activation and inhibition on Wnt and its inhibitors were analyzed using western blot and real-time quantitative PCR. The results showed that SMSP positively regulated Wnt/β-catenin signaling by increasing the expression of β-catenin and p-GSK3β protein, which resulted from the weakened expression of the Wnt inhibitor Dickkopf-1 (DKK1). On the contrary, L-733060 lowered the expression of β-catenin and p-GSK3β protein through the up-regulation of DKK1 expression. Furthermore, in L-733060-treated mice, it was found that the pigmentation level as well as the melanogenic proteins and β-catenin protein expression were down-regulated, while the expression of DKK1 was up-regulated. These results showed the interaction between NK-1R and Wnt in human melanocytes in vitro and C57BL/6J mice in vivo, indicating that NK-1R may positively regulate melanogenesis through Wnt/β-catenin signaling pathway. PMID:27835606

  15. Social anxiety and emotion regulation in daily life: spillover effects on positive and negative social events.

    PubMed

    Farmer, Antonina Savostyanova; Kashdan, Todd B

    2012-01-01

    To minimize the possibility of scrutiny, people with social anxiety difficulties exert great effort to manage their emotions, particularly during social interactions. We examined how the use of two emotion regulation strategies, emotion suppression and cognitive reappraisal, predict the generation of emotions and social events in daily life. Over 14 consecutive days, 89 participants completed daily diary entries on emotions, positive and negative social events, and their regulation of emotions. Using multilevel modeling, we found that when people high in social anxiety relied more on positive emotion suppression, they reported fewer positive social events and less positive emotion on the subsequent day. In contrast, people low in social anxiety reported fewer negative social events on days subsequent to using cognitive reappraisal to reduce distress; the use of cognitive reappraisal did not influence the daily lives of people high in social anxiety. Our findings support theories of emotion regulation difficulties associated with social anxiety. In particular, for people high in social anxiety, maladaptive strategy use contributed to diminished reward responsiveness.

  16. Interaction with the Yes-associated protein (YAP) allows TEAD1 to positively regulate NAIP expression.

    PubMed

    Landin Malt, André; Georges, Adrien; Silber, Joël; Zider, Alain; Flagiello, Domenico

    2013-10-01

    Although the expression of the neuronal apoptosis inhibitory protein (NAIP) gene is considered involved in apoptosis suppression as well as in inflammatory response, the molecular basis of the NAIP gene expression is poorly understood. Here we show that the TEA domain protein 1 (TEAD1) is able to positively activate the transcription of NAIP. We further demonstrate that this regulation is mediated by the presence of the endogenous Yes associated protein (YAP) cofactor, and requires the interaction with YAP. We finally identified an intronic region of the NAIP gene responding to TEAD1/YAP activity, suggesting that regulation of NAIP by TEAD1/YAP is at the transcriptional level.

  17. CTCF regulates positioning of the human cystic fibrosis gene in association with a histone deacetylase

    PubMed Central

    Muck, Joscha

    2014-01-01

    The nuclear positioning of mammalian genes often correlates with their functional state. For instance, the human cystic fibrosis transmembrane conductance regulator (CFTR) gene associates with the nuclear periphery in its inactive state, but occupies interior positions when active. Treatment with the histone deacetylase inhibitor trichostatin a (TSA) changes the radial positioning of the CFTR gene in HeLa S3 cells. The gene relocates from the nuclear periphery to the nuclear interior. In Calu-3 cells the gene is located in the nuclear interior. To identify potential regulatory elements for the positioning of CFTR, the histone H3 and H4 acetylation patterns of untreated and TSA-treated HeLa S3 and untreated Calu-3 cells were determined by ChIP–chip. Here is a detailed description of the datasets associated with the study by Muck et al. published in the Journal of Cellular Biochemistry in 2012. PMID:26484076

  18. Hedgehog is a positive regulator of FGF signalling during embryonic tracheal cell migration.

    PubMed

    Butí, Elisenda; Mesquita, Duarte; Araújo, Sofia J

    2014-01-01

    Cell migration is a widespread and complex process that is crucial for morphogenesis and for the underlying invasion and metastasis of human cancers. During migration, cells are steered toward target sites by guidance molecules that induce cell direction and movement through complex intracellular mechanisms. The spatio-temporal regulation of the expression of these guidance molecules is of extreme importance for both normal morphogenesis and human disease. One way to achieve this precise regulation is by combinatorial inputs of different transcription factors. Here we used Drosophila melanogaster mutants with migration defects in the ganglionic branches of the tracheal system to further clarify guidance regulation during cell migration. By studying the cellular consequences of overactivated Hh signalling, using ptc mutants, we found that Hh positively regulates Bnl/FGF levels during embryonic stages. Our results show that Hh modulates cell migration non-autonomously in the tissues surrounding the action of its activity. We further demonstrate that the Hh signalling pathway regulates bnl expression via Stripe (Sr), a zinc-finger transcription factor with homology to the Early Growth Response (EGR) family of vertebrate transcription factors. We propose that Hh modulates embryonic cell migration by participating in the spatio-temporal regulation of bnl expression in a permissive mode. By doing so, we provide a molecular link between the activation of Hh signalling and increased chemotactic responses during cell migration.

  19. JMJD8 is a positive regulator of TNF-induced NF-κB signaling.

    PubMed

    Yeo, Kok Siong; Tan, Ming Cheang; Wong, Wan Ying; Loh, Sheng Wei; Lam, Yi Lyn; Tan, Chin Leng; Lim, Yat-Yuen; Ea, Chee-Kwee

    2016-09-27

    TNF-induced signaling mediates pleiotropic biological consequences including inflammation, immunity, cell proliferation and apoptosis. Misregulation of TNF signaling has been attributed as a major cause of chronic inflammatory diseases and cancer. Jumonji domain-containing protein 8 (JMJD8) belongs to the JmjC family. However, only part of the family members has been described as hydroxylase enzymes that function as histone demethylases. Here, we report that JMJD8 positively regulates TNF-induced NF-κB signaling. Silencing the expression of JMJD8 using RNA interference (RNAi) greatly suppresses TNF-induced expression of several NF-κB-dependent genes. Furthermore, knockdown of JMJD8 expression reduces RIP ubiquitination, IKK kinase activity, delays IκBα degradation and subsequently blocks nuclear translocation of p65. In addition, JMJD8 deficiency enhances TNF-induced apoptosis. Taken together, these findings indicate that JMJD8 functions as a positive regulator of TNF-induced NF-κB signaling.

  20. Ellis Englesberg and the discovery of positive control in gene regulation.

    PubMed

    Hahn, Steven

    2014-10-01

    Based on his work with the Escherichia coli l-arabinose operon, Ellis Englesberg proposed in 1965 that the regulatory gene araC was an "activator gene" required for positive control of the ara operon. This challenged the widely held belief in a universal mechanism of negative regulation proposed earlier by Jacob and Monod. For years, Englesberg's model was met with deep skepticism. Despite much frustration with complex ad hoc explanations used to challenge his model, Englesberg persisted until the evidence for positive control in ara and other systems became overwhelming. Englesberg's pioneering work enriched the original operon model and had a lasting impact in opening new and exciting ways of thinking about transcriptional regulation.

  1. Light is a positive regulator of strigolactone levels in tomato roots.

    PubMed

    Koltai, Hinanit; Cohen, Maja; Chesin, Ori; Mayzlish-Gati, Einav; Bécard, Guillaume; Puech, Virginie; Ben Dor, Bruria; Resnick, Natalie; Wininger, Smadar; Kapulnik, Yoram

    2011-11-01

    Strigolactones (SLs) or closely related molecules were recently identified as phytohormones, acting as long-distance branching factors that suppress growth of pre-formed axillary buds in the shoot. The SL signaling pathways and light appear to be connected, as SLs were shown to induce light-regulated pathways and to mimic light-adapted plant growth. However, it is not yet clear how light affects SL levels. Here, we examined the effect of different light intensities on SL levels in tomato roots. The results show that light intensity, above a certain threshold, is a positive regulator of SL levels and of Sl-CCD7 transcription; Sl-CCD7 is involved in SLs biosynthesis in tomato. Moreover, SL accumulation in plant roots is shown to be a time-dependent process. At least some of the similar effects of light and SLs on plant responses might result from a positive effect of light on SL levels.

  2. The silent information regulator 1 (Sirt1) is a positive regulator of the Notch pathway in Drosophila.

    PubMed

    Horvath, Matej; Mihajlovic, Zorana; Slaninova, Vera; Perez-Gomez, Raquel; Moshkin, Yuri; Krejci, Alena

    2016-11-15

    The silent information regulator 1 (Sirt1) has been shown to have negative effects on the Notch pathway in several contexts. We bring evidence that Sirt1 has a positive effect on Notch activation in Drosophila, in the context of sensory organ precursor specification and during wing development. The phenotype of Sirt1 mutant resembles weak Notch loss-of-function phenotypes, and genetic interactions of Sirt1 with the components of the Notch pathway also suggest a positive role for Sirt1 in Notch signalling. Sirt1 is necessary for the efficient activation of enhancer of split [E(spl)] genes by Notch in S2N cells. Additionally, the Notch-dependent response of several E(spl) genes is sensitive to metabolic stress caused by 2-deoxy-d-glucose treatment, in a Sirt1-dependent manner. We found Sirt1 associated with several proteins involved in Notch repression as well as activation, including the cofactor exchange factor Ebi (TBL1), the RLAF/LAF histone chaperone complex and the Tip60 acetylation complex. Moreover, Sirt1 participates in the deacetylation of the CSL transcription factor Suppressor of Hairless. The role of Sirt1 in Notch signalling is, therefore, more complex than previously recognized, and its diverse effects may be explained by a plethora of Sirt1 substrates involved in the regulation of Notch signalling.

  3. Transcription factor Fli-1 positively regulates lipopolysaccharide-induced interleukin-27 production in macrophages.

    PubMed

    Gao, Peng; Yuan, Ming; Ma, Xianwei; Jiang, Wei; Zhu, Lingxi; Wen, Mingyue; Xu, Jing; Liu, Qiuyan; An, Huazhang

    2016-03-01

    IL-27 is an important regulator of TLR4-activated innate immune. The mechanism by which IL-27 production is regulated in TLR4-activated innate immune remains largely unclear. Here we show that expression of transcription factor Fli-1 at protein level is increased in macrophages following LPS stimulation. Fli-1 overexpression increases LPS-activated IL-27 production in macrophages. Consistently, Fli-1 knockdown inhibits LPS-induced IL-27 production in macrophages. Chromatin immunoprecipitation (ChIP) assay reveals that Fli-1 binds the promoter of IL-27 p28 subunit. Further experiments manifest that Fli-1 binds the region between -250 and -150 bp upstream of the transcriptional start site of p28 gene and increases p28 gene promoter-controlled transcription. These results demonstrate that Fli-1 positively regulates IL-27 production in TLR4-activated immune response by promoting transcription of IL-27 p28 gene.

  4. Cytokinin as a positional cue regulating lateral root spacing in Arabidopsis.

    PubMed

    Chang, Ling; Ramireddy, Eswarayya; Schmülling, Thomas

    2015-08-01

    The root systems of plants have developed adaptive architectures to exploit soil resources. The formation of lateral roots (LRs) contributes to root system architecture. Roots of plants with a lower cytokinin status form LR primordia (LRP) in unusually close proximity, indicating a role for the hormone in regulating the positioning of LRs along the main root axis. Data obtained from cytokinin-synthesis mutants of Arabidopsis thaliana combined with gene expression analysis indicate that cytokinin synthesis by IPT5 and LOG4 occurring early during LRP initiation generates a local cytokinin signal abbreviating LRP formation in neighbouring pericycle cells. In addition, IPT3, IPT5, and IPT7 contribute to cytokinin synthesis in the vicinity of existing LRP, thus suppressing initiation of new LRs. Interestingly, mutation of CYP735A genes required for trans-zeatin biosynthesis caused strong defects in LR positioning, indicating an important role for this cytokinin metabolite in regulating LR spacing. Further it is shown that cytokinin and a known regulator of LR spacing, the receptor-like kinase ARABIDOPSIS CRINKLY4 (ACR4), operate in a non-hierarchical manner but might exert reciprocal control at the transcript level. Taken together, the results suggest that cytokinin acts as a paracrine hormonal signal in regulating root system architecture.

  5. Positive and negative regulation of a SNARE protein by control of intracellular localization.

    PubMed

    Nakanishi, Hideki; de los Santos, Pablo; Neiman, Aaron M

    2004-04-01

    In Saccharomyces cerevisiae, the developmentally regulated Soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) protein Spo20p mediates the fusion of vesicles with the prospore membrane, which is required for the formation of spores. Spo20p is subject to both positive and negative regulation by separate sequences in its aminoterminal domain. We report that the positive activity is conferred by a short, amphipathic helix that is sufficient to confer plasma membrane or prospore membrane localization to green fluorescent protein. In vitro, this helix binds to acidic phospholipids, and mutations that reduce or eliminate phospholipid binding in vitro inactivate Spo20p in vivo. Genetic manipulation of phospholipid pools indicates that the likely in vivo ligand of this domain is phosphatidic acid. The inhibitory activity is a nuclear targeting signal, which confers nuclear localization in vegetative cells and in cells entering meiosis. However, as cells initiate spore formation, fusions containing the inhibitory domain exit the nucleus and localize to the nascent prospore membrane. Thus, the SNARE Spo20p is both positively and negatively regulated by control of its intracellular localization.

  6. OsGF14b Positively Regulates Panicle Blast Resistance but Negatively Regulates Leaf Blast Resistance in Rice.

    PubMed

    Liu, Qing; Yang, Jianyuan; Zhang, Shaohong; Zhao, Junliang; Feng, Aiqing; Yang, Tifeng; Wang, Xiaofei; Mao, Xinxue; Dong, Jingfang; Zhu, Xiaoyuan; Leung, Hei; Leach, Jan E; Liu, Bin

    2016-01-01

    Although 14-3-3 proteins have been reported to be involved in responses to biotic stresses in plants, their functions in rice blast, the most destructive disease in rice, are largely unknown. Only GF14e has been confirmed to negatively regulate leaf blast. We report that GF14b is highly expressed in seedlings and panicles during blast infection. Rice plants overexpressing GF14b show enhanced resistance to panicle blast but are susceptible to leaf blast. In contrast, GF14b-silenced plants show increased susceptibility to panicle blast but enhanced resistance to leaf blast. Yeast one-hybrid assays demonstrate that WRKY71 binds to the promoter of GF14b and modulates its expression. Overexpression of GF14b induces expression of jasmonic acid (JA) synthesis-related genes but suppresses expression of salicylic acid (SA) synthesis-related genes. In contrast, suppressed GF14b expression causes decreased expression of JA synthesis-related genes but activation of SA synthesis-related genes. These results suggest that GF14b positively regulates panicle blast resistance but negatively regulates leaf blast resistance, and that GF14b-mediated disease resistance is associated with the JA- and SA-dependent pathway. The different functions for 14-3-3 proteins in leaf and panicle blast provide new evidence that leaf and panicle blast resistance are controlled by different mechanisms.

  7. Nitric oxide acts as a positive regulator to induce metamorphosis of the ascidian Herdmania momus.

    PubMed

    Ueda, Nobuo; Degnan, Sandie M

    2013-01-01

    Marine invertebrates commonly have a biphasic life cycle in which the metamorphic transition from a pelagic larva to a benthic post-larva is mediated by the nitric oxide signalling pathway. Nitric oxide (NO) is synthesised by nitric oxide synthase (NOS), which is a client protein of the molecular chaperon heat shock protein 90 (HSP90). It is notable, then, that both NO and HSP90 have been implicated in regulating metamorphosis in marine invertebrates as diverse as urochordates, echinoderms, molluscs, annelids, and crustaceans. Specifically, the suppression of NOS activity by the application of either NOS- or HSP90-inhibiting pharmacological agents has been shown consistently to induce the initiation of metamorphosis, leading to the hypothesis that a negative regulatory role of NO is widely conserved in biphasic life cycles. Further, the induction of metamorphosis by heat-shock has been demonstrated for multiple species. Here, we investigate the regulatory role of NO in induction of metamorphosis of the solitary tropical ascidian, Herdmania momus. By coupling pharmacological treatments with analysis of HmNOS and HmHSP90 gene expression, we present compelling evidence of a positive regulatory role for NO in metamorphosis of this species, in contrast to all existing ascidian data that supports the hypothesis of NO as a conserved negative regulator of metamorphosis. The exposure of competent H. momus larvae to a NOS inhibitor or an NO donor results in an up-regulation of NOS and HSP90 genes. Heat shock of competent larvae induces metamorphosis in a temperature dependent manner, up to a thermal tolerance that approaches 35°C. Both larval/post-larval survival and the appearance of abnormal morphologies in H. momus post-larvae reflect the magnitude of up-regulation of the HSP90 gene in response to heat-shock. The demonstrated role of NO as a positive metamorphic regulator in H. momus suggests the existence of inter-specific adaptations of NO regulation in ascidian

  8. Nitric Oxide Acts as a Positive Regulator to Induce Metamorphosis of the Ascidian Herdmania momus

    PubMed Central

    Ueda, Nobuo; Degnan, Sandie M.

    2013-01-01

    Marine invertebrates commonly have a biphasic life cycle in which the metamorphic transition from a pelagic larva to a benthic post-larva is mediated by the nitric oxide signalling pathway. Nitric oxide (NO) is synthesised by nitric oxide synthase (NOS), which is a client protein of the molecular chaperon heat shock protein 90 (HSP90). It is notable, then, that both NO and HSP90 have been implicated in regulating metamorphosis in marine invertebrates as diverse as urochordates, echinoderms, molluscs, annelids, and crustaceans. Specifically, the suppression of NOS activity by the application of either NOS- or HSP90-inhibiting pharmacological agents has been shown consistently to induce the initiation of metamorphosis, leading to the hypothesis that a negative regulatory role of NO is widely conserved in biphasic life cycles. Further, the induction of metamorphosis by heat-shock has been demonstrated for multiple species. Here, we investigate the regulatory role of NO in induction of metamorphosis of the solitary tropical ascidian, Herdmania momus. By coupling pharmacological treatments with analysis of HmNOS and HmHSP90 gene expression, we present compelling evidence of a positive regulatory role for NO in metamorphosis of this species, in contrast to all existing ascidian data that supports the hypothesis of NO as a conserved negative regulator of metamorphosis. The exposure of competent H. momus larvae to a NOS inhibitor or an NO donor results in an up-regulation of NOS and HSP90 genes. Heat shock of competent larvae induces metamorphosis in a temperature dependent manner, up to a thermal tolerance that approaches 35°C. Both larval/post-larval survival and the appearance of abnormal morphologies in H. momus post-larvae reflect the magnitude of up-regulation of the HSP90 gene in response to heat-shock. The demonstrated role of NO as a positive metamorphic regulator in H. momus suggests the existence of inter-specific adaptations of NO regulation in ascidian

  9. Soybean Homologs of MPK4 Negatively Regulate Defense Responses and Positively Regulate Growth and Development1[W][OA

    PubMed Central

    Liu, Jian-Zhong; Horstman, Heidi D.; Braun, Edward; Graham, Michelle A.; Zhang, Chunquan; Navarre, Duroy; Qiu, Wen-Li; Lee, Yeunsook; Nettleton, Dan; Hill, John H.; Whitham, Steven A.

    2011-01-01

    Mitogen-activated protein kinase (MAPK) cascades play important roles in disease resistance in model plant species such as Arabidopsis (Arabidopsis thaliana) and tobacco (Nicotiana tabacum). However, the importance of MAPK signaling pathways in the disease resistance of crops is still largely uninvestigated. To better understand the role of MAPK signaling pathways in disease resistance in soybean (Glycine max), 13, nine, and 10 genes encoding distinct MAPKs, MAPKKs, and MAPKKKs, respectively, were silenced using virus-induced gene silencing mediated by Bean pod mottle virus. Among the plants silenced for various MAPKs, MAPKKs, and MAPKKKs, those in which GmMAPK4 homologs (GmMPK4s) were silenced displayed strong phenotypes including stunted stature and spontaneous cell death on the leaves and stems, the characteristic hallmarks of activated defense responses. Microarray analysis showed that genes involved in defense responses, such as those in salicylic acid (SA) signaling pathways, were significantly up-regulated in GmMPK4-silenced plants, whereas genes involved in growth and development, such as those in auxin signaling pathways and in cell cycle and proliferation, were significantly down-regulated. As expected, SA and hydrogen peroxide accumulation was significantly increased in GmMPK4-silenced plants. Accordingly, GmMPK4-silenced plants were more resistant to downy mildew and Soybean mosaic virus compared with vector control plants. Using bimolecular fluorescence complementation analysis and in vitro kinase assays, we determined that GmMKK1 and GmMKK2 might function upstream of GmMPK4. Taken together, our results indicate that GmMPK4s negatively regulate SA accumulation and defense response but positively regulate plant growth and development, and their functions are conserved across plant species. PMID:21878550

  10. Spatial organization of the extracellular matrix regulates cell–cell junction positioning

    PubMed Central

    Tseng, Qingzong; Duchemin-Pelletier, Eve; Deshiere, Alexandre; Balland, Martial; Guillou, Hervé; Filhol, Odile; Théry, Manuel

    2012-01-01

    The organization of cells into epithelium depends on cell interaction with both the extracellular matrix (ECM) and adjacent cells. The role of cell–cell adhesion in the regulation of epithelial topology is well-described. ECM is better known to promote cell migration and provide a structural scaffold for cell anchoring, but its contribution to multicellular morphogenesis is less well-understood. We developed a minimal model system to investigate how ECM affects the spatial organization of intercellular junctions. Fibronectin micropatterns were used to constrain the location of cell–ECM adhesion. We found that ECM affects the degree of stability of intercellular junction positioning and the magnitude of intra- and intercellular forces. Intercellular junctions were permanently displaced, and experienced large perpendicular tensional forces as long as they were positioned close to ECM. They remained stable solely in regions deprived of ECM, where they were submitted to lower tensional forces. The heterogeneity of the spatial organization of ECM induced anisotropic distribution of mechanical constraints in cells, which seemed to adapt their position to minimize both intra- and intercellular forces. These results uncover a morphogenetic role for ECM in the mechanical regulation of cells and intercellular junction positioning. PMID:22307605

  11. Positive and negative regulation of T-cell activation through kinases and phosphatases.

    PubMed Central

    Mustelin, Tomas; Taskén, Kjetil

    2003-01-01

    The sequence of events in T-cell antigen receptor (TCR) signalling leading to T-cell activation involves regulation of a number of protein tyrosine kinases (PTKs) and the phosphorylation status of many of their substrates. Proximal signalling pathways involve PTKs of the Src, Syk, Csk and Tec families, adapter proteins and effector enzymes in a highly organized tyrosine-phosphorylation cascade. In intact cells, tyrosine phosphorylation is rapidly reversible and generally of a very low stoichiometry even under induced conditions due to the fact that the enzymes removing phosphate from tyrosine-phosphorylated substrates, the protein tyrosine phosphatases (PTPases), have a capacity that is several orders of magnitude higher than that of the PTKs. It follows that a relatively minor change in the PTK/PTPase balance can have a major impact on net tyrosine phosphorylation and thereby on activation and proliferation of T-cells. This review focuses on the involvement of PTKs and PTPases in positive and negative regulation of T-cell activation, the emerging theme of reciprocal regulation of each type of enzyme by the other, as well as regulation of phosphotyrosine turnover by Ser/Thr phosphorylation and regulation of localization of signal components. PMID:12485116

  12. Expression of POEM, a positive regulator of osteoblast differentiation, is suppressed by TNF-{alpha}

    SciTech Connect

    Tsukasaki, Masayuki; Yamada, Atsushi; Suzuki, Dai; Aizawa, Ryo; Miyazono, Agasa; Miyamoto, Yoichi; Suzawa, Tetsuo; Takami, Masamichi; Yoshimura, Kentaro; Morimura, Naoko; Yamamoto, Matsuo; Kamijo, Ryutaro

    2011-07-15

    Highlights: {yields} TNF-{alpha} inhibits POEM gene expression. {yields} Inhibition of POEM gene expression is caused by NF-{kappa}B activation by TNF-{alpha}. {yields} Over-expression of POEM recovers inhibition of osteoblast differentiation by TNF-{alpha}. -- Abstract: POEM, also known as nephronectin, is an extracellular matrix protein considered to be a positive regulator of osteoblast differentiation. In the present study, we found that tumor necrosis factor-{alpha} (TNF-{alpha}), a key regulator of bone matrix properties and composition that also inhibits terminal osteoblast differentiation, strongly inhibited POEM expression in the mouse osteoblastic cell line MC3T3-E1. TNF-{alpha}-induced down-regulation of POEM gene expression occurred in both time- and dose-dependent manners through the nuclear factor kappa B (NF-{kappa}B) pathway. In addition, expressions of marker genes in differentiated osteoblasts were down-regulated by TNF-{alpha} in a manner consistent with our findings for POEM, while over-expression of POEM recovered TNF-{alpha}-induced inhibition of osteoblast differentiation. These results suggest that TNF-{alpha} inhibits POEM expression through the NF-{kappa}B signaling pathway and down-regulation of POEM influences the inhibition of osteoblast differentiation by TNF-{alpha}.

  13. Identification of E2F1 as a positive transcriptional regulator for {delta}-catenin

    SciTech Connect

    Kim, Kwonseop; Oh, Minsoo; Ki, Hyunkyoung; Wang Tao; Bareiss, Sonja; Fini, M. Elizabeth.; Li Dawei; Lu Qun

    2008-05-02

    {delta}-Catenin is upregulated in human carcinomas. However, little is known about the potential transcriptional factors that regulate {delta}-catenin expression in cancer. Using a human {delta}-catenin reporter system, we have screened several nuclear signaling modulators to test whether they can affect {delta}-catenin transcription. Among {beta}-catenin/LEF-1, Notch1, and E2F1, E2F1 dramatically increased {delta}-catenin-luciferase activities while {beta}-catenin/LEF-1 induced only a marginal increase. Rb suppressed the upregulation of {delta}-catenin-luciferase activities induced by E2F1 but did not interact with {delta}-catenin. RT-PCR and Western blot analyses in 4 different prostate cancer cell lines revealed that regulation of {delta}-catenin expression is controlled mainly at the transcriptional level. Interestingly, the effects of E2F1 on {delta}-catenin expression were observed only in human cancer cells expressing abundant endogenous {delta}-catenin. These studies identify E2F1 as a positive transcriptional regulator for {delta}-catenin, but further suggest the presence of strong negative regulator(s) for {delta}-catenin in prostate cancer cells with minimal endogenous {delta}-catenin expression.

  14. Protein kinase D regulates positive selection of CD4+ thymocytes through phosphorylation of SHP-1

    PubMed Central

    Ishikawa, Eri; Kosako, Hidetaka; Yasuda, Tomoharu; Ohmuraya, Masaki; Araki, Kimi; Kurosaki, Tomohiro; Saito, Takashi; Yamasaki, Sho

    2016-01-01

    Thymic selection shapes an appropriate T cell antigen receptor (TCR) repertoire during T cell development. Here, we show that a serine/threonine kinase, protein kinase D (PKD), is crucial for thymocyte positive selection. In T cell-specific PKD-deficient (PKD2/PKD3 double-deficient) mice, the generation of CD4 single positive thymocytes is abrogated. This defect is likely caused by attenuated TCR signalling during positive selection and incomplete CD4 lineage specification in PKD-deficient thymocytes; however, TCR-proximal tyrosine phosphorylation is not affected. PKD is activated in CD4+CD8+ double positive (DP) thymocytes on stimulation with positively selecting peptides. By phosphoproteomic analysis, we identify SH2-containing protein tyrosine phosphatase-1 (SHP-1) as a direct substrate of PKD. Substitution of wild-type SHP-1 by phosphorylation-defective mutant (SHP-1S557A) impairs generation of CD4+ thymocytes. These results suggest that the PKD–SHP-1 axis positively regulates TCR signalling to promote CD4+ T cell development. PMID:27670070

  15. A positive feedback regulation of ISL-1 in DLBCL but not in pancreatic β-cells

    SciTech Connect

    Zhang, Qiao; Yang, Zhe; Wang, Weiping; Guo, Ting; Jia, Zhuqing; Ma, Kangtao; Zhou, Chunyan

    2014-07-04

    Highlights: • ISL-1 is highly expressed in human pancreatic β-cells and DLBCL. • ISL-1 accelerates the tumorigenesis of DLBCL in vivo. • c-Myc positively regulates ISL-1 expression in DLBCL but not in pancreatic β-cells. • ISL-1 and c-Myc forms an ISL-1/c-Myc transcriptional complex only in DLBCL. • Positive feedback regulation of ISL-1 does not exist in normal pancreatic β-cell. - Abstract: Insulin enhancer binding protein-1 (ISL-1), a LIM-homeodomain transcription factor, has been reported to play essential roles in promoting adult pancreatic β-cells proliferation. Recent studies indicate that ISL-1 may also involve in the occurrence of a variety of tumors. However, whether ISL-1 has any functional effect on tumorigenesis, and what are the differences on ISL-1 function in distinct conditions, are completely unknown. In this study, we found that ISL-1 was highly expressed in human pancreatic β-cells, as well as in diffuse large B cell lymphoma (DLBCL), but to a much less extent in other normal tissues or tumor specimens. Further study revealed that ISL-1 promoted the proliferation of pancreatic β-cells and DLBCL cells, and also accelerated the tumorigenesis of DLBCL in vivo. We also found that ISL-1 could activate c-Myc transcription not only in pancreatic β-cells but also in DLBCL cells. However, a cell-specific feedback regulation was detectable only in DLBCL cells. This auto-regulatory loop was established by the interaction of ISL-1 and c-Myc to form an ISL-1/c-Myc transcriptional complex, and synergistically to promote ISL-1 transcription through binding on the ISL-1 promoter. Taken together, our results demonstrate a positive feedback regulation of ISL-1 in DLBCL but not in pancreatic β-cells, which might result in the functional diversities of ISL-1 in different physiological and pathological processes.

  16. The Role of Positive Personality Traits in Emotion Regulation of Patients with Irritable Bowel Syndrome (IBS)

    PubMed Central

    MAZAHERI, Mina; NIKNESHAN, Shekoufeh; DAGHAGHZADEH, Hamed; AFSHAR, Hamid

    2015-01-01

    Background: Personality traits and emotion regulation processes play an important role in human health. The purpose of this study was to investigate the role of positive personality traits (psychological hardiness and interpersonal forgiveness) in emotion regulation of patients with Irritable Bowel Syndrome. Methods: The research was a cross-sectional study. Statistical population included all of IBS patients referred to the Subspecialty Center of Psychiatry in Isfahan in 2013. Overall, 123 subjects (100 women, 83.3%, and 30 men, 16.7%) were selected by census method, according to criteria of research and during a particular period. To collect data, the Difficulties in Emotion Regulation Scale (DERS), Lang and Goulet Hardiness Scale (LGHS) and Interpersonal forgiveness Inventory (IFI) were used. Data was analyzed using Pearson’s correlation coefficient and Multivariate and Binary Logistic regression analyses. Results: Mean age of patients was 33.82±10.45 years and 83.3% (100) of them were female. Regression analyses showed that both personality traits of hardiness and forgiveness were as protective factors for emotional dysregulation with OR, 95% CI: 0.93 and 0.96 sequentially, with adjusting demographic variables (age, gender, and education level and disease duration). Conclusion: Patients who are more hardy and forgiving toward others, are likely more successful at adaptive emotion regulation. It emphasizes the positive and beneficial role of the personality traits in regulating of emotional problems of IBS patients. Hence, these variables should be considered as effective factors in the treatment process of the patients. PMID:26056675

  17. The unique regulation of iron-sulfur cluster biogenesis in a Gram-positive bacterium

    PubMed Central

    Santos, Joana A.; Alonso-García, Noelia; Macedo-Ribeiro, Sandra; Pereira, Pedro José Barbosa

    2014-01-01

    Iron-sulfur clusters function as cofactors of a wide range of proteins, with diverse molecular roles in both prokaryotic and eukaryotic cells. Dedicated machineries assemble the clusters and deliver them to the final acceptor molecules in a tightly regulated process. In the prototypical Gram-negative bacterium Escherichia coli, the two existing iron-sulfur cluster assembly systems, iron-sulfur cluster (ISC) and sulfur assimilation (SUF) pathways, are closely interconnected. The ISC pathway regulator, IscR, is a transcription factor of the helix-turn-helix type that can coordinate a [2Fe-2S] cluster. Redox conditions and iron or sulfur availability modulate the ligation status of the labile IscR cluster, which in turn determines a switch in DNA sequence specificity of the regulator: cluster-containing IscR can bind to a family of gene promoters (type-1) whereas the clusterless form recognizes only a second group of sequences (type-2). However, iron-sulfur cluster biogenesis in Gram-positive bacteria is not so well characterized, and most organisms of this group display only one of the iron-sulfur cluster assembly systems. A notable exception is the unique Gram-positive dissimilatory metal reducing bacterium Thermincola potens, where genes from both systems could be identified, albeit with a diverging organization from that of Gram-negative bacteria. We demonstrated that one of these genes encodes a functional IscR homolog and is likely involved in the regulation of iron-sulfur cluster biogenesis in T. potens. Structural and biochemical characterization of T. potens and E. coli IscR revealed a strikingly similar architecture and unveiled an unforeseen conservation of the unique mechanism of sequence discrimination characteristic of this distinctive group of transcription regulators. PMID:24847070

  18. Identification of HEXIM1 as a Positive Regulator of p53*

    PubMed Central

    Lew, Qiao Jing; Chia, Yi Ling; Chu, Kai Ling; Lam, Yuen Ting; Gurumurthy, Meera; Xu, Shengli; Lam, Kong Peng; Cheong, Nge; Chao, Sheng-Hao

    2012-01-01

    Hexamethylene bisacetamide-inducible protein 1 (HEXIM1) is best known as the inhibitor of positive transcription elongation factor b (P-TEFb), which regulates the transcription elongation of RNA polymerase II and controls 60–70% of mRNA synthesis. Our previous studies show that HEXIM1 interacts with two key p53 regulators, nucleophosmin and human double minute-2 protein (HDM2), implying a possible connection between HEXIM1 and the p53 signaling pathway. Here we report the interaction between p53 and HEXIM1 in breast cancer, acute myeloid leukemia, and colorectal carcinoma cells. The C-terminal regions of p53 and HEXIM1 are required for the protein-protein interaction. Overexpression of HEXIM1 prevents the ubiquitination of p53 by HDM2 and enhances the protein stability of p53, resulting in up-regulation of p53 target genes, such as Puma and p21. Induction of p53 can be achieved by several means, such as UV radiation and treatment with anti-cancer agents (including doxorubicin, etoposide, roscovitine, flavopiridol, and nutlin-3). Under all the conditions examined, elevated protein levels of p53 are found to associate with the increased p53-HEXIM1 interaction. In addition, knockdown of HEXIM1 significantly inhibits the induction of p53 and releases the cell cycle arrest caused by p53. Finally, the transcription of the p53 target genes is regulated by HEXIM1 in a p53-dependent fashion. Our results not only identify HEXIM1 as a positive regulator of p53, but also propose a novel molecular mechanism of p53 activation caused by the anti-cancer drugs and compounds. PMID:22948151

  19. Phosphorylation of human Jak3 at tyrosines 904 and 939 positively regulates its activity.

    PubMed

    Cheng, Hanyin; Ross, Jeremy A; Frost, Jeffrey A; Kirken, Robert A

    2008-04-01

    Janus tyrosine kinase 3 (Jak3) is essential for signaling by interleukin-2 (IL-2) family cytokines and proper immune function. Dysfunctional regulation of Jak3 may result in certain disease states. However, the molecular mechanisms governing Jak3 activation are not fully understood. In this study, we used a functional-proteomics approach to identify two novel tyrosine phosphorylation sites within Jak3, Y904 and Y939, which are conserved among Jak family proteins. By using phosphospecific antibodies, both residues were observed to be rapidly induced by stimulation of cells with IL-2 or other gammac cytokines. Mechanistic studies indicated that Y904 and Y939 regulate Jak3 activities. A phenylalanine substitution at either site greatly reduced Jak3 kinase activity in vitro and its ability to phosphorylate signal transducer and activator of transcription 5 (Stat5) in vivo, suggesting that phosphorylation of these previously unrecognized residues positively regulates Jak3 activity. Y904 and Y939 were required for optimal ATP usage by Jak3, while phosphorylation of Y939 preferentially promoted Stat5 activity in intact cells. Together, these findings demonstrate positive functional roles for two novel Jak3 phosphoregulatory sites which may be similarly important for other Jak family members. Identification of these sites also provides new therapeutic opportunities to modulate Jak3 function.

  20. [Regulation of Positive and Negative Emotions as Mediator between Maternal Emotion Socialization and Child Problem Behavior].

    PubMed

    Fäsche, Anika; Gunzenhauser, Catherine; Friedlmeier, Wolfgang; von Suchodoletz, Antje

    2015-01-01

    The present study investigated five to six year old children's ability to regulate negative and positive emotions in relation to psychosocial problem behavior (N=53). It was explored, whether mothers' supportive and nonsupportive strategies of emotion socialization influence children's problem behavior by shaping their emotion regulation ability. Mothers reported on children's emotion regulation and internalizing and externalizing problem behavior via questionnaire, and were interviewed about their preferences for socialization strategies in response to children's expression of negative affect. Results showed that children with more adaptive expression of adequate positive emotions had less internalizing behavior problems. When children showed more control of inadequate negative emotions, children were less internalizing as well as externalizing in their behavior. Furthermore, results indicated indirect relations of mothers' socialization strategies with children's problem behavior. Control of inadequate negative emotions mediated the link between non-supportive strategies on externalizing problem behavior. Results suggest that emotion regulatory processes should be part of interventions to reduce the development of problematic behavior in young children. Parents should be trained in dealing with children's emotions in a constructive way.

  1. The MYB107 Transcription Factor Positively Regulates Suberin Biosynthesis1[OPEN

    PubMed Central

    Yang, Huijun; Cai, Yuanheng; Kai, Guoyin

    2017-01-01

    Suberin, a lipophilic polymer deposited in the outer integument of the Arabidopsis (Arabidopsis thaliana) seed coat, represents an essential sealing component controlling water and solute movement and protecting seed from pathogenic infection. Although many genes responsible for suberin synthesis are identified, the regulatory components controlling its biosynthesis have not been definitively determined. Here, we show that the Arabidopsis MYB107 transcription factor acts as a positive regulator controlling suberin biosynthetic gene expression in the seed coat. MYB107 coexpresses with suberin biosynthetic genes in a temporal manner during seed development. Disrupting MYB107 particularly suppresses the expression of genes involved in suberin but not cutin biosynthesis, lowers seed coat suberin accumulation, alters suberin lamellar structure, and consequently renders higher seed coat permeability and susceptibility to abiotic stresses. Furthermore, MYB107 directly binds to the promoters of suberin biosynthetic genes, verifying its primary role in regulating their expression. Identifying MYB107 as a positive regulator for seed coat suberin synthesis offers a basis for discovering the potential transcriptional network behind one of the most abundant lipid-based polymers in nature. PMID:27965303

  2. Flattop regulates basal body docking and positioning in mono- and multiciliated cells

    PubMed Central

    Gegg, Moritz; Böttcher, Anika; Burtscher, Ingo; Hasenoeder, Stefan; Van Campenhout, Claude; Aichler, Michaela; Walch, Axel; Grant, Seth G N; Lickert, Heiko

    2014-01-01

    Planar cell polarity (PCP) regulates basal body (BB) docking and positioning during cilia formation, but the underlying mechanisms remain elusive. In this study, we investigate the uncharacterized gene Flattop (Fltp) that is transcriptionally activated during PCP acquisition in ciliated tissues. Fltp knock-out mice show BB docking and ciliogenesis defects in multiciliated lung cells. Furthermore, Fltp is necessary for kinocilium positioning in monociliated inner ear hair cells. In these cells, the core PCP molecule Dishevelled 2, the BB/spindle positioning protein Dlg3, and Fltp localize directly adjacent to the apical plasma membrane, physically interact and surround the BB at the interface of the microtubule and actin cytoskeleton. Dlg3 and Fltp knock-outs suggest that both cooperatively translate PCP cues for BB positioning in the inner ear. Taken together, the identification of novel BB/spindle positioning components as potential mediators of PCP signaling might have broader implications for other cell types, ciliary disease, and asymmetric cell division. DOI: http://dx.doi.org/10.7554/eLife.03842.001 PMID:25296022

  3. microRNAs are differentially regulated between MDM2-positive and negative malignant pleural mesothelioma

    PubMed Central

    Walter, Robert Fred Henry; Vollbrecht, Claudia; Werner, Robert; Wohlschlaeger, Jeremias; Christoph, Daniel Christian; Schmid, Kurt Werner; Mairinger, Fabian Dominik

    2016-01-01

    Background Malignant pleural mesothelioma (MPM) is a highly aggressive tumour first-line treated with a combination of cisplatin and pemetrexed. MDM2 and P14/ARF (CDKN2A) are upstream regulators of TP53 and may contribute to its inactivation. In the present study, we now aimed to define the impact of miRNA expression on this mechanism. Material and Methods 24 formalin-fixed paraffin-embedded (FFPE) tumour specimens were used for miRNA expression analysis of the 800 most important miRNAs using the nCounter technique (NanoString). Significantly deregulated miRNAs were identified before a KEGG-pathway analysis was performed. Results 17 miRNAs regulating TP53, 18 miRNAs regulating MDM2, and 11 miRNAs directly regulating CDKN2A are significantly downregulated in MDM2-expressing mesotheliomas. TP53 is downregulated in MDM2-negative tumours through miRNAs with a miSVR prediction score of 11.67, RB1 with a prediction score of 8.02, MDM2 with a prediction score of 4.50 and CDKN2A with a prediction score of 1.27. Conclusion MDM2 expression seems to impact miRNA expression levels in MPM. Especially, miRNAs involved in TP53-signaling are strongly decreased in MDM2-positive mesotheliomas. A better understanding of its tumour biology may open the chance for new therapeutic approaches and thereby augment patients' outcome. PMID:26918730

  4. msaABCR operon positively regulates biofilm development by repressing proteases and autolysis in Staphylococcus aureus.

    PubMed

    Sahukhal, Gyan S; Batte, Justin L; Elasri, Mohamed O

    2015-02-01

    Staphylococcus aureus is an important human pathogen that causes nosocomial and community-acquired infections. One of the most important aspects of staphylococcal infections is biofilm development within the host, which renders the bacterium resistant to the host's immune response and antimicrobial agents. Biofilm development is very complex and involves several regulators that ensure cell survival on surfaces within the extracellular polymeric matrix. Previously, we identified the msaABCR operon as an additional positive regulator of biofilm formation. In this study, we define the regulatory pathway by which msaABCR controls biofilm formation. We demonstrate that the msaABCR operon is a negative regulator of proteases. The control of protease production mediates the processing of the major autolysin, Atl, and thus regulates the rate of autolysis. In the absence of the msaABCR operon, Atl is processed by proteases at a high rate, leading to increased cell death and a defect in biofilm maturation. We conclude that the msaABCR operon plays a key role in maintaining the balance between autolysis and growth within the staphylococcal biofilm.

  5. Dlx5 is a positive regulator of chondrocyte differentiation during endochondral ossification.

    PubMed

    Ferrari, Deborah; Kosher, Robert A

    2002-12-15

    The process of endochondral ossification in which the bones of the limb are formed after generation of cartilage models is dependent on a precisely regulated program of chondrocyte maturation. Here, we show that the homeobox-containing gene Dlx5 is expressed at the onset of chondrocyte maturation during the conversion of immature proliferating chondrocytes into postmitotic hypertrophying chondrocytes, a critical step in the maturation process. Moreover, retroviral misexpression of Dlx5 during differentiation of the skeletal elements of the chick limb in vivo results in the formation of severely shortened skeletal elements that contain excessive numbers of hypertrophying chondrocytes which extend into ectopic regions, including sites normally occupied by immature chondrocytes. The expansion in the extent of hypertrophic maturation detectable histologically is accompanied by expanded and upregulated domains of expression of molecular markers of chondrocyte maturation, particularly type X collagen and osteopontin, and by expansion of mineralized cartilage matrix, which is characteristic of terminal hypertrophic differentiation. Furthermore, Dlx5 misexpression markedly reduces chondrocyte proliferation concomitant with promoting hypertrophic maturation. Taken together, these results indicate that Dlx5 is a positive regulator of chondrocyte maturation and suggest that it regulates the process at least in part by promoting conversion of immature proliferating chondrocytes into hypertrophying chondrocytes. Retroviral misexpression of Dlx5 also enhances formation of periosteal bone, which is derived from the Dlx5-expressing perichondrium that surrounds the diaphyses of the cartilage models. This suggests that Dlx5 may be involved in regulating osteoblast differentiation, as well as chondrocyte maturation, during endochondral ossification.

  6. Effects of continuous positive airway pressure on energy balance regulation: a systematic review

    PubMed Central

    Shechter, Ari

    2016-01-01

    Obesity is both a cause and a possible consequence of obstructive sleep apnoea (OSA), as OSA seems to affect parameters involved in energy balance regulation, including food intake, hormonal regulation of hunger/satiety, energy metabolism and physical activity. It is known that weight loss improves OSA, yet it remains unclear why continuous positive airway pressure (CPAP) often results in weight gain. The goal of this systematic review is to explore if and how CPAP affects the behaviour and/or metabolism involved in regulating energy balance. CPAP appears to correct for a hormonal profile characterised by abnormally high leptin and ghrelin levels in OSA, by reducing the circulating levels of each. This is expected to reduce excess food intake. However, reliable measures of food intake are lacking, and not yet sufficient to make conclusions. Although studies are limited and inconsistent, CPAP may alter energy metabolism, with reports of reductions in resting metabolic rate or sleeping metabolic rate. CPAP appears to not have an appreciable effect on altering physical activity levels. More work is needed to characterise how CPAP affects energy balance regulation. It is clear that promoting CPAP in conjunction with other weight loss approaches should be used to encourage optimal outcomes in OSA patients. PMID:27824596

  7. Arabidopsis CBF3 and DELLAs positively regulate each other in response to low temperature

    PubMed Central

    Zhou, Mingqi; Chen, Hu; Wei, Donghui; Ma, Hong; Lin, Juan

    2017-01-01

    The C-repeat binding factor (CBF) is crucial for regulation of cold response in higher plants. In Arabidopsis, the mechanism of CBF3-caused growth retardation is still unclear. Our present work shows that CBF3 shares the similar repression of bioactive gibberellin (GA) as well as upregulation of DELLA proteins with CBF1 and -2. Genetic analysis reveals that DELLAs play an essential role in growth reduction mediated by CBF1, -2, -3 genes. The in vivo and in vitro evidences demonstrate that GA2-oxidase 7 gene is a novel CBF3 regulon. Meanwhile, DELLAs contribute to cold induction of CBF1, -2, -3 genes through interaction with jasmonate (JA) signaling. We conclude that CBF3 promotes DELLAs accumulation through repressing GA biosynthesis and DELLAs positively regulate CBF3 involving JA signaling. CBFs and DELLAs collaborate to retard plant growth in response to low temperature. PMID:28051152

  8. Positive Gene Regulation by a Natural Protective miRNA Enables Arbuscular Mycorrhizal Symbiosis.

    PubMed

    Couzigou, Jean-Malo; Lauressergues, Dominique; André, Olivier; Gutjahr, Caroline; Guillotin, Bruno; Bécard, Guillaume; Combier, Jean-Philippe

    2017-01-11

    Arbuscular mycorrhizal (AM) symbiosis associates most plants with fungi of the phylum Glomeromycota. The fungus penetrates into roots and forms within cortical cell branched structures called arbuscules for nutrient exchange. We discovered that miR171b has a mismatched cleavage site and is unable to downregulate the miR171 family target gene, LOM1 (LOST MERISTEMS 1). This mismatched cleavage site is conserved among plants that establish AM symbiosis, but not in non-mycotrophic plants. Unlike other members of the miR171 family, miR171b stimulates AM symbiosis and is expressed specifically in root cells that contain arbuscules. MiR171b protects LOM1 from negative regulation by other miR171 family members. These findings uncover a unique mechanism of positive post-transcriptional regulation of gene expression by miRNAs and demonstrate its relevance for the establishment of AM symbiosis.

  9. OsGF14e positively regulates panicle blast resistance in rice.

    PubMed

    Liu, Qing; Yang, Jianyuan; Zhang, Shaohong; Zhao, Junliang; Feng, Aiqing; Yang, Tifeng; Wang, Xiaofei; Mao, Xingxue; Dong, Jingfang; Zhu, Xiaoyuan; Leung, Hei; Leach, Jan E; Liu, Bin

    2016-02-26

    Though GF14e has been reported to negatively regulate bacterial blight and sheath blight resistance in rice, its effect on panicle blast, the most destructive disease in rice is still unknown. In the present study, we identified that GF14e was highly expressed in panicles and was induced in panicles infected by blast pathogen. Overexpression of GF14e enhances resistance to panicle blast whereas silencing GF14e results in increased susceptibility to panicle blast, suggesting that GF14e plays a positive role in quantitative panicle blast resistance in rice. Our results also demonstrate that GF14e is regulated by WRKY71 and GF14e-mediated panicle blast resistance is related to activation of SA-dependent pathway and suppression of JA-dependent pathway. The functional confirmation of GF14e in panicle blast resistance makes it to be a promising target in molecular rice breeding.

  10. Regulating and facilitating: the role of emotional intelligence in maintaining and using positive affect for creativity.

    PubMed

    Parke, Michael R; Seo, Myeong-Gu; Sherf, Elad N

    2015-05-01

    Although past research has identified the effects of emotional intelligence on numerous employee outcomes, the relationship between emotional intelligence and creativity has not been well established. We draw upon affective information processing theory to explain how two facets of emotional intelligence-emotion regulation and emotion facilitation-shape employee creativity. Specifically, we propose that emotion regulation ability enables employees to maintain higher positive affect (PA) when faced with unique knowledge processing requirements, while emotion facilitation ability enables employees to use their PA to enhance their creativity. We find support for our hypotheses using a multimethod (ability test, experience sampling, survey) and multisource (archival, self-reported, supervisor-reported) research design of early career managers across a wide range of jobs.

  11. The Role of Nucleosome Positioning in the Evolution of Gene Regulation

    PubMed Central

    Tsankov, Alexander M.; Thompson, Dawn Anne; Socha, Amanda

    2010-01-01

    Chromatin organization plays a major role in gene regulation and can affect the function and evolution of new transcriptional programs. However, it can be difficult to decipher the basis of changes in chromatin organization and their functional effect on gene expression. Here, we present a large-scale comparative genomic analysis of the relationship between chromatin organization and gene expression, by measuring mRNA abundance and nucleosome positions genome-wide in 12 Hemiascomycota yeast species. We found substantial conservation of global and functional chromatin organization in all species, including prominent nucleosome-free regions (NFRs) at gene promoters, and distinct chromatin architecture in growth and stress genes. Chromatin organization has also substantially diverged in both global quantitative features, such as spacing between adjacent nucleosomes, and in functional groups of genes. Expression levels, intrinsic anti-nucleosomal sequences, and trans-acting chromatin modifiers all play important, complementary, and evolvable roles in determining NFRs. We identify five mechanisms that couple chromatin organization to evolution of gene regulation and have contributed to the evolution of respiro-fermentation and other key systems, including (1) compensatory evolution of alternative modifiers associated with conserved chromatin organization, (2) a gradual transition from constitutive to trans-regulated NFRs, (3) a loss of intrinsic anti-nucleosomal sequences accompanying changes in chromatin organization and gene expression, (4) re-positioning of motifs from NFRs to nucleosome-occluded regions, and (5) the expanded use of NFRs by paralogous activator-repressor pairs. Our study sheds light on the molecular basis of chromatin organization, and on the role of chromatin organization in the evolution of gene regulation. PMID:20625544

  12. Regulation of dynein localization and centrosome positioning by Lis-1 and asunder during Drosophila spermatogenesis

    PubMed Central

    Sitaram, Poojitha; Anderson, Michael A.; Jodoin, Jeanne N.; Lee, Ethan; Lee, Laura A.

    2012-01-01

    Dynein, a microtubule motor complex, plays crucial roles in cell-cycle progression in many systems. The LIS1 accessory protein directly binds dynein, although its precise role in regulating dynein remains unclear. Mutation of human LIS1 causes lissencephaly, a developmental brain disorder. To gain insight into the in vivo functions of LIS1, we characterized a male-sterile allele of the Drosophila homolog of human LIS1. We found that centrosomes do not properly detach from the cell cortex at the onset of meiosis in most Lis-1 spermatocytes; centrosomes that do break cortical associations fail to attach to the nucleus. In Lis-1 spermatids, we observed loss of attachments between the nucleus, basal body and mitochondria. The localization pattern of LIS-1 protein throughout Drosophila spermatogenesis mirrors that of dynein. We show that dynein recruitment to the nuclear surface and spindle poles is severely reduced in Lis-1 male germ cells. We propose that Lis-1 spermatogenesis phenotypes are due to loss of dynein regulation, as we observed similar phenotypes in flies null for Tctex-1, a dynein light chain. We have previously identified asunder (asun) as another regulator of dynein localization and centrosome positioning during Drosophila spermatogenesis. We now report that Lis-1 is a strong dominant enhancer of asun and that localization of LIS-1 in male germ cells is ASUN dependent. We found that Drosophila LIS-1 and ASUN colocalize and coimmunoprecipitate from transfected cells, suggesting that they function within a common complex. We present a model in which Lis-1 and asun cooperate to regulate dynein localization and centrosome positioning during Drosophila spermatogenesis. PMID:22764052

  13. Positional effects of AAN motifs in rpoS regulation by sRNAs and Hfq

    PubMed Central

    Peng, Yi; Soper, Toby J.; Woodson, Sarah A.

    2013-01-01

    The E. coli stationary phase transcription factor RpoS is translated in response to small noncoding RNAs (sRNAs), which base pair with the rpoS mRNA leader. The bacterial Sm-like protein Hfq anneals sRNAs with their mRNA targets by simultaneously binding the mRNA and sRNA. Intriguingly, Hfq is recruited to the rpoS leader via AAN motifs far upstream of the sRNA. SHAPE chemical footprinting showed that the rpoS leader is divided into a far upstream domain, an Hfq binding domain, and a downstream inhibitory stem-loop containing the sRNA and ribosome binding sites. To investigate how Hfq promotes sRNA-mRNA base pairing from a distance, the natural AAN Hfq binding site was deleted, and artificial AAN binding sites were inserted at various positions in the rpoS leader. All the relocated AAN motifs restored tight Hfq binding in vitro, but only insertion at the natural position restored Hfq-dependent sRNA annealing in vitro and sRNA regulation of rpoS translation in vivo. Furthermore, U-rich motifs in the downstream inhibitory domain stabilized the rpoS mRNA-Hfq complex and contributed to regulation of rpoS expression. We propose that the natural Hfq binding domain is optimal for positive regulation because it recruits Hfq to the mRNA and allows it to act on incoming sRNAs without opening the inhibitory stem-loop when sRNA are absent. PMID:24051417

  14. Long-distance signals positively regulate the expression of iron uptake genes in tobacco roots.

    PubMed

    Enomoto, Yusuke; Hodoshima, Hirotaka; Shimada, Hiroaki; Shoji, Kazuhiro; Yoshihara, Toshihiro; Goto, Fumiyuki

    2007-12-01

    Long-distance signals generated in shoots are thought to be associated with the regulation of iron uptake from roots; however, the signaling mechanism is still unknown. To elucidate whether the signal regulates iron uptake genes in roots positively or negatively, we analyzed the expressions of two representative iron uptake genes: NtIRT1 and NtFRO1 in tobacco (Nicotiana tabacum L.) roots, after shoots were manipulated in vitro. When iron-deficient leaves were treated with Fe(II)-EDTA, the expressions of both genes were significantly reduced; nevertheless iron concentration in the roots maintained a similar level to that in roots grown under iron-deficient conditions. Next, all leaves from tobacco plants grown under the iron-deficient condition were excised. The expression of two genes were quickly reduced below half within 2 h after the leaf excision and gradually disappeared by the end of a 24-h period. The NtIRT1 expression was compared among the plants whose leaves were cut off in various patterns. The expression increased in proportion to the dry weight of iron-deficient leaves, although no relation was observed between the gene expression and the position of excised leaves. Interestingly, the NtIRT1 expression in hairy roots increased under the iron-deficient condition, suggesting that roots also have the signaling mechanism of iron status as well as shoots. Taken together, these results indicate that the long-distance signal generated in iron-deficient tissues including roots is a major factor in positive regulation of the expression of NtIRT1 and NtFRO1 in roots, and that the strength of the signal depends on the size of plants.

  15. Mindfulness Broadens Awareness and Builds Eudaimonic Meaning: A Process Model of Mindful Positive Emotion Regulation

    PubMed Central

    Garland, Eric L.; Farb, Norman A.; Goldin, Philippe; Fredrickson, Barbara L.

    2015-01-01

    Contemporary scholarship on mindfulness casts it as a form of purely non-evaluative engagement with experience. Yet, traditionally mindfulness was not intended to operate in a vacuum of dispassionate observation, but was seen as facilitative of eudaimonic mental states. In spite of this historical context, modern psychological research has neglected to ask the question of how the practice of mindfulness affects downstream emotion regulatory processes to impact the sense of meaning in life. To fill this lacuna, here we describe the Mindfulness-to-Meaning Theory, from which we derive a novel process model of mindful positive emotion regulation informed by affective science, in which mindfulness is proposed to introduce flexibility in the generation of cognitive appraisals by enhancing interoceptive attention, thereby expanding the scope of cognition to facilitate reappraisal of adversity and savoring of positive experience. This process is proposed to culminate in a deepened capacity for meaning-making and greater engagement with life. PMID:27087765

  16. Mindfulness Broadens Awareness and Builds Eudaimonic Meaning: A Process Model of Mindful Positive Emotion Regulation.

    PubMed

    Garland, Eric L; Farb, Norman A; Goldin, Philippe; Fredrickson, Barbara L

    2015-10-01

    Contemporary scholarship on mindfulness casts it as a form of purely non-evaluative engagement with experience. Yet, traditionally mindfulness was not intended to operate in a vacuum of dispassionate observation, but was seen as facilitative of eudaimonic mental states. In spite of this historical context, modern psychological research has neglected to ask the question of how the practice of mindfulness affects downstream emotion regulatory processes to impact the sense of meaning in life. To fill this lacuna, here we describe the Mindfulness-to-Meaning Theory, from which we derive a novel process model of mindful positive emotion regulation informed by affective science, in which mindfulness is proposed to introduce flexibility in the generation of cognitive appraisals by enhancing interoceptive attention, thereby expanding the scope of cognition to facilitate reappraisal of adversity and savoring of positive experience. This process is proposed to culminate in a deepened capacity for meaning-making and greater engagement with life.

  17. Effect of upper torso inclination in Fowler's position on autonomic cardiovascular regulation.

    PubMed

    Kubota, Satoshi; Endo, Yutaka; Kubota, Mitsue

    2013-09-01

    The present study investigates autonomic cardiovascular regulation during postural changes while in Fowler's position. Respiratory sinus arrhythmia (RSA) and sequence baroreflex sensitivity (sBRS) were measured in 12 healthy individuals in three positions (Experiment 1). We also measured RSA, sBRS, tidal volume (TV), lung volume spectrum (LV spectrum), and transfer gain and phase between lung volume and RR interval (RSA-TF, RSATF-phase) in 11 healthy individuals in two positions (Experiment 2). All participants maintained respiratory frequency at 15 breaths/min. The three positions in Experiment 1 were 30°, 45°, and 60° of upper torso inclination with a lower torso inclination of 30° throughout all evaluations. The two positions in Experiment 2 were 30° and 60° of upper torso backrest inclination with a lower torso inclination of 30° throughout all evaluations. The results of Experiment 1 showed significantly higher RSA and sBRS at 60° and 45° than at 30°, whereas RR interval (RRI), systolic blood pressure (SBP), and diastolic blood pressure (DBP) did not differ significantly under any condition. The results of Experiment 2 showed that RSA, RSA-TF, sBRS, TV, and LV spectrum were significantly higher at 60° than at 30°, and that RRI, SBP, DBP, and the RSATF phase did not significantly differ under any condition. These findings suggested that slight flexion of the upper torso in Fowler's position activates respiratory function and increases the contribution of vagal nerve activity to the cardiovascular system in young participants under conditions of a fixed respiratory rate.

  18. Autoantibody‐Positive Healthy Individuals Display Unique Immune Profiles That May Regulate Autoimmunity

    PubMed Central

    Slight‐Webb, Samantha; Lu, Rufei; Ritterhouse, Lauren L.; Munroe, Melissa E.; Maecker, Holden T.; Fathman, Charles G.; Utz, Paul J.; Merrill, Joan T.; Guthridge, Joel M.

    2016-01-01

    Objective Antinuclear antibodies (ANAs) are detected in ∼18% of females, yet autoimmune disease develops in only 5–8%. Immunologic differences between ANA‐positive healthy individuals and patients with systemic lupus erythematosus (SLE) may elucidate the regulatory mechanisms by which ANA‐positive individuals avoid transition to clinical autoimmune disease. Methods Healthy individuals (n = 790) were screened for autoantibodies specific for 11 antigens associated with lupus, systemic sclerosis, and Sjögren's syndrome. From this screening, 31 European American ANA‐positive healthy individuals were selected and demographically matched to ANA‐negative controls and SLE patients. Serum cytokine profiles, leukocyte subset frequency, and reactivity were analyzed by multiplex assays, immunophenotyping, and phosphospecific flow cytometry. Results Of 790 individuals screened, 57 (7%) were ANA‐positive. The majority of proinflammatory cytokines, including interferon‐γ (IFNγ), tumor necrosis factor, interleukin‐17 (IL‐17), and granulocyte colony‐stimulating factor, exhibited a stepwise increase in serum levels from ANA‐negative controls to ANA‐positive healthy individuals to SLE patients (P < 0.0001). IFNα, IFNβ, IL‐12p40, and stem cell factor/c‐Kit ligand were increased in SLE patients only (P < 0.05). B lymphocyte stimulator (BlyS) was elevated in SLE patients but decreased in ANA‐positive individuals (P < 0.001). Further, IL‐1 receptor antagonist (IL‐1Ra) was down‐regulated in SLE patients only (P < 0.0001). ANA‐positive individuals had increased frequencies of monocytes, memory B cells, and plasmablasts and increased levels of pSTAT‐1 and pSTAT‐3 following IFNα stimulation compared with ANA‐negative controls (P < 0.05). Conclusion ANA‐positive healthy individuals exhibit dysregulation in multiple immune pathways yet differ from SLE patients by the absence of elevated IFNs, BLyS, IL‐12p40, and stem cell factor

  19. Patterned Contractile Forces Promote Epidermal Spreading and Regulate Segment Positioning during Drosophila Head Involution.

    PubMed

    Czerniak, Natalia Dorota; Dierkes, Kai; D'Angelo, Arturo; Colombelli, Julien; Solon, Jérôme

    2016-07-25

    Epithelial spreading is a fundamental mode of tissue rearrangement occurring during animal development and wound closure. It has been associated either with the collective migration of cells [1, 2] or with actomyosin-generated forces acting at the leading edge (LE) and pulling the epithelial tissue [3, 4]. During the process of Drosophila head involution (HI), the epidermis spreads anteriorly to envelope the head tissues and fully cover the embryo [5]. This results in epidermal segments of equal width that will give rise to the different organs of the fly [6]. Here we perform a quantitative analysis of tissue spreading during HI. Combining high-resolution live microscopy with laser microsurgery and genetic perturbations, we show that epidermal movement is in part, but not solely, driven by a contractile actomyosin cable at the LE. Additional driving forces are generated within each segment by a gradient of actomyosin-based circumferential tension. Interfering with Hedgehog (Hh) signaling can modulate this gradient, thus suggesting the involvement of polarity genes in the regulation of HI. In particular, we show that disruption of these contractile forces alters segment widths and leads to a mispositioning of segments. Within the framework of a physical description, we confirm that given the geometry of the embryo, a patterned profile of active circumferential tensions can indeed generate propelling forces and control final segment position. Our study thus unravels a mechanism by which patterned tensile forces can regulate spreading and positioning of epithelial tissues.

  20. Reciprocal positive regulation between TRPV6 and NUMB in PTEN-deficient prostate cancer cells

    SciTech Connect

    Kim, Sung-Young; Hong, Chansik; Wie, Jinhong; Kim, Euiyong; Kim, Byung Joo; Ha, Kotdaji; Cho, Nam-Hyuk; Kim, In-Gyu; Jeon, Ju-Hong; So, Insuk

    2014-04-25

    Highlights: • TRPV6 interacts with tumor suppressor proteins. • Numb has a selective effect on TRPV6, depending on the prostate cancer cell line. • PTEN is a novel regulator of TRPV6–Numb complex. - Abstract: Calcium acts as a second messenger and plays a crucial role in signaling pathways involved in cell proliferation. Recently, calcium channels related to calcium influx into the cytosol of epithelial cells have attracted attention as a cancer therapy target. Of these calcium channels, TRPV6 is overexpressed in prostate cancer and is considered an important molecule in the process of metastasis. However, its exact role and mechanism is unclear. NUMB, well-known tumor suppressor gene, is a novel interacting partner of TRPV6. We show that NUMB and TRPV6 have a reciprocal positive regulatory relationship in PC-3 cells. We repeated this experiment in two other prostate cancer cell lines, DU145 and LNCaP. Interestingly, there were no significant changes in TRPV6 expression following NUMB knockdown in DU145. We revealed that the presence or absence of PTEN was the cause of NUMB–TRPV6 function. Loss of PTEN caused a positive correlation of TRPV6–NUMB expression. Collectively, we determined that PTEN is a novel interacting partner of TRPV6 and NUMB. These results demonstrated a novel relationship of NUMB–TRPV6 in prostate cancer cells, and show that PTEN is a novel regulator of this complex.

  1. Ethylene positively regulates cold tolerance in grapevine by modulating the expression of ETHYLENE RESPONSE FACTOR 057

    PubMed Central

    Sun, Xiaoming; Zhao, Tingting; Gan, Shuheng; Ren, Xiaodie; Fang, Linchuan; Karungo, Sospeter Karanja; Wang, Yi; Chen, Liang; Li, Shaohua; Xin, Haiping

    2016-01-01

    Ethylene (ET) is a gaseous plant hormone that plays essential roles in biotic and abiotic stress responses in plants. However, the role of ET in cold tolerance varies in different species. This study revealed that low temperature promotes the release of ET in grapevine. The treatment of exogenous 1-aminocyclopropane-1-carboxylate increased the cold tolerance of grapevine. By contrast, the application of the ET biosynthesis inhibitor aminoethoxyvinylglycine reduced the cold tolerance of grapevine. This finding suggested that ET positively affected cold stress responses in grapevine. The expression of VaERF057, an ET signaling downstream gene, was strongly induced by low temperature. The overexpression of VaERF057 also enhanced the cold tolerance of Arabidopsis. Under cold treatment, malondialdehyde content was lower and superoxide dismutase, peroxidase, and catalase activities were higher in transgenic lines than in wild-type plants. RNA-Seq results showed that 32 stress-related genes, such as CBF1-3, were upregulated in VaERF057-overexpressing transgenic line. Yeast one-hybrid results further demonstrated that VaERF057 specifically binds to GCC-box and DRE motifs. Thus, VaERF057 may directly regulate the expression of its target stress-responsive genes by interacting with a GCC-box or a DRE element. Our work confirmed that ET positively regulates cold tolerance in grapevine by modulating the expression of VaERF057. PMID:27039848

  2. PLEKHM1/DEF8/RAB7 complex regulates lysosome positioning and bone homeostasis

    PubMed Central

    Fujiwara, Toshifumi; Ye, Shiqiao; Winchell, Caylin G.; Andrews, Norma W.; Voth, Daniel E.; Varughese, Kottayil I.; Mackintosh, Samuel G.; Feng, Yunfeng; Nakamura, Takashi; Manolagas, Stavros C.

    2016-01-01

    Mutations of the Plekhm1 gene in humans and rats cause osteopetrosis, an inherited bone disease characterized by diminished bone resorption by osteoclasts. PLEKHM1 binds to RAB7 and is critical for lysosome trafficking. However, the molecular mechanisms by which PLEKHM1 regulates lysosomal pathways remain unknown. Here, we generated germline and conditional Plekhm1-deficient mice. These mice displayed no overt abnormalities in major organs, except for an increase in trabecular bone mass. Furthermore, loss of PLEKHM1 abrogated the peripheral distribution of lysosomes and bone resorption in osteoclasts. Mechanistically, we indicated that DEF8 interacts with PLEKHM1 and promotes its binding to RAB7, whereas the binding of FAM98A and NDEL1 with PLEKHM1 connects lysosomes to microtubules. Importantly, suppression of these proteins results in lysosome positioning and bone resorption defects similar to those of Plekhm1-null osteoclasts. Thus, PLHKEM1, DEF8, FAM98A, and NDEL1 constitute a molecular complex that regulates lysosome positioning and secretion through RAB7. PMID:27777970

  3. JMJD8 is a positive regulator of TNF-induced NF-κB signaling

    PubMed Central

    Yeo, Kok Siong; Tan, Ming Cheang; Wong, Wan Ying; Loh, Sheng Wei; Lam, Yi Lyn; Tan, Chin Leng; Lim, Yat-Yuen; Ea, Chee-Kwee

    2016-01-01

    TNF-induced signaling mediates pleiotropic biological consequences including inflammation, immunity, cell proliferation and apoptosis. Misregulation of TNF signaling has been attributed as a major cause of chronic inflammatory diseases and cancer. Jumonji domain-containing protein 8 (JMJD8) belongs to the JmjC family. However, only part of the family members has been described as hydroxylase enzymes that function as histone demethylases. Here, we report that JMJD8 positively regulates TNF-induced NF-κB signaling. Silencing the expression of JMJD8 using RNA interference (RNAi) greatly suppresses TNF-induced expression of several NF-κB-dependent genes. Furthermore, knockdown of JMJD8 expression reduces RIP ubiquitination, IKK kinase activity, delays IκBα degradation and subsequently blocks nuclear translocation of p65. In addition, JMJD8 deficiency enhances TNF-induced apoptosis. Taken together, these findings indicate that JMJD8 functions as a positive regulator of TNF-induced NF-κB signaling. PMID:27671354

  4. Phosphorylation acts positively and negatively to regulate MRTF-A subcellular localisation and activity

    PubMed Central

    Panayiotou, Richard; Miralles, Francesc; Pawlowski, Rafal; Diring, Jessica; Flynn, Helen R; Skehel, Mark; Treisman, Richard

    2016-01-01

    The myocardin-related transcription factors (MRTF-A and MRTF-B) regulate cytoskeletal genes through their partner transcription factor SRF. The MRTFs bind G-actin, and signal-regulated changes in cellular G-actin concentration control their nuclear accumulation. The MRTFs also undergo Rho- and ERK-dependent phosphorylation, but the function of MRTF phosphorylation, and the elements and signals involved in MRTF-A nuclear export are largely unexplored. We show that Rho-dependent MRTF-A phosphorylation reflects relief from an inhibitory function of nuclear actin. We map multiple sites of serum-induced phosphorylation, most of which are S/T-P motifs and show that S/T-P phosphorylation is required for transcriptional activation. ERK-mediated S98 phosphorylation inhibits assembly of G-actin complexes on the MRTF-A regulatory RPEL domain, promoting nuclear import. In contrast, S33 phosphorylation potentiates the activity of an autonomous Crm1-dependent N-terminal NES, which cooperates with five other NES elements to exclude MRTF-A from the nucleus. Phosphorylation thus plays positive and negative roles in the regulation of MRTF-A. DOI: http://dx.doi.org/10.7554/eLife.15460.001 PMID:27304076

  5. Dlx5, a Positive Regulator of Osteoblastogenesis, is Essential for Osteoblast-Osteoclast Coupling

    PubMed Central

    Samee, Nadeem; Geoffroy, Valerie; Marty, Caroline; Schiltz, Corinne; Vieux-Rochas, Maxence; Levi, Giovanni; de Vernejoul, Marie-Christine

    2008-01-01

    The homeodomain protein Dlx5 is an activator of Runx2 (a key regulator of osteogenesis) and is thought to be an important regulator of bone formation. At present, however, the perinatal lethality of Dlx5-null mice has hampered the elucidation of its function in osteogenesis. Here we provide the first analysis of the effects of Dlx5 inactivation on bone development. Femurs of Dlx5-null mouse embryos at the end of gestation exhibit a reduction in both total and trabecular bone volume associated with increased trabecular separation and reduced trabecular number. These parameters are often associated with pathological conditions characterized by reduced osteoblast activity and increased bone resorption. Dlx5−/− osteoblasts in culture display reduced proliferation and differentiation rate and reduction of Runx2, Osx, Osteocalcin and Bone Sialoprotein expression. In addition to impaired osteoblast function, Dlx5−/− femurs exhibit significant increases in osteoclast number. As Dlx5 is not expressed by osteoclasts, we suggest that its osteoblastic expression might control osteoblast/osteoclast coupling. Cultured Dlx5−/− osteoblasts displayed a higher RANKL/OPG ratio. Furthermore, Dlx5−/− osteoblasts induced a higher number of TRAP-positive multinucleated cells in normal spleen cultures with a globally increased resorption activity. These findings suggest that Dlx5 is a central regulator of bone turnover as it activates bone formation directly and bone resorption indirectly. PMID:18669617

  6. Sorghum DW1 positively regulates brassinosteroid signaling by inhibiting the nuclear localization of BRASSINOSTEROID INSENSITIVE 2.

    PubMed

    Hirano, Ko; Kawamura, Mayuko; Araki-Nakamura, Satoko; Fujimoto, Haruka; Ohmae-Shinohara, Kozue; Yamaguchi, Miki; Fujii, Akihiro; Sasaki, Hiroaki; Kasuga, Shigemitsu; Sazuka, Takashi

    2017-12-01

    Semi-dwarf traits have been widely introgressed into cereal crops to improve lodging resistance. In sorghum (Sorghum bicolor L. Moench), four major unlinked dwarfing genes, Dw1-Dw4, have been introduced to reduce plant height, and among them, Dw3 and Dw1 have been cloned. Dw3 encodes a gene involved in auxin transport, whereas, Dw1 was recently isolated and identified as a gene encoding a protein of unknown function. In this study, we show that DW1 is a novel component of brassinosteroid (BR) signaling. Sorghum possessing the mutated allele of Dw1 (dw1), showed similar phenotypes to rice BR-deficient mutants, such as reduced lamina joint bending, attenuated skotomorphogenesis, and insensitivity against feedback regulation of BR-related genes. Furthermore, DW1 interacted with a negative regulator of BR signaling, BRASSINOSTEROID INSENSITIVE 2 (BIN2), and inhibited its nuclear localization, indicating that DW1 positively regulates BR signaling by inhibiting the function of BIN2. In contrast to rice and wheat breeding which used gibberellin (GA) deficiency to reduce plant height, sorghum breeding modified auxin and BR signaling. This difference may result from GA deficiency in rice and wheat does not cause deleterious side effects on plant morphology, whereas in sorghum it leads to abnormal culm bending.

  7. TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    PubMed Central

    Kitajima, Naoyuki; Numaga-Tomita, Takuro; Watanabe, Masahiko; Kuroda, Takuya; Nishimura, Akiyuki; Miyano, Kei; Yasuda, Satoshi; Kuwahara, Koichiro; Sato, Yoji; Ide, Tomomi; Birnbaumer, Lutz; Sumimoto, Hideki; Mori, Yasuo; Nishida, Motohiro

    2016-01-01

    Reactive oxygen species (ROS) produced by NADPH oxidase 2 (Nox2) function as key mediators of mechanotransduction during both physiological adaptation to mechanical load and maladaptive remodeling of the heart. This is despite low levels of cardiac Nox2 expression. The mechanism underlying the transition from adaptation to maladaptation remains obscure, however. We demonstrate that transient receptor potential canonical 3 (TRPC3), a Ca2+-permeable channel, acts as a positive regulator of ROS (PRROS) in cardiomyocytes, and specifically regulates pressure overload-induced maladaptive cardiac remodeling in mice. TRPC3 physically interacts with Nox2 at specific C-terminal sites, thereby protecting Nox2 from proteasome-dependent degradation and amplifying Ca2+-dependent Nox2 activation through TRPC3-mediated background Ca2+ entry. Nox2 also stabilizes TRPC3 proteins to enhance TRPC3 channel activity. Expression of TRPC3 C-terminal polypeptide abolished TRPC3-regulated ROS production by disrupting TRPC3-Nox2 interaction, without affecting TRPC3-mediated Ca2+ influx. The novel TRPC3 function as a PRROS provides a mechanistic explanation for how diastolic Ca2+ influx specifically encodes signals to induce ROS-mediated maladaptive remodeling and offers new therapeutic possibilities. PMID:27833156

  8. SO2426 is a positive regulator of siderophore expression in Shewanella oneidensis MR-1

    PubMed Central

    2011-01-01

    Background The Shewanella oneidensis MR-1 genome encodes a predicted orphan DNA-binding response regulator, SO2426. Previous studies with a SO2426-deficient MR-1 strain suggested a putative functional role for SO2426 in the regulation of iron acquisition genes, in particular, the siderophore (hydroxamate) biosynthesis operon so3030-3031-3032. To further investigate the functional role of SO2426 in iron homeostasis, we employed computational strategies to identify putative gene targets of SO2426 regulation and biochemical approaches to validate the participation of SO2426 in the control of siderophore biosynthesis in S. oneidensis MR-1. Results In silico prediction analyses revealed a single 14-bp consensus motif consisting of two tandem conserved pentamers (5'-CAAAA-3') in the upstream regulatory regions of 46 genes, which were shown previously to be significantly down-regulated in a so2426 deletion mutant. These genes included so3030 and so3032, members of an annotated siderophore biosynthetic operon in MR-1. Electrophoretic mobility shift assays demonstrated that the SO2426 protein binds to its motif in the operator region of so3030. A "short" form of SO2426, beginning with a methionine at position 11 (M11) of the originally annotated coding sequence for SO2426, was also functional in binding to its consensus motif, confirming previous 5' RACE results that suggested that amino acid M11 is the actual translation start codon for SO2426. Alignment of SO2426 orthologs from all sequenced Shewanella spp. showed a high degree of sequence conservation beginning at M11, in addition to conservation of a putative aspartyl phosphorylation residue and the helix-turn-helix (HTH) DNA-binding domain. Finally, the so2426 deletion mutant was unable to synthesize siderophores at wild-type rates upon exposure to the iron chelator 2,2'-dipyridyl. Conclusions Collectively, these data support the functional characterization of SO2426 as a positive regulator of siderophore-mediated iron

  9. SIRT1 positively regulates breast cancer associated human aromatase (CYP19A1) expression.

    PubMed

    Holloway, Kimberly R; Barbieri, Andreia; Malyarchuk, Svitlana; Saxena, Madhurima; Nedeljkovic-Kurepa, Ana; Cameron Mehl, Mathieu; Wang, Allison; Gu, Xin; Pruitt, Kevin

    2013-03-01

    Breast cancer remains one of the leading causes of death in women diagnosed with cancer. In breast cancer, aberrant expression of the CYP19A1 gene, which encodes the aromatase enzyme, contributes to increased intratumoral levels of estradiol. Regardless of whether this estrogen is produced by peripheral tissues or within specific subpopulations of cells within the breast tumor, it is clear that the aromatase enzymatic activity is critical for the growth of estrogen-dependent tumors. Currently, aromatase inhibitors have proven to be highly effective in blocking the growth of estrogen-dependent forms of breast cancer. CYP19A1 transcription is tightly controlled by 10 tissue-specific promoters. In breast cancer, however, aromatase transcription is driven by multiple promoters that somehow override the tissue-specific regulation of normal tissue. Here, we explore the role that the deacetylase, sirtuin-1 (SIRT1), plays in positively regulating aromatase in breast cancer. We demonstrate that the use of cambinol and the SIRT1/2 inhibitor VII, 2 small molecule inhibitors of SIRT1 and SIRT2, as well as small molecule inhibitors and small interfering RNA specific to SIRT1, all reduce the levels of aromatase mRNA. We further demonstrate that pharmacologic inhibition causes a marked reduction in aromatase protein levels. Additionally, by chromatin immunoprecipitation, we demonstrate that SIRT1 occupies the promoter regions PI.3/PII and PI.4, and its inhibition leads to increased acetylation of estrogen-related receptorα, a transcription factor that positively regulates CYP19A1 transcription in epithelial cells. Finally, we demonstrate by immunohistochemistry that SIRT1 is significantly up-regulated in invasive ductal carcinoma relative to normal tissue adjacent to tumor, further suggesting a role of SIRT1 in breast cancer. This work uncovers a new mechanism for the regulation of aromatase and provides rationale for further investigation of how the inhibition of specific

  10. Cardiomyogenesis in the Developing Heart Is Regulated by c-kit-Positive Cardiac Stem Cells

    PubMed Central

    Ferreira-Martins, João; Ogórek, Barbara; Cappetta, Donato; Matsuda, Alex; Signore, Sergio; D'Amario, Domenico; Kostyla, James; Steadman, Elisabeth; Ide-Iwata, Noriko; Sanada, Fumihiro; Iaffaldano, Grazia; Ottolenghi, Sergio; Hosoda, Toru; Leri, Annarosa; Kajstura, Jan; Anversa, Piero; Rota, Marcello

    2012-01-01

    Rationale Embryonic and fetal myocardial growth is characterized by a dramatic increase in myocyte number, but whether the expansion of the myocyte compartment is dictated by activation and commitment of resident cardiac stem cells (CSCs), division of immature myocytes or both is currently unknown. Objectives In this study, we tested whether prenatal cardiac development is controlled by activation and differentiation of CSCs and whether division of c-kit-positive CSCs in the mouse heart is triggered by spontaneous Ca2+ oscillations. Results We report that embryonic-fetal c-kit-positive CSCs are self-renewing, clonogenic and multipotent in vitro and in vivo. The growth and commitment of c-kit-positive CSCs is responsible for the generation of the myocyte progeny of the developing heart. The close correspondence between values computed by mathematical modeling and direct measurements of myocyte number at E9, E14, E19 and one day after birth strongly suggests that the organogenesis of the embryonic heart is dependent on a hierarchical model of cell differentiation regulated by resident CSCs. The growth promoting effects of c-kit-positive CSCs are triggered by spontaneous oscillations in intracellular Ca2+, mediated by IP3 receptor activation, which condition asymmetric stem cell division and myocyte lineage specification. Conclusions Myocyte formation derived from CSC differentiation is the major determinant of cardiac growth during development. Division of c-kit-positive CSCs in the mouse is promoted by spontaneous Ca2+ spikes, which dictate the pattern of stem cell replication and the generation of a myocyte progeny at all phases of prenatal life and up to one day after birth. PMID:22275487

  11. Overexpression of 14-3-3σ counteracts tumorigenicity by positively regulating p73 in vivo

    PubMed Central

    GENG, CUIZHI; SANG, MEIXIANG; YANG, RUILING; GAO, WEI; ZHOU, TAO; WANG, SHIJIE

    2011-01-01

    14-3-3σ, one of the 14-3-3 family members, was initially identified as a human mammary epithelium-specific marker 1. The expression of 14-3-3σ is directly regulated by p53. It has been demonstrated that 14-3-3σ stabilizes p53 and enhances its transcriptional activity through the interaction with p53, suggesting that 14-3-3σ has a positive feedback effect on p53. Our previous study showed that 14-3-3σ is a direct transcriptional target of p73 and enhances the p73-mediated transcriptional as well as pro-apoptotic activity in vitro. In the present study, we explored the tumor-suppressive effect of 14-3-3σ by establishing a breast cancer xenograft nude mouse model with an inducible expression of 14-3-3σ or with an inducible expression of p53/p73 plus 14-3-3σ with ADR treatment. Tumor formation was then assayed. Moreover, 66 primary breast cancer specimens and paired tumor-free breast specimens obtained from the female patients were examined. Results showed that the expression of p73 and 14-3-3σ in breast cancer specimens was significantly lower than the tumor-free breast specimens and that 14-3-3σ expression was positively correlated with the expression of p73. Furthermore, overexpression of 14-3-3σ counteracts tumorigenicity by positively regulating p73 in p53-mutated or -deficient cancers in vivo. Therefore, our results may lead to the use of 14-3-3σ in the therapeutic application for the p53-mutated and p73-expressed breast cancer patients. PMID:22848285

  12. Expansion of a 12-kb VNTR containing the REXO1L1 gene cluster underlies the microscopically visible euchromatic variant of 8q21.2

    PubMed Central

    Tyson, Christine; Sharp, Andrew J; Hrynchak, Monica; Yong, Siu L; Hollox, Edward J; Warburton, Peter; Barber, John CK

    2014-01-01

    Copy number variants visible with the light microscope have been described as euchromatic variants (EVs) and EVs with extra G-light material at 8q21.2 have been reported only once before. We report four further patients with EVs of 8q21.2 ascertained for clinical (3) or reproductive reasons (1). Enhanced signal strength from two overlapping bacterial artificial chromosomes (BACs) and microarray analysis mapped the EV to a 284-kb interval in the reference genome. This interval consists of a sequence gap flanked by segmental duplications that contain the 12-kb components of one of the largest Variable Number Tandem Repeat arrays in the human genome. Using digital NanoString technology with a custom probe for the RNA exonuclease 1 homologue (S. cerevisiae)-like 1 (REXO1L1) gene within each 12-kb repeat, significantly enhanced diploid copy numbers of 270 and 265 were found in an EV family and a median diploid copy number of 166 copies in 216 controls. These 8q21.2 EVs are not thought to have clinical consequences as the phenotypes of the probands were inconsistent, those referred for reproductive reasons were otherwise phenotypically normal and the REXO1L1 gene has no known disease association. This EV was found in 4/3078 (1 in 770) consecutive referrals for chromosome analysis and needs to be distinguished from pathogenic imbalances of medial 8q. The REXO1L1 gene product is a marker of hepatitis C virus (HCV) infection and a possible association between REXO1L1 copy number and susceptibility to HCV infection, progression or response to treatment has not yet been excluded. PMID:24045839

  13. Positive regulation of myoblast differentiation by medaka Neu3b sialidase through gangliosides desialylation.

    PubMed

    Shiozaki, Kazuhiro; Harasaki, Yusuke; Fukuda, Midori; Yoshinaga, Ayana; Ryuzono, Sena; Chigwechokha, Petros Kingstone; Komatsu, Masaharu; Miyagi, Taeko

    2016-04-01

    Sialidase Neu3b is an unique enzyme conserved in medaka and tilapia, but not in mammals. Previous study revealed that medaka Neu3b is localized at cytosol and is a ganglioside-specific sialidase. Neu3b functions, however, have not been understood, while Neu3a sialidase, which is widely conserved from human to fish, is known as a regulator of neurite formation. Here, we investigated the biological function of Neu3b for C2C12 myoblast cell differentiation. Bioinformatics analysis using genome browser revealed the presence of neu3b gene in some orders of fish species such as Beloniformes, Perciformes and Cyprinodontiformes. With the treatment of 2% horse serum, Neu3b-overexpression accelerated myoblast cell differentiation to myotubes accompanied with up-regulation of myogenesis biomarkers mRNA, myod and myog. Neu3b altered ganglioside composition in C2C12 cells results showing a decrease in GM2, and the increase of Lac-Cer, while desialylation of glycoproteins were not detected. Contrary to cell differentiation, Neu3b cell proliferation was suppressed in normal growth medium. To understand the mechanism of the alteration of cell differentiation and proliferation, phosphorylation of signal molecules in EGFR/ERK pathway was investigated. Neu3b induced a decline in phosphorylation of ERK and EGFR. Surprisingly, immuno-blot and real-time PCR analysis revealed that down-regulation of egfr gene could be involved in the acceleration of cell differentiation by Neu3b. These results suggested that Neu3b sialidase is a positive regulator for myoblast differentiation, similar with mammalian cytosolic sialidase Neu2.

  14. Rasputin Functions as a Positive Regulator of Orb in Drosophila Oogenesis

    PubMed Central

    Sinsimer, Kristina S.; Wong, Li Chin; McLeod, Ian; Yates, John; Haynes, Susan; Schedl, Paul

    2013-01-01

    The determination of cell fate and the establishment of polarity axes during Drosophila oogenesis depend upon pathways that localize mRNAs within the egg chamber and control their on-site translation. One factor that plays a central role in regulating on-site translation of mRNAs is Orb. Orb is a founding member of the conserved CPEB family of RNA-binding proteins. These proteins bind to target sequences in 3′ UTRs and regulate mRNA translation by modulating poly(A) tail length. In addition to controlling the translation of axis-determining mRNAs like grk, fs(1)K10, and osk, Orb protein autoregulates its own synthesis by binding to orb mRNA and activating its translation. We have previously shown that Rasputin (Rin), the Drosophila homologue of Ras-GAP SH3 Binding Protein (G3BP), associates with Orb in a messenger ribonucleoprotein (mRNP) complex. Rin is an evolutionarily conserved RNA-binding protein believed to function as a link between Ras signaling and RNA metabolism. Here we show that Orb and Rin form a complex in the female germline. Characterization of a new rin allele shows that rin is essential for oogenesis. Co-localization studies suggest that Orb and Rin form a complex in the oocyte at different stages of oogenesis. This is supported by genetic and biochemical analyses showing that rin functions as a positive regulator in the orb autoregulatory pathway by increasing Orb protein expression. Tandem Mass Spectrometry analysis shows that several canonical stress granule proteins are associated with the Orb-Rin complex suggesting that a conserved mRNP complex regulates localized translation during oogenesis in Drosophila. PMID:24069162

  15. Rasputin functions as a positive regulator of orb in Drosophila oogenesis.

    PubMed

    Costa, Alexandre; Pazman, Cecilia; Sinsimer, Kristina S; Wong, Li Chin; McLeod, Ian; Yates, John; Haynes, Susan; Schedl, Paul

    2013-01-01

    The determination of cell fate and the establishment of polarity axes during Drosophila oogenesis depend upon pathways that localize mRNAs within the egg chamber and control their on-site translation. One factor that plays a central role in regulating on-site translation of mRNAs is Orb. Orb is a founding member of the conserved CPEB family of RNA-binding proteins. These proteins bind to target sequences in 3' UTRs and regulate mRNA translation by modulating poly(A) tail length. In addition to controlling the translation of axis-determining mRNAs like grk, fs(1)K10, and osk, Orb protein autoregulates its own synthesis by binding to orb mRNA and activating its translation. We have previously shown that Rasputin (Rin), the Drosophila homologue of Ras-GAP SH3 Binding Protein (G3BP), associates with Orb in a messenger ribonucleoprotein (mRNP) complex. Rin is an evolutionarily conserved RNA-binding protein believed to function as a link between Ras signaling and RNA metabolism. Here we show that Orb and Rin form a complex in the female germline. Characterization of a new rin allele shows that rin is essential for oogenesis. Co-localization studies suggest that Orb and Rin form a complex in the oocyte at different stages of oogenesis. This is supported by genetic and biochemical analyses showing that rin functions as a positive regulator in the orb autoregulatory pathway by increasing Orb protein expression. Tandem Mass Spectrometry analysis shows that several canonical stress granule proteins are associated with the Orb-Rin complex suggesting that a conserved mRNP complex regulates localized translation during oogenesis in Drosophila.

  16. Nuclear dynamical deformation induced hetero- and euchromatin positioning

    NASA Astrophysics Data System (ADS)

    Awazu, Akinori

    2015-09-01

    We studied the role of active deformation dynamics in cell nuclei in chromatin positioning. Model chains containing two types of regions, with high (euchromatic) or low (heterochromatic) mobility, were confined in a pulsating container simulating a nucleus showing dynamic deformations. Brownian dynamic simulations show that the positioning of low mobility regions changes from sites near the periphery to the center if the affinity between these regions and the container periphery disappears. The former and latter positionings are similar to the "conventional" and "inverted" chromatin positionings in nuclei of normal differentiated cells and cells lacking Lamin-related proteins. Additionally, nuclear dynamical deformation played essential roles in "inverted" chromatin positioning.

  17. Nitric oxide is a positive regulator of the Warburg effect in ovarian cancer cells

    PubMed Central

    Caneba, C A; Yang, L; Baddour, J; Curtis, R; Win, J; Hartig, S; Marini, J; Nagrath, D

    2014-01-01

    Ovarian cancer (OVCA) is among the most lethal gynecological cancers leading to high mortality rates among women. Increasing evidence indicate that cancer cells undergo metabolic transformation during tumorigenesis and growth through nutrients and growth factors available in tumor microenvironment. This altered metabolic rewiring further enhances tumor progression. Recent studies have begun to unravel the role of amino acids in the tumor microenvironment on the proliferation of cancer cells. One critically important, yet often overlooked, component to tumor growth is the metabolic reprogramming of nitric oxide (NO) pathways in cancer cells. Multiple lines of evidence support the link between NO and tumor growth in some cancers, including pancreas, breast and ovarian. However, the multifaceted role of NO in the metabolism of OVCA is unclear and direct demonstration of NO's role in modulating OVCA cells' metabolism is lacking. This study aims at indentifying the mechanistic links between NO and OVCA metabolism. We uncover a role of NO in modulating OVCA metabolism: NO positively regulates the Warburg effect, which postulates increased glycolysis along with reduced mitochondrial activity under aerobic conditions in cancer cells. Through both NO synthesis inhibition (using L-arginine deprivation, arginine is a substrate for NO synthase (NOS), which catalyzes NO synthesis; using L-Name, a NOS inhibitor) and NO donor (using DETA-NONOate) analysis, we show that NO not only positively regulates tumor growth but also inhibits mitochondrial respiration in OVCA cells, shifting these cells towards glycolysis to maintain their ATP production. Additionally, NO led to an increase in TCA cycle flux and glutaminolysis, suggesting that NO decreases ROS levels by increasing NADPH and glutathione levels. Our results place NO as a central player in the metabolism of OVCA cells. Understanding the effects of NO on cancer cell metabolism can lead to the development of NO targeting drugs

  18. Ly49Q, an ITIM-bearing NK receptor, positively regulates osteoclast differentiation

    SciTech Connect

    Hayashi, Mikihito; Nakashima, Tomoki; Kodama, Tatsuhiko; Makrigiannis, Andrew P.; Toyama-Sorimachi, Noriko; Takayanagi, Hiroshi

    2010-03-12

    Osteoclasts, multinucleated cells that resorb bone, play a key role in bone remodeling. Although immunoreceptor tyrosine-based activation motif (ITAM)-mediated signaling is critical for osteoclast differentiation, the significance of immunoreceptor tyrosine-based inhibitory motif (ITIM) has not been well understood. Here we report the function of Ly49Q, an Ly49 family member possessing an ITIM motif, in osteoclastogenesis. Ly49Q is selectively induced by receptor activator of nuclear factor-{kappa}B (NF-{kappa}B) ligand (RANKL) stimulation in bone marrow-derived monocyte/macrophage precursor cells (BMMs) among the Ly49 family of NK receptors. The knockdown of Ly49Q resulted in a significant reduction in the RANKL-induced formation of tartrate-resistance acid phosphatase (TRAP)-positive multinucleated cells, accompanied by a decreased expression of osteoclast-specific genes such as Nfatc1, Tm7sf4, Oscar, Ctsk, and Acp5. Osteoclastogenesis was also significantly impaired in Ly49Q-deficient cells in vitro. The inhibitory effect of Ly49Q-deficiency may be explained by the finding that Ly49Q competed for the association of Src-homology domain-2 phosphatase-1 (SHP-1) with paired immunoglobulin-like receptor-B (PIR-B), an ITIM-bearing receptor which negatively regulates osteoclast differentiation. Unexpectedly, Ly49Q deficiency did not lead to impaired osteoclast formation in vivo, suggesting the existence of a compensatory mechanism. This study provides an example in which an ITIM-bearing receptor functions as a positive regulator of osteoclast differentiation.

  19. Atrophin Protein RERE Positively Regulates Notch Targets in the Developing Vertebrate Spinal Cord.

    PubMed

    Wang, Hui; Gui, Hongxing; Rallo, Michael S; Xu, Zhiyan; Matise, Michael P

    2017-01-31

    The Notch signaling pathway controls cell fate decision, proliferation and other biological functions in both vertebrates and invertebrates. Precise regulation of the canonical Notch pathway ensures robustness of the signal throughout development and adult tissue homeostasis. Aberrant Notch signaling results in profound developmental defects and is linked to many human diseases. In this study, we identified the Atrophin family protein RERE (also called Atro2) as a positive regulator of Notch target Hes genes in the developing vertebrate spinal cord. Prior studies have shown that during early embryogenesis in mouse and zebrafish, deficit of RERE causes various patterning defects in multiple organs including the neural tube. Here, we detected the expression of RERE in the developing chick spinal cord, and found that normal RERE activity is needed for proper neural progenitor proliferation and neuronal differentiation possibly by affecting Notch mediated Hes expression. In mammalian cells, RERE co-immunoprecipitates with CBF1 and Notch intracellular domain (NICD), and is recruited to nuclear foci formed by overexpressed NICD1. RERE is also necessary for NICD to activate the expression of Notch target genes. Our findings suggest that RERE stimulates Notch target gene expression by preventing degradation of NICD protein, thereby facilitating the assembly of a transcriptional activating complex containing NICD, CSL and other coactivators. This article is protected by copyright. All rights reserved.

  20. HY5, a positive regulator of light signaling, negatively controls the unfolded protein response in Arabidopsis

    PubMed Central

    Nawkar, Ganesh M.; Kang, Chang Ho; Maibam, Punyakishore; Park, Joung Hun; Jung, Young Jun; Chae, Ho Byoung; Chi, Yong Hun; Jung, In Jung; Kim, Woe Yeon; Yun, Dae-Jin; Lee, Sang Yeol

    2017-01-01

    Light influences essentially all aspects of plant growth and development. Integration of light signaling with different stress response results in improvement of plant survival rates in ever changing environmental conditions. Diverse environmental stresses affect the protein-folding capacity of the endoplasmic reticulum (ER), thus evoking ER stress in plants. Consequently, the unfolded protein response (UPR), in which a set of molecular chaperones is expressed, is initiated in the ER to alleviate this stress. Although its underlying molecular mechanism remains unknown, light is believed to be required for the ER stress response. In this study, we demonstrate that increasing light intensity elevates the ER stress sensitivity of plants. Moreover, mutation of the ELONGATED HYPOCOTYL 5 (HY5), a key component of light signaling, leads to tolerance to ER stress. This enhanced tolerance of hy5 plants can be attributed to higher expression of UPR genes. HY5 negatively regulates the UPR by competing with basic leucine zipper 28 (bZIP28) to bind to the G-box–like element present in the ER stress response element (ERSE). Furthermore, we found that HY5 undergoes 26S proteasome-mediated degradation under ER stress conditions. Conclusively, we propose a molecular mechanism of crosstalk between the UPR and light signaling, mediated by HY5, which positively mediates light signaling, but negatively regulates UPR gene expression. PMID:28167764

  1. SOCS1 Mimetics and Antagonists: A Complementary Approach to Positive and Negative Regulation of Immune Function

    PubMed Central

    Ahmed, Chulbul M. I.; Larkin, Joseph; Johnson, Howard M.

    2015-01-01

    Suppressors of cytokine signaling (SOCS) are inducible intracellular proteins that play essential regulatory roles in both immune and non-immune function. Of the eight known members, SOCS1 and SOCS3 in conjunction with regulatory T cells play key roles in regulation of the immune system. Molecular tools such as gene transfections and siRNA have played a major role in our functional understanding of the SOCS proteins where a key functional domain of 12-amino acid residues called the kinase inhibitory region (KIR) has been identified on SOCS1 and SOCS3. KIR plays a key role in inhibition of the JAK2 tyrosine kinase, which in turn plays a key role in cytokine signaling. A peptide corresponding to KIR (SOCS1-KIR) bound to the activation loop of JAK2 and inhibited tyrosine phosphorylation of STAT1α transcription factor by JAK2. Cell internalized SOCS1-KIR is a potent therapeutic in the experimental allergic encephalomyelitis (EAE) mouse model of multiple sclerosis and showed promise in a psoriasis model and a model of diabetes-associated cardiovascular disease. By contrast, a peptide, pJAK2(1001–1013), that corresponds to the activation loop of JAK2 is a SOCS1 antagonist. The antagonist enhanced innate and adaptive immune response against a broad range of viruses including herpes simplex virus, vaccinia virus, and an EMC picornavirus. SOCS mimetics and antagonists are thus potential therapeutics for negative and positive regulation of the immune system. PMID:25954276

  2. Positive roles for negative regulators in the mating response of yeast

    PubMed Central

    Houser, John R; Ford, Eintou; Nagiec, Michal J; Errede, Beverly; Elston, Timothy C

    2012-01-01

    All cells must detect and respond to changes in their environment, often through changes in gene expression. The yeast pheromone pathway has been extensively characterized, and is an ideal system for studying transcriptional regulation. Here we combine computational and experimental approaches to study transcriptional regulation mediated by Ste12, the key transcription factor in the pheromone response. Our mathematical model is able to explain multiple counterintuitive experimental results and led to several novel findings. First, we found that the transcriptional repressors Dig1 and Dig2 positively affect transcription by stabilizing Ste12. This stabilization through protein–protein interactions creates a large pool of Ste12 that is rapidly activated following pheromone stimulation. Second, we found that protein degradation follows saturating kinetics, explaining the long half-life of Ste12 in mutants expressing elevated amounts of Ste12. Finally, our model reveals a novel mechanism for robust perfect adaptation through protein–protein interactions that enhance complex stability. This mechanism allows the transcriptional response to act on a shorter time scale than upstream pathway activity. PMID:22669614

  3. Heparan sulfate regulates the number and centrosome positioning of Drosophila male germline stem cells.

    PubMed

    Levings, Daniel C; Arashiro, Takeshi; Nakato, Hiroshi

    2016-03-15

    Stem cell division is tightly controlled via secreted signaling factors and cell adhesion molecules provided from local niche structures. Molecular mechanisms by which each niche component regulates stem cell behaviors remain to be elucidated. Here we show that heparan sulfate (HS), a class of glycosaminoglycan chains, regulates the number and asymmetric division of germline stem cells (GSCs) in the Drosophila testis. We found that GSC number is sensitive to the levels of 6-O sulfate groups on HS. Loss of 6-O sulfation also disrupted normal positioning of centrosomes, a process required for asymmetric division of GSCs. Blocking HS sulfation specifically in the niche, termed the hub, led to increased GSC numbers and mispositioning of centrosomes. The same treatment also perturbed the enrichment of Apc2, a component of the centrosome-anchoring machinery, at the hub-GSC interface. This perturbation of the centrosome-anchoring process ultimately led to an increase in the rate of spindle misorientation and symmetric GSC division. This study shows that specific HS modifications provide a novel regulatory mechanism for stem cell asymmetric division. The results also suggest that HS-mediated niche signaling acts upstream of GSC division orientation control.

  4. Positive Regulation of Interleukin-2 Expression by a Pseudokinase, Tribbles 1, in Activated T Cells.

    PubMed

    Miyajima, Chiharu; Itoh, Yuka; Inoue, Yasumichi; Hayashi, Hidetoshi

    2015-01-01

    Tribbles 1 (TRB1), a member of the Tribbles family, is a pseudokinase that is conserved among species and implicated in various human diseases including leukemia, cardiovascular diseases, and metabolic disorders. However, the role of TRB1 in the immune response is not understood. To evaluate this role, we examined regulation of TRB1 expression and the function of TRB1 in interleukin-2 (IL-2) induction in Jurkat cells, a human acute T cell leukemia cell line. We found that TRB1 was strongly induced by phorbol 12-myristate 13-acetate (PMA) and ionomycin in these cells. IL-2 expression was induced in Jurkat cells activated by PMA and ionomycin; however, knockdown of TRB1 resulted in decreased induction of IL-2. TRB1 null Jurkat cells established using the CRISPR/Cas9 system also showed reduction of IL-2 expression on PMA/ionomycin stimulation. TRB1 knockdown also markedly inhibited IL-2 promoter activation. To determine the mechanism of the stimulatory effect on IL-2 induction, we focused on histone deacetylases (HDACs), and found that HDAC1 preferentially interacts with TRB1. TRB1 suppressed the interaction of HDAC1 with nuclear factor of activated T cells 2 (NFAT2), which is a crucial transcription factor for IL-2 induction. These results indicate that TRB1 is a positive regulator of IL-2 induction in activated T cells.

  5. LncRNA AK023948 is a positive regulator of AKT

    PubMed Central

    Koirala, Pratirodh; Huang, Jianguo; Ho, Tsui-Ting; Wu, Fangting; Ding, Xianfeng; Mo, Yin-Yuan

    2017-01-01

    Despite the overwhelming number of human long non-coding RNAs (lncRNAs) reported so far, little is known about their physiological functions for the majority of them. The present study uses a CRISPR/Cas9-based synergistic activation mediator (SAM) system to identify potential lncRNAs capable of regulating AKT activity. Among lncRNAs identified from this screen, we demonstrate that AK023948 is a positive regulator for AKT. Knockout of AK023948 suppresses, whereas rescue with AK023948 restores the AKT activity. Mechanistically, AK023948 functionally interacts with DHX9 and p85. Importantly, AK023948 is required for the interaction between DHX9 and p85 to hence the p85 stability and promote AKT activity. Finally, AK023948 is upregulated in breast cancer; interrogation of TCGA data set indicates that upregulation of DHX9 in breast cancer is associated with poor survival. Together, this study demonstrates two previously uncharacterized factors AK023948 and DHX9 as important players in the AKT pathway, and that their upregulation may contribute to breast tumour progression. PMID:28176758

  6. Tomato MAPKKKε is a positive regulator of cell-death signaling networks associated with plant immunity.

    PubMed

    Melech-Bonfil, Shiri; Sessa, Guido

    2010-11-01

    Mitogen-activated protein (MAP) kinase cascades are fundamental components of the signaling pathways associated with plant immunity. Despite the large number of MAP kinase kinase kinases (MAPKKK) encoded in the plant genome, only very few of them have an assigned function. Here, we identified MAPKKK gene of tomato (Solanum lycopersicum), SIMAPKKKε, which is required for hypersensitive response cell death and disease resistance against Gram-negative bacterial pathogens. Silencing of SIMAPKKKε compromised tomato resistance to Xanthomonas campestris and Pseudomonas syringae strains, resulting in the appearance of disease symptoms and enhanced bacterial growth. In addition, silencing of NbMAPKKKε in Nicotiana benthamiana plants significantly inhibited the cell death triggered by expression of different R gene/effector gene pairs. Conversely, overexpression of either the full-length SIMAPKKKε gene or its kinase domain in N. benthamiana leaves caused pathogen-independent activation of cell death that required an intact kinase catalytic domain. Moreover, by suppressing the expression of various MAPKK and MAPK genes and overexpressing the SIMAPKKKε kinase domain, we identified a signaling cascade acting downstream of SIMAPKKKε that includes MEK2, WIPK and SIPK. Additional epistasis experiments revealed that SIPKK functions as a negative regulator of SIMAPKKKε-mediated cell death. Our results provide evidence that SIMAPKKKε is a signaling molecule that positively regulates cell death networks associated with plant immunity.

  7. cAMP-response-element-binding protein positively regulates breast cancer metastasis and subsequent bone destruction

    SciTech Connect

    Son, Jieun; Lee, Jong-Ho; Kim, Ha-Neui; Ha, Hyunil Lee, Zang Hee

    2010-07-23

    Research highlights: {yields} CREB is highly expressed in advanced breast cancer cells. {yields} Tumor-related factors such as TGF-{beta} further elevate CREB expression. {yields} CREB upregulation stimulates metastatic potential of breast cancer cells. {yields} CREB signaling is required for breast cancer-induced bone destruction. -- Abstract: cAMP-response-element-binding protein (CREB) signaling has been reported to be associated with cancer development and poor clinical outcome in various types of cancer. However, it remains to be elucidated whether CREB is involved in breast cancer development and osteotropism. Here, we found that metastatic MDA-MB-231 breast cancer cells exhibited higher CREB expression than did non-metastatic MCF-7 cells and that CREB expression was further increased by several soluble factors linked to cancer progression, such as IL-1, IGF-1, and TGF-{beta}. Using wild-type CREB and a dominant-negative form (K-CREB), we found that CREB signaling positively regulated the proliferation, migration, and invasion of MDA-MB-231 cells. In addition, K-CREB prevented MDA-MB-231 cell-induced osteolytic lesions in a mouse model of cancer metastasis. Furthermore, CREB signaling in cancer cells regulated the gene expression of PTHrP, MMPs, and OPG, which are closely involved in cancer metastasis and bone destruction. These results indicate that breast cancer cells acquire CREB overexpression during their development and that this CREB upregulation plays an important role in multiple steps of breast cancer bone metastasis.

  8. Notch and TGFβ form a positive regulatory loop and regulate EMT in epithelial ovarian cancer cells.

    PubMed

    Zhou, Jiesi; Jain, Saket; Azad, Abul K; Xu, Xia; Yu, Hai Chuan; Xu, Zhihua; Godbout, Roseline; Fu, YangXin

    2016-08-01

    Epithelial-mesenchymal transition (EMT) plays a critical role in the progression of epithelial ovarian cancer (EOC). However, the mechanisms that regulate EMT in EOC are not fully understood. Here, we report that activation of Notch1 induces EMT in EOC cells as evidenced by downregulation of E-cadherin and cytokeratins, upregulation of Slug and Snail, as well as morphological changes. Interestingly, activation of Notch1 increases TGFβ/Smad signaling by upregulating the expression of TGFβ and TGFβ type 1 receptor. Time course experiments demonstrate that inhibition of Notch by DAPT (a γ-secretase inhibitor) decreases TGFβ-induced phosphorylation of receptor Smads at late, but not at early, timepoints. These results suggest that Notch activation plays a role in sustaining TGFβ/Smad signaling in EOC cells. Furthermore, inhibition of Notch by DAPT decreases TGFβ induction of Slug and repression of E-cadherin and knockdown of Notch1 decreases TGFβ-induced repression of E-cadherin, indicating that Notch is required, at least in part, for TGFβ-induced EMT in EOC cells. On the other hand, TGFβ treatment increases the expression of Notch ligand Jagged1 and Notch target gene HES1 in EOC cells. Functionally, the combination of Notch1 activation and TGFβ treatment is more potent in promoting motility and migration of EOC cells than either stimulation alone. Taken together, our results indicate that Notch and TGFβ form a reciprocal positive regulatory loop and cooperatively regulate EMT and promote EOC cell motility and migration.

  9. HY5, a positive regulator of light signaling, negatively controls the unfolded protein response in Arabidopsis.

    PubMed

    Nawkar, Ganesh M; Kang, Chang Ho; Maibam, Punyakishore; Park, Joung Hun; Jung, Young Jun; Chae, Ho Byoung; Chi, Yong Hun; Jung, In Jung; Kim, Woe Yeon; Yun, Dae-Jin; Lee, Sang Yeol

    2017-02-21

    Light influences essentially all aspects of plant growth and development. Integration of light signaling with different stress response results in improvement of plant survival rates in ever changing environmental conditions. Diverse environmental stresses affect the protein-folding capacity of the endoplasmic reticulum (ER), thus evoking ER stress in plants. Consequently, the unfolded protein response (UPR), in which a set of molecular chaperones is expressed, is initiated in the ER to alleviate this stress. Although its underlying molecular mechanism remains unknown, light is believed to be required for the ER stress response. In this study, we demonstrate that increasing light intensity elevates the ER stress sensitivity of plants. Moreover, mutation of the ELONGATED HYPOCOTYL 5 (HY5), a key component of light signaling, leads to tolerance to ER stress. This enhanced tolerance of hy5 plants can be attributed to higher expression of UPR genes. HY5 negatively regulates the UPR by competing with basic leucine zipper 28 (bZIP28) to bind to the G-box-like element present in the ER stress response element (ERSE). Furthermore, we found that HY5 undergoes 26S proteasome-mediated degradation under ER stress conditions. Conclusively, we propose a molecular mechanism of crosstalk between the UPR and light signaling, mediated by HY5, which positively mediates light signaling, but negatively regulates UPR gene expression.

  10. Heparan sulfate regulates the number and centrosome positioning of Drosophila male germline stem cells

    PubMed Central

    Levings, Daniel C.; Arashiro, Takeshi; Nakato, Hiroshi

    2016-01-01

    Stem cell division is tightly controlled via secreted signaling factors and cell adhesion molecules provided from local niche structures. Molecular mechanisms by which each niche component regulates stem cell behaviors remain to be elucidated. Here we show that heparan sulfate (HS), a class of glycosaminoglycan chains, regulates the number and asymmetric division of germline stem cells (GSCs) in the Drosophila testis. We found that GSC number is sensitive to the levels of 6-O sulfate groups on HS. Loss of 6-O sulfation also disrupted normal positioning of centrosomes, a process required for asymmetric division of GSCs. Blocking HS sulfation specifically in the niche, termed the hub, led to increased GSC numbers and mispositioning of centrosomes. The same treatment also perturbed the enrichment of Apc2, a component of the centrosome-anchoring machinery, at the hub–GSC interface. This perturbation of the centrosome-anchoring process ultimately led to an increase in the rate of spindle misorientation and symmetric GSC division. This study shows that specific HS modifications provide a novel regulatory mechanism for stem cell asymmetric division. The results also suggest that HS-mediated niche signaling acts upstream of GSC division orientation control. PMID:26792837

  11. The F-box protein MAX2 functions as a positive regulator of photomorphogenesis in Arabidopsis.

    PubMed

    Shen, Hui; Luong, Phi; Huq, Enamul

    2007-12-01

    Light is vital for plant growth and development. To respond to ambient light signals, plants are equipped with an array of photoreceptors, including phytochromes that sense red (R)/far-R (FR) regions and cryptochromes and phototropins that respond to the ultraviolet-A/blue (B) region of the light spectrum, respectively. Several positively and negatively acting components in light-signaling pathways have been identified using genetic approaches; however, the pathways are not saturated. Here, we characterize a new mutant named pleiotropic photosignaling (pps), isolated from a genetic screen under continuous R light. pps has longer hypocotyls and slightly smaller cotyledons under continuous R, FR, and B light compared to that of the wild type. pps is also hyposensitive to both R and FR light-induced seed germination. Although photosynthetic marker genes are constitutively expressed in pps in the dark at high levels, the expression of early light-regulated genes is reduced in the pps seedlings compared to wild-type seedlings under R light. PPS encodes MAX2/ORE9 (for MORE AXILLARY BRANCHES2/ORESARA9), an F-box protein involved in inflorescence architecture and senescence. MAX2 is expressed ubiquitously in the seedling stage. However, its expression is restricted to vascular tissues and meristems at adult stages. MAX2 is also localized to the nucleus. As an F-box protein, MAX2 is predicted to be a component of the SCF (for SKP, Cullin, and F-box protein) complex involved in regulated proteolysis. These results suggest that SCF(MAX2) plays critical roles in R, FR, and B light-signaling pathways. In addition, MAX2 might regulate multiple targets at different developmental stages to optimize plant growth and development.

  12. Quantitative trait gene Slit2 positively regulates murine hematopoietic stem cell numbers

    PubMed Central

    Waterstrat, Amanda; Rector, Kyle; Geiger, Hartmut; Liang, Ying

    2016-01-01

    Hematopoietic stem cells (HSC) demonstrate natural variation in number and function. The genetic factors responsible for the variations (or quantitative traits) are largely unknown. We previously identified a gene whose differential expression underlies the natural variation of HSC numbers in C57BL/6 (B6) and DBA/2 (D2) mice. We now report the finding of another gene, Slit2, on chromosome 5 that also accounts for variation in HSC number. In reciprocal chromosome 5 congenic mice, introgressed D2 alleles increased HSC numbers, whereas B6 alleles had the opposite effect. Using gene array and quantitative polymerase chain reaction, we identified Slit2 as a quantitative trait gene whose expression was positively correlated with the number of HSCs. Ectopic expression of Slit2 not only increased the number of the long-term colony forming HSCs, but also enhanced their repopulation capacity upon transplantation. Therefore, Slit2 is a novel quantitative trait gene and a positive regulator of the number and function of murine HSCs. This finding suggests that Slit2 may be a potential therapeutic target for the effective in vitro and in vivo expansion of HSCs without compromising normal hematopoiesis. PMID:27503415

  13. Youth development program participation and intentional self-regulation skills: contextual and individual bases of pathways to positive youth development.

    PubMed

    Mueller, Megan Kiely; Phelps, Erin; Bowers, Edmond P; Agans, Jennifer P; Urban, Jennifer Brown; Lerner, Richard M

    2011-12-01

    The present research used data from Grades 8, 9, and 10 of the 4-H Study of Positive Youth Development, a longitudinal study involving U.S. adolescents, in order to better elucidate the process through which the strengths of youth and the ecological resources promoting healthy development (such as out-of-school-time programs) may contribute to thriving. We examined the relationship between adolescents' self-regulation skills (selection, optimization, and compensation) and their participation in youth development (YD) programs across Grades 8 and 9 in predicting Grade 10 PYD and Contribution. Results indicated that while self-regulation skills alone predicted PYD, self regulation and YD program participation both predicted Contribution. In addition, Grade 8 YD participation positively predicted Grade 9 self regulation, which, in turn, predicted Grade 10 PYD and Contribution. We discuss how the alignment of youth strengths and resources within the environment may promote positive youth development.

  14. Identification of ATP Citrate Lyase as a Positive Regulator of Glycolytic Function in Glioblastomas

    PubMed Central

    Beckner, Marie E.; Fellows-Mayle, Wendy; Zhang, Zhe; Agostino, Naomi R.; Kant, Jeffrey A.; Day, Billy W.; Pollack, Ian F.

    2009-01-01

    Glioblastomas, the most malignant type of glioma, are more glycolytic than normal brain tissue. Robust migration of glioblastoma cells has been previously demonstrated under glycolytic conditions and their pseudopodia contain increased glycolytic and decreased mitochondrial enzymes. Glycolysis is suppressed by metabolic acids, including citric acid which is excluded from mitochondria during hypoxia. We postulated that glioma cells maintain glycolysis by regulating metabolic acids, especially in their pseudopodia. The enzyme that breaks down cytosolic citric acid is ATP citrate lyase (ACLY). Our identification of increased ACLY in pseudopodia of U87 glioblastoma cells on 1D gels and immunoblots prompted investigation of ACLY gene expression in gliomas for survival data and correlation with expression of ENO1, that encodes enolase 1. Queries of the NIH’s REMBRANDT brain tumor database based on Affymetrix data indicated that decreased survival correlated with increased gene expression of ACLY in gliomas. Queries of gliomas and glioblastomas found an association of upregulated ACLY and ENO1 expression by chi square for all probe sets (reporters) combined and correlation for numbers of probe sets indicating shared upregulation of these genes. Real-time quantitative PCR confirmed correlation between ACLY and ENO1 in 21 glioblastomas (p < 0.001). Inhibition of ACLY with hydroxycitrate suppressed (p < 0.05) in vitro glioblastoma cell migration, clonogenicity and brain invasion under glycolytic conditions and enhanced the suppressive effects of a Met inhibitor on cell migration. In summary, gene expression data, proteomics and functional assays support ACLY as a positive regulator of glycolysis in glioblastomas. PMID:19795461

  15. SlnR is a positive pathway-specific regulator for salinomycin biosynthesis in Streptomyces albus.

    PubMed

    Zhu, Zhenhong; Li, Han; Yu, Pin; Guo, Yuanyang; Luo, Shuai; Chen, Zhongbin; Mao, Xuming; Guan, Wenjun; Li, Yongquan

    2017-02-01

    Salinomycin, a polyether antibiotic produced by Streptomyces albus, is widely used in animal husbandry as an anticoccidial drug and growth promoter. Situated within the salinomycin biosynthetic gene cluster, slnR encodes a LAL-family transcriptional regulator. The role of slnR in salinomycin production in S. albus was investigated by gene deletion, complementation, and overexpression. Gene replacement of slnR from S. albus chromosome results in almost loss of salinomycin production. Complementation of slnR restored salinomycin production, suggesting that SlnR is a positive regulator of salinomycin biosynthesis. Overexpression of slnR in S. albus led to about 25 % increase in salinomycin production compared to wild type. Quantitative RT-PCR analysis revealed that the expression of most sal structural genes was downregulated in the ΔslnR mutant but upregulated in the slnR overexpression strain. Electrophoretic mobility gel shift assays (EMSAs) also revealed that SlnR(DBD) binds directly to the three intergenic regions of slnQ-slnA1, slnF-slnT1, and slnC-slnB3. The SlnR binding sites within the three intergenic regions were determined by footprinting analysis and identified a consensus-directed repeat sequence 5'-ACCCCT-3'. These results indicated that SlnR modulated salinomycin biosynthesis as an enhancer via interaction with the promoters of slnA1, slnQ, slnF, slnT1, slnC, and slnB3 and activates the transcription of most of the genes belonging to the salinomycin gene cluster but not its own transcription.

  16. PEG-3, a nontransforming cancer progression gene, is a positive regulator of cancer aggressiveness and angiogenesis.

    PubMed

    Su, Z Z; Goldstein, N I; Jiang, H; Wang, M N; Duigou, G J; Young, C S; Fisher, P B

    1999-12-21

    Cancer is a progressive disease culminating in acquisition of metastatic potential by a subset of evolving tumor cells. Generation of an adequate blood supply in tumors by production of new blood vessels, angiogenesis, is a defining element in this process. Although extensively investigated, the precise molecular events underlying tumor development, cancer progression, and angiogenesis remain unclear. Subtraction hybridization identified a genetic element, progression elevated gene-3 (PEG-3), whose expression directly correlates with cancer progression and acquisition of oncogenic potential by transformed rodent cells. We presently demonstrate that forced expression of PEG-3 in tumorigenic rodent cells, and in human cancer cells, increases their oncogenic potential in nude mice as reflected by a shorter tumor latency time and the production of larger tumors with increased vascularization. Moreover, inhibiting endogenous PEG-3 expression in progressed rodent cancer cells by stable expression of an antisense expression vector extinguishes the progressed cancer phenotype. Cancer aggressiveness of PEG-3 expressing rodent cells correlates directly with increased RNA transcription, elevated mRNA levels, and augmented secretion of vascular endothelial growth factor (VEGF). Furthermore, transient ectopic expression of PEG-3 transcriptionally activates VEGF in transformed rodent and human cancer cells. Taken together these data demonstrate that PEG-3 is a positive regulator of cancer aggressiveness, a process regulated by augmented VEGF production. These studies also support an association between expression of a single nontransforming cancer progression-inducing gene, PEG-3, and the processes of cancer aggressiveness and angiogenesis. In these contexts, PEG-3 may represent an important target molecule for developing cancer therapeutics and inhibitors of angiogenesis.

  17. TcpP protein is a positive regulator of virulence gene expression in Vibrio cholerae.

    PubMed

    Häse, C C; Mekalanos, J J

    1998-01-20

    The production of several virulence factors in Vibrio cholerae O1, including cholera toxin and the pilus colonization factor TCP (toxin-coregulated pilus), is strongly influenced by environmental conditions. To specifically identify membrane proteins involved in these signal transduction events, we examined a transposon library of V. cholerae generated by Tnbla mutagenesis for cells that produce TCP when grown under various nonpermissive conditions. To select for TCP-producing cells we used the recently described bacteriophage CTX phi-Kan, which uses TCP as its receptor and carries a gene encoding resistance to kanamycin. Among the isolated mutants was a transposon insertion in a gene homologous to nqrB from Vibrio alginolyticus, which encodes a subunit of a Na(+)-translocating NADH:ubiquinone oxidoreductase, and tcpI, encoding a chemo-receptor previously implicated in the negative regulation of TCP production. A third transposon mutant had an insertion in tcpP, which is in an operon with tcpH, a known positive regulator of TCP production. However, TcpP was shown to be essential for TCP production in V. cholerae, as a tcpP-deletion strain was deficient in pili production. The amino-terminal region of TcpP shows sequence homology to the DNA-binding domains of several regulatory proteins, including ToxR from V. cholerae and PsaE from Yersinia pestis. Like ToxR, TcpP activates transcription of the toxT gene, an essential activator of tcp operon transcription. Furthermore, TcpH, with its large periplasmic domain and inner membrane anchor, has a structure similar to that of ToxS and was shown to enhance the activity of TcpP. We propose that TcpP/TcpH constitute a pair of regulatory proteins functionally similar to ToxR/ToxS and PsaE/PsaF that are required for toxT transcription in V. cholerae.

  18. Crystal Structure of Human Cyclin K, a Positive Regulator of Cyclin-dependent Kinase 9

    PubMed Central

    Baek, Kyuwon; Brown, Raymond S.; Birrane, Gabriel; Ladias, John A.A.

    2007-01-01

    Summary Cyclin K and the closely related cyclins T1, T2a, and T2b interact with cyclin-dependent kinase 9 (CDK9) forming multiple nuclear complexes, collectively referred to as positive transcription elongation factor b (P-TEFb). Through phosphorylation of the C-terminal domain of the RNA polymerase II largest subunit, distinct P-TEFb species regulate the transcriptional elongation of specific genes that play central roles in human physiology and disease development, including cardiac hypertrophy and human immunodeficiency virus-1 pathogenesis. We have determined the crystal structure of human cyclin K (residues 11-267) at 1.5 Å resolution, which represents the first atomic structure of a P-TEFb subunit. The cyclin K fold comprises two typical cyclin boxes with two short helices preceding the N-terminal box. A prominent feature of cyclin K is an additional helix (H4a) in the first cyclin box that obstructs the binding pocket for the cell cycle inhibitor p27Kip1. Modeling of CDK9 bound to cyclin K provides insights into the structural determinants underlying the formation and regulation of this complex. A homology model of human cyclin T1 generated using the cyclin K as a template reveals that the two proteins have similar structures, as expected from their high sequence identity. Nevertheless, their CDK9-interacting surfaces display significant structural differences, which could potentially be exploited for the design of cyclin-targeted inhibitors of the CDK9–cyclin K and CDK9–cyclin T1 complexes. PMID:17169370

  19. Developmental silencing of the AtTERT gene is associated with increased H3K27me3 loading and maintenance of its euchromatic environment

    PubMed Central

    Ogrocká, Anna; Sýkorová, Eva; Fajkus, Jiří; Fojtová, Miloslava

    2012-01-01

    Telomerase, an enzyme responsible for the maintenance of linear chromosome ends, is precisely regulated during plant development. In animals, involvement of the epigenetic state of the telomerase reverse transcriptase (TERT) gene in the complex regulation of telomerase activity has been reported. To reveal whether epigenetic mechanisms participate in the regulation of plant telomerase, the relationship between telomerase activity in tissues of Arabidopsis thaliana and DNA methylation and histone modifications in the A. thaliana TERT (AtTERT) upstream region was studied. As expected, a gradual decrease of telomerase activity during leaf maturation was observed. A different pattern with a more progressive loss of telomerase activity and AtTERT transcription during leaf development was revealed in MET1 gene-knockout mutants. Analysis of DNA methylation in the AtTERT upstream region showed low levels of methylated cytosines without notable differences between telomerase-positive and telomerase-negative wild-type tissues. Surprisingly, a high level of CG methylation was found in the AtTERT coding region, although this type of methylation is a characteristic attribute of constitutively expressed genes. Analysis of chromatin modifications in the AtTERT upstream region and in exon 5 showed increased loading of the H3K27me3 mark in the telomerase-negative mature leaf compared to telomerase-positive seedlings, whereas H3K4me3, H3K9Ac, and H3K9me2 were approximately at the same level. Consistently, the chromatin structure of the AtTERT gene was maintained. These results are discussed in the context of the general involvement of epigenetic mechanisms in the regulation of gene expression and with respect to similar studies performed in animal models. PMID:22511802

  20. Positioning of extracellular loop 1 affects pore gating of the cystic fibrosis transmembrane conductance regulator

    PubMed Central

    Infield, Daniel T.; Cui, Guiying; Kuang, Christopher

    2015-01-01

    The cystic fibrosis (CF) transmembrane conductance regulator (CFTR) is a chloride ion channel, the dysfunction of which directly leads to the life-shortening disease CF. Extracellular loop 1 (ECL1) of CFTR contains several residues involved in stabilizing the open state of the channel; some, including D110, are sites of disease-associated gating mutations. Structures from related proteins suggest that the position of CFTR's extracellular loops may change considerably during gating. To better understand the roles of ECL1 in CFTR function, we utilized functional cysteine cross-linking to determine the effects of modulation of D110C-CFTR and of a double mutant of D110C with K892C in extracellular loop 4 (ECL4). The reducing agent DTT elicited a large potentiation of the macroscopic conductance of D110C/K892C-CFTR, likely due to breakage of a spontaneous disulfide bond between C110 and C892. DTT-reduced D110C/K892C-CFTR was rapidly inhibited by binding cadmium ions with high affinity, suggesting that these residues frequently come in close proximity in actively gating channels. Effects of DTT and cadmium on modulation of pore gating were demonstrated at the single-channel level. Finally, disulfided D110C/K892C-CFTR channels were found to be less sensitive than wild-type or DTT-treated D110C/K892C-CFTR channels to stimulation by IBMX, suggesting an impact of this conformational restriction on channel activation by phosphorylation. The results are best explained in the context of a model of CFTR gating wherein stable channel opening requires correct positioning of functional elements structurally influenced by ECL1. PMID:26684250

  1. DasR positively controls monensin production at two-level regulation in Streptomyces cinnamonensis.

    PubMed

    Zhang, Yue; Lin, Chun-Yan; Li, Xiao-Mei; Tang, Zheng-Kun; Qiao, Jianjun; Zhao, Guang-Rong

    2016-12-01

    The polyether ionophore antibiotic monensin is produced by Streptomyces cinnamonensis and is used as a coccidiostat for chickens and growth-promoting agent for cattle. Monensin biosynthetic gene cluster has been cloned and partially characterized. The GntR-family transcription factor DasR regulates antibiotic production and morphological development in Streptomyces coelicolor and Saccharopolyspora erythraea. In this study, we identified and characterized the two-level regulatory cascade of DasR to monensin production in S. cinnamonensis. Forward and reverse genetics by overexpression and antisense RNA silence of dasR revealed that DasR positively controls monensin production under nutrient-rich condition. Electrophoresis mobility shift assay (EMSA) showed that DasR protein specifically binds to the promoter regions of both pathway-specific regulatory gene monRII and biosynthetic genes monAIX, monE and monT. Semi-quantitative RT-PCR further confirmed that DasR upregulates the transcriptional levels of these genes during monensin fermentation. Subsequently, co-overexpressed dasR with pathway-specific regulatory genes monRI, monRII or monH greatly improved monensin production.

  2. Properdin, the positive regulator of complement, is highly C-mannosylated.

    PubMed

    Hartmann, S; Hofsteenge, J

    2000-09-15

    Properdin is the positive regulator of the alternative pathway of complement activation. The 53-kDa protein is essentially composed of six thrombospondin type 1 repeats, all of which contain the WXXW motif, the recognition sequence for C-mannosylation. C-Mannosylation is a post-translational modification of tryptophan residues in which, in contrast to the well known N- and O-glycosylation, the carbohydrate is attached via a C-C bond to C-2 of the indole moiety of tryptophan. C-Mannosylation was first found in human RNase 2 and interleukin-12. The terminal complement proteins C6-C9 also carry this modification as part of their thrombospondin type 1 repeats. We studied the C-mannosylation pattern of human properdin by mass spectrometry and Edman degradation. Properdin contains 20 tryptophans of which 17 are part of a WXXW motif. Fourteen tryptophans were found to be modified 100%. This is the first example of a protein in which the majority of tryptophan residues occurs in the C-mannosylated form. These results show that C-mannosylated proteins occur at several steps along the complement activation cascade. Therefore, this system would be ideal to investigate the function of C-mannosylation.

  3. A Molecular Mechanism to Regulate Lysosome Motility for Lysosome Positioning and Tubulation

    PubMed Central

    Li, Xinran; Rydzewski, Nicholas; Hider, Ahmad; Zhang, Xiaoli; Yang, Junsheng; Wang, Wuyang; Gao, Qiong; Cheng, Xiping; Xu, Haoxing

    2016-01-01

    To mediate the degradation of bio-macromolecules, lysosomes must traffic towards cargo-carrying vesicles for subsequent membrane fusion or fission. Mutations of the lysosomal Ca2+ channel TRPML1 cause lysosome storage disease (LSD) characterized by disordered lysosomal membrane trafficking in cells. Here we show that TRPML1 activity is required to promote Ca2+-dependent centripetal movement of lysosomes towards the perinuclear region, where autophagosomes accumulate, upon autophagy induction. ALG-2, an EF-hand-containing protein, serves as a lysosomal Ca2+ sensor that associates physically with the minus-end directed dynactin-dynein motor, while PI(3,5)P2, a lysosome-localized phosphoinositide, acts upstream of TRPML1. Furthermore, the PI(3,5)P2-TRPML1-ALG-2-dynein signaling is necessary for lysosome tubulation and reformation. In contrast, the TRPML1 pathway is not required for the perinuclear accumulation of lysosomes observed in many LSDs, which is instead likely caused by secondary cholesterol accumulation that constitutively activates Rab7-RILP-dependent retrograde transport. Collectively, Ca2+ release from lysosomes provides an on-demand mechanism regulating lysosome motility, positioning, and tubulation. PMID:26950892

  4. Toutatis, a TIP5-related protein, positively regulates Pannier function during Drosophila neural development.

    PubMed

    Vanolst, Luc; Fromental-Ramain, Catherine; Ramain, Philippe

    2005-10-01

    The GATA factor Pannier (Pnr) activates proneural expression through binding to a remote enhancer of the achaete-scute (ac-sc) complex. Chip associates both with Pnr and with the (Ac-Sc)-Daughterless heterodimer bound to the ac-sc promoters to give a proneural complex that facilitates enhancer-promoter communication during development. Using a yeast two-hybrid screening, we have identified Toutatis (Tou), which physically interacts with both Pnr and Chip. Loss-of-function and gain-of-function experiments indicate that Tou cooperates with Pnr and Chip during neural development. Tou shares functional domains with chromatin remodelling proteins, including TIP5 (termination factor TTFI-interacting protein 5) of NoRC (nucleolar remodelling complex), which mediates repression of RNA polymerase 1 transcription. In contrast, Tou acts positively to activate proneural gene expression. Moreover, we show that Iswi associates with Tou, Pnr and Chip, and is also required during Pnr-driven neural development. The results suggest that Tou and Iswi may belong to a complex that directly regulates the activity of Pnr and Chip during enhancer-promoter communication, possibly through chromatin remodelling.

  5. Regulation of alkane degradation pathway by a TetR family repressor via an autoregulation positive feedback mechanism in a Gram-positive Dietzia bacterium.

    PubMed

    Liang, Jie-Liang; Nie, Yong; Wang, Miaoxiao; Xiong, Guangming; Wang, Yi-Ping; Maser, Edmund; Wu, Xiao-Lei

    2016-01-01

    n-Alkanes are ubiquitous in nature and serve as important carbon sources for both Gram-positive and Gram-negative bacteria. Hydroxylation of n-alkanes by alkane monooxygenases is the first and most critical step in n-alkane metabolism. However, regulation of alkane degradation genes in Gram-positive bacteria remains poorly characterized. We therefore explored the transcriptional regulation of an alkB-type alkane hydroxylase-rubredoxin fusion gene, alkW1, from Dietzia sp. DQ12-45-1b. The alkW1 promoter was characterized and so was the putative TetR family regulator, AlkX, located downstream of alkW1 gene. We further identified an unusually long 48 bp inverted repeat upstream of alkW1 and demonstrated the binding of AlkX to this operator. Analytical ultracentrifugation and microcalorimetric results indicated that AlkX formed stable dimers in solution and two dimers bound to one operator in a positive cooperative fashion characterized by a Hill coefficient of 1.64 (± 0.03) [k(D)  = 1.06 (± 0.16) μM, k(D) ' = 0.05 (± 0.01) μM]. However, the DNA-binding affinity was disrupted in the presence of long-chain fatty acids (C10-C24), suggesting that AlkX can sense the concentrations of n-alkane degradation metabolites. A model was therefore proposed where AlkX controls alkW1 expression in a metabolite-dependent manner. Bioinformatic analysis revealed that the alkane hydroxylase gene regulation mechanism may be common among Actinobacteria.

  6. Genome-Wide Nucleosome Occupancy and Positioning and Their Impact on Gene Expression and Evolution in Plants.

    PubMed

    Zhang, Tao; Zhang, Wenli; Jiang, Jiming

    2015-08-01

    The fundamental unit of chromatin is the nucleosome that consists of a protein octamer composed of the four core histones (Hs; H3, H4, H2A, and H2B) wrapped by 147 bp of DNA. Nucleosome occupancy and positioning have proven to be dynamic and have a critical impact on expression, regulation, and evolution of eukaryotic genes. We developed nucleosome occupancy and positioning data sets using leaf tissue of rice (Oryza sativa) and both leaf and flower tissues of Arabidopsis (Arabidopsis thaliana). We show that model plant and animal species share the fundamental characteristics associated with nucleosome dynamics. Only 12% and 16% of the Arabidopsis and rice genomes, respectively, were occupied by well-positioned nucleosomes. The cores of positioned nucleosomes were enriched with G/C dinucleotides and showed a lower C→T mutation rate than the linker sequences. We discovered that nucleosomes associated with heterochromatic regions were more spaced with longer linkers than those in euchromatic regions in both plant species. Surprisingly, different nucleosome densities were found to be associated with chromatin in leaf and flower tissues in Arabidopsis. We show that deep MNase-seq data sets can be used to map nucleosome occupancy of specific genomic loci and reveal gene expression patterns correlated with chromatin dynamics in plant genomes.

  7. The RNA-binding protein CsrA plays a central role in positively regulating virulence factors in Erwinia amylovora.

    PubMed

    Ancona, Veronica; Lee, Jae Hoon; Zhao, Youfu

    2016-11-15

    The GacS/GacA two-component system (also called GrrS/GrrA) is a global regulatory system which is highly conserved among gamma-proteobacteria. This system positively regulates non-coding small regulatory RNA csrB, which in turn binds to the RNA-binding protein CsrA. However, how GacS/GacA-Csr system regulates virulence traits in E. amylovora remains unknown. Results from mutant characterization showed that the csrB mutant was hypermotile, produced higher amount of exopolysaccharide amylovoran, and had increased expression of type III secretion (T3SS) genes in vitro. In contrast, the csrA mutant exhibited complete opposite phenotypes, including non-motile, reduced amylovoran production and expression of T3SS genes. Furthermore, the csrA mutant did not induce hypersensitive response on tobacco or cause disease on immature pear fruits, indicating that CsrA is a positive regulator of virulence factors. These findings demonstrated that CsrA plays a critical role in E. amylovora virulence and suggested that negative regulation of virulence by GacS/GacA acts through csrB sRNA, which binds to CsrA and neutralizes its positive effect on T3SS gene expression, flagellar formation and amylovoran production. Future research will be focused on determining the molecular mechanism underlying the positive regulation of virulence traits by CsrA.

  8. The protective role of attachment security for adolescent borderline personality disorder features via enhanced positive emotion regulation strategies.

    PubMed

    Kim, Sohye; Sharp, Carla; Carbone, Crystal

    2014-04-01

    While studies have documented significant associations between insecure attachment, emotion dysregulation, and borderline personality disorder (BPD) features, no research to date has empirically delineated the specific mechanisms by which these constructs are related. The present study brings together 2 lines of research that have hitherto separately examined attachment disturbance and emotion dysregulation as they respectively manifest in the pathogenesis of BPD, and explores the complex relations between the 2 well-established correlates of borderline traits in a clinical sample of adolescents (N = 228). We examined the adolescents' use of positive and negative emotion regulation strategies, along with their maternal and paternal attachment security. Results indicated that positive and negative emotion regulation strategies were differentially implicated in the link between attachment insecurity and BPD features. Attachment security functioned as a buffer against adolescent BPD by enhancing positive emotion regulation strategies, while negative emotion regulation strategies served to dilute the protective effect of attachment and positive regulation strategies, culminating in clinically significant levels of borderline traits. Findings are discussed with regard to interventions in the developmental trajectory of BPD as it unfolds during adolescence.

  9. The RNA-binding protein CsrA plays a central role in positively regulating virulence factors in Erwinia amylovora

    PubMed Central

    Ancona, Veronica; Lee, Jae Hoon; Zhao, Youfu

    2016-01-01

    The GacS/GacA two-component system (also called GrrS/GrrA) is a global regulatory system which is highly conserved among gamma-proteobacteria. This system positively regulates non-coding small regulatory RNA csrB, which in turn binds to the RNA-binding protein CsrA. However, how GacS/GacA-Csr system regulates virulence traits in E. amylovora remains unknown. Results from mutant characterization showed that the csrB mutant was hypermotile, produced higher amount of exopolysaccharide amylovoran, and had increased expression of type III secretion (T3SS) genes in vitro. In contrast, the csrA mutant exhibited complete opposite phenotypes, including non-motile, reduced amylovoran production and expression of T3SS genes. Furthermore, the csrA mutant did not induce hypersensitive response on tobacco or cause disease on immature pear fruits, indicating that CsrA is a positive regulator of virulence factors. These findings demonstrated that CsrA plays a critical role in E. amylovora virulence and suggested that negative regulation of virulence by GacS/GacA acts through csrB sRNA, which binds to CsrA and neutralizes its positive effect on T3SS gene expression, flagellar formation and amylovoran production. Future research will be focused on determining the molecular mechanism underlying the positive regulation of virulence traits by CsrA. PMID:27845410

  10. Genes Regulated by Vitamin D in Bone Cells Are Positively Selected in East Asians

    PubMed Central

    Chen, Yuan; Xue, Yali; Luiselli, Donata; Tyler-Smith, Chris; Pagani, Luca; Ayub, Qasim

    2015-01-01

    Vitamin D and folate are activated and degraded by sunlight, respectively, and the physiological processes they control are likely to have been targets of selection as humans expanded from Africa into Eurasia. We investigated signals of positive selection in gene sets involved in the metabolism, regulation and action of these two vitamins in worldwide populations sequenced by Phase I of the 1000 Genomes Project. Comparing allele frequency-spectrum-based summary statistics between these gene sets and matched control genes, we observed a selection signal specific to East Asians for a gene set associated with vitamin D action in bones. The selection signal was mainly driven by three genes CXXC finger protein 1 (CXXC1), low density lipoprotein receptor-related protein 5 (LRP5) and runt-related transcription factor 2 (RUNX2). Examination of population differentiation and haplotypes allowed us to identify several candidate causal regulatory variants in each gene. Four of these candidate variants (one each in CXXC1 and RUNX2 and two in LRP5) had a >70% derived allele frequency in East Asians, but were present at lower (20–60%) frequency in Europeans as well, suggesting that the adaptation might have been part of a common response to climatic and dietary changes as humans expanded out of Africa, with implications for their role in vitamin D-dependent bone mineralization and osteoporosis insurgence. We also observed haplotype sharing between East Asians, Finns and an extinct archaic human (Denisovan) sample at the CXXC1 locus, which is best explained by incomplete lineage sorting. PMID:26719974

  11. Positive regulation of the Egr-1/osteopontin positive feedback loop in rat vascular smooth muscle cells by TGF-{beta}, ERK, JNK, and p38 MAPK signaling

    SciTech Connect

    Yu, Hong-Wei; Liu, Qi-Feng; Liu, Gui-Nan

    2010-05-28

    Previous studies identified a positive feedback loop in rat vascular smooth muscle cells (VSMCs) in which early growth response factor-1 (Egr-1) binds to the osteopontin (OPN) promoter and upregulates OPN expression, and OPN upregulates Egr-1 expression via the extracellular signal-regulated protein kinase (ERK) signaling pathway. The current study examined whether transforming growth factor-{beta} (TGF-{beta}) activity contributes to Egr-1 binding to the OPN promoter, and whether other signaling pathways act downstream of OPN to regulate Egr-1 expression. ChIP assays using an anti-Egr-1 antibody showed that amplification of the OPN promoter sequence decreased in TGF-{beta} DNA enzyme-transfected VSMCs relative to control VSMCs. Treatment of VSMCs with PD98059 (ERK inhibitor), SP600125 (JNK inhibitor), or SB203580 (p38 MAPK inhibitor) significantly inhibited OPN-induced Egr-1 expression, and PD98059 treatment was associated with the most significant decrease in Egr-1 expression. OPN-stimulated VSMC cell migration was inhibited by SP600125 or SB203580, but not by PD98059. Furthermore, MTT assays showed that OPN-mediated cell proliferation was inhibited by PD98059, but not by SP600125 or SB203580. Taken together, the results of the current study show that Egr-1 binding to the OPN promoter is positively regulated by TGF-{beta}, and that the p38 MAPK, JNK, and ERK pathways are involved in OPN-mediated Egr-1 upregulation.

  12. DepR1, a TetR Family Transcriptional Regulator, Positively Regulates Daptomycin Production in an Industrial Producer, Streptomyces roseosporus SW0702.

    PubMed

    Yuan, Peng-Hui; Zhou, Ri-Cheng; Chen, Xuepeng; Luo, Shuai; Wang, Feng; Mao, Xu-Ming; Li, Yong-Quan

    2016-01-15

    Daptomycin is a potent cyclic lipopeptide antibiotic. It is widely used against various Gram-positive bacterial pathogens. Historically, a poor understanding of the transcriptional regulation of daptomycin biosynthesis has limited the options for targeted genetic engineering toward titer improvement. Here, we isolated a TetR family transcriptional regulator, DepR1, from the industrial producer Streptomyces roseosporus SW0702 using a biotinylated dptE promoter (dptEp) as a probe. The direct interaction between DepR1 and dptEp then was confirmed by electrophoretic mobility shift assays and DNase I footprinting assays. The deletion of depR1 led to a reduction in dptEp activity and the cessation of daptomycin production. The ΔdepR1 mutant produced less red pigment and failed to sporulate on R5 medium. This suggests that DepR1 plays a positive role in the control of morphological differentiation. Moreover, DepR1 was positively autoregulated by directly binding to its own promoter. This might account for the positive feedback regulation of daptomycin production. Based on these positive effects, genetic engineering by overexpression of depR1 raised daptomycin production and shortened the fermentation period both in flask and in fermentor.

  13. Longitudinal Study of Self-Regulation, Positive Parenting, and Adjustment Problems among Physically Abused Children

    ERIC Educational Resources Information Center

    Kim-Spoon, Jungmeen.; Haskett, Mary E.; Longo, Gregory S.; Nice, Rachel.

    2012-01-01

    Objective: Research using normative and high-risk samples indicates a significant link between problems with self-regulation and child maladjustment. Nevertheless, little is known about the processes that may modify the link between self-regulation and maladjustment. This longitudinal study examined the joint contributions of child self-regulation…

  14. Maternal Positive and Negative Interaction Behaviors and Early Adolescents' Depressive Symptoms: Adolescent Emotion Regulation as a Mediator

    ERIC Educational Resources Information Center

    Yap, Marie B. H.; Schwartz, Orli S.; Byrne, Michelle L.; Simmons, Julian G.; Allen, Nicholas B.

    2010-01-01

    This study examined the relation between mothers' positive and negative interaction behaviors during mother-child interactions and the emotion regulation (ER) and depressive symptoms of their adolescent offspring. Event-planning (EPI) and problem-solving interactions (PSI) were observed in 163 mother-adolescent dyads, and adolescents also provided…

  15. The protein phosphatase 2A functions in the spindle position checkpoint by regulating the checkpoint kinase Kin4

    PubMed Central

    Chan, Leon Y.; Amon, Angelika

    2009-01-01

    In budding yeast, a surveillance mechanism known as the spindle position checkpoint (SPOC) ensures accurate genome partitioning. In the event of spindle misposition, the checkpoint delays exit from mitosis by restraining the activity of the mitotic exit network (MEN). To date, the only component of the checkpoint to be identified is the protein kinase Kin4. Furthermore, how the kinase is regulated by spindle position is not known. Here, we identify the protein phosphatase 2A (PP2A) in complex with the regulatory subunit Rts1 as a component of the SPOC. Loss of PP2A-Rts1 function abrogates the SPOC but not other mitotic checkpoints. We further show that the protein phosphatase functions upstream of Kin4, regulating the kinase's phosphorylation and localization during an unperturbed cell cycle and during SPOC activation, thus defining the phosphatase as a key regulator of SPOC function. PMID:19605686

  16. The protein phosphatase 2A functions in the spindle position checkpoint by regulating the checkpoint kinase Kin4.

    PubMed

    Chan, Leon Y; Amon, Angelika

    2009-07-15

    In budding yeast, a surveillance mechanism known as the spindle position checkpoint (SPOC) ensures accurate genome partitioning. In the event of spindle misposition, the checkpoint delays exit from mitosis by restraining the activity of the mitotic exit network (MEN). To date, the only component of the checkpoint to be identified is the protein kinase Kin4. Furthermore, how the kinase is regulated by spindle position is not known. Here, we identify the protein phosphatase 2A (PP2A) in complex with the regulatory subunit Rts1 as a component of the SPOC. Loss of PP2A-Rts1 function abrogates the SPOC but not other mitotic checkpoints. We further show that the protein phosphatase functions upstream of Kin4, regulating the kinase's phosphorylation and localization during an unperturbed cell cycle and during SPOC activation, thus defining the phosphatase as a key regulator of SPOC function.

  17. Mothers' attachment styles and their children's self-reported security, as related to maternal socialization of children's positive affect regulation.

    PubMed

    Gentzler, Amy L; Ramsey, Meagan A; Black, Katelyn R

    2015-01-01

    This study investigated how mothers' attachment was related to their responses to their own and their children's positive events and positive affect (PA). Ninety-seven mothers reported on their attachment and their responses to their own and their 7-12-year-old children's positive events and emotions. Children reported on their mothers' responses to the children's positive events and their attachment security with their mothers. The results indicated that more avoidant mothers reported less intense PA in response to their own and their children's positive events. More avoidant mothers also were less likely to encourage their children to savor positive events (through expressing PA, reflecting on PA or themselves, giving rewards, and affectionate responses). Mothers higher on anxiety reported greater likelihood of dampening (e.g., minimizing the event's importance) their own positive events and reported being more likely to feel discomfort and to reprimand their children for expressing PA. Children's security was predicted by mothers' lower likelihood of encouraging children's dampening and of reprimanding children for PA displays. This study advances the literature on how mothers' attachment is related to the ways in which they regulate their own and their children's PA, which may have implications for children's attachment and developing PA regulation.

  18. Evaluation of STMicroelectronics RH-L49 1 3 positive hardened low drop voltage regulator

    NASA Astrophysics Data System (ADS)

    Briand, P.; Vadrot, J.-F.; Belasic, M.

    2002-12-01

    An evaluation of the RH-L4913 from the manufacturer STMicroelectronics, total dose radiation hardened, has been done. This evaluation has consisted of an electrical characterization, a physical analysis, endurance tests (silicon & package) and radiations tests, in order to assess the specific failure modes and the reliability of this regulator in space environment. The results are presented in this paper and demonstrate the RH-L4913 regulator ability to be used in space applications.

  19. 75 FR 4143 - Federal Speculative Position Limits for Referenced Energy Contracts and Associated Regulations

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-01-26

    ...The Commodity Futures Trading Commission (``CFTC'' or ``Commission'') is proposing to implement speculative position limits for futures and option contracts in certain energy commodities. The Commodity Exchange Act of 1936 (``CEA'' or ``Act'') gives the Commission the authority to establish limits on positions to diminish, eliminate or prevent excessive speculation causing sudden or......

  20. Effects of Regulating Positive Emotions through Reappraisal and Suppression on Verbal and Non-Verbal Recognition Memory

    PubMed Central

    Ortner, Catherine N. M.; de Koning, Monica

    2013-01-01

    Previous research has suggested that regulating emotions through reappraisal does not incur cognitive costs. However, in those experiments, cognitive costs were often assessed by recognition memory for information that was contextually related to the emotionally evocative stimuli and may have been incorporated into the reappraisal script, facilitating memory. Furthermore, there is little research on the cognitive correlates of regulating positive emotions. In the current experiment, we tested memory for information that was contextually unrelated to the emotional stimuli and could not easily be related to the reappraisal. Participants viewed neutral and mildly positive slides and either reappraised, suppressed their emotions, or viewed the images with no emotion regulation instruction. At the same time, they heard abstract words that were unrelated to the picture stimuli. Subsequent verbal recognition memory was lower after reappraising than viewing, whereas non-verbal recognition memory (of the slides) was higher after reappraising, but only for positive pictures and when participants viewed the positive pictures first. Suppression had no significant effect on either verbal or non-verbal recognition scores, although there was a trend towards poorer recognition of verbal information. The findings support the notion that reappraisal is effortful and draws on limited cognitive resources, causing decrements in performance in a concurrent memory task. PMID:23658647

  1. Regulation of emotion in ADHD: can children with ADHD override the natural tendency to approach positive and avoid negative pictures?

    PubMed

    Van Cauwenberge, Valerie; Sonuga-Barke, Edmund J S; Hoppenbrouwers, Karel; Van Leeuwen, Karla; Wiersema, Jan R

    2017-03-01

    Studies have demonstrated inefficient use of antecedent-focused emotion regulation strategies in children with ADHD attention-deficit/hyperactivity disorder (ADHD). In the current study we tested for the first time if ADHD is also associated with difficulties in response-focused strategies by measuring the ability to override action tendencies induced by emotional information. Performance data on a computer-based approach-avoidance paradigm of 28 children with ADHD and 38 typically developing children between 8 and 15 years of age were analyzed, by comparing a congruent condition in which they were instructed to approach positive and avoid negative pictures and an incongruent condition where they had to override these automatic reactions and approach negative and avoid positive pictures. Children also rated the valence and salience of the pictures. Children with ADHD and typically developing children rated the emotional valence of the pictures appropriately and similarly, while positive pictures were rated as more arousing by children with ADHD. Solid congruency effects were found indicating that the task measured response-focused emotion regulation; however groups did not differ in this respect. Our findings do not support a deficit in emotion regulation in ADHD in terms of the ability to override natural tendencies to approach positive and avoid negative pictures.

  2. Effects of regulating positive emotions through reappraisal and suppression on verbal and non-verbal recognition memory.

    PubMed

    Ortner, Catherine N M; de Koning, Monica

    2013-01-01

    Previous research has suggested that regulating emotions through reappraisal does not incur cognitive costs. However, in those experiments, cognitive costs were often assessed by recognition memory for information that was contextually related to the emotionally evocative stimuli and may have been incorporated into the reappraisal script, facilitating memory. Furthermore, there is little research on the cognitive correlates of regulating positive emotions. In the current experiment, we tested memory for information that was contextually unrelated to the emotional stimuli and could not easily be related to the reappraisal. Participants viewed neutral and mildly positive slides and either reappraised, suppressed their emotions, or viewed the images with no emotion regulation instruction. At the same time, they heard abstract words that were unrelated to the picture stimuli. Subsequent verbal recognition memory was lower after reappraising than viewing, whereas non-verbal recognition memory (of the slides) was higher after reappraising, but only for positive pictures and when participants viewed the positive pictures first. Suppression had no significant effect on either verbal or non-verbal recognition scores, although there was a trend towards poorer recognition of verbal information. The findings support the notion that reappraisal is effortful and draws on limited cognitive resources, causing decrements in performance in a concurrent memory task.

  3. Biological functions of miR-29b contribute to positive regulation of osteoblast differentiation.

    PubMed

    Li, Zhaoyong; Hassan, Mohammad Q; Jafferji, Mohammed; Aqeilan, Rami I; Garzon, Ramiro; Croce, Carlo M; van Wijnen, Andre J; Stein, Janet L; Stein, Gary S; Lian, Jane B

    2009-06-05

    Bone tissue arises from mesenchymal cells induced into the osteoblast lineage by essential transcription factors and signaling cascades. MicroRNAs regulate biological processes by binding to mRNA 3'-untranslated region (UTR) sequences to attenuate protein synthesis. Here we performed microRNA profiling and identified miRs that are up-regulated through stages of osteoblast differentiation. Among these are the miR-29, miR-let-7, and miR-26 families that target many collagens and extracellular matrix proteins. We find that miR-29b supports osteoblast differentiation through several mechanisms. miR-29b decreased and anti-miR-29b increased activity of COL1A1, COL5A3, and COL4A2 3'-UTR sequences in reporter assays, as well as endogenous gene expression. These results support a mechanism for regulating collagen protein accumulation during the mineralization stage when miR-29b reaches peak levels. We propose that this mechanism prevents fibrosis and facilitates mineral deposition. Our studies further demonstrate that miR-29b promotes osteogenesis by directly down-regulating known inhibitors of osteoblast differentiation, HDAC4, TGFbeta3, ACVR2A, CTNNBIP1, and DUSP2 proteins through binding to target 3'-UTR sequences in their mRNAs. Thus, miR-29b is a key regulator of development of the osteoblast phenotype by targeting anti-osteogenic factors and modulating bone extracellular matrix proteins.

  4. MYB98 positively regulates a battery of synergid-expressed genes encoding filiform apparatus localized proteins.

    PubMed

    Punwani, Jayson A; Rabiger, David S; Drews, Gary N

    2007-08-01

    The synergid cells within the female gametophyte are essential for reproduction in angiosperms. MYB98 encodes an R2R3-MYB protein required for pollen tube guidance and filiform apparatus formation by the synergid cells. To test the predicted function of MYB98 as a transcriptional regulator, we determined its subcellular localization and examined its DNA binding properties. We show that MYB98 binds to a specific DNA sequence (TAAC) and that a MYB98-green fluorescent protein fusion protein localizes to the nucleus, consistent with a role in transcriptional regulation. To identify genes regulated by MYB98, we tested previously identified synergid-expressed genes for reduced expression in myb98 female gametophytes and identified 16 such genes. We dissected the promoter of one of the downstream genes, DD11, and show that it contains a MYB98 binding site required for synergid expression, suggesting that DD11 is regulated directly by MYB98. To gain insight into the functions of the downstream genes, we chose five genes and determined the subcellular localization of the encoded proteins. We show that these five proteins are secreted into the filiform apparatus, suggesting that they play a role in either the formation or the function of this unique structure. Together, these data suggest that MYB98 functions as a transcriptional regulator in the synergid cells and activates the expression of genes required for pollen tube guidance and filiform apparatus formation.

  5. The development of intentional self-regulation in adolescence: describing, explaining, and optimizing its link to positive youth development.

    PubMed

    Napolitano, Christopher M; Bowers, Edmond P; Gestsdóttir, Steinunn; Chase, Paul A

    2011-01-01

    Intentional self-regulation (ISR) skills are key assets promoting healthy and positive development across the life span. In this chapter, we describe the development of ISR in adolescence, offer explanations for the development of these skills and their relation to positive youth development among diverse youth in diverse contexts, and provide suggestions for future research and programs seeking to optimize youth outcomes through the promotion of ISR skills. Primarily drawing from data from the 4-H Study of Positive Youth Development, we discuss research using the Selection, Optimization, and Compensation model of Baltes, Freund and colleagues, measures of which have been linked to a variety of positive developmental outcomes in adolescence. In addition to providing a review of the literature and relevant recent research, an applied program designed to promote ISR--termed Project GPS--is also discussed.

  6. Regulation of nucleosome positioning by a CHD Type III chromatin remodeler and its relationship to developmental gene expression in Dictyostelium

    PubMed Central

    Platt, James L.; Kent, Nicholas A.; Kimmel, Alan R.

    2017-01-01

    Nucleosome placement and repositioning can direct transcription of individual genes; however, the precise interactions of these events are complex and largely unresolved at the whole-genome level. The Chromodomain-Helicase-DNA binding (CHD) Type III proteins are a subfamily of SWI2/SNF2 proteins that control nucleosome positioning and are associated with several complex human disorders, including CHARGE syndrome and autism. Type III CHDs are required for multicellular development of animals and Dictyostelium but are absent in plants and yeast. These CHDs can mediate nucleosome translocation in vitro, but their in vivo mechanism is unknown. Here, we use genome-wide analysis of nucleosome positioning and transcription profiling to investigate the in vivo relationship between nucleosome positioning and gene expression during development of wild-type (WT) Dictyostelium and mutant cells lacking ChdC, a Type III CHD protein ortholog. We demonstrate major nucleosome positional changes associated with developmental gene regulation in WT. Loss of chdC caused an increase of intragenic nucleosome spacing and misregulation of gene expression, affecting ∼50% of the genes that are repositioned during WT development. These analyses demonstrate active nucleosome repositioning during Dictyostelium multicellular development, establish an in vivo function of CHD Type III chromatin remodeling proteins in this process, and reveal the detailed relationship between nucleosome positioning and gene regulation, as cells transition between developmental states. PMID:28330902

  7. 78 FR 70191 - Post-Employment Conflict of Interest Regulations; Exempted Senior Employee Positions; Withdrawal...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-25

    ... / Monday, November 25, 2013 / Rules and Regulations#0;#0; ] OFFICE OF GOVERNMENT ETHICS 5 CFR Part 2641 RIN...; Withdrawal of Final Rule AGENCY: Office of Government Ethics (OGE). ACTION: Withdrawal of final rule. SUMMARY: The Office of Government Ethics is withdrawing the final rule ``Post-Employment Conflict of...

  8. Glucocorticoid receptor positively regulates transcription of FNDC5 in the liver

    PubMed Central

    Kim, Hyoung Kyu; Jeong, Yu Jeong; Song, In-Sung; Noh, Yeon Hee; Seo, Kyo Won; Kim, Min; Han, Jin

    2017-01-01

    Irisin is secreted by skeletal muscle during exercise and influences energy and metabolic homeostasis. This hormone is a cleaved and secreted fragment of fibronectin type III domain-containing 5 (FNDC5). Elucidation of the FNDC5 gene regulation mechanism is necessary to clarify the function of irisin as a potential therapeutic target in human metabolic diseases. Thus, we investigated the genetic and epigenetic mechanisms that regulate expression of the FNDC5 gene. FNDC5 mRNA was strong expressed in major energy-dependent human tissues, including heart, brain, liver, and skeletal muscle. Promoter analysis of the FNDC5 gene revealed that the core promoter region of the FNDC5 gene contained one CpG island that was located just upstream of the transcriptional start site for variants 2 and 3. Treatment with the histone deacetylase inhibitor sodium butyrate and the demethylating agent 5-azacytidine increased mRNA expression of FNDC5 in Huh7 cells. Prediction of transcription factor binding sites suggested that the glucocorticoid receptor was involved in the regulation of FNDC5 expression, and indeed, cortisol treatment increased mRNA expression of FNDC5 in Huh7 cells. Collectively, these findings offer insight into the genetic and epigenetic regulation of FNDC5, providing the initial steps required for understanding the role of irisin in the metabolic homeostasis. PMID:28240298

  9. Hope, Self-Esteem, and Self-Regulation: Positive Characteristics among Men and Women in Recovery

    ERIC Educational Resources Information Center

    Ferrari, Joseph R.; Stevens, Edward B.; Legler, Raymond; Jason, Leonard A.

    2012-01-01

    Hopefulness remains unclear in relation to aspects of self-control and self-esteem among adults in substance abuse recovery. The present study explored the relationship between dispositional hope (agency and pathway) with self-esteem (self-liking, self-competency, and self-confidence) and self-regulation (impulse control and self-discipline),…

  10. Salmonella enterica serovar Typhimurium BaeSR two-component system positively regulates sodA in response to ciprofloxacin.

    PubMed

    Guerrero, P; Collao, B; Álvarez, R; Salinas, H; Morales, E H; Calderón, I L; Saavedra, C P; Gil, F

    2013-10-01

    In response to antibiotics, bacteria activate regulatory systems that control the expression of genes that participate in detoxifying these compounds, like multidrug efflux systems. We previously demonstrated that the BaeSR two-component system from Salmonella enterica serovar Typhimurium (S. Typhimurium) participates in the detection of ciprofloxacin, a bactericidal antibiotic, and in the positive regulation of mdtA, an efflux pump implicated in antibiotic resistance. In the present work, we provide further evidence for a role of the S. Typhimurium BaeSR two-component system in response to ciprofloxacin treatment and show that it regulates sodA expression. We demonstrate that, in the absence of BaeSR, the transcript levels of sodA and the activity of its gene product are lower. Using electrophoretic mobility shift assays and transcriptional fusions, we demonstrate that BaeR regulates sodA by a direct interaction with the promoter region.

  11. A histone methylation network regulates transgenerational epigenetic memory in C. elegans.

    PubMed

    Greer, Eric L; Beese-Sims, Sara E; Brookes, Emily; Spadafora, Ruggero; Zhu, Yun; Rothbart, Scott B; Aristizábal-Corrales, David; Chen, Shuzhen; Badeaux, Aimee I; Jin, Qiuye; Wang, Wei; Strahl, Brian D; Colaiácovo, Monica P; Shi, Yang

    2014-04-10

    How epigenetic information is transmitted from generation to generation remains largely unknown. Deletion of the C. elegans histone H3 lysine 4 dimethyl (H3K4me2) demethylase spr-5 leads to inherited accumulation of the euchromatic H3K4me2 mark and progressive decline in fertility. Here, we identified multiple chromatin-modifying factors, including H3K4me1/me2 and H3K9me3 methyltransferases, an H3K9me3 demethylase, and an H3K9me reader, which either suppress or accelerate the progressive transgenerational phenotypes of spr-5 mutant worms. Our findings uncover a network of chromatin regulators that control the transgenerational flow of epigenetic information and suggest that the balance between euchromatic H3K4 and heterochromatic H3K9 methylation regulates transgenerational effects on fertility.

  12. A histone methylation network regulates transgenerational epigenetic memory in C. elegans

    PubMed Central

    Greer, Eric L.; Beese-Sims, Sara E.; Brookes, Emily; Spadafora, Ruggero; Zhu, Yun; Rothbart, Scott B.; Aristizábal-Corrales, David; Chen, Shuzhen; Badeaux, Aimee I.; Jin, Qiuye; Wang, Wei; Strahl, Brian D.; Colaiácovo, Monica P.; Shi, Yang

    2014-01-01

    Summary How epigenetic information is transmitted from generation to generation remains largely unknown. Deletion of the C. elegans Histone H3 lysine 4 dimethyl (H3K4me2) demethylase spr-5 leads to inherited accumulation of the euchromatic H3K4me2 mark and progressive decline in fertility. Here we identified multiple chromatin-modifying factors, including novel H3K4me1/me2 and H3K9me3 methyltransferases, an H3K9me3 demethylase and an H3K9me reader, which either suppress or accelerate the progressive transgenerational phenotypes of spr-5 mutant worms. Our findings uncover a network of chromatin regulators that control the trans-generational flow of epigenetic information, and suggest that the balance between euchromatic H3K4 and heterochromatic H3K9 methylation regulates trans-generational effects on fertility. PMID:24685137

  13. Spatial landmarks regulate a Cdc42-dependent MAPK pathway to control differentiation and the response to positional compromise

    PubMed Central

    Basu, Sukanya; Vadaie, Nadia; Prabhakar, Aditi; Li, Boyang; Adhikari, Hema; Pitoniak, Andrew; Chow, Jacky; Chavel, Colin A.; Cullen, Paul J.

    2016-01-01

    A fundamental problem in cell biology is to understand how spatial information is recognized and integrated into morphogenetic responses. Budding yeast undergoes differentiation to filamentous growth, which involves changes in cell polarity through mechanisms that remain obscure. Here we define a regulatory input where spatial landmarks (bud-site–selection proteins) regulate the MAPK pathway that controls filamentous growth (fMAPK pathway). The bud-site GTPase Rsr1p regulated the fMAPK pathway through Cdc24p, the guanine nucleotide exchange factor for the polarity establishment GTPase Cdc42p. Positional landmarks that direct Rsr1p to bud sites conditionally regulated the fMAPK pathway, corresponding to their roles in regulating bud-site selection. Therefore, cell differentiation is achieved in part by the reorganization of polarity at bud sites. In line with this conclusion, dynamic changes in budding pattern during filamentous growth induced corresponding changes in fMAPK activity. Intrinsic compromise of bud-site selection also impacted fMAPK activity. Therefore, a surveillance mechanism monitors spatial position in response to extrinsic and intrinsic stress and modulates the response through a differentiation MAPK pathway. PMID:27001830

  14. AMPK: positive and negative regulation, and its role in whole-body energy homeostasis.

    PubMed

    Hardie, D Grahame

    2015-04-01

    The AMP-activated protein kinase (AMPK) is a sensor of energy status that, when activated by metabolic stress, maintains cellular energy homeostasis by switching on catabolic pathways and switching off ATP-consuming processes. Recent results suggest that activation of AMPK by the upstream kinase LKB1 in response to nutrient lack occurs at the surface of the lysosome. AMPK is also crucial in regulation of whole body energy balance, particularly by mediating effects of hormones acting on the hypothalamus. Recent crystal structures of complete AMPK heterotrimers have illuminated its complex mechanisms of activation, involving both allosteric activation and increased net phosphorylation mediated by effects on phosphorylation and dephosphorylation. Finally, AMPK is negatively regulated by phosphorylation of the 'ST loop' within the catalytic subunit.

  15. The tyrosine phosphatase CD148 is an essential positive regulator of platelet activation and thrombosis.

    PubMed

    Senis, Yotis A; Tomlinson, Michael G; Ellison, Stuart; Mazharian, Alexandra; Lim, Jenson; Zhao, Yan; Kornerup, Kristin N; Auger, Jocelyn M; Thomas, Steve G; Dhanjal, Tarvinder; Kalia, Neena; Zhu, Jing W; Weiss, Arthur; Watson, Steve P

    2009-05-14

    Platelets play a fundamental role in hemostasis and thrombosis. They are also involved in pathologic conditions resulting from blocked blood vessels, including myocardial infarction and ischemic stroke. Platelet adhesion, activation, and aggregation at sites of vascular injury are regulated by a diverse repertoire of tyrosine kinase-linked and G protein-coupled receptors. Src family kinases (SFKs) play a central role in initiating and propagating signaling from several platelet surface receptors; however, the underlying mechanism of how SFK activity is regulated in platelets remains unclear. CD148 is the only receptor-like protein tyrosine phosphatase identified in platelets to date. In the present study, we show that mutant mice lacking CD148 exhibited a bleeding tendency and defective arterial thrombosis. Basal SFK activity was found to be markedly reduced in CD148-deficient platelets, resulting in a global hyporesponsiveness to agonists that signal through SFKs, including collagen and fibrinogen. G protein-coupled receptor responses to thrombin and other agonists were also marginally reduced. These results highlight CD148 as a global regulator of platelet activation and a novel antithrombotic drug target.

  16. The tyrosine phosphatase CD148 is an essential positive regulator of platelet activation and thrombosis

    PubMed Central

    Tomlinson, Michael G.; Ellison, Stuart; Mazharian, Alexandra; Lim, Jenson; Zhao, Yan; Kornerup, Kristin N.; Auger, Jocelyn M.; Thomas, Steve G.; Dhanjal, Tarvinder; Kalia, Neena; Zhu, Jing W.; Weiss, Arthur; Watson, Steve P.

    2009-01-01

    Platelets play a fundamental role in hemostasis and thrombosis. They are also involved in pathologic conditions resulting from blocked blood vessels, including myocardial infarction and ischemic stroke. Platelet adhesion, activation, and aggregation at sites of vascular injury are regulated by a diverse repertoire of tyrosine kinase–linked and G protein–coupled receptors. Src family kinases (SFKs) play a central role in initiating and propagating signaling from several platelet surface receptors; however, the underlying mechanism of how SFK activity is regulated in platelets remains unclear. CD148 is the only receptor-like protein tyrosine phosphatase identified in platelets to date. In the present study, we show that mutant mice lacking CD148 exhibited a bleeding tendency and defective arterial thrombosis. Basal SFK activity was found to be markedly reduced in CD148-deficient platelets, resulting in a global hyporesponsiveness to agonists that signal through SFKs, including collagen and fibrinogen. G protein–coupled receptor responses to thrombin and other agonists were also marginally reduced. These results highlight CD148 as a global regulator of platelet activation and a novel antithrombotic drug target. PMID:19246339

  17. Tumor necrosis factor receptor-associated factor 3 is a positive regulator of pathological cardiac hypertrophy.

    PubMed

    Jiang, Xi; Deng, Ke-Qiong; Luo, Yuxuan; Jiang, Ding-Sheng; Gao, Lu; Zhang, Xiao-Fei; Zhang, Peng; Zhao, Guang-Nian; Zhu, Xueyong; Li, Hongliang

    2015-08-01

    Cardiac hypertrophy, a common early symptom of heart failure, is regulated by numerous signaling pathways. Here, we identified tumor necrosis factor receptor-associated factor 3 (TRAF3), an adaptor protein in tumor necrosis factor-related signaling cascades, as a key regulator of cardiac hypertrophy in response to pressure overload. TRAF3 expression was upregulated in hypertrophied mice hearts and failing human hearts. Four weeks after aortic banding, cardiac-specific conditional TRAF3-knockout mice exhibited significantly reduced cardiac hypertrophy, fibrosis, and dysfunction. Conversely, transgenic mice overexpressing TRAF3 in the heart developed exaggerated cardiac hypertrophy in response to pressure overload. TRAF3 also promoted an angiotensin II- or phenylephrine-induced hypertrophic response in isolated cardiomyocytes. Mechanistically, TRAF3 directly bound to TANK-binding kinase 1 (TBK1), causing increased TBK1 phosphorylation in response to hypertrophic stimuli. This interaction between TRAF3 and TBK1 further activated AKT signaling, which ultimately promoted the development of cardiac hypertrophy. Our findings not only reveal a key role of TRAF3 in regulating the hypertrophic response but also uncover TRAF3-TBK1-AKT as a novel signaling pathway in the development of cardiac hypertrophy and heart failure. This pathway may represent a potential therapeutic target for this pathological process.

  18. Fibrocytes Regulate Wilms’ Tumor 1-Positive Cell Accumulation in Severe Fibrotic Lung Disease

    PubMed Central

    Sontake, Vishwaraj; Shanmukhappa, Shiva K.; DiPasquale, Betsy A.; Reddy, Geereddy B.; Medvedovic, Mario; Hardie, William D.; White, Eric S.; Madala, Satish K.

    2015-01-01

    Collagen-producing myofibroblast transdifferentiation is considered a crucial determinant in the formation of scar tissue in the lungs of patients with idiopathic pulmonary fibrosis (IPF). Multiple resident pulmonary cell types and bone marrow-derived fibrocytes have been implicated as contributors to fibrotic lesions due to the transdifferentiation potential of these cells into myofibroblasts. In this study, we assessed the expression of Wilms’ tumor 1 (WT1), a known marker of mesothelial cells, in various cell types in normal and fibrotic lungs. We demonstrate that WT1 is expressed by both mesothelial and mesenchymal cells in IPF lungs, but has limited or no expression in normal human lungs. We also demonstrate that WT1-positive cells accumulate in fibrotic lung lesions, using two different mouse models of pulmonary fibrosis and WT1 promoter-driven fluorescent reporter mice. Reconstitution of bone-marrow cells into a transforming growth factor-α transgenic-mouse model demonstrated that fibrocytes do not transform into WT1-positive mesenchymal cells, but do augment accumulation of WT1-positive cells in severe fibrotic lung disease. Importantly, the number of WT1-positive cells in fibrotic lesions were correlated with severity of lung disease as assessed by changes in lung function, histology, and hydroxyproline levels in mice. Finally, inhibition of WT1 expression was sufficient to attenuate collagen and other extracellular-matrix gene production by mesenchymal cells from both murine and human fibrotic lungs. Thus, the results of this study demonstrate a novel association between fibrocyte-driven WT1-positive cell accumulation and severe fibrotic lung disease. PMID:26371248

  19. Activation and repression functions of an SR splicing regulator depend on exonic versus intronic-binding position.

    PubMed

    Shen, Manli; Mattox, William

    2012-01-01

    SR proteins and related factors play widespread roles in alternative pre-mRNA splicing and are known to promote splice site recognition through their Arg-Ser-rich effector domains. However, binding of SR regulators to some targets results in repression of splice sites through a distinct mechanism. Here, we investigate how activated and repressed targets of the Drosophila SR regulator Transformer2 elicit its differing effects on splicing. We find that, like activation, repression affects early steps in the recognition of splice sites and spliceosome assembly. Repositioning of regulatory elements reveals that Tra2 complexes that normally repress splicing from intronic positions activate splicing when located in an exon. Protein tethering experiments demonstrate that this position dependence is an intrinsic property of Tra2 and further show that repression and activation are mediated by separate effector domains of this protein. When other Drosophila SR factors (SF2 and Rbp1) that activate splicing from exonic positions were tethered intronically they failed to either activate or repress splicing. Interestingly, both activities of Tra2 favor the exonic identity of the RNA sequences that encompass its binding sites. This suggests a model in which these two opposite functions act in concert to define both the position and extent of alternatively spliced exons.

  20. TOR complex 2-Ypk1 signaling is an essential positive regulator of the general amino acid control response and autophagy.

    PubMed

    Vlahakis, Ariadne; Graef, Martin; Nunnari, Jodi; Powers, Ted

    2014-07-22

    The highly conserved Target of Rapamycin (TOR) kinase is a central regulator of cell growth and metabolism in response to nutrient availability. TOR functions in two structurally and functionally distinct complexes, TOR Complex 1 (TORC1) and TOR Complex 2 (TORC2). Through TORC1, TOR negatively regulates autophagy, a conserved process that functions in quality control and cellular homeostasis and, in this capacity, is part of an adaptive nutrient deprivation response. Here we demonstrate that during amino acid starvation TOR also operates independently as a positive regulator of autophagy through the conserved TORC2 and its downstream target protein kinase, Ypk1. Under these conditions, TORC2-Ypk1 signaling negatively regulates the Ca(2+)/calmodulin-dependent phosphatase, calcineurin, to enable the activation of the amino acid-sensing eIF2α kinase, Gcn2, and to promote autophagy. Our work reveals that the TORC2 pathway regulates autophagy in an opposing manner to TORC1 to provide a tunable response to cellular metabolic status.

  1. Identification of positive and negative regulators of disease resistance to rice blast fungus using constitutive gene expression patterns.

    PubMed

    Grand, Xavier; Espinoza, Rocio; Michel, Corinne; Cros, Sandrine; Chalvon, Véronique; Jacobs, John; Morel, Jean-Benoit

    2012-09-01

    Elevated constitutive expression of components of the defence arsenal is associated with quantitative resistance to the rice blast fungus, a phenomenon called preformed defence. While the role of many disease regulators in inducible defence systems has been extensively studied, little attention has been paid so far to genes that regulate preformed defence. In this study, we show by microarray analysis across rice diversity that the preformed defence phenomenon impacts on a large number of defence-related genes without apparently affecting other biological processes. Using a guilt-by-association strategy, we identified two positive regulators that promote constitutive expression of known defence markers and partial resistance to rice blast. The HSF23 gene encodes for a putative member of the heat shock transcription factor family, while CaMBP encodes for a putative Calmodulin-binding protein. Both HSF23 and CaMBP strongly affect preformed defence and also plant growth. Additionally, we identified the OB-fold gene as a negative regulator of blast resistance, which could be involved in RNA stabilization. The OB-fold mutants do not suffer from obvious developmental defects. Taken together, our results prove that our strategy of combining analysis of gene expression diversity with guilt-by-association is a powerful way to identify disease resistance regulators in rice.

  2. Rice open beak is a negative regulator of class 1 knox genes and a positive regulator of class B floral homeotic gene.

    PubMed

    Horigome, Ayako; Nagasawa, Nobuhiro; Ikeda, Kyoko; Ito, Momoyo; Itoh, Jun-Ichi; Nagato, Yasuo

    2009-06-01

    Numerous genes are involved in the regulation of plant development, including those that regulate floral homeotic genes, We identified two recessive allelic rice mutants, open beak-1 (opb-1) and opb-2, which exhibited pleiotropic defects in leaf morphogenesis, inflorescence architecture, and floral organ identity. Abnormal cell proliferation was observed in the leaves and spikelets, and ectopic or overexpression of several class 1 knox genes was detected; thus, the abnormal cell proliferation in opb mutants is probably caused by ectopic class 1 knox gene expression. The opb mutants also had defects in floral organ identity, resulting in the development of mosaic organs, including gluminous lodicules, staminoid lodicules, and pistiloid stamens. These results, together with the reduced expression of a class B gene, indicate that OPB positively regulates the expression of class B genes. Map-based cloning revealed that OPB encodes a transcription factor that is orthologous to the Arabidopsis JAGGED gene and is expressed in leaf primordia, inflorescence meristem, rachis branch meristems, floral meristem, and floral organ primordia. Taken together, our data suggest that the OPB gene affects cellular proliferation and floral organ identity through the regulation of class 1 knox genes and floral homeotic genes.

  3. Cyclin G Functions as a Positive Regulator of Growth and Metabolism in Drosophila

    PubMed Central

    Schulz, Adriana; Preiss, Anette; Nagel, Anja C.

    2015-01-01

    In multicellular organisms, growth and proliferation is adjusted to nutritional conditions by a complex signaling network. The Insulin receptor/target of rapamycin (InR/TOR) signaling cascade plays a pivotal role in nutrient dependent growth regulation in Drosophila and mammals alike. Here we identify Cyclin G (CycG) as a regulator of growth and metabolism in Drosophila. CycG mutants have a reduced body size and weight and show signs of starvation accompanied by a disturbed fat metabolism. InR/TOR signaling activity is impaired in cycG mutants, combined with a reduced phosphorylation status of the kinase Akt1 and the downstream factors S6-kinase and eukaryotic translation initiation factor 4E binding protein (4E-BP). Moreover, the expression and accumulation of Drosophila insulin like peptides (dILPs) is disturbed in cycG mutant brains. Using a reporter assay, we show that the activity of one of the first effectors of InR signaling, Phosphoinositide 3-kinase (PI3K92E), is unaffected in cycG mutants. However, the metabolic defects and weight loss in cycG mutants were rescued by overexpression of Akt1 specifically in the fat body and by mutants in widerborst (wdb), the B'-subunit of the phosphatase PP2A, known to downregulate Akt1 by dephosphorylation. Together, our data suggest that CycG acts at the level of Akt1 to regulate growth and metabolism via PP2A in Drosophila. PMID:26274446

  4. The Coordinated Positive Regulation of Topoisomerase Genes Maintains Topological Homeostasis in Streptomyces coelicolor

    PubMed Central

    Gongerowska, Martyna; Gutkowski, Paweł; Zakrzewska-Czerwińska, Jolanta; Jakimowicz, Dagmara

    2016-01-01

    ABSTRACT Maintaining an optimal level of chromosomal supercoiling is critical for the progression of DNA replication and transcription. Moreover, changes in global supercoiling affect the expression of a large number of genes and play a fundamental role in adapting to stress. Topoisomerase I (TopA) and gyrase are key players in the regulation of bacterial chromosomal topology through their respective abilities to relax and compact DNA. Soil bacteria such as Streptomyces species, which grow as branched, multigenomic hyphae, are subject to environmental stresses that are associated with changes in chromosomal topology. The topological fluctuations modulate the transcriptional activity of a large number of genes and in Streptomyces are related to the production of antibiotics. To better understand the regulation of topological homeostasis in Streptomyces coelicolor, we investigated the interplay between the activities of the topoisomerase-encoding genes topA and gyrBA. We show that the expression of both genes is supercoiling sensitive. Remarkably, increased chromosomal supercoiling induces the topA promoter but only slightly influences gyrBA transcription, while DNA relaxation affects the topA promoter only marginally but strongly activates the gyrBA operon. Moreover, we showed that exposure to elevated temperatures induces rapid relaxation, which results in changes in the levels of both topoisomerases. We therefore propose a unique mechanism of S. coelicolor chromosomal topology maintenance based on the supercoiling-dependent stimulation, rather than repression, of the transcription of both topoisomerase genes. These findings provide important insight into the maintenance of topological homeostasis in an industrially important antibiotic producer. IMPORTANCE We describe the unique regulation of genes encoding two topoisomerases, topoisomerase I (TopA) and gyrase, in a model Streptomyces species. Our studies demonstrate the coordination of topoisomerase gene

  5. LPXRFa, the piscine ortholog of GnIH, and LPXRF receptor positively regulate gonadotropin secretion in Tilapia (Oreochromis niloticus).

    PubMed

    Biran, Jakob; Golan, Matan; Mizrahi, Naama; Ogawa, Satoshi; Parhar, Ishwar S; Levavi-Sivan, Berta

    2014-11-01

    LPXRFamide (LPXRFa) peptides have been characterized for their ability to inhibit gonadotropin (GTH) release in birds and stimulate growth hormone (GH) release in frogs. However, their involvement in regulating the reproductive hypothalamo-pituitary-gonadal axis in mammals and fish is inconclusive. To study the role of LPXRFa peptides in the regulation of GTH secretion, we cloned tilapia LPXRFa and LPXRF receptor (LPXRF-R). Processing of the tilapia preproLPXRFa liberated three mature LPXRFa peptides that varied in size and post-translational modifications. Phylogenetic analysis of LPXRFa and the closely related RFamide peptide PQRFa showed clear clustering of each peptide sequence with its orthologs from various vertebrates. Signal-transduction analysis of the tilapia LPXRF-R in COS-7 cells showed clear stimulation of CRE-dependent luciferase activity, whereas the human NPFFR1 showed suppression of forskolin-induced CRE-dependent activity in this system. Administration of the tilapia pyroglutaminated LPXRFa-2 peptide to primary cell culture of tilapia pituitaries, or to reproductive female tilapia by ip injection, positively regulated both LH and FSH release in vivo and in vitro. Using double-labeled fluorescent in-situ hybridization and immunofluorescence, βLH cells were found to co-express both tilapia lpxrf and tilapia lpxrf-r mRNA, whereas some of the βFSH cells coexpressed only lpxrf-r mRNA. No coexpression of tilapia lpxrf-r was identified in GH-positive cells. These findings suggest that the LPXRFa system is a potent positive regulator of the reproductive neuroendocrine axis of tilapia.

  6. Positive correlation between ADAR expression and its targets suggests a complex regulation mediated by RNA editing in the human brain.

    PubMed

    Liscovitch, Noa; Bazak, Lily; Levanon, Erez Y; Chechik, Gal

    2014-01-01

    A-to-I RNA editing by adenosine deaminases acting on RNA is a post-transcriptional modification that is crucial for normal life and development in vertebrates. RNA editing has been shown to be very abundant in the human transcriptome, specifically at the primate-specific Alu elements. The functional role of this wide-spread effect is still not clear; it is believed that editing of transcripts is a mechanism for their down-regulation via processes such as nuclear retention or RNA degradation. Here we combine 2 neural gene expression datasets with genome-level editing information to examine the relation between the expression of ADAR genes with the expression of their target genes. Specifically, we computed the spatial correlation across structures of post-mortem human brains between ADAR and a large set of targets that were found to be edited in their Alu repeats. Surprisingly, we found that a large fraction of the edited genes are positively correlated with ADAR, opposing the assumption that editing would reduce expression. When considering the correlations between ADAR and its targets over development, 2 gene subsets emerge, positively correlated and negatively correlated with ADAR expression. Specifically, in embryonic time points, ADAR is positively correlated with many genes related to RNA processing and regulation of gene expression. These findings imply that the suggested mechanism of regulation of expression by editing is probably not a global one; ADAR expression does not have a genome wide effect reducing the expression of editing targets. It is possible, however, that RNA editing by ADAR in non-coding regions of the gene might be a part of a more complex expression regulation mechanism.

  7. A Flexible Binding Site Architecture Provides New Insights into CcpA Global Regulation in Gram-Positive Bacteria

    PubMed Central

    Yang, Yunpeng; Zhang, Lu; Huang, He; Yang, Chen; Yang, Sheng

    2017-01-01

    ABSTRACT Catabolite control protein A (CcpA) is the master regulator in Gram-positive bacteria that mediates carbon catabolite repression (CCR) and carbon catabolite activation (CCA), two fundamental regulatory mechanisms that enable competitive advantages in carbon catabolism. It is generally regarded that CcpA exerts its regulatory role by binding to a typical 14- to 16-nucleotide (nt) consensus site that is called a catabolite response element (cre) within the target regions. However, here we report a previously unknown noncanonical flexible architecture of the CcpA-binding site in solventogenic clostridia, providing new mechanistic insights into catabolite regulation. This novel CcpA-binding site, named crevar, has a unique architecture that consists of two inverted repeats and an intervening spacer, all of which are variable in nucleotide composition and length, except for a 6-bp core palindromic sequence (TGTAAA/TTTACA). It was found that the length of the intervening spacer of crevar can affect CcpA binding affinity, and moreover, the core palindromic sequence of crevar is the key structure for regulation. Such a variable architecture of crevar shows potential importance for CcpA’s diverse and fine regulation. A total of 103 potential crevar sites were discovered in solventogenic Clostridium acetobutylicum, of which 42 sites were picked out for electrophoretic mobility shift assays (EMSAs), and 30 sites were confirmed to be bound by CcpA. These 30 crevar sites are associated with 27 genes involved in many important pathways. Also of significance, the crevar sites are found to be widespread and function in a great number of taxonomically different Gram-positive bacteria, including pathogens, suggesting their global role in Gram-positive bacteria. PMID:28119470

  8. Influence of sleep state and position on cardio-respiratory regulation in newborn babies.

    PubMed

    Lucchini, Maristella; Fifer, William P; Perez, Albany; Signorini, Maria G

    2015-01-01

    The paper presents results of a sleep study on 60 newborn infants and 22 one-month infants, in quiet and active sleep and in prone and supine position. During the study, HRV and respiration were acquired and then analyzed with a multi-parametric approach. Time, Frequency Domain and Non-Linear parameters were calculated, also encompassing indices from the adult and fetal field. The novelty of this study is the introduction of innovative measurements in a thorough investigation to characterize the effect of sleep state and position on the cardio-respiratory control in newborns. Results show that most parameters succeed in classifying different sleep states, while differences between positions were found in the one-month population only. This study comes as a continuation of previous analysis with the addition of respiratory signal. These results are encouraging for the aim of defining a set of parameters that could help characterizing the autonomic control of infants and early detect the onset of distress or particular pathologies.

  9. Extracellular signal-related kinase positively regulates ataxia telangiectasia mutated, homologous recombination repair, and the DNA damage response.

    PubMed

    Golding, Sarah E; Rosenberg, Elizabeth; Neill, Steven; Dent, Paul; Povirk, Lawrence F; Valerie, Kristoffer

    2007-02-01

    The accurate joining of DNA double-strand breaks by homologous recombination repair (HRR) is critical to the long-term survival of the cell. The three major mitogen-activated protein (MAP) kinase (MAPK) signaling pathways, extracellular signal-regulated kinase (ERK), p38, and c-Jun-NH(2)-kinase (JNK), regulate cell growth, survival, and apoptosis. To determine the role of MAPK signaling in HRR, we used a human in vivo I-SceI-based repair system. First, we verified that this repair platform is amenable to pharmacologic manipulation and show that the ataxia telangiectasia mutated (ATM) kinase is critical for HRR. The ATM-specific inhibitor KU-55933 compromised HRR up to 90% in growth-arrested cells, whereas this effect was less pronounced in cycling cells. Then, using well-characterized MAPK small-molecule inhibitors, we show that ERK1/2 and JNK signaling are important positive regulators of HRR in growth-arrested cells. On the other hand, inhibition of the p38 MAPK pathway generated an almost 2-fold stimulation of HRR. When ERK1/2 signaling was stimulated by oncogenic RAF-1, an approximately 2-fold increase in HRR was observed. KU-55933 partly blocked radiation-induced ERK1/2 phosphorylation, suggesting that ATM regulates ERK1/2 signaling. Furthermore, inhibition of MAP/ERK kinase (MEK)/ERK signaling resulted in severely reduced levels of phosphorylated (S1981) ATM foci but not gamma-H2AX foci, and suppressed ATM phosphorylation levels >85% throughout the cell cycle. Collectively, these results show that MAPK signaling positively and negatively regulates HRR in human cells. More specifically, ATM-dependent signaling through the RAF/MEK/ERK pathway is critical for efficient HRR and for radiation-induced ATM activation, suggestive of a regulatory feedback loop between ERK and ATM.

  10. The transcription factor Pitx2 positions the embryonic axis and regulates twinning

    PubMed Central

    Torlopp, Angela; Khan, Mohsin A F; Oliveira, Nidia M M; Lekk, Ingrid; Soto-Jiménez, Luz Mayela; Sosinsky, Alona; Stern, Claudio D

    2014-01-01

    Embryonic polarity of invertebrates, amphibians and fish is specified largely by maternal determinants, which fixes cell fates early in development. In contrast, amniote embryos remain plastic and can form multiple individuals until gastrulation. How is their polarity determined? In the chick embryo, the earliest known factor is cVg1 (homologous to mammalian growth differentiation factor 1, GDF1), a transforming growth factor beta (TGFβ) signal expressed posteriorly before gastrulation. A molecular screen to find upstream regulators of cVg1 in normal embryos and in embryos manipulated to form twins now uncovers the transcription factor Pitx2 as a candidate. We show that Pitx2 is essential for axis formation, and that it acts as a direct regulator of cVg1 expression by binding to enhancers within neighbouring genes. Pitx2, Vg1/GDF1 and Nodal are also key actors in left–right asymmetry, suggesting that the same ancient polarity determination mechanism has been co-opted to different functions during evolution. DOI: http://dx.doi.org/10.7554/eLife.03743.001 PMID:25496870

  11. O-GlcNAcylation of ATG4B positively regulates autophagy by increasing its hydroxylase activity.

    PubMed

    Jo, Yoon Kyung; Park, Na Yeon; Park, So Jung; Kim, Byung-Gyu; Shin, Ji Hyun; Jo, Doo Sin; Bae, Dong-Jun; Suh, Young-Ah; Chang, Jeong Ho; Lee, Eun Kyung; Kim, Sang-Yeob; Kim, Jin Cheon; Cho, Dong-Hyung

    2016-08-30

    Autophagy is a catabolic degradation process and maintains cellular homeostasis. And autophagy is activated in response to various stress conditions. Although O-GlcNAcylation functions a sensor for nutrient and stress, the relationship between O-GlcNAcylation and autophagy is largely unknown. Here, we identified that ATG4B is novel target for O-GlcNAcylation under metabolic stress condition. Treatment with PugNAc, an O-GlcNAcase inhibitor increased activation of autophagy in SH-SY5Y cells. Both bimolecular fluorescence complementation and immunoprecipitation assay indicated that OGT directly interacts with ATG4B in SH-SY5Y cells. We also found that the O-GlcNAcylated ATG4B was increased in autophagy activation conditions, and down-regulation of OGT reduces O-GlcNAcylation of ATG4B under low glucose condition. Furthermore, the proteolytic activity of ATG4B for LC3 cleavage was enhanced in PugNAc-treated cells. Taken together, these results imply that O-GlcNAcylation of ATG4B regulates autophagy activation by increasing its proteolytic activity under metabolic stress condition.

  12. Baicalin positively regulates osteoclast function by activating MAPK/Mitf signalling.

    PubMed

    Lu, Li; Rao, Li; Jia, Huanhuan; Chen, Jun; Lu, Xingyan; Yang, Guozhu; Li, Qingnan; Lee, Kenneth Ka Ho; Yang, Li

    2017-02-03

    Activation of osteoblasts in bone formation and osteoclasts in bone resorption is important during the bone fracture healing process. There has been a long interest in identifying and developing a natural therapy for bone fracture healing. In this study, we investigated the regulation of osteoclast differentiation by baicalin, which is a natural molecule extracted from Eucommiaulmoides (small tree native to China). It was determined that baicalin enhanced osteoclast maturation and bone resorption activity in a dose-dependent manner. Moreover, this involves the activation of MAPK, increased Mitf nuclear translocation and up-regulation of downstream osteoclast-related target genes expression. The baicalin-induced effect on osteoclast differentiation can be mimicked by specific inhibitors of p-ERK (U0126) and the Mitf-specific siRNA, respectively. Protein-ligand docking prediction identified that baicalin might bind to RANK, which is the upstream receptor of p-ERK/Mitf signalling in osteoclasts. This indicated that RANK might be the binding target of baicalin. In sum, our findings revealed baicalin increased osteoclast maturation and function via p-ERK/Mitf signalling. In addition, the results suggest that baicalin can potentially be used as a natural product for the treatment of bone fracture.

  13. O-GlcNAcylation of ATG4B positively regulates autophagy by increasing its hydroxylase activity

    PubMed Central

    Jo, Yoon Kyung; Park, Na Yeon; Park, So Jung; Kim, Byung-Gyu; Shin, Ji Hyun; Jo, Doo Sin; Bae, Dong-Jun; Suh, Young-Ah; Chang, Jeong Ho; Lee, Eun Kyung; Kim, Sang-Yeob; Kim, Jin Cheon; Cho, Dong-Hyung

    2016-01-01

    Autophagy is a catabolic degradation process and maintains cellular homeostasis. And autophagy is activated in response to various stress conditions. Although O-GlcNAcylation functions a sensor for nutrient and stress, the relationship between O-GlcNAcylation and autophagy is largely unknown. Here, we identified that ATG4B is novel target for O-GlcNAcylation under metabolic stress condition. Treatment with PugNAc, an O-GlcNAcase inhibitor increased activation of autophagy in SH-SY5Y cells. Both bimolecular fluorescence complementation and immunoprecipitation assay indicated that OGT directly interacts with ATG4B in SH-SY5Y cells. We also found that the O-GlcNAcylated ATG4B was increased in autophagy activation conditions, and down-regulation of OGT reduces O-GlcNAcylation of ATG4B under low glucose condition. Furthermore, the proteolytic activity of ATG4B for LC3 cleavage was enhanced in PugNAc-treated cells. Taken together, these results imply that O-GlcNAcylation of ATG4B regulates autophagy activation by increasing its proteolytic activity under metabolic stress condition. PMID:27527864

  14. Glucose oxidation positively regulates glucose uptake and improves cardiac function recovery after myocardial reperfusion.

    PubMed

    Li, Tingting; Xu, Jie; Qin, Xinghua; Hou, Zuoxu; Guo, Yongzheng; Liu, Zhenhua; Wu, Jianjiang; Zheng, Hong; Zhang, Xing; Gao, Feng

    2017-03-21

    Myocardial reperfusion decreases glucose oxidation and uncouples glucose oxidation from glycolysis. Therapies that increase glucose oxidation lessen myocardial ischemia/reperfusion injury. However, the regulation of glucose uptake during reperfusion remains poorly understood. Here we found that glucose uptake was remarkably diminished in myocardium following reperfusion in Sprague-Dawley rats as detected by 18F-labeled and fluorescent-labeled glucose analogs, even though GLUT1 was upregulated by 3 folds and GLUT4 translocation remained unchanged compared with those of sham rats. The decreased glucose uptake was accompanied by suppressed glucose oxidation. Interestingly, stimulating glucose oxidation by inhibition of pyruvate dehydrogenase kinase 4 (PDK4), a rate-limiting enzyme for glucose oxidation, increased glucose uptake and alleviated ischemia/reperfusion injury. In vitro data in neonatal myocytes showed that PDK4 overexpression decreased glucose uptake, while its knockdown increased glucose uptake, suggesting a role of PDK4 in regulating glucose uptake. Moreover, inhibition of PDK4 increased myocardial glucose uptake with concomitant enhancement of cardiac insulin sensitivity following myocardial ischemia/reperfusion. These results showed that the suppressed glucose oxidation mediated by PDK4 contributes to the reduced glucose uptake in myocardium following reperfusion, and enhancement of glucose uptake exerts cardioprotection. The findings suggest that stimulating glucose oxidation via PDK4 could be an efficient approach to improve recovery from myocardial ischemia/reperfusion injury.

  15. Direct evidence that PKCα positively regulates wound re-epithelialization: correlation with changes in desmosomal adhesiveness.

    PubMed

    Thomason, Helen A; Cooper, Nichola H; Ansell, David M; Chiu, Maybo; Merrit, Anita J; Hardman, Matthew J; Garrod, David R

    2012-07-01

    Non-healing wounds cause considerable patient morbidity and represent a significant economic burden. Central to wound repair is re-epithelialization, a crucial process involving the modulation of cell adhesion to allow keratinocyte migration to cover the exposed underlying tissues. The cellular mechanisms regulating the earliest stages of re-epithelialization are unclear. We present the first direct evidence that protein kinase Cα (PKCα) plays an important role in regulating wound re-epithelialization. In PKCα(-/-) mice re-epithelialization is delayed, while in novel bitransgenic mice over-expressing constitutively active PKCα it is accelerated. These effects are not due to changes in keratinocyte proliferation, apoptosis or intrinsic cell motility. Instead, they correlate with changes in desmosomal adhesiveness, delay being preceded by retained desmosomal hyper-adhesiveness and acceleration with a rapid switch to desmosomal Ca(2+) -dependence. We demonstrate mechanistic conservation in acute human wounds where PKCα localizes to wound edge desmosomes, which become Ca(2+) -dependent. However, in chronic wounds PKCα remains cytoplasmic and desmosomes fail to switch from the hyper-adhesive state. These results throw new mechanistic light on the earliest stages of wound re-epithelialization and suggest activation of PKCα as a new therapeutic strategy for non-healing wounds.

  16. Evaluation of intramitochondrial ATP levels identifies G0/G1 switch gene 2 as a positive regulator of oxidative phosphorylation

    PubMed Central

    Kioka, Hidetaka; Kato, Hisakazu; Fujikawa, Makoto; Tsukamoto, Osamu; Suzuki, Toshiharu; Imamura, Hiromi; Nakano, Atsushi; Higo, Shuichiro; Yamazaki, Satoru; Matsuzaki, Takashi; Takafuji, Kazuaki; Asanuma, Hiroshi; Asakura, Masanori; Minamino, Tetsuo; Shintani, Yasunori; Yoshida, Masasuke; Noji, Hiroyuki; Kitakaze, Masafumi; Komuro, Issei; Asano, Yoshihiro; Takashima, Seiji

    2014-01-01

    The oxidative phosphorylation (OXPHOS) system generates most of the ATP in respiring cells. ATP-depleting conditions, such as hypoxia, trigger responses that promote ATP production. However, how OXPHOS is regulated during hypoxia has yet to be elucidated. In this study, selective measurement of intramitochondrial ATP levels identified the hypoxia-inducible protein G0/G1 switch gene 2 (G0s2) as a positive regulator of OXPHOS. A mitochondria-targeted, FRET-based ATP biosensor enabled us to assess OXPHOS activity in living cells. Mitochondria-targeted, FRET-based ATP biosensor and ATP production assay in a semiintact cell system revealed that G0s2 increases mitochondrial ATP production. The expression of G0s2 was rapidly and transiently induced by hypoxic stimuli, and G0s2 interacts with OXPHOS complex V (FoF1-ATP synthase). Furthermore, physiological enhancement of G0s2 expression prevented cells from ATP depletion and induced a cellular tolerance for hypoxic stress. These results show that G0s2 positively regulates OXPHOS activity by interacting with FoF1-ATP synthase, which causes an increase in ATP production in response to hypoxic stress and protects cells from a critical energy crisis. These findings contribute to the understanding of a unique stress response to energy depletion. Additionally, this study shows the importance of assessing intramitochondrial ATP levels to evaluate OXPHOS activity in living cells. PMID:24344269

  17. NUCLEAR FACTOR Y, Subunit C (NF-YC) Transcription Factors Are Positive Regulators of Photomorphogenesis in Arabidopsis thaliana

    PubMed Central

    Siriwardana, Chamindika L.; Holt III, Ben F.

    2016-01-01

    Recent reports suggest that NF-Y transcription factors are positive regulators of skotomorphogenesis in Arabidopsis thaliana. Three NF-YC genes (NF-YC3, NF-YC4, and NF-YC9) are known to have overlapping functions in photoperiod dependent flowering and previous studies demonstrated that they interact with basic leucine zipper (bZIP) transcription factors. This included ELONGATED HYPOCOTYL 5 (HY5), which has well-demonstrated roles in photomorphogenesis. Similar to hy5 mutants, we report that nf-yc3 nf-yc4 nf-yc9 triple mutants failed to inhibit hypocotyl elongation in all tested light wavelengths. Surprisingly, nf-yc3 nf-yc4 nf-yc9 hy5 mutants had synergistic defects in light perception, suggesting that NF-Ys represent a parallel light signaling pathway. As with other photomorphogenic transcription factors, nf-yc3 nf-yc4 nf-yc9 triple mutants also partially suppressed the short hypocotyl and dwarf rosette phenotypes of CONSTITUTIVE PHOTOMORPHOGENIC 1 (cop1) mutants. Thus, our data strongly suggest that NF-Y transcription factors have important roles as positive regulators of photomorphogenesis, and in conjunction with other recent reports, implies that the NF-Y are multifaceted regulators of early seedling development. PMID:27685091

  18. Applying High-Speed Vision Sensing to an Industrial Robot for High-Performance Position Regulation under Uncertainties.

    PubMed

    Huang, Shouren; Bergström, Niklas; Yamakawa, Yuji; Senoo, Taku; Ishikawa, Masatoshi

    2016-07-29

    It is traditionally difficult to implement fast and accurate position regulation on an industrial robot in the presence of uncertainties. The uncertain factors can be attributed either to the industrial robot itself (e.g., a mismatch of dynamics, mechanical defects such as backlash, etc.) or to the external environment (e.g., calibration errors, misalignment or perturbations of a workpiece, etc.). This paper proposes a systematic approach to implement high-performance position regulation under uncertainties on a general industrial robot (referred to as the main robot) with minimal or no manual teaching. The method is based on a coarse-to-fine strategy that involves configuring an add-on module for the main robot's end effector. The add-on module consists of a 1000 Hz vision sensor and a high-speed actuator to compensate for accumulated uncertainties. The main robot only focuses on fast and coarse motion, with its trajectories automatically planned by image information from a static low-cost camera. Fast and accurate peg-and-hole alignment in one dimension was implemented as an application scenario by using a commercial parallel-link robot and an add-on compensation module with one degree of freedom (DoF). Experimental results yielded an almost 100% success rate for fast peg-in-hole manipulation (with regulation accuracy at about 0.1 mm) when the workpiece was randomly placed.

  19. Applying High-Speed Vision Sensing to an Industrial Robot for High-Performance Position Regulation under Uncertainties

    PubMed Central

    Huang, Shouren; Bergström, Niklas; Yamakawa, Yuji; Senoo, Taku; Ishikawa, Masatoshi

    2016-01-01

    It is traditionally difficult to implement fast and accurate position regulation on an industrial robot in the presence of uncertainties. The uncertain factors can be attributed either to the industrial robot itself (e.g., a mismatch of dynamics, mechanical defects such as backlash, etc.) or to the external environment (e.g., calibration errors, misalignment or perturbations of a workpiece, etc.). This paper proposes a systematic approach to implement high-performance position regulation under uncertainties on a general industrial robot (referred to as the main robot) with minimal or no manual teaching. The method is based on a coarse-to-fine strategy that involves configuring an add-on module for the main robot’s end effector. The add-on module consists of a 1000 Hz vision sensor and a high-speed actuator to compensate for accumulated uncertainties. The main robot only focuses on fast and coarse motion, with its trajectories automatically planned by image information from a static low-cost camera. Fast and accurate peg-and-hole alignment in one dimension was implemented as an application scenario by using a commercial parallel-link robot and an add-on compensation module with one degree of freedom (DoF). Experimental results yielded an almost 100% success rate for fast peg-in-hole manipulation (with regulation accuracy at about 0.1 mm) when the workpiece was randomly placed. PMID:27483274

  20. Small Antisense RNA RblR Positively Regulates RuBisCo in Synechocystis sp. PCC 6803

    PubMed Central

    Hu, Jinlu; Li, Tianpei; Xu, Wen; Zhan, Jiao; Chen, Hui; He, Chenliu; Wang, Qiang

    2017-01-01

    Small regulatory RNAs (sRNAs) function as transcriptional and post-transcriptional regulators of gene expression in organisms from all domains of life. Cyanobacteria are thought to have developed a complex RNA-based regulatory mechanism. In the current study, by genome-wide analysis of differentially expressed small RNAs in Synechocystis sp. PCC 6803 under high light conditions, we discovered an asRNA (RblR) that is 113nt in length and completely complementary to its target gene rbcL, which encodes the large chain of RuBisCO, the enzyme that catalyzes carbon fixation. Further analysis of the RblR(+)/(−) mutants revealed that RblR acts as a positive regulator of rbcL under various stress conditions; Suppressing RblR adversely affects carbon assimilation and thus the yield, and those phenotypes of both the wild type and the overexpressor could be downgraded to the suppressor level by carbonate depletion, indicated a regulatory role of RblR in CO2 assimilation. In addition, a real-time expression platform in Escherichia coli was setup and which confirmed that RblR promoted the translation of the rbcL mRNA into the RbcL protein. The present study is the first report of a regulatory RNA that targets RbcL in Synechocystis sp. PCC 6803, and provides strong evidence that RblR regulates photosynthesis by positively modulating rbcL expression in Synechocystis. PMID:28261186

  1. CD44 regulates dendrite morphogenesis through Src tyrosine kinase-dependent positioning of the Golgi.

    PubMed

    Skupien, Anna; Konopka, Anna; Trzaskoma, PaweI; Labus, Josephine; Gorlewicz, Adam; Swiech, Lukasz; Babraj, Matylda; Dolezyczek, Hubert; Figiel, Izabela; Ponimaskin, Evgeni; Wlodarczyk, Jakub; Jaworski, Jacek; Wilczynski, Grzegorz M; Dzwonek, Joanna

    2014-12-01

    The acquisition of proper dendrite morphology is a crucial aspect of neuronal development towards the formation of a functional network. The role of the extracellular matrix and its cellular receptors in this process has remained enigmatic. We report that the CD44 adhesion molecule, the main hyaluronan receptor, is localized in dendrites and plays a crucial inhibitory role in dendritic tree arborization in vitro and in vivo. This novel function is exerted by the activation of Src tyrosine kinase, leading to the alteration of Golgi morphology. The mechanism operates during normal brain development, but its inhibition might have a protective influence on dendritic trees under toxic conditions, during which the silencing of CD44 expression prevents dendritic shortening induced by glutamate exposure. Overall, our results indicate a novel role for CD44 as an essential regulator of dendritic arbor complexity in both health and disease.

  2. Paxillin mediates sensing of physical cues and regulates directional cell motility by controlling lamellipodia positioning.

    PubMed

    Sero, Julia E; Thodeti, Charles K; Mammoto, Akiko; Bakal, Chris; Thomas, Sheila; Ingber, Donald E

    2011-01-01

    Physical interactions between cells and the extracellular matrix (ECM) guide directional migration by spatially controlling where cells form focal adhesions (FAs), which in turn regulate the extension of motile processes. Here we show that physical control of directional migration requires the FA scaffold protein paxillin. Using single-cell sized ECM islands to constrain cell shape, we found that fibroblasts cultured on square islands preferentially activated Rac and extended lamellipodia from corner, rather than side regions after 30 min stimulation with PDGF, but that cells lacking paxillin failed to restrict Rac activity to corners and formed small lamellipodia along their entire peripheries. This spatial preference was preceded by non-spatially constrained formation of both dorsal and lateral membrane ruffles from 5-10 min. Expression of paxillin N-terminal (paxN) or C-terminal (paxC) truncation mutants produced opposite, but complementary, effects on lamellipodia formation. Surprisingly, pax-/- and paxN cells also formed more circular dorsal ruffles (CDRs) than pax+ cells, while paxC cells formed fewer CDRs and extended larger lamellipodia even in the absence of PDGF. In a two-dimensional (2D) wound assay, pax-/- cells migrated at similar speeds to controls but lost directional persistence. Directional motility was rescued by expressing full-length paxillin or the N-terminus alone, but paxN cells migrated more slowly. In contrast, pax-/- and paxN cells exhibited increased migration in a three-dimensional (3D) invasion assay, with paxN cells invading Matrigel even in the absence of PDGF. These studies indicate that paxillin integrates physical and chemical motility signals by spatially constraining where cells will form motile processes, and thereby regulates directional migration both in 2D and 3D. These findings also suggest that CDRs may correspond to invasive protrusions that drive cell migration through 3D extracellular matrices.

  3. Multilayered Regulation of Ethylene Induction Plays a Positive Role in Arabidopsis Resistance against Pseudomonas syringae.

    PubMed

    Guan, Rongxia; Su, Jianbin; Meng, Xiangzong; Li, Sen; Liu, Yidong; Xu, Juan; Zhang, Shuqun

    2015-09-01

    Ethylene, a key phytohormone involved in plant-pathogen interaction, plays a positive role in plant resistance against fungal pathogens. However, its function in plant bacterial resistance remains unclear. Here, we report a detailed analysis of ethylene induction in Arabidopsis (Arabidopsis thaliana) in response to Pseudomonas syringae pv tomato DC3000 (Pst). Ethylene biosynthesis is highly induced in both pathogen/microbe-associated molecular pattern (PAMP)-triggered immunity and effector-triggered immunity (ETI), and the induction is potentiated by salicylic acid (SA) pretreatment. In addition, Pst actively suppresses PAMP-triggered ethylene induction in a type III secretion system-dependent manner. SA potentiation of ethylene induction is dependent mostly on MITOGEN-ACTIVATED PROTEIN KINASE6 (MPK6) and MPK3 and their downstream ACS2 and ACS6, two type I isoforms of 1-aminocyclopropane-1-carboxylic acid synthases (ACSs). ACS7, a type III ACS whose expression is enhanced by SA pretreatment, is also involved. Pst expressing the avrRpt2 effector gene (Pst-avrRpt2), which is capable of triggering ETI, induces a higher level of ethylene production, and the elevated portion is dependent on SALICYLIC ACID INDUCTION DEFICIENT2 and NONEXPRESSER OF PATHOGENESIS-RELATED GENE1, two key players in SA biosynthesis and signaling. High-order ACS mutants with reduced ethylene induction are more susceptible to both Pst and Pst-avrRpt2, demonstrating a positive role of ethylene in plant bacterial resistance mediated by both PAMP-triggered immunity and ETI.

  4. Glia-derived ATP inversely regulates excitability of pyramidal and CCK-positive neurons

    PubMed Central

    Tan, Zhibing; Liu, Yu; Xi, Wang; Lou, Hui-fang; Zhu, Liya; Guo, Zhifei; Mei, Lin; Duan, Shumin

    2017-01-01

    Astrocyte responds to neuronal activity with calcium waves and modulates synaptic transmission through the release of gliotransmitters. However, little is known about the direct effect of gliotransmitters on the excitability of neuronal networks beyond synapses. Here we show that selective stimulation of astrocytes expressing channelrhodopsin-2 in the CA1 area specifically increases the firing frequency of CCK-positive but not parvalbumin-positive interneurons and decreases the firing rate of pyramidal neurons, phenomena mimicked by exogenously applied ATP. Further evidences indicate that ATP-induced increase and decrease of excitability are caused, respectively, by P2Y1 receptor-mediated inhibition of a two-pore domain potassium channel and A1 receptor-mediated opening of a G-protein-coupled inwardly rectifying potassium channel. Moreover, the activation of ChR2-expressing astrocytes reduces the power of kainate-induced hippocampal ex vivo gamma oscillation. Thus, through distinct receptor subtypes coupled with different K+ channels, astrocyte-derived ATP differentially modulates the excitability of different types of neurons and efficiently controls the activity of neuronal network. PMID:28128211

  5. CaCDPK15 positively regulates pepper responses to Ralstonia solanacearum inoculation and forms a positive-feedback loop with CaWRKY40 to amplify defense signaling

    PubMed Central

    Shen, Lei; Yang, Sheng; Yang, Tong; Liang, Jiaqi; Cheng, Wei; Wen, Jiayu; Liu, Yanyan; Li, Jiazhi; Shi, Lanping; Tang, Qian; Shi, Wei; Hu, Jiong; Liu, Cailing; Zhang, Yangwen; Mou, Shaoliang; Liu, Zhiqin; Cai, Hanyang; He, Li; Guan, Deyi; Wu, Yang; He, Shuilin

    2016-01-01

    CaWRKY40 is a positive regulator of pepper (Capsicum annum) response to Ralstonia solanacearum inoculation (RSI), but the underlying mechanism remains largely unknown. Here, we functionally characterize CaCDPK15 in the defense signaling mediated by CaWRKY40. Pathogen-responsive TGA, W, and ERE boxes were identified in the CaCDPK15 promoter (pCaCDPK15), and pCaCDPK15-driven GUS expression was significantly enhanced in response to RSI and exogenously applied salicylic acid, methyl jasmonate, abscisic acid, and ethephon. Virus-induced gene silencing (VIGS) of CaCDPK15 significantly increased the susceptibility of pepper to RSI and downregulated the immunity-associated markers CaNPR1, CaPR1, and CaDEF1. By contrast, transient CaCDPK15 overexpression significantly activated hypersensitive response associated cell death, upregulated the immunity-associated marker genes, upregulated CaWRKY40 expression, and enriched CaWRKY40 at the promoters of its targets genes. Although CaCDPK15 failed to interact with CaWRKY40, the direct binding of CaWRKY40 to pCaCDPK15 was detected by chromatin immunoprecipitation, which was significantly potentiated by RSI in pepper plants. These combined results suggest that RSI in pepper induces CaCDPK15 and indirectly activates downstream CaWRKY40, which in turn potentiates CaCDPK15 expression. This positive-feedback loop would amplify defense signaling against RSI and efficiently activate strong plant immunity. PMID:26928570

  6. Cell type-specific expression of JC virus early promoter is determined by positive and negative regulation.

    PubMed

    Tada, H; Lashgari, M; Rappaport, J; Khalili, K

    1989-01-01

    We analyzed control sequences of the human papovavirus JC virus (JCV) to define the cis-acting elements that regulate specific expression of the viral early region genes in glial cells. Nuclear run-on transcription, S1 analysis, and chloramphenicol acetyltransferase enzyme activity in a transient transfection assay established that the cell type-specific expression of JCV early genes is determined at the transcriptional level. Using DNase footprinting analysis of nuclear proteins prepared from glial and nonglial cells, we located four regions within the JCV control sequences that specifically interacted with the proteins. In glial cells, all four domains contributed to the specific expression of a heterologous promoter, whereas in nonglial cells, two protein-binding regions showed no effect on basal transcriptional activity and the other two domains significantly downregulated transcription of the promoter. We conclude that cell type-specific transcription of the JCV early promoter is under both positive and negative regulation in eucaryotic cells.

  7. Salvianolic acid A positively regulates PTEN protein level and inhibits growth of A549 lung cancer cells

    PubMed Central

    BI, LEI; CHEN, JIANPING; YUAN, XIAOJING; JIANG, ZEQUN; CHEN, WEIPING

    2013-01-01

    Salvianolic acid A (Sal A) is an effective compound extracted from Salvia miltiorrhiza which has been used in the treatment of various diseases. Preliminary data indicate that Sal A treatment has a specific anti-lung cancer effect. However, the manner in which Sal A regulates cancer growth remains unknown. In this study, the A549 lung cancer cell line and its response to Sal A treatment was examined. Results showed that Sal A treatment significantly decreased A549 cell growth, promoted partial apoptosis and increased mitochondrial membrane permeability. Western blot analysis showed that Sal A upregulated the phosphatase and tensin homolog (PTEN) protein level, while consistently downregulating Akt phosphorylation. These results indicate that Sal A negatively mediates A549 lung cancer cell line growth or apoptosis, most likely by positively regulating PTEN protein level. PMID:24648921

  8. TRIM52: A nuclear TRIM protein that positively regulates the nuclear factor-kappa B signaling pathway.

    PubMed

    Fan, Wenchun; Liu, Tingting; Li, Xiangmin; Zhou, Yun; Wu, Mengge; Cui, Xiaofang; Chen, Huanchun; Qian, Ping

    2017-02-01

    Emerging evidence suggests that TRIM family proteins play a crucial role in regulating the NF-κB signaling pathway. TRIM52 is a novel noncanonical antiviral TRIM gene with a unique expanded RING domain. Information on the biological function of TRIM52 is limited. Herein, we demonstrated TRIM52 involvement in NF-κB activation. We found that TRIM52 overexpression specifically activated the NF-κB signal. TRIM52 overexpression can significantly induce TNFα and IL-6 expression. We also found that the RING domain of TRIM52 was essential for its activation of the NF-κB signal. Further study showed that TRIM52 overexpression did not affect the protein level of IκBα and phosphorylated p65 protein. We found that the pro-inflammatory cytokines TNFα and IL-6 could induce TRIM52 expression. Overall, these data suggested that TRIM52 was a positive regulator of the NF-κB pathway.

  9. Constitutively expressed COX-2 in osteoblasts positively regulates Akt signal transduction via suppression of PTEN activity.

    PubMed

    Li, Ching-Ju; Chang, Je-Ken; Wang, Gwo-Jaw; Ho, Mei-Ling

    2011-02-01

    Cyclooxygenase-2 (COX-2) is thought to be an inducible enzyme, but increasing reports indicate that COX-2 is constitutively expressed in several organs. The status of COX-2 expression in bone and its physiological role remains undefined. Non-selective non-steroidal anti-inflammatory drugs (NSAIDs) and selective COX-2 inhibitors, which commonly suppress COX-2 activity, were reported to suppress osteoblast proliferation via Akt/FOXO3a/p27(Kip1) signaling, suggesting that COX-2 may be the key factor of the suppressive effects of NSAIDs on proliferation. Although Akt activation correlates with PTEN deficiency and cell viability, the role of COX-2 on PTEN/Akt regulation remains unclear. In this study, we hypothesized that COX-2 may be constitutively expressed in osteoblasts and regulate PTEN/Akt-related proliferation. We examined the localization and co-expression of COX-2 and p-Akt in normal mouse femurs and in cultured mouse (mOBs) and human osteoblasts (hOBs). Our results showed that osteoblasts adjacent to the trabeculae, periosteum and endosteum in mouse femurs constitutively expressed COX-2, while COX-2 co-expressed with p-Akt in osteoblasts sitting adjacent to trabeculae in vivo, and in mOBs and hOBs in vitro. We further used COX-2 siRNA to test the role of COX-2 in Akt signaling in hOBs; COX-2 silencing significantly inhibited PTEN phosphorylation, enhanced PTEN activity, and suppressed p-Akt level and proliferation. However, replenishment of the COX-2 enzymatic product, PGE2, failed to reverse COX-2-dependent Akt phosphorylation. Furthermore, transfection with recombinant human COX-2 (rhCOX-2) significantly reversed COX-2 siRNA-suppressed PTEN phosphorylation, but this effect was reduced when the enzymatic activity of rhCOX-2 was blocked. This finding indicated that the effect of COX-2 on PTEN/Akt signaling is not related to PGE2 but still dependent on COX-2 enzymatic activity. Conversely, COX-1 silencing did not affect PTEN/Akt signaling. Our findings provide

  10. Chromosome position at the spindle equator is regulated by chromokinesin and a bipolar microtubule array.

    PubMed

    Takagi, Jun; Itabashi, Takeshi; Suzuki, Kazuya; Ishiwata, Shin'ichi

    2013-09-30

    The chromosome alignment is mediated by polar ejection and poleward forces acting on the chromosome arm and kinetochores, respectively. Although components of the motile machinery such as chromokinesin have been characterized, their dynamics within the spindle is poorly understood. Here we show that a quantum dot (Qdot) binding up to four Xenopus chromokinesin (Xkid) molecules behaved like a nanosize chromosome arm in the meiotic spindle, which is self-organized in cytoplasmic egg extracts. Xkid-Qdots travelled long distances along microtubules by changing several tracks, resulting in their accumulation toward and distribution around the metaphase plate. The analysis indicated that the direction of motion and velocity depend on the distribution of microtubule polarity within the spindle. Thus, this mechanism is governed by chromokinesin motors, which is dependent on symmetrical microtubule orientation that may allow chromosomes to maintain their position around the spindle equator until correct microtubule-kinetochore attachment is established.

  11. The regulation of positive and negative social feedback: A psychophysiological study.

    PubMed

    Vanderhasselt, Marie-Anne; Remue, Jonathan; Ng, Kwun Kei; Mueller, Sven C; De Raedt, Rudi

    2015-09-01

    Everyday social evaluations are psychologically potent and trigger self-reflective thoughts and feelings. The present study sought to examine the psychophysiological impact of such evaluations using eye tracking, pupillometry, and heart-rate variability. Fifty-nine healthy adult volunteers received rigged social feedback (criticism and praise) based on their photograph. Gaze data were collected to investigate processes of attentional deployment/allocation toward the self or the evaluator expressing criticism or praise. Whereas voluntary attention was directed to evaluators who expressed praise, attention was drawn to one's own picture after criticism. Pupil dilation and heart-rate variability were larger in response to criticism as compared to praise, suggesting a flexible and adaptive emotion regulatory effort in response to social information that triggers an affective response. Altogether, healthy individuals recruited more regulatory resources to cope with negative (as compared to positive) social feedback, and this processing of social feedback was associated with adjustments in self-focused attention.

  12. Temporally Distinct Six2-Positive Second Heart Field Progenitors Regulate Mammalian Heart Development and Disease.

    PubMed

    Zhou, Zhengfang; Wang, Jingying; Guo, Chaoshe; Chang, Weiting; Zhuang, Jian; Zhu, Ping; Li, Xue

    2017-01-24

    The embryonic process of forming a complex structure such as the heart remains poorly understood. Here, we show that Six2 marks a dynamic subset of second heart field progenitors. Six2-positive (Six2(+)) progenitors are rapidly recruited and assigned, and their descendants are allocated successively to regions of the heart from the right ventricle (RV) to the pulmonary trunk. Global ablation of Six2(+) progenitors resulted in RV hypoplasia and pulmonary atresia. An early stage-specific ablation of a small subset of Six2(+) progenitors did not cause any apparent structural defect at birth but rather resulted in adult-onset cardiac hypertrophy and dysfunction. Furthermore, Six2 expression depends in part on Shh signaling, and Shh deletion resulted in severe deficiency of Six2(+) progenitors. Collectively, these findings unveil the chronological features of cardiogenesis, in which the mammalian heart is built sequentially by temporally distinct populations of cardiac progenitors, and provide insights into late-onset congenital heart disease.

  13. Vps33b regulates Vwf-positive vesicular trafficking in megakaryocytes.

    PubMed

    Dai, Jing; Lu, Yeling; Wang, Conghui; Chen, Xue; Fan, Xuemei; Gu, Hao; Wu, Xiaolin; Wang, Kemin; Gartner, T Kent; Zheng, Junke; Chen, Guoqiang; Wang, Xuefeng; Liu, Junling

    2016-09-01

    Mutations of vacuolar protein sorting-associated protein 33b (VPS33B) cause arthrogryposis, renal dysfunction, and cholestasis syndrome, and a lack of platelet α-granules in the affected patients. Conditional Vps33b knockout mice were developed to investigate the function(s) of Vps33b in platelet α-granule formation. We found that early embryonic deletion of Vps33b was lethal. PF4-Cre-driven megakaryocyte-targeted Vps33b gene deletion greatly diminished Vps33b expression in platelets, but had no effect on platelet α-granule formation and protein content. Tamoxifen-induced, haematopoietic stem cell (HSC)-specific Vps33b deletion completely depleted Vps33b in platelets, caused the absence of α-granules, and increased the number of vacuoles in platelets and megakaryocytes. VPS33B association with VIPAS39, α-tubulin, and SEC22B was identified by co-immunoprecipitation, mass spectra, and immunoblotting in human embryonic kidney 293T (HEK293T) cells. Also, pull-down experiments revealed that VIPAS39 bound to intact VPS33B; in contrast, α-tubulin and SEC22B separately interacted with the sec1-like domains of VPS33B. Vps33b deficiency in megakaryocytes disturbs the redistribution of Vipas39 and Sec22b to proplatelets, and interrupted the co-localization of Sec22b with Vwf-positive vesicles. The data presented in this study suggest that Vps33b is involved in α-granule formation possibly by facilitating the Vwf-positive vesicular trafficking to α-granule-related vacuoles in megakaryocytes. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  14. Reelin is a platelet protein and functions as a positive regulator of platelet spreading on fibrinogen.

    PubMed

    Tseng, Wei-Lien; Huang, Chien-Ling; Chong, Kowit-Yu; Liao, Chang-Huei; Stern, Arnold; Cheng, Ju-Chien; Tseng, Ching-Ping

    2010-02-01

    Abnormalities of platelet functions have been linked to reelin-impaired neuronal disorders. However, little attention has been given to understanding the interplay between reelin and platelet. In this study, reelin was found to present in the human platelets and megakaryocyte-like leukemic cells. Reelin-binding assays revealed that extracellular reelin can interact with platelets through the receptor belonging to the low density lipoprotein receptor gene family. The reelin-to-platelet interactions enhance platelet spreading on fibrinogen concomitant with the augmentation of lamellipodia formation and F-actin bundling. In contrast, reelin has no effect on integrin alphaIIbbeta3 activation and agonist-induced platelet aggregation. Molecular analysis revealed that the up-regulation of Rac1 activity and the inhibition of protein kinase C delta-Thr505 phosphorylation are important for reelin-mediated enhancement of platelet spreading on fibrinogen. These findings demonstrate for the first time that reelin is present in platelets and the reelin-to-platelet interactions play a novel role in platelet signaling and functions.

  15. Glycogen synthase kinase-3β positively regulates protein synthesis and cell proliferation through the regulation of translation initiation factor 4E-binding protein 1.

    PubMed

    Shin, S; Wolgamott, L; Tcherkezian, J; Vallabhapurapu, S; Yu, Y; Roux, P P; Yoon, S-O

    2014-03-27

    Protein synthesis has a key role in the control of cell proliferation, and its deregulation is associated with pathological conditions, notably cancer. Rapamycin, an inhibitor of mammalian target of rapamycin complex 1 (mTORC1), was known to inhibit protein synthesis. However, it does not substantially inhibit protein synthesis and cell proliferation in many cancer types. We were interested in finding a novel target in rapamycin-resistant cancer. The rate-limiting factor for translation is eukaryotic translation initiation factor 4E (eIF4E), which is negatively regulated by eIF4E-binding protein 1 (4E-BP1). Here, we provide evidence that glycogen synthase kinase (GSK)-3β promotes cell proliferation through positive regulation of protein synthesis. We found that GSK-3β phosphorylates and inactivates 4E-BP1, thereby increasing eIF4E-dependent protein synthesis. Considering the clinical relevance of pathways regulating protein synthesis, our study provides a promising new strategy and target for cancer therapy.

  16. Os2 MAP kinase-mediated osmostress tolerance in Penicillium digitatum is associated with its positive regulation on glycerol synthesis and negative regulation on ergosterol synthesis.

    PubMed

    Wang, Mingshuang; Chen, Changsheng; Zhu, Congyi; Sun, Xuepeng; Ruan, Ruoxin; Li, Hongye

    2014-01-01

    High osmolarity glycerol (HOG) pathway is ubiquitously distributed among eukaryotic organisms and plays an important role in adaptation to changes in the environment. In this study, the Hog1 ortholog in Penicillium digitatum, designated Pdos2, was identified and characterized using a gene knock-out strategy. The ΔPdos2 mutant showed a considerably increased sensitivity to salt stress and cell wall-disturbing agents and a slightly increased resistance to fungicides iprodione and fludioxonil, indicating that Pdos2 is involved in response to hyperosmotic stress, regulation of cell wall integrity and sensitivity to fungicides iprodione and fludioxonil. Surprisingly, the mutant was not affected in response to oxidative stress caused by H2O2. The average lesion size in citrus fruits caused by ΔPdos2 mutant was smaller (approximately 25.0% reduction) than that caused by the wild-type strain of P. digitatum at 4 days post inoculation, which suggests that Pdos2 is needed for full virulence of P. digitatum. Interestingly, in the presence of 0.7 M NaCl, the glycerol content was remarkably increased and the ergosterol was decreased in mycelia of the wide-type P. digitatum, whereas the glycerol content was only slightly increased and the ergosterol content remained stable in the ΔPdos2 mutant, suggesting that Pdos2-mediated osmotic adaption is associated with its positive regulation on glycerol synthesis and negative regulation on ergosterol synthesis.

  17. Processing of alpha4 integrin by the proprotein convertases: histidine at position P6 regulates cleavage.

    PubMed Central

    Bergeron, Eric; Basak, Ajoy; Decroly, Etienne; Seidah, Nabil G

    2003-01-01

    The proprotein convertases (PCs) participate in the limited proteolysis of integrin alpha4 subunit at the H(592)VISKR(597) downward arrow ST site (where underlined residues indicate positively charged amino acids important for PC-mediated cleavage and downward arrow indicates the cleavage site), since this cleavage is inhibited by the serpin alpha1-PDX (alpha1-antitrypsin Portland). Co-expression of alpha4 with each convertase in LoVo (furin-deficient human colon carcinoma) cells revealed that furin and proprotein convertase 5A (PC5A) are the best pro-alpha4 convertases. In agreement, processing of endogenous pro-alpha4 in human lymphoblastoid CEM-T4 cells was enhanced greatly in stable transfectants overexpressing either enzyme. In many leucocyte cell lines, the expression of furin closely correlated with the endogenous processing efficacy, suggesting that furin is a candidate pro-alpha4 convertase. Mutational analysis showed that replacement of P1 Arg(597) with alanine (R597A) abrogated cleavage, whereas the P6 mutant H592R is even better processed by the endogenous convertases of Chinese-hamster ovary CHO-K1 cells. In vitro kinetic studies using synthetic peptides confirmed the importance of a positively charged residue at P6 and showed that wild-type alpha4 processing is performed best by furin and PC5A at acidic and neutral pHs, respectively. Biosynthetic analysis of pro-alpha4 and its H592R and H592K mutants in the presence or absence of the weak base, NH(4)Cl, revealed that the P6 histidine residue renders its processing by furin sensitive to cellular pH. This suggests that pro-alpha4 cleavage occurs preferentially in acidic compartments. In conclusion, although the accepted furin processing motif is Arg-Xaa-(Lys/Arg)-Arg downward arrow, our data further extend it to include a regulatory histidine residue at P6 in precursors that lack a basic residue at P4. PMID:12691605

  18. E3 ubiquitin ligase NKLAM positively regulates macrophage inducible nitric oxide synthase expression.

    PubMed

    Lawrence, Donald W; Gullickson, Gail; Kornbluth, Jacki

    2015-01-01

    Stimulated macrophages generate potent anti-microbial reactive oxygen and nitrogen species within their phagosomes. Previous studies have shown that the E3 ubiquitin ligase natural killer lytic-associated molecule (NKLAM) is a macrophage phagosomal protein that plays a role in macrophage anti-bacterial activity. In vivo, NKLAM-knockout (KO) mice produce less nitric oxide (NO) upon exposure to lipopolysaccharide (LPS) than wild type (WT) mice. In vitro, we found that NO production and inducible nitric oxide synthase (iNOS) protein were diminished in LPS-stimulated NKLAM-KO bone marrow-derived and splenic macrophages. Additionally, LPS-stimulated NKLAM-KO macrophages displayed defects in STAT1 tyrosine phosphorylation and production of interferon beta (IFNβ). The JAK/STAT pathway is critical for the production of IFNβ, which augments iNOS protein expression in mice. iNOS protein expression is also regulated by the transcription factor NFκB, thus we investigated whether NKLAM influences NFκB function. LPS-stimulated NKLAM-KO macrophages showed evidence of delayed nuclear translocation of the NFκB subunit p65. This was associated with a reduction in p65/DNA colocalization. The defect in p65 translocation was independent of IKBα degradation. NKLAM-KO macrophages also expressed less p65 and showed evidence of defective p65 phosphorylation at serine 536. Importantly, LPS-stimulated NKLAM-KO macrophages have diminished NFκB transcriptional activity as assessed by transfection of a luciferase reporter plasmid. Collectively, our data implicate NKLAM as a novel modulator of macrophage iNOS expression.

  19. Proteasome Modulates Positive and Negative Translational Regulators in Long-Term Synaptic Plasticity

    PubMed Central

    Dong, Chenghai; Bach, Svitlana V.; Haynes, Kathryn A.

    2014-01-01

    Proteolysis by the ubiquitin-proteasome pathway appears to have a complex role in synaptic plasticity, but its various functions remain to be elucidated. Using late phase long-term potentiation (L-LTP) in the hippocampus of the mouse as a model for long-term synaptic plasticity, we previously showed that inhibition of the proteasome enhances induction but blocks maintenance of L-LTP. In this study, we investigated the possible mechanisms by which proteasome inhibition has opposite effects on L-LTP induction and maintenance. Our results show that inhibiting phosphatidyl inositol-3 kinase or blocking the interaction between eukaryotic initiation factors 4E (eIF4E) and 4G (eIF4G) reduces the enhancement of L-LTP induction brought about by proteasome inhibition suggesting interplay between proteolysis and the signaling pathway mediated by mammalian target of rapamycin (mTOR). Also, proteasome inhibition leads to accumulation of translational activators in the mTOR pathway such as eIF4E and eukaryotic elongation factor 1A (eEF1A) early during L-LTP causing increased induction. Furthermore, inhibition of the proteasome causes a buildup of translational repressors, such as polyadenylate-binding protein interacting protein 2 (Paip2) and eukaryotic initiation factor 4E-binding protein 2 (4E-BP2), during late stages of L-LTP contributing to the blockade of L-LTP maintenance. Thus, the proteasome plays a critical role in regulating protein synthesis during L-LTP by tightly controlling translation. Our results provide novel mechanistic insights into the interplay between protein degradation and protein synthesis in long-term synaptic plasticity. PMID:24573276

  20. Positive regulation of osteoclastic differentiation by growth differentiation factor 15 upregulated in osteocytic cells under hypoxia.

    PubMed

    Hinoi, Eiichi; Ochi, Hiroki; Takarada, Takeshi; Nakatani, Eri; Iezaki, Takashi; Nakajima, Hiroko; Fujita, Hiroyuki; Takahata, Yoshifumi; Hidano, Shinya; Kobayashi, Takashi; Takeda, Shu; Yoneda, Yukio

    2012-04-01

    Osteocytes are thought to play a role as a mechanical sensor through their communication network in bone. Although osteocytes are the most abundant cells in bone, little attention has been paid to their physiological and pathological functions in skeletogenesis. Here, we have attempted to delineate the pivotal functional role of osteocytes in regulation of bone remodeling under pathological conditions. We first found markedly increased osteoclastic differentiation by conditioned media (CM) from osteocytic MLO-Y4 cells previously exposed to hypoxia in vitro. Using microarray and real-time PCR analyses, we identified growth differentiation factor 15 (GDF15) as a key candidate factor secreted from osteocytes under hypoxia. Recombinant GDF15 significantly promoted osteoclastic differentiation in a concentration-dependent manner, with concomitant facilitation of phosphorylation of both p65 and inhibitory-κB in the presence of receptor activator of nuclear factor-κB ligand. To examine the possible functional significance of GDF15 in vivo, mice were subjected to ligation of the right femoral artery as a hypoxic model. A significant increase in GDF15 expression was specifically observed in tibias of the ligated limb but not in tibias of the normally perfused limb. Under these experimental conditions, in cancellous bone of proximal tibias in the ligated limb, a significant reduction was observed in bone volume, whereas a significant increase was seen in the extent of osteoclast surface/bone surface when determined by bone histomorphometric analysis. Finally, the anti-GDF15 antibody prevented bone loss through inhibiting osteoclastic activation in tibias from mice with femoral artery ligation in vivo, in addition to suppressing osteoclastic activity enhanced by CM from osteocytes exposed to hypoxia in vitro. These findings suggest that GDF15 could play a pivotal role in the pathogenesis of bone loss relevant to hypoxia through promotion of osteoclastogenesis after

  1. Pellino protein from pacific white shrimp Litopenaeus vannamei positively regulates NF-κB activation.

    PubMed

    Li, Chaozheng; Chai, Jiaoting; Li, Haoyang; Zuo, Hongliang; Wang, Sheng; Qiu, Wei; Weng, Shaoping; He, Jianguo; Xu, Xiaopeng

    2014-06-01

    Pellino, named after its property that binds Pelle (the Drosophila melanogaster homolog of IRAK1), is a highly conserved E3 class ubiquitin ligase in both vertebrates and invertebrates. Pellino interacts with phosphorylated IRAK1, causing polyubiquitination of IRAK1, and plays a critical upstream role in the toll-like receptor (TLR) pathway. In this study, we firstly cloned and identified a crustacean Pellino from pacific white shrimp Litopenaeus vannamei (LvPellino). LvPellino contains a putative N-terminal forkhead-associated (FHA) domain and a C-terminal ring finger (RING) domain with a potential E3 ubiquitin-protein ligase activity, and shows a high similarity with D. melanogaster Pellino. LvPellino could interact with L. vannamei Pelle (LvPelle) and over-expression of LvPellino could increase the activity of LvDorsal (a L. vannamei homolog of NF-κB) on promoters containing NF-κB binding motifs and enhance the expression of arthropod antimicrobial peptides (AMPs). The LvPellino protein was located in the cytoplasm and nucleus and LvPellino mRNA was detected in all the tissues examined and could be up-regulated after lipopolysaccharides, white spot syndrome virus (WSSV), Vibrio parahaemolyticus, and Staphylococcus aureus challenges, suggesting a stimulation response of LvPellino to bacterial and immune stimulant challenges. Knockdown of LvPellino in vivo could significantly decrease the expression of AMPs and increase the mortality of shrimps caused by V. parahaemolyticus challenge. However, suppression of the LvPellino expression could not change the mortality caused by WSSV infection, and dual-luciferase reporter assays demonstrated that over-expression of LvPellino could enhance the promoters of WSSV genes wsv069 (ie1), wsv303, and wsv371, indicating a complex role of LvPellino in WSSV pathogenesis and shrimp antiviral mechanisms.

  2. Maize acetylcholinesterase is a positive regulator of heat tolerance in plants.

    PubMed

    Yamamoto, Kosuke; Sakamoto, Hikaru; Momonoki, Yoshie S

    2011-11-01

    We previously reported that native tropical zone plants showed high acetylcholinesterase (AChE) activity during heat stress, and that AChE activity in endodermal cells of maize seedlings was increased by heat treatment. However, the physiological role of AChE in heat stressed plants is still unclear. Here we report (1) tissue-specific expression and subcellular localization of maize AChE, (2) elevation of AChE activity and possible post-translational modifications of this enzyme under heat stress, and (3) involvement of AChE in plant heat stress tolerance. Maize AChE was mainly expressed in coleoptile nodes and seeds. Maize AChE fused with green fluorescent protein (GFP) was localized in extracellular spaces of transgenic rice plants. Therefore, in maize coleoptile nodes and seeds AChE mainly functions in the cell wall matrix. After heat treatment, enhanced maize AChE activity was observed by in vitro activity measurement and by in situ cytochemical staining; transcript and protein levels, however, were not changed. Protein gel blot analysis revealed two AChE isoforms (upper and lower); the upper-form gradually disappeared after heat treatment. Thus, maize AChE activity might be enhanced through a post-translational modification response to heat stress. Finally, we found that overexpression of maize AChE in transgenic tobacco plants enhanced heat tolerance relative to that of non-transgenic plants, suggesting AChE plays a positive role in maize heat tolerance.

  3. ATG7 regulates energy metabolism, differentiation and survival of Philadelphia-chromosome-positive cells

    PubMed Central

    Karvela, Maria; Baquero, Pablo; Kuntz, Elodie M.; Mukhopadhyay, Arunima; Mitchell, Rebecca; Allan, Elaine K.; Chan, Edmond; Kranc, Kamil R.; Calabretta, Bruno; Salomoni, Paolo; Gottlieb, Eyal; Holyoake, Tessa L.; Helgason, G. Vignir

    2016-01-01

    ABSTRACT A major drawback of tyrosine kinase inhibitor (TKI) treatment in chronic myeloid leukemia (CML) is that primitive CML cells are able to survive TKI-mediated BCR-ABL inhibition, leading to disease persistence in patients. Investigation of strategies aiming to inhibit alternative survival pathways in CML is therefore critical. We have previously shown that a nonspecific pharmacological inhibition of autophagy potentiates TKI-induced death in Philadelphia chromosome-positive cells. Here we provide further understanding of how specific and pharmacological autophagy inhibition affects nonmitochondrial and mitochondrial energy metabolism and reactive oxygen species (ROS)-mediated differentiation of CML cells and highlight ATG7 (a critical component of the LC3 conjugation system) as a potential specific therapeutic target. By combining extra- and intracellular steady state metabolite measurements by liquid chromatography-mass spectrometry with metabolic flux assays using labeled glucose and functional assays, we demonstrate that knockdown of ATG7 results in decreased glycolysis and increased flux of labeled carbons through the mitochondrial tricarboxylic acid cycle. This leads to increased oxidative phosphorylation and mitochondrial ROS accumulation. Furthermore, following ROS accumulation, CML cells, including primary CML CD34+ progenitor cells, differentiate toward the erythroid lineage. Finally, ATG7 knockdown sensitizes CML progenitor cells to TKI-induced death, without affecting survival of normal cells, suggesting that specific inhibitors of ATG7 in combination with TKI would provide a novel therapeutic approach for CML patients exhibiting persistent disease. PMID:27168493

  4. Protection of Armadillo/β-Catenin by Armless, a Novel Positive Regulator of Wingless Signaling

    PubMed Central

    Reim, Gerlinde; Hruzova, Martina; Goetze, Sandra; Basler, Konrad

    2014-01-01

    The Wingless (Wg/Wnt) signaling pathway is essential for metazoan development, where it is central to tissue growth and cellular differentiation. Deregulated Wg pathway activation underlies severe developmental abnormalities, as well as carcinogenesis. Armadillo/β-Catenin plays a key role in the Wg transduction cascade; its cytoplasmic and nuclear levels directly determine the output activity of Wg signaling and are thus tightly controlled. In all current models, once Arm is targeted for degradation by the Arm/β-Catenin destruction complex, its fate is viewed as set. We identified a novel Wg/Wnt pathway component, Armless (Als), which is required for Wg target gene expression in a cell-autonomous manner. We found by genetic and biochemical analyses that Als functions downstream of the destruction complex, at the level of the SCF/Slimb/βTRCP E3 Ub ligase. In the absence of Als, Arm levels are severely reduced. We show by biochemical and in vivo studies that Als interacts directly with Ter94, an AAA ATPase known to associate with E3 ligases and to drive protein turnover. We suggest that Als antagonizes Ter94's positive effect on E3 ligase function and propose that Als promotes Wg signaling by rescuing Arm from proteolytic degradation, spotlighting an unexpected step where the Wg pathway signal is modulated. PMID:25369031

  5. CSR-1 RNAi pathway positively regulates histone expression in C. elegans.

    PubMed

    Avgousti, Daphne C; Palani, Santhosh; Sherman, Yekaterina; Grishok, Alla

    2012-10-03

    Endogenous small interfering RNAs (endo-siRNAs) have been discovered in many organisms, including mammals. In C. elegans, depletion of germline-enriched endo-siRNAs found in complex with the CSR-1 Argonaute protein causes sterility and defects in chromosome segregation in early embryos. We discovered that knockdown of either csr-1, the RNA-dependent RNA polymerase (RdRP) ego-1, or the dicer-related helicase drh-3, leads to defects in histone mRNA processing, resulting in severe depletion of core histone proteins. The maturation of replication-dependent histone mRNAs, unlike that of other mRNAs, requires processing of their 3'UTRs through an endonucleolytic cleavage guided by the U7 snRNA, which is lacking in C. elegans. We found that CSR-1-bound antisense endo-siRNAs match histone mRNAs and mRNA precursors. Consistently, we demonstrate that CSR-1 directly binds to histone mRNA in an ego-1-dependent manner using biotinylated 2'-O-methyl RNA oligonucleotides. Moreover, we demonstrate that increasing the dosage of histone genes rescues the lethality associated with depletion of CSR-1 and EGO-1. These results support a positive and direct effect of RNAi on histone gene expression.

  6. The transcription factor CREBZF is a novel positive regulator of p53

    PubMed Central

    López-Mateo, Irene; Villaronga, M. Ángeles; Llanos, Susana; Belandia, Borja

    2012-01-01

    CREBZF is a member of the mammalian ATF/CREB family of transcription factors. Here, we describe a novel functional interaction between CREBZF and the tumor suppressor p53. CREBZF was identified in a yeast two-hybrid screen using HEY1, recently characterized as an indirect p53 activator, as bait. CREBZF interacts in vitro with both HEY1 and p53, and CREBZF expression stabilizes and activates p53. Moreover, CREBZF cooperates synergistically with HEY1 to enhance p53 transcriptional activity. On the other hand, partial depletion of endogenous CREBZF diminishes p53 protein levels and inhibits HEY1-mediated activation of p53. CREBZF-positive effects on p53 signaling may reflect, at least in part, an observed induction of posttranslational modifications in p53 known to prevent its degradation. CREBZF expression protects HCT116 cells from UV radiation-induced cell death. In addition, CREBZF expression confers sensitivity to 5-fluorouracil, a p53-activating chemotherapeutic drug. Our study suggests that CREBZF may participate in the modulation of p53 tumor suppressor function. PMID:22983008

  7. ATR controls cellular adaptation to hypoxia through positive regulation of hypoxia-inducible factor 1 (HIF-1) expression.

    PubMed

    Fallone, F; Britton, S; Nieto, L; Salles, B; Muller, C

    2013-09-12

    Tumor cells adaptation to severe oxygen deprivation (hypoxia) plays a major role in tumor progression. The transcription factor HIF-1 (hypoxia-inducible factor 1), whose α-subunit is stabilized under hypoxic conditions is a key component of this process. Recent studies showed that two members of the phosphoinositide 3-kinase-related kinases (PIKKs) family, ATM (ataxia telangiectasia mutated) and DNA-PK (DNA-dependent protein kinase), regulate the hypoxic-dependent accumulation of HIF-1. These proteins initiate cellular stress responses when DNA damage occurs. In addition, it has been demonstrated that extreme hypoxia induces a replicative stress resulting in regions of single-stranded DNA at stalled replication forks and the activation of ATR (ataxia telangiectasia and Rad3 related protein), another member of the PIKKs family. Here, we show that even less severe hypoxia (0.1% O2) also induces activation of ATR through replicative stress. Importantly, in using either transiently silenced ATR cells, cells expressing an inactive form of ATR or cells exposed to an ATR inhibitor (CGK733), we demonstrate that hypoxic ATR activation positively regulates the key transcription factor HIF-1 independently of the checkpoint kinase Chk1. We show that ATR kinase activity regulates HIF-1α at the translational level and we find that the elements necessary for the regulation of HIF-1α translation are located within the coding region of HIF-1α mRNA. Finally, by using three independent cellular models, we clearly show that the loss of ATR expression and/or kinase activity results in the decrease of HIF-1 DNA binding under hypoxia and consequently affects protein expression levels of two HIF-1 target genes, GLUT-1 and CAIX. Taken together, our data show a new function for ATR in cellular adaptation to hypoxia through regulation of HIF-1α translation. Our work offers new prospect for cancer therapy using ATR inhibitors with the potential to decrease cellular adaptation in hypoxic

  8. Lysosome-associated membrane proteins (LAMPs) regulate intracellular positioning of mitochondria in MC3T3-E1 cells.

    PubMed

    Rajapakshe, Anupama R; Podyma-Inoue, Katarzyna A; Terasawa, Kazue; Hasegawa, Katsuya; Namba, Toshimitsu; Kumei, Yasuhiro; Yanagishita, Masaki; Hara-Yokoyama, Miki

    2015-02-01

    The intracellular positioning of both lysosomes and mitochondria meets the requirements of degradation and energy supply, which are respectively the two major functions for cellular maintenance. The positioning of both lysosomes and mitochondria is apparently affected by the nutrient status of the cells. However, the mechanism coordinating the positioning of the organelles has not been sufficiently elucidated. Lysosome-associated membrane proteins-1 and -2 (LAMP-1 and LAMP-2) are highly glycosylated proteins that are abundant in lysosomal membranes. In the present study, we demonstrated that the siRNA-mediated downregulation of LAMP-1, LAMP-2 or their combination enhanced the perinuclear localization of mitochondria, in the pre-osteoblastic cell line MC3T3-E1. On the other hand, in the osteocytic cell line MLO-Y4, in which both the lysosomes and mitochondria originally accumulate in the perinuclear region and mitochondria also fill dendrites, the effect of siRNA of LAMP-1 or LAMP-2 was barely observed. LAMPs are not directly associated with mitochondria, and there do not seem to be any accessory molecules commonly required to recruit the motor proteins to lysosomes and mitochondria. Our results suggest that LAMPs may regulate the positioning of lysosomes and mitochondria. A possible mechanism involving the indirect and context-dependent action of LAMPs is discussed.

  9. Positive density-dependent reproduction regulated by local kinship and size in an understorey tropical tree

    PubMed Central

    Castilla, Antonio R.; Pope, Nathaniel; Jha, Shalene

    2016-01-01

    Background and Aims Global pollinator declines and continued habitat fragmentation highlight the critical need to understand reproduction and gene flow across plant populations. Plant size, conspecific density and local kinship (i.e. neighbourhood genetic relatedness) have been proposed as important mechanisms influencing the reproductive success of flowering plants, but have rarely been simultaneously investigated. Methods We conducted this study on a continuous population of the understorey tree Miconia affinis in the Forest Dynamics Plot on Barro Colorado Island in central Panama. We used spatial, reproductive and population genetic data to investigate the effects of tree size, conspecific neighbourhood density and local kinship on maternal and paternal reproductive success. We used a Bayesian framework to simultaneously model the effects of our explanatory variables on the mean and variance of maternal viable seed set and siring success. Key Results Our results reveal that large trees had lower proportions of viable seeds in their fruits but sired more seeds. We documented differential effects of neighbourhood density and local kinship on both maternal and paternal reproductive components. Trees in more dense neighbourhoods produced on average more viable seeds, although this positive density effect was influenced by variance-inflation with increasing local kinship. Neighbourhood density did not have significant effects on siring success. Conclusions This study is one of the first to reveal an interaction among tree size, conspecific density and local kinship as critical factors differentially influencing maternal and paternal reproductive success. We show that both maternal and paternal reproductive success should be evaluated to determine the population-level and individual traits most essential for plant reproduction. In addition to conserving large trees, we suggest the inclusion of small trees and the conservation of dense patches with low kinship as

  10. Sox2 regulates cholinergic amacrine cell positioning and dendritic stratification in the retina.

    PubMed

    Whitney, Irene E; Keeley, Patrick W; St John, Ace J; Kautzman, Amanda G; Kay, Jeremy N; Reese, Benjamin E

    2014-07-23

    The retina contains two populations of cholinergic amacrine cells, one positioned in the ganglion cell layer (GCL) and the other in the inner nuclear layer (INL), that together comprise ∼1/2 of a percent of all retinal neurons. The present study examined the genetic control of cholinergic amacrine cell number and distribution between these two layers. The total number of cholinergic amacrine cells was quantified in the C57BL/6J and A/J inbred mouse strains, and in 25 recombinant inbred strains derived from them, and variations in their number and ratio (GCL/INL) across these strains were mapped to genomic loci. The total cholinergic amacrine cell number was found to vary across the strains, from 27,000 to 40,000 cells, despite little variation within individual strains. The number of cells was always lower within the GCL relative to the INL, and the sizes of the two populations were strongly correlated, yet there was variation in their ratio between the strains. Approximately 1/3 of that variation in cell ratio was mapped to a locus on chromosome 3, where Sex determining region Y box 2 (Sox2) was identified as a candidate gene due to the presence of a 6-nucleotide insertion in the protein-coding sequence in C57BL/6J and because of robust and selective expression in cholinergic amacrine cells. Conditionally deleting Sox2 from the population of nascent cholinergic amacrine cells perturbed the normal ratio of cells situated in the GCL versus the INL and induced a bistratifying morphology, with dendrites distributed to both ON and OFF strata within the inner plexiform layer.

  11. A super-family of transcriptional activators regulates bacteriophage packaging and lysis in Gram-positive bacteria.

    PubMed

    Quiles-Puchalt, Nuria; Tormo-Más, María Ángeles; Campoy, Susana; Toledo-Arana, Alejandro; Monedero, Vicente; Lasa, Iñigo; Novick, Richard P; Christie, Gail E; Penadés, José R

    2013-08-01

    The propagation of bacteriophages and other mobile genetic elements requires exploitation of the phage mechanisms involved in virion assembly and DNA packaging. Here, we identified and characterized four different families of phage-encoded proteins that function as activators required for transcription of the late operons (morphogenetic and lysis genes) in a large group of phages infecting Gram-positive bacteria. These regulators constitute a super-family of proteins, here named late transcriptional regulators (Ltr), which share common structural, biochemical and functional characteristics and are unique to this group of phages. They are all small basic proteins, encoded by genes present at the end of the early gene cluster in their respective phage genomes and expressed under cI repressor control. To control expression of the late operon, the Ltr proteins bind to a DNA repeat region situated upstream of the terS gene, activating its transcription. This involves the C-terminal part of the Ltr proteins, which control specificity for the DNA repeat region. Finally, we show that the Ltr proteins are the only phage-encoded proteins required for the activation of the packaging and lysis modules. In summary, we provide evidence that phage packaging and lysis is a conserved mechanism in Siphoviridae infecting a wide variety of Gram-positive bacteria.

  12. A super-family of transcriptional activators regulates bacteriophage packaging and lysis in Gram-positive bacteria

    PubMed Central

    Quiles-Puchalt, Nuria; Tormo-Más, María Ángeles; Campoy, Susana; Toledo-Arana, Alejandro; Monedero, Vicente; Lasa, Íñigo; Novick, Richard P.; Christie, Gail E.; Penadés, José R.

    2013-01-01

    The propagation of bacteriophages and other mobile genetic elements requires exploitation of the phage mechanisms involved in virion assembly and DNA packaging. Here, we identified and characterized four different families of phage-encoded proteins that function as activators required for transcription of the late operons (morphogenetic and lysis genes) in a large group of phages infecting Gram-positive bacteria. These regulators constitute a super-family of proteins, here named late transcriptional regulators (Ltr), which share common structural, biochemical and functional characteristics and are unique to this group of phages. They are all small basic proteins, encoded by genes present at the end of the early gene cluster in their respective phage genomes and expressed under cI repressor control. To control expression of the late operon, the Ltr proteins bind to a DNA repeat region situated upstream of the terS gene, activating its transcription. This involves the C-terminal part of the Ltr proteins, which control specificity for the DNA repeat region. Finally, we show that the Ltr proteins are the only phage-encoded proteins required for the activation of the packaging and lysis modules. In summary, we provide evidence that phage packaging and lysis is a conserved mechanism in Siphoviridae infecting a wide variety of Gram-positive bacteria. PMID:23771138

  13. NUCLEAR FACTOR Y, Subunit A (NF-YA) Proteins Positively Regulate Flowering and Act Through FLOWERING LOCUS T

    PubMed Central

    Siriwardana, Chamindika L.; Kumimoto, Roderick W.; Mantovani, Roberto

    2016-01-01

    Photoperiod dependent flowering is one of several mechanisms used by plants to initiate the developmental transition from vegetative growth to reproductive growth. The NUCLEAR FACTOR Y (NF-Y) transcription factors are heterotrimeric complexes composed of NF-YA and histone-fold domain (HFD) containing NF-YB/NF-YC, that initiate photoperiod-dependent flowering by cooperatively interacting with CONSTANS (CO) to drive the expression of FLOWERING LOCUS T (FT). This involves NF-Y and CO binding at distal CCAAT and proximal “CORE” elements, respectively, in the FT promoter. While this is well established for the HFD subunits, there remains some question over the potential role of NF-YA as either positive or negative regulators of this process. Here we provide strong support, in the form of genetic and biochemical analyses, that NF-YA, in complex with NF-YB/NF-YC proteins, can directly bind the distal CCAAT box in the FT promoter and are positive regulators of flowering in an FT-dependent manner. PMID:27977687

  14. The Late Positive Potential Predicts Emotion Regulation Strategy Use in School-Aged Children Concurrently and Two Years Later

    PubMed Central

    Babkirk, Sarah; Rios, Victor; Dennis, Tracy A.

    2014-01-01

    The ability to use cognitive emotion regulation strategies such as reappraisal may be a core component of emotional competence across development, but due to methodological challenges in measuring such strategies, they are rarely studied in young children. One neurophysiological measure, the late positive potential (LPP) has been examined in response to reappraisal as a potential neurosignature for emotion regulatory capacity in adults. The association between the LPP and emotion regulatory capacity in children is unknown. The present study examined whether the LPP during reappraisal could predict greater observed adaptive emotion regulation strategy use in school-aged children over a two-year period. Thirty-two five- to seven- year- olds participated in two identical lab visits spaced two years apart. EEG was continuously recorded during a computerized reappraisal task in which children viewed unpleasant images paired with either reappraisal or negative stories. Next they completed a disappointing and a frustrating task during which emotion regulation strategies was observed. As predicted, children who showed reappraisal-induced reductions in the LPP at the first assessment used significantly more adaptive ER strategies concurrently and two years later. These findings provide observation-based evidence that the LPP may be a viable neurosignature for emotion regulatory capacity in children. PMID:25438825

  15. PHYTOCHROME-INTERACTING FACTOR 5 (PIF5) positively regulates dark-induced senescence and chlorophyll degradation in Arabidopsis.

    PubMed

    Zhang, Yongqiang; Liu, Zhongjuan; Chen, Yadi; He, Jun-Xian; Bi, Yurong

    2015-08-01

    Darkness is a known environmental factor that induces plant senescence. Here, Phytochrome-Interacting Factors (PIFs), several bHLH transcription factors involved in plant skotomorphogenesis, were examined for their roles in the regulation of dark-induced senescence and chlorophyll breakdown in Arabidopsis thaliana. After light-grown seedlings were transferred to darkness, green leaves turned yellow, and chlorophyll contents decreased, but membrane lipid peroxidation and cell death increased in wild-type Col-0. These responses were enhanced in overexpression line PIF5OX but decreased in mutant pif5-3. Darkness significantly induced expression of several genes involved in chlorophyll breakdown, including SGR, NYC1, NOL, and PAO, as well as genes encoding for transcription factors that have been shown to be required for dark-induced senescence, including WRKY22, NAP, EIN3, EIL1, and ORE1. These effects on gene expression were also enhanced in PIF5OX but decreased in pif5-3 relative to Col-0. Further analyses using ChIP-qPCR, EMSA, and protoplast transient assays indicated that PIF5 binds to the G-box motifs in the promoters of SGR, NYC1, and ORE1 genes and stimulate their expression. Collectively, our data indicate that PIF5 is a key factor that positively regulates dark-induced senescence upstream of ORE1 and regulates chlorophyll breakdown upstream of SGR and NYC1.

  16. OsSIDP366, a DUF1644 gene, positively regulates responses to drought and salt stresses in rice.

    PubMed

    Guo, Chiming; Luo, Chengke; Guo, Lijia; Li, Min; Guo, Xiaoling; Zhang, Yuxia; Wang, Liangjiang; Chen, Liang

    2016-05-01

    Domain of unknown function 1644 (DUF1644) is a highly conserved amino acid sequence motif present only in plants. Analysis of expression data of the family of DUF1644-containing genes indicated that they may regulate responses to abiotic stress in rice. Here we present our discovery of the role of OsSIDP366, a member of the DUF1644 gene family, in response to drought and salinity stresses in rice. Transgenic rice plants overexpressing OsSIDP366 showed enhanced drought and salinity tolerance and reduced water loss as compared to that in the control, whereas plants with downregulated OsSIDP366 expression levels using RNA interference (RNAi) were more sensitive to salinity and drought treatments. The sensitivity to abscisic acid (ABA) treatment was not changed in OsSIDP366-overexpressing plants, and OsSIDP366 expression was not affected in ABA-deficient mutants. Subcellular localization analysis revealed that OsSIDP366 is presented in the cytoplasmic foci that colocalized with protein markers for both processing bodies (PBs) and stress granules (SGs) in rice protoplasts. Digital gene expression (DGE) profile analysis indicated that stress-related genes such as SNAC1, OsHAK5 and PRs were upregulated in OsSIDP366-overexpressing plants. These results suggest that OsSIDP366 may function as a regulator of the PBs/SGs and positively regulate salt and drought resistance in rice.

  17. Arabidopsis DE-ETIOLATED1 represses photomorphogenesis by positively regulating phytochrome-interacting factors in the dark.

    PubMed

    Dong, Jie; Tang, Dafang; Gao, Zhaoxu; Yu, Renbo; Li, Kunlun; He, Hang; Terzaghi, William; Deng, Xing Wang; Chen, Haodong

    2014-09-01

    Arabidopsis thaliana seedlings undergo photomorphogenic development even in darkness when the function of DE-ETIOLATED1 (DET1), a repressor of photomorphogenesis, is disrupted. However, the mechanism by which DET1 represses photomorphogenesis remains unclear. Our results indicate that DET1 directly interacts with a group of transcription factors known as the phytochrome-interacting factors (PIFs). Furthermore, our results suggest that DET1 positively regulates PIF protein levels primarily by stabilizing PIF proteins in the dark. Genetic analysis showed that each pif single mutant could enhance the det1-1 phenotype, and ectopic expression of each PIF in det1-1 partially suppressed the det1-1 phenotype, based on hypocotyl elongation and cotyledon opening angles observed in darkness. Genomic analysis also revealed that DET1 may modulate the expression of light-regulated genes to mediate photomorphogenesis partially through PIFs. The observed interaction and regulation between DET1 and PIFs not only reveal how DET1 represses photomorphogenesis, but also suggest a possible mechanism by which two groups of photomorphogenic repressors, CONSTITUTIVE PHOTOMORPHOGENESIS/DET/FUSCA and PIFs, work in concert to repress photomorphogenesis in darkness.

  18. The chromatin remodeling complex NuRD establishes the poised state of rRNA genes characterized by bivalent histone modifications and altered nucleosome positions.

    PubMed

    Xie, Wenbing; Ling, Te; Zhou, Yonggang; Feng, Weijun; Zhu, Qiaoyun; Stunnenberg, Henk G; Grummt, Ingrid; Tao, Wei

    2012-05-22

    rRNA genes (rDNA) exist in two distinct epigenetic states, active promoters being unmethylated and marked by euchromatic histone modifications, whereas silent ones are methylated and exhibit heterochromatic features. Here we show that the nucleosome remodeling and deacetylation (NuRD) complex establishes a specific chromatin structure at rRNA genes that are poised for transcription activation. The promoter of poised rRNA genes is unmethylated, associated with components of the preinitiation complex, marked by bivalent histone modifications and covered by a nucleosome in the "off" position, which is refractory to transcription initiation. Repression of rDNA transcription in growth-arrested and differentiated cells correlates with elevated association of NuRD and increased levels of poised rRNA genes. Reactivation of transcription requires resetting the promoter-bound nucleosome into the "on" position by the DNA-dependent ATPase CSB (Cockayne syndrome protein B). The results uncover a unique mechanism by which ATP-dependent chromatin remodeling complexes with opposing activities establish a specific chromatin state and regulate transcription.

  19. A Positive Regulatory Loop Controls Expression of the Locus of Enterocyte Effacement-Encoded Regulators Ler and GrlA

    PubMed Central

    Barba, Jeannette; Bustamante, Víctor H.; Flores-Valdez, Mario A.; Deng, Wanyin; Finlay, B. Brett; Puente, José L.

    2005-01-01

    The formation of attaching and effacing (A/E) lesions on intestinal epithelial cells is an essential step in the pathogenesis of human enteropathogenic and enterohemorrhagic Escherichia coli and of the mouse pathogen Citrobacter rodentium. The genes required for the development of the A/E phenotype are located within a pathogenicity island known as the locus of enterocyte effacement (LEE). The LEE-encoded transcriptional regulators Ler, an H-NS-like protein, and GrlA, a member of a novel family of transcriptional activators, positively control the expression of the genes located in the LEE and their corresponding virulence. In this study, we used C. rodentium as a model to study the mechanisms controlling the expression of Ler and GrlA. By deletion analysis of the ler and grlRA regulatory regions and complementation experiments, negative and positive cis-acting regulatory motifs were identified that are essential for the regulation of both genes. This analysis confirmed that GrlA is required for the activation of ler, but it also showed that Ler is required for the expression of grlRA, revealing a novel regulatory loop controlling the optimal expression of virulence genes in A/E pathogens. Furthermore, our results indicate that Ler and GrlA induce the expression of each other by, at least in part, counteracting the repression mediated by H-NS. However, whereas GrlA is still required for the optimal expression of ler even in the absence of H-NS, Ler is not needed for the expression of grlRA in the absence of H-NS. This type of transcriptional positive regulatory loop represents a novel mechanism in pathogenic bacteria that is likely required to maintain an appropriate spatiotemporal transcriptional response during infection. PMID:16291665

  20. Engagement of the T-cell receptor during positive selection in the thymus down-regulates RAG-1 expression.

    PubMed Central

    Brändle, D; Müller, C; Rülicke, T; Hengartner, H; Pircher, H

    1992-01-01

    We have examined the expression of the recombination activating gene RAG-1 by in situ hybridization to thymi from mice bearing transgenes for the T-cell receptor (TCR) alpha chain, TCR beta chain, or both TCR alpha and beta chains. RAG-1 transcription was found in the thymic cortex of transgenic mice carrying a single TCR alpha- or TCR beta-chain transgene, comparable to normal mice. However, RAG-1 transcription was strikingly reduced in the thymic cortex from transgenic mice carrying both TCR alpha- and beta-chain genes and expressing major histocompatibility complex (MHC) class I (H-2b) molecules necessary for positive selection of the transgenic TCR. In contrast, thymi of transgenic mice also carrying both TCR alpha- and beta-chain genes but expressing MHC molecules (H-2d) that did not positively select the transgenic TCR displayed high levels of RAG-1 transcription. The low thymic RAG-1 expression coincided with high transgenic TCR alpha-chain surface expression and with inhibition of endogenous TCR alpha-chain rearrangement. Our findings suggest that binding of the TCR to self MHC molecules during positive selection down-regulates RAG-1 transcription in cortical thymocytes and thereby prevents further TCR alpha-chain rearrangements. Images PMID:1329099

  1. Regulation of nitrogen fixation in Azorhizobium caulinodans: identification of a fixK-like gene, a positive regulator of nifA.

    PubMed

    Kaminski, P A; Mandon, K; Arigoni, F; Desnoues, N; Elmerich, C

    1991-08-01

    The nucleotide sequence of a 1 kb fragment upstream of Azorhizobium caulinodans fixL was established. An open reading frame of 744 bp was identified as a fixK homologue. A kanamycin cartridge was inserted into the cloned fixK-like gene and recombined into the host genome. The resulting mutant was Nif-Fix-, suggesting that FixK was required for nitrogen fixation both in symbiotic conditions and in the free-living state. Using a pfixK-lacZ fusion, the FixLJ products were shown to control the expression of fixK. Using a pnifA-lacZ fusion, the FixK product was shown to regulate positively the transcription of nifA in bacteria grown in the free-living state. In addition, a double ntrC-fixL mutant was constructed and was shown to be completely devoid of nitrogenase activity. A model of regulation, based on these data, is presented and might explain the unusual ability of A. caulinodans to fix nitrogen both under symbiotic conditions and in the free-living state.

  2. Iron-Regulated Protein HupB of Mycobacterium tuberculosis Positively Regulates Siderophore Biosynthesis and Is Essential for Growth in Macrophages

    PubMed Central

    Pandey, Satya Deo; Choudhury, Mitali; Yousuf, Suhail; Wheeler, Paul R.; Gordon, Stephen V.; Ranjan, Akash

    2014-01-01

    Mycobacterium tuberculosis expresses the 28-kDa protein HupB (Rv2986c) and the Fe3+-specific high-affinity siderophores mycobactin and carboxymycobactin upon iron limitation. The objective of this study was to understand the functional role of HupB in iron acquisition. A hupB mutant strain of M. tuberculosis, subjected to growth in low-iron medium (0.02 μg Fe ml−1), showed a marked reduction of both siderophores with low transcript levels of the mbt genes encoding the MB biosynthetic machinery. Complementation of the mutant strain with hupB restored siderophore production to levels comparable to that of the wild type. We demonstrated the binding of HupB to the mbtB promoter by both electrophoretic mobility shift assays and DNA footprinting. The latter revealed the HupB binding site to be a 10-bp AT-rich region. While negative regulation of the mbt machinery by IdeR is known, this is the first report of positive regulation of the mbt operon by HupB. Interestingly, the mutant strain failed to survive inside macrophages, suggesting that HupB plays an important role in vivo. PMID:24610707

  3. Expression of the alaE gene is positively regulated by the global regulator Lrp in response to intracellular accumulation of l-alanine in Escherichia coli.

    PubMed

    Ihara, Kohei; Sato, Kazuki; Hori, Hatsuhiro; Makino, Yumiko; Shigenobu, Shuji; Ando, Tasuke; Isogai, Emiko; Yoneyama, Hiroshi

    2017-04-01

    The alaE gene in Escherichia coli encodes an l-alanine exporter that catalyzes the active export of l-alanine using proton electrochemical potential. In our previous study, alaE expression was shown to increase in the presence of l-alanyl-l-alanine (Ala-Ala). In this study, the global regulator leucine-responsive regulatory protein (Lrp) was identified as an activator of the alaE gene. A promoter less β-galactosidase gene was fused to an alaE upstream region (240 nucleotides). Cells that were lacZ-deficient and harbored this reporter plasmid showed significant induction of β-galactosidase activity (approximately 17-fold) in the presence of 6 mM l-alanine, l-leucine, and Ala-Ala. However, a reporter plasmid possessing a smaller alaE upstream region (180 nucleotides) yielded transformants with strikingly low enzyme activity under the same conditions. In contrast, lrp-deficient cells showed almost no β-galactosidase induction, indicating that Lrp positively regulates alaE expression. We next performed an electrophoretic mobility shift assay (EMSA) and a DNase I footprinting assay using purified hexahistidine-tagged Lrp (Lrp-His). Consequently, we found that Lrp-His binds to the alaE upstream region spanning nucleotide -161 to -83 with a physiologically relevant affinity (apparent KD, 288.7 ± 83.8 nM). Furthermore, the binding affinity of Lrp-His toward its cis-element was increased by l-alanine and l-leucine, but not by Ala-Ala and d-alanine. Based on these results, we concluded that the gene expression of the alaE is regulated by Lrp in response to intracellular levels of l-alanine, which eventually leads to intracellular homeostasis of l-alanine concentrations.

  4. Position-dependent activity of CELF2 in the regulation of splicing and implications for signal-responsive regulation in T cells

    PubMed Central

    Ajith, Sandya; Gazzara, Matthew R.; Cole, Brian S.; Shankarling, Ganesh; Martinez, Nicole M.; Mallory, Michael J.; Lynch, Kristen W.

    2016-01-01

    ABSTRACT CELF2 is an RNA binding protein that has been implicated in developmental and signal-dependent splicing in the heart, brain and T cells. In the heart, CELF2 expression decreases during development, while in T cells CELF2 expression increases both during development and in response to antigen-induced signaling events. Although hundreds of CELF2-responsive splicing events have been identified in both heart and T cells, the way in which CELF2 functions has not been broadly investigated. Here we use CLIP-Seq to identified physical targets of CELF2 in a cultured human T cell line. By comparing the results with known functional targets of CELF2 splicing regulation from the same cell line we demonstrate a generalizable position-dependence of CELF2 activity that is consistent with previous mechanistic studies of individual CELF2 target genes in heart and brain. Strikingly, this general position-dependence is sufficient to explain the bi-directional activity of CELF2 on 2 T cell targets recently reported. Therefore, we propose that the location of CELF2 binding around an exon is a primary predictor of CELF2 function in a broad range of cellular contexts. PMID:27096301

  5. Positive and Negative Regulation of Muscle Cell Identity by Members of the hedgehog and TGF-β Gene Families

    PubMed Central

    Du, Shao Jun; Devoto, Stephen H.; Westerfield, Monte; Moon, Randall T.

    1997-01-01

    We have examined whether the development of embryonic muscle fiber type is regulated by competing influences between Hedgehog and TGF-β signals, as previously shown for development of neuronal cell identity in the neural tube. We found that ectopic expression of Hedgehogs or inhibition of protein kinase A in zebrafish embryos induces slow muscle precursors throughout the somite but muscle pioneer cells only in the middle of the somite. Ectopic expression in the notochord of Dorsalin-1, a member of the TGF-β superfamily, inhibits the formation of muscle pioneer cells, demonstrating that TGF-β signals can antagonize the induction of muscle pioneer cells by Hedgehog. We propose that a Hedgehog signal first induces the formation of slow muscle precursor cells, and subsequent Hedgehog and TGF-β signals exert competing positive and negative influences on the development of muscle pioneer cells. PMID:9314535

  6. miR-21-3p is a positive regulator of L1CAM in several human carcinomas.

    PubMed

    Doberstein, Kai; Bretz, Niko P; Schirmer, Uwe; Fiegl, Heidi; Blaheta, Roman; Breunig, Christian; Müller-Holzner, Elisabeth; Reimer, Dan; Zeimet, Alain G; Altevogt, Peter

    2014-11-28

    Expression of L1 cell adhesion molecule (L1CAM) occurs frequently in human cancers and is associated with poor prognosis in cancers such as ovarian, endometrial, breast, renal cell carcinoma and pancreatic ductal adenocarcinoma. L1CAM promotes cell motility, invasion, chemoresistance and metastasis formation. Elucidating genetic processes involved in the expression of L1CAM in cancers is of considerable importance. Transcription factors such as SLUG, β-catenin/TCF-LEF, PAX8 and VHL have been implicated in the re-activation of L1CAM in various types of cancers. There is increasing evidence that micro-RNAs can also have strong effects on gene expression. Here we have identified miR-21-3p as a positive regulator of L1CAM expression. Over-expression of miR-21-3p (miR-21*) but not the complementary sequence miR-21-5p (miR-21) could strongly augment L1CAM expression in renal, endometrial and ovarian carcinoma derived cell lines by an unknown mechanism involving transcriptional activation of the L1CAM gene. In patient cohorts from renal, endometrial and ovarian cancers we observed a strong positive correlation of L1CAM and miR-21-3p expressions. Although L1CAM alone was a reliable marker for overall and disease free survival, the combination of L1CAM and miR-21-3p expressions strongly enhanced the predictive power. Our findings shed new light on the complex regulation of L1CAM in cancers and advocate the use of L1CAM/miR-21-3p for diagnostic application.

  7. NF90 Binds the Dengue Virus RNA 3′ Terminus and Is a Positive Regulator of Dengue Virus Replication

    PubMed Central

    Gehrke, Lee

    2011-01-01

    Background Viral RNA translation and replication are regulated by sequence and structural elements in the 5′ and 3′ untranslated regions (UTR) and by host cell and/or viral proteins that bind them. Dengue virus has a single-stranded RNA genome with positive polarity, a 5′ m7GpppG cap, and a conserved 3′-terminal stem loop (SL) that is linked to proposed functions in viral RNA transcription and translation. Mechanisms explaining the contributions of host proteins to viral RNA translation and replication are poorly defined, yet understanding host protein-viral RNA interactions may identify new targets for therapeutic intervention. This study was directed at identifying functionally significant host proteins that bind the conserved dengue virus RNA 3′ terminus. Methodology/Principal Findings Proteins eluted from a dengue 3′ SL RNA affinity column at increasing ionic strength included two with double-strand RNA binding motifs (NF90/DRBP76 and DEAH box polypeptide 9/RNA helicase A (RHA)), in addition to NF45, which forms a heterodimer with NF90. Although detectable NF90 and RHA proteins localized to the nucleus of uninfected cells, immunofluorescence revealed cytoplasmic NF90 in dengue virus-infected cells, leading us to hypothesize that NF90 has a functional role(s) in dengue infections. Cells depleted of NF90 were used to quantify viral RNA transcript levels and production of infectious dengue virus. NF90 depletion was accompanied by a 50%-70% decrease in dengue RNA levels and in production of infectious viral progeny. Conclusions/Significance The results indicate that NF90 interacts with the 3′ SL structure of the dengue RNA and is a positive regulator of dengue virus replication. NF90 depletion diminished the production of infectious dengue virus by more than 50%, which may have important significance for identifying therapeutic targets to limit a virus that threatens more than a billion people worldwide. PMID:21386893

  8. Ataxin 2-binding protein 1 is a context-specific positive regulator of Notch signaling during neurogenesis in Drosophila melanogaster.

    PubMed

    Shukla, Jay Prakash; Deshpande, Girish; Shashidhara, L S

    2017-03-01

    The role of the Notch pathway during the lateral inhibition that underlies binary cell fate choice is extensively studied, but the context specificity that generates diverse outcomes is less well understood. In the peripheral nervous system of Drosophila melanogaster, differential Notch signaling between cells of the proneural cluster orchestrates sensory organ specification. Here we report functional analysis of Drosophila Ataxin 2-binding protein 1 (A2BP1) during this process. Its human ortholog is linked to type 2 spinocerebellar ataxia and other complex neuronal disorders. Downregulation of Drosophila A2BP1 in the proneural cluster increases adult sensory bristle number, whereas its overexpression results in loss of bristles. We show that A2BP1 regulates sensory organ specification by potentiating Notch signaling. Supporting its direct involvement, biochemical analysis shows that A2BP1 is part of the Suppressor of Hairless [Su(H)] complex in the presence and absence of Notch. However, in the absence of Notch signaling, the A2BP1 interacting fraction of Su(H) does not associate with the repressor proteins Groucho and CtBP. We propose a model explaining the requirement of A2BP1 as a positive regulator of context-specific Notch activity.

  9. Nel positively regulates the genesis of retinal ganglion cells by promoting their differentiation and survival during development.

    PubMed

    Nakamoto, Chizu; Kuan, Soh-Leh; Findlay, Amy S; Durward, Elaine; Ouyang, Zhufeng; Zakrzewska, Ewa D; Endo, Takuma; Nakamoto, Masaru

    2014-01-01

    For correct functioning of the nervous system, the appropriate number and complement of neuronal cell types must be produced during development. However, the molecular mechanisms that regulate the production of individual classes of neurons are poorly understood. In this study, we investigate the function of the thrombospondin-1-like glycoprotein, Nel (neural epidermal growth factor [EGF]-like), in the generation of retinal ganglion cells (RGCs) in chicks. During eye development, Nel is strongly expressed in the presumptive retinal pigment epithelium and RGCs. Nel overexpression in the developing retina by in ovo electroporation increases the number of RGCs, whereas the number of displaced amacrine cells decreases. Conversely, knockdown of Nel expression by transposon-mediated introduction of RNA interference constructs results in decrease in RGC number and increase in the number of displaced amacrine cells. Modifications of Nel expression levels do not appear to affect proliferation of retinal progenitor cells, but they significantly alter the progression rate of RGC differentiation from the central retina to the periphery. Furthermore, Nel protects RGCs from apoptosis during retinal development. These results indicate that Nel positively regulates RGC production by promoting their differentiation and survival during development.

  10. Characterisation of SalRAB a salicylic acid inducible positively regulated efflux system of Rhizobium leguminosarum bv viciae 3841.

    PubMed

    Tett, Adrian J; Karunakaran, Ramakrishnan; Poole, Philip S

    2014-01-01

    Salicylic acid is an important signalling molecule in plant-microbe defence and symbiosis. We analysed the transcriptional responses of the nitrogen fixing plant symbiont, Rhizobium leguminosarum bv viciae 3841 to salicylic acid. Two MFS-type multicomponent efflux systems were induced in response to salicylic acid, rmrAB and the hitherto undescribed system salRAB. Based on sequence similarity salA and salB encode a membrane fusion and inner membrane protein respectively. salAB are positively regulated by the LysR regulator SalR. Disruption of salA significantly increased the sensitivity of the mutant to salicylic acid, while disruption of rmrA did not. A salA/rmrA double mutation did not have increased sensitivity relative to the salA mutant. Pea plants nodulated by salA or rmrA strains did not have altered nodule number or nitrogen fixation rates, consistent with weak expression of salA in the rhizosphere and in nodule bacteria. However, BLAST analysis revealed seventeen putative efflux systems in Rlv3841 and several of these were highly differentially expressed during rhizosphere colonisation, host infection and bacteroid differentiation. This suggests they have an integral role in symbiosis with host plants.

  11. Positive and negative regulation of T cell responses by fibroblastic reticular cells within paracortical regions of lymph nodes

    PubMed Central

    Siegert, Stefanie; Luther, Sanjiv A.

    2012-01-01

    Fibroblastic reticular cells (FRC) form the structural backbone of the T cell rich zones in secondary lymphoid organs (SLO), but also actively influence the adaptive immune response. They provide a guidance path for immigrating T lymphocytes and dendritic cells (DC) and are the main local source of the cytokines CCL19, CCL21, and IL-7, all of which are thought to positively regulate T cell homeostasis and T cell interactions with DC. Recently, FRC in lymph nodes (LN) were also described to negatively regulate T cell responses in two distinct ways. During homeostasis they express and present a range of peripheral tissue antigens, thereby participating in peripheral tolerance induction of self-reactive CD8+ T cells. During acute inflammation T cells responding to foreign antigens presented on DC very quickly release pro-inflammatory cytokines such as interferon γ. These cytokines are sensed by FRC which transiently produce nitric oxide (NO) gas dampening the proliferation of neighboring T cells in a non-cognate fashion. In summary, we propose a model in which FRC engage in a bidirectional crosstalk with both DC and T cells to increase the efficiency of the T cell response. However, during an acute response, FRC limit excessive expansion and inflammatory activity of antigen-specific T cells. This negative feedback loop may help to maintain tissue integrity and function during rapid organ growth. PMID:22973278

  12. Positive and negative regulation of type II TGF-beta receptor signal transduction by autophosphorylation on multiple serine residues.

    PubMed Central

    Luo, K; Lodish, H F

    1997-01-01

    The type II transforming growth factor-beta (TGF-beta) receptor Ser/Thr kinase (TbetaRII) is responsible for the initiation of multiple TGF-beta signaling pathways, and loss of its function is associated with many types of human cancer. Here we show that TbetaRII kinase is regulated intricately by autophosphorylation on at least three serine residues. Ser213, in the membrane-proximal segment outside the kinase domain, undergoes intra-molecular autophosphorylation which is essential for the activation of TbetaRII kinase activity, activation of TbetaRI and TGF-beta-induced growth inhibition. In contrast, phosphorylation of Ser409 and Ser416, located in a segment corresponding to the substrate recognition T-loop region in a three-dimensional structural model of protein kinases, is enhanced by receptor dimerization and can occur via an intermolecular mechanism. Phosphorylation of Ser409 is essential for TbetaRII kinase signaling, while phosphorylation of Ser416 inhibits receptor function. Mutation of Ser416 to alanine results in a hyperactive receptor that is better able than wild-type to induce TbetaRI activation and subsequent cell cycle arrest. Since on a single receptor either Ser409 or Ser416, but not both simultaneously, can become autophosphorylated, our results show that TbetaRII phosphorylation is regulated intricately and affects TGF-beta receptor signal transduction both positively and negatively. PMID:9155023

  13. TREM-1 is a positive regulator of TNF-α and IL-8 production in U937 foam cells.

    PubMed

    Wang, Yu-Shi; Li, Xiang-Jun; Zhao, Wai-Ou

    2012-05-01

    The purpose of our study was to investigate the expression levels of TREM-1 (triggering receptor expressed on myeloid cells-1) in U937 foam cells and determine whether TREM-1 regulates the production of tumor necrosis factor-alpha and interleukin-8 in these cells. Human U937 cells were incubated with phorbol 12-myristate 13-acetate and then oxidized human low-density lipoprotein to induce foam cell formation. Oil red O staining was used to identify the foam cells. The production of IL-8 and TNF-α by U937 foam cells was assayed by enzyme-linked immunosorbent assay. The expression of TREM-1 mRNA in U937 foam cells was detected by reverse transcription-polymerase chain reaction. Moreover, U937 foam cells were transfected by small interfering RNA using Lipofectamine 2000 to knockdown TREM-1. Western blot was performed to assay protein expression of TREM-1 and ELISA was used to examine the effect of TREM-1 knockdown on IL-8 and TNF-α production. PMA and ox-LDL induced U937 cells to form foam cells. The production of TNF-α and IL-8 was found to be significantly elevated in U937 foam cells, concomitant with a significant up-regulation of TREM-1 mRNA. TREM-1 siRNA was able to partially silence the expression of TREM-1 protein and remarkably inhibited TNF-α and IL-8 production in U937 foam cells, suggesting that TREM-1 is a positive regulator of TNF-α and IL-8 production in U937 foam cells. Our finding that TREM-1 controls the production of IL-8 and TNF-α in U937 foam cells defines a potentially critical role of TREM-1 in the pathogenesis of atherosclerosis and implicates TREM-1 as a potential therapeutic target for the disease.

  14. Predicting Short-Term Positive Affect in Individuals with Social Anxiety Disorder: The Role of Selected Personality Traits and Emotion Regulation Strategies

    PubMed Central

    Weisman, Jaclyn S.; Rodebaugh, Thomas L.; Lim, Michelle H.; Fernandez, Katya C.

    2015-01-01

    Recently, research has provided support for a moderate, inverse relationship between social anxiety and dispositional positive affect. However, the dynamics of this relationship remain poorly understood. The present study evaluates whether certain personality traits and emotion regulation variables predict short-term positive affect for individuals with social anxiety disorder and healthy controls. Positive affect as measured by two self-report instruments was assessed before and after two tasks in which the participant conversed with either a friend or a romantic partner. Tests of models examining the hypothesized prospective predictors revealed that the paths did not differ significantly across diagnostic group and both groups showed the hypothesized patterns of endorsement for the emotion regulation variables. Further, a variable reflecting difficulty redirecting oneself when distressed prospectively predicted one measure of positive affect. Additional research is needed to explore further the role of emotion regulation strategies on positive emotions for individuals higher in social anxiety. PMID:26119140

  15. Predicting short-term positive affect in individuals with social anxiety disorder: The role of selected personality traits and emotion regulation strategies.

    PubMed

    Weisman, Jaclyn S; Rodebaugh, Thomas L; Lim, Michelle H; Fernandez, Katya C

    2015-08-01

    Recently, research has provided support for a moderate, inverse relationship between social anxiety and dispositional positive affect. However, the dynamics of this relationship remain poorly understood. The present study evaluates whether certain personality traits and emotion regulation variables predict short-term positive affect for individuals with social anxiety disorder and healthy controls. Positive affect as measured by two self-report instruments was assessed before and after two tasks in which the participant conversed with either a friend or a romantic partner. Tests of models examining the hypothesized prospective predictors revealed that the paths did not differ significantly across diagnostic group and both groups showed the hypothesized patterns of endorsement for the emotion regulation variables. Further, a variable reflecting difficulty redirecting oneself when distressed prospectively predicted one measure of positive affect. Additional research is needed to explore further the role of emotion regulation strategies on positive emotions for individuals higher in social anxiety.

  16. Type III secretion system 1 genes in Vibrio parahaemolyticus are positively regulated by ExsA and negatively regulated by ExsD

    PubMed Central

    Zhou, Xiaohui; Shah, Devendra H; Konkel, Michael E; Call, Douglas R

    2008-01-01

    Vibrio parahaemolyticus harbours two distinct type III secretion systems (T3SS1 and T3SS2). A subset of 10 T3SS1 genes are transcribed when V. parahaemolyticus is grown in tissue culture medium [Dulbecco's modified Eagle's medium (DMEM)], while transcription of these genes (except exsD) is minimal upon growth in Luria–Bertani-Salt (LB-S). Transcription of T3SS1 genes and cytotoxicity towards HeLa cells was prevented by deletion of exsA while complementation with exsA restored these traits. Overexpression of ExsA in the wild-type strain, NY-4, activated the transcription of T3SS1 genes when bacteria were grown in LB-S. Thus, ExsA is necessary and sufficient to induce the transcription of T3SS1 genes. Deletion of the exsD permitted the transcription of T3SS1 genes when bacteria were grown in the LB-S medium and complementation with the wild-type exsD gene-blocked transcription of T3SS1 genes. Overexpression of ExsD in NY-4 prevented the transcription of T3SS1 gene when bacteria were grown in DMEM. A gel mobility shift assay demonstrated that purified ExsA protein binds a novel motif in the upstream region of vp1668 and vp1687, indicating that ExsA interacts directly with the promoter sequences of T3SS1 genes. ExsA positively regulates the expression and secretion of Vp1656 while ExsD negatively regulates the expression and secretion of Vp1656. PMID:18554322

  17. p32, a novel binding partner of Mcl-1, positively regulates mitochondrial Ca{sup 2+} uptake and apoptosis

    SciTech Connect

    Xiao, Kang; Wang, Yinyin; Chang, Zhijie; Lao, Yuanzhi; Chang, Donald C.

    2014-08-22

    Highlights: • p32 binds to Mcl-1. • p32 affects apoptosis. • p32 and Mcl-1 regulate mitochondrial Ca{sup 2+}. - Abstract: Mcl-1 is a major anti-apoptotic Bcl-2 family protein. It is well known that Mcl-1 can interact with certain pro-apoptotic Bcl-2 family proteins in normal cells to neutralize their pro-apoptotic functions, thus prevent apoptosis. In addition, it was recently found that Mcl-1 can also inhibit mitochondrial calcium uptake. The detailed mechanism, however, is still not clear. Based on Yeast Two-Hybrid screening and co-immunoprecipitation, we identified a mitochondrial protein p32 (C1qbp) as a novel binding partner of Mcl-1. We found that p32 had a number of interesting properties: (1) p32 can positively regulate UV-induced apoptosis in HeLa cells. (2) Over-expressing p32 could significantly promote mitochondrial calcium uptake, while silencing p32 by siRNA suppressed it. (3) In p32 knockdown cells, Ruthenium Red treatment (an inhibitor of mitochondrial calcium uniporter) showed no further suppressive effect on mitochondrial calcium uptake. In addition, in Ruthenium Red treated cells, Mcl-1 also failed to suppress mitochondrial calcium uptake. Taken together, our findings suggest that p32 is part of the putative mitochondrial uniporter that facilitates mitochondrial calcium uptake. By binding to p32, Mcl-1 can interfere with the uniporter function, thus inhibit the mitochondrial Ca{sup 2+} uploading. This may provide a novel mechanism to explain the anti-apoptotic function of Mcl-1.

  18. Grape Ripening Is Regulated by Deficit Irrigation/Elevated Temperatures According to Cluster Position in the Canopy

    PubMed Central

    Zarrouk, Olfa; Brunetti, Cecilia; Egipto, Ricardo; Pinheiro, Carla; Genebra, Tânia; Gori, Antonella; Lopes, Carlos M.; Tattini, Massimiliano; Chaves, M. Manuela

    2016-01-01

    The impact of water deficit on berry quality has been extensively investigated during the last decades. Nonetheless, there is a scarcity of knowledge on the performance of varieties exposed to a combination of high temperatures/water stress during the growing season and under vineyard conditions. The objective of this research was to investigate the effects of two irrigation regimes, sustained deficit irrigation (SDI, 30% ETc) and regulated deficit irrigation (RDI, 15% ETc) and of two cluster positions within the canopy (east- and west-exposed sides) on berry ripening in red Aragonez (Tempranillo) grapevines. The study was undertaken for two successive years in a commercial vineyard in South Portugal, monitoring the following parameters: pre-dawn leaf water potential, berry temperature, sugars, polyphenols, abscisic acid (ABA) and related metabolites. Additionally, expression patterns for different transcripts encoding for enzymes responsible for anthocyanin and ABA biosynthesis (VviUFGT, VvNCED1, VvβG1, VviHyd1, VviHyd2) were analyzed. In both years anthocyanin concentration was lower in RDI at the west side (RDIW- the hottest one) from véraison onwards, suggesting that the most severe water stress conditions exacerbated the negative impact of high temperature on anthocyanin. The down-regulation of VviUFGT expression revealed a repression of the anthocyanin synthesis in berries of RDIW, at early stages of berry ripening. At full-maturation, anthocyanin degradation products were detected, being highest at RDIW. This suggests that the negative impact of water stress and high temperature on anthocyanins results from the repression of biosynthesis at the onset of ripening and from degradation at later stages. On the other hand, berries grown under SDI displayed a higher content in phenolics than those under RDI, pointing out for the attenuation of the negative temperature effects under SDI. Irrigation regime and berry position had small effect on free

  19. Nuclear Protein Sam68 Interacts with the Enterovirus 71 Internal Ribosome Entry Site and Positively Regulates Viral Protein Translation

    PubMed Central

    Zhang, Hua; Song, Lei; Cong, Haolong

    2015-01-01

    ABSTRACT Enterovirus 71 (EV71) recruits various cellular factors to assist in the replication and translation of its genome. Identification of the host factors involved in the EV71 life cycle not only will enable a better understanding of the infection mechanism but also has the potential to be of use in the development of antiviral therapeutics. In this study, we demonstrated that the cellular factor 68-kDa Src-associated protein in mitosis (Sam68) acts as an internal ribosome entry site (IRES) trans-acting factor (ITAF) that binds specifically to the EV71 5′ untranslated region (5′UTR). Interaction sites in both the viral IRES (stem-loops IV and V) and the heterogeneous nuclear ribonucleoprotein K homology (KH) domain of Sam68 protein were further mapped using an electrophoretic mobility shift assay (EMSA) and biotin RNA pulldown assay. More importantly, dual-luciferase (firefly) reporter analysis suggested that overexpression of Sam68 positively regulated IRES-dependent translation of virus proteins. In contrast, both IRES activity and viral protein translation significantly decreased in Sam68 knockdown cells compared with the negative-control cells treated with short hairpin RNA (shRNA). However, downregulation of Sam68 did not have a significant inhibitory effect on the accumulation of the EV71 genome. Moreover, Sam68 was redistributed from the nucleus to the cytoplasm and interacts with cellular factors, such as poly(rC)-binding protein 2 (PCBP2) and poly(A)-binding protein (PABP), during EV71 infection. The cytoplasmic relocalization of Sam68 in EV71-infected cells may be involved in the enhancement of EV71 IRES-mediated translation. Since Sam68 is known to be a RNA-binding protein, these results provide direct evidence that Sam68 is a novel ITAF that interacts with EV71 IRES and positively regulates viral protein translation. IMPORTANCE The nuclear protein Sam68 is found as an additional new host factor that interacts with the EV71 IRES during infection

  20. Positive Regulation of psbA Gene Expression by cis-Encoded Antisense RNAs in Synechocystis sp. PCC 68031[OA

    PubMed Central

    Sakurai, Isamu; Stazic, Damir; Eisenhut, Marion; Vuorio, Eerika; Steglich, Claudia; Hess, Wolfgang R.; Aro, Eva-Mari

    2012-01-01

    The D1 protein of photosystem II in the thylakoid membrane of photosynthetic organisms is encoded by psbA genes, which in cyanobacteria occur in the form of a small gene family. Light-dependent up-regulation of psbA gene expression is crucial to ensure the proper replacement of the D1 protein. To gain a high level of gene expression, psbA transcription can be enhanced by several orders of magnitude. Recent transcriptome analyses demonstrated a high number of cis-encoded antisense RNAs (asRNAs) in bacteria, but very little is known about their possible functions. Here, we show the presence of two cis-encoded asRNAs (PsbA2R and PsbA3R) of psbA2 and psbA3 from Synechocystis sp. PCC 6803. These asRNAs are located in the 5′ untranslated region of psbA2 and psbA3 genes. Their expression becomes up-regulated by light and down-regulated by darkness, similar to their target mRNAs. In the PsbA2R-suppressing strain [PsbA2R(−)], the amount of psbA2 mRNA was only about 50% compared with the control strain. Likewise, we identified a 15% lowered activity of photosystem II and a reduced amount of the D1 protein in PsbA2R(−) compared with the control strain. The function of PsbA2R in the stabilization of psbA2 mRNA was shown from in vitro RNase E assay when the AU box and the ribosome-binding site in the 5′ untranslated region of psbA2 mRNA were both covered by PsbA2R. These results add another layer of complexity to the mechanisms that contribute to psbA gene expression and show PsbA2R as a positively acting factor to achieve a maximum level of D1 synthesis. PMID:22858634

  1. A Novel Modulator of Kv3 Potassium Channels Regulates the Firing of Parvalbumin-Positive Cortical Interneurons.

    PubMed

    Rosato-Siri, Marcelo D; Zambello, Erika; Mutinelli, Chiara; Garbati, Nicoletta; Benedetti, Roberto; Aldegheri, Laura; Graziani, Francesca; Virginio, Caterina; Alvaro, Giuseppe; Large, Charles H

    2015-09-01

    Kv3.1 and Kv3.2 high voltage-activated potassium channels, which display fast activation and deactivation kinetics, are known to make a crucial contribution to the fast-spiking phenotype of certain neurons. Pharmacological experiments show that the blockade of native Kv3 currents with low concentrations of tetraethylammonium or 4-aminopyridine impairs the expression of this firing phenotype. In particular, Kv3 channels are highly expressed by fast-spiking, parvalbumin-positive interneurons in corticolimbic brain circuits, which modulate the synchronization of cortical circuits and the generation of brain rhythms. Here, we describe a novel small molecule, (5R)-5-ethyl-3-(6-{[4-methyl-3-(methyloxy)phenyl]oxy}-3-pyridinyl)-2,4-imidazolidinedione (AUT1), which modulates Kv3.1 and Kv3.2 channels in human recombinant and rodent native neurons. AUT1 increased whole currents mediated by human Kv3.1b and Kv3.2a channels, with a concomitant leftward shift in the voltage dependence of activation. A less potent effect was observed on hKv3.3 currents. In mouse somatosensory cortex slices in vitro, AUT1 rescued the fast-spiking phenotype of parvalbumin-positive-fast-spiking interneurons following an impairment of their firing capacity by blocking a proportion of Kv3 channels with a low concentration of tetraethylammonium. Notably, AUT1 had no effect on interneuron firing when applied alone. Together, these data confirm the role played by Kv3 channels in the regulation of the firing phenotype of somatosensory interneurons and suggest that AUT1 and other Kv3 modulators could represent a new and promising therapeutic approach to the treatment of disorders associated with dysfunction of inhibitory feedback in corticolimbic circuits, such as schizophrenia.

  2. ErbB4 in parvalbumin-positive interneurons is critical for neuregulin 1 regulation of long-term potentiation.

    PubMed

    Chen, Yong-Jun; Zhang, Meng; Yin, Dong-Min; Wen, Lei; Ting, Annie; Wang, Pu; Lu, Yi-Sheng; Zhu, Xin-Hong; Li, Shu-Ji; Wu, Cui-Ying; Wang, Xue-Ming; Lai, Cary; Xiong, Wen-Cheng; Mei, Lin; Gao, Tian-Ming

    2010-12-14

    Neuregulin 1 (NRG1) is a trophic factor that acts by stimulating ErbB receptor tyrosine kinases and has been implicated in neural development and synaptic plasticity. In this study, we investigated mechanisms of its suppression of long-term potentiation (LTP) in the hippocampus. We found that NRG1 did not alter glutamatergic transmission at SC-CA1 synapses but increased the GABA(A) receptor-mediated synaptic currents in CA1 pyramidal cells via a presynaptic mechanism. Inhibition of GABA(A) receptors blocked the suppressing effect of NRG1 on LTP and prevented ecto-ErbB4 from enhancing LTP, implicating a role of GABAergic transmission. To test this hypothesis further, we generated parvalbumin (PV)-Cre;ErbB4(-/-) mice in which ErbB4, an NRG1 receptor in the brain, is ablated specifically in PV-positive interneurons. NRG1 was no longer able to increase inhibitory postsynaptic currents and to suppress LTP in PV-Cre;ErbB4(-/-) hippocampus. Accordingly, contextual fear conditioning, a hippocampus-dependent test, was impaired in PV-Cre;ErbB4(-/-) mice. In contrast, ablation of ErbB4 in pyramidal neurons had no effect on NRG1 regulation of hippocampal LTP or contextual fear conditioning. These results demonstrate a critical role of ErbB4 in PV-positive interneurons but not in pyramidal neurons in synaptic plasticity and support a working model that NRG1 suppresses LTP by enhancing GABA release. Considering that NRG1 and ErbB4 are susceptibility genes of schizophrenia, these observations contribute to a better understanding of how abnormal NRG1/ErbB4 signaling may be involved in the pathogenesis of schizophrenia.

  3. SUMOylation by the E3 ligase TbSIZ1/PIAS1 positively regulates VSG expression in Trypanosoma brucei.

    PubMed

    López-Farfán, Diana; Bart, Jean-Mathieu; Rojas-Barros, Domingo I; Navarro, Miguel

    2014-12-01

    Bloodstream form trypanosomes avoid the host immune response by switching the expression of their surface proteins between Variant Surface Glycoproteins (VSG), only one of which is expressed at any given time. Monoallelic transcription of the telomeric VSG Expression Site (ES) by RNA polymerase I (RNA pol I) localizes to a unique nuclear body named the ESB. Most work has focused on silencing mechanisms of inactive VSG-ESs, but the mechanisms involved in transcriptional activation of a single VSG-ES remain largely unknown. Here, we identify a highly SUMOylated focus (HSF) in the nucleus of the bloodstream form that partially colocalizes with the ESB and the active VSG-ES locus. SUMOylation of chromatin-associated proteins was enriched along the active VSG-ES transcriptional unit, in contrast to silent VSG-ES or rDNA, suggesting that it is a distinct feature of VSG-ES monoallelic expression. In addition, sequences upstream of the active VSG-ES promoter were highly enriched in SUMOylated proteins. We identified TbSIZ1/PIAS1 as the SUMO E3 ligase responsible for SUMOylation in the active VSG-ES chromatin. Reduction of SUMO-conjugated proteins by TbSIZ1 knockdown decreased the recruitment of RNA pol I to the VSG-ES and the VSG-ES-derived transcripts. Furthermore, cells depleted of SUMO conjugated proteins by TbUBC9 and TbSUMO knockdown confirmed the positive function of SUMO for VSG-ES expression. In addition, the largest subunit of RNA pol I TbRPA1 was SUMOylated in a TbSIZ-dependent manner. Our results show a positive mechanism associated with active VSG-ES expression via post-translational modification, and indicate that chromatin SUMOylation plays an important role in the regulation of VSG-ES. Thus, protein SUMOylation is linked to active gene expression in this protozoan parasite that diverged early in evolution.

  4. Intentional Self-Regulation and Positive Youth Development in Early Adolescence: Findings from the 4-H Study of Positive Youth Development

    ERIC Educational Resources Information Center

    Gestsdottir, Steinunn; Lerner, Richard M.

    2007-01-01

    In this research, the authors examined the development of intentional self-regulation in early adolescence, which was operationalized through the use of a measure derived from the model of selection, optimization, and compensation (SOC). This model describes the individual's contributions to mutually influential relations between the person and…

  5. 5-HT3a Receptors Modulate Hippocampal Gamma Oscillations by Regulating Synchrony of Parvalbumin-Positive Interneurons.

    PubMed

    Huang, Ying; Yoon, Kristopher; Ko, Ho; Jiao, Song; Ito, Wataru; Wu, Jian-Young; Yung, Wing-Ho; Lu, Bai; Morozov, Alexei

    2016-02-01

    Gamma-frequency oscillatory activity plays an important role in information integration across brain areas. Disruption in gamma oscillations is implicated in cognitive impairments in psychiatric disorders, and 5-HT3 receptors (5-HT3Rs) are suggested as therapeutic targets for cognitive dysfunction in psychiatric disorders. Using a 5-HT3aR-EGFP transgenic mouse line and inducing gamma oscillations by carbachol in hippocampal slices, we show that activation of 5-HT3aRs, which are exclusively expressed in cholecystokinin (CCK)-containing interneurons, selectively suppressed and desynchronized firings in these interneurons by enhancing spike-frequency accommodation in a small conductance potassium (SK)-channel-dependent manner. Parvalbumin-positive interneurons therefore received diminished inhibitory input leading to increased but desynchronized firings of PV cells. As a consequence, the firing of pyramidal neurons was desynchronized and gamma oscillations were impaired. These effects were independent of 5-HT3aR-mediated CCK release. Our results therefore revealed an important role of 5-HT3aRs in gamma oscillations and identified a novel crosstalk among different types of interneurons for regulation of network oscillations. The functional link between 5-HT3aR and gamma oscillations may have implications for understanding the cognitive impairments in psychiatric disorders.

  6. Regulation of transcription by eukaryotic-like serine-threonine kinases and phosphatases in Gram-positive bacterial pathogens

    PubMed Central

    Wright, David P; Ulijasz, Andrew T

    2014-01-01

    Bacterial eukaryotic-like serine threonine kinases (eSTKs) and serine threonine phosphatases (eSTPs) have emerged as important signaling elements that are indispensable for pathogenesis. Differing considerably from their histidine kinase counterparts, few eSTK genes are encoded within the average bacterial genome, and their targets are pleiotropic in nature instead of exclusive. The growing list of important eSTK/P substrates includes proteins involved in translation, cell division, peptidoglycan synthesis, antibiotic tolerance, resistance to innate immunity and control of virulence factors. Recently it has come to light that eSTK/Ps also directly modulate transcriptional machinery in many microbial pathogens. This novel form of regulation is now emerging as an additional means by which bacteria can alter their transcriptomes in response to host-specific environmental stimuli. Here we focus on the ability of eSTKs and eSTPs in Gram-positive bacterial pathogens to directly modulate transcription, the known mechanistic outcomes of these modifications, and their roles as an added layer of complexity in controlling targeted RNA synthesis to enhance virulence potential. PMID:25603430

  7. Histone demethylase KDM5A is regulated by its reader domain through a positive-feedback mechanism

    PubMed Central

    Torres, Idelisse Ortiz; Kuchenbecker, Kristopher M.; Nnadi, Chimno I.; Fletterick, Robert J.; Kelly, Mark J.S.; Fujimori, Danica Galonić

    2016-01-01

    The retinoblastoma binding protein KDM5A removes methyl marks from lysine 4 of histone H3 (H3K4). Misregulation of KDM5A contributes to the pathogenesis of lung and gastric cancers. In addition to its catalytic jumonji C domain, KDM5A contains three PHD reader domains, commonly recognized as chromatin recruitment modules. It is unknown whether any of these domains in KDM5A have functions beyond recruitment and whether they regulate the catalytic activity of the demethylase. Here using biochemical and nuclear magnetic resonance (NMR)-based structural studies, we show that the PHD1 preferentially recognizes unmethylated H3K4 histone tail, product of KDM5A-mediated demethylation of tri-methylated H3K4 (H3K4me3). Binding of unmodified H3 peptide to the PHD1 stimulates catalytic domain-mediated removal of methyl marks from H3K4me3 peptide and nucleosome substrates. This positive-feedback mechanism—enabled by the functional coupling between a reader and a catalytic domain in KDM5A—suggests a model for the spread of demethylation on chromatin. PMID:25686748

  8. Integrin CD11b positively regulates TLR4-induced signalling pathways in dendritic cells but not in macrophages

    NASA Astrophysics Data System (ADS)

    Ling, Guang Sheng; Bennett, Jason; Woollard, Kevin J.; Szajna, Marta; Fossati-Jimack, Liliane; Taylor, Philip R.; Scott, Diane; Franzoso, Guido; Cook, H. Terence; Botto, Marina

    2014-01-01

    Tuned and distinct responses of macrophages and dendritic cells to Toll-like receptor 4 (TLR4) activation induced by lipopolysaccharide (LPS) underpin the balance between innate and adaptive immunity. However, the molecule(s) that confer these cell-type-specific LPS-induced effects remain poorly understood. Here we report that the integrin αM (CD11b) positively regulates LPS-induced signalling pathways selectively in myeloid dendritic cells but not in macrophages. In dendritic cells, which express lower levels of CD14 and TLR4 than macrophages, CD11b promotes MyD88-dependent and MyD88-independent signalling pathways. In particular, in dendritic cells CD11b facilitates LPS-induced TLR4 endocytosis and is required for the subsequent signalling in the endosomes. Consistent with this, CD11b deficiency dampens dendritic cell-mediated TLR4-triggered responses in vivo leading to impaired T-cell activation. Thus, by modulating the trafficking and signalling functions of TLR4 in a cell-type-specific manner CD11b fine tunes the balance between adaptive and innate immune responses initiated by LPS.

  9. Plk1-mediated stabilization of 53BP1 through USP7 regulates centrosome positioning to maintain bipolarity.

    PubMed

    Yim, H; Shin, S-B; Woo, S U; Lee, P C-W; Erikson, R L

    2017-02-16

    Although 53BP1 has been established well as a mediator in DNA damage response, its function in mitosis is not clearly understood. We found that 53BP1 is a mitotic-binding partner of the kinases Plk1 and AuroraA, and that the binding with Plk1 increases the stability of 53BP1 by accelerating its interaction with the deubiquitinase USP7. Depletion of 53BP1 induces mitotic defects such as chromosomal missegregation, misorientation of spindle poles and the generation of extra centrosomes, which is similar phenotype to USP7-knockdown cells. In addition, 53BP1 depletion reduces the levels of p53 and centromere protein F (CENPF), interacting proteins of 53BP1. These phenotypes induced by 53BP1 depletion were rescued by expression of wild-type or phosphomimic mutant 53BP1 but not by expression of a dephosphomimic mutant. We propose that phosphorylation of 53BP1 at S380 accelerates complex formation with USP7 and CENPF to regulate their stability, thus having a crucial role in proper centrosome positioning, chromosomal alignment, and centrosome number.

  10. Histone demethylase KDM5A is regulated by its reader domain through a positive-feedback mechanism

    NASA Astrophysics Data System (ADS)

    Torres, Idelisse Ortiz; Kuchenbecker, Kristopher M.; Nnadi, Chimno I.; Fletterick, Robert J.; Kelly, Mark J. S.; Fujimori, Danica Galonić

    2015-02-01

    The retinoblastoma binding protein KDM5A removes methyl marks from lysine 4 of histone H3 (H3K4). Misregulation of KDM5A contributes to the pathogenesis of lung and gastric cancers. In addition to its catalytic jumonji C domain, KDM5A contains three PHD reader domains, commonly recognized as chromatin recruitment modules. It is unknown whether any of these domains in KDM5A have functions beyond recruitment and whether they regulate the catalytic activity of the demethylase. Here using biochemical and nuclear magnetic resonance (NMR)-based structural studies, we show that the PHD1 preferentially recognizes unmethylated H3K4 histone tail, product of KDM5A-mediated demethylation of tri-methylated H3K4 (H3K4me3). Binding of unmodified H3 peptide to the PHD1 stimulates catalytic domain-mediated removal of methyl marks from H3K4me3 peptide and nucleosome substrates. This positive-feedback mechanism—enabled by the functional coupling between a reader and a catalytic domain in KDM5A—suggests a model for the spread of demethylation on chromatin.

  11. Bid is a positive regulator for donor-derived lymphoid cell regeneration in γ-irradiated recipients

    PubMed Central

    Shen, Hongmei; Yu, Hui; Liang, Paulina H.; XuFeng, Richard; Song, Yifang; Hu, Xiaoxia; Chen, Xiaoyun; Yin, Xiao-Ming; Cheng, Tao

    2011-01-01

    Objective Hematopoietic regeneration is regulated by cell survival proteins such as the Bcl-2 family. Bid, a BH3-only protein of the Bcl-2 family, has multiple cellular functions and is involved in a variety of physiological or pathological conditions. We attempted to define its role in hematopoietic cell repopulation under the stress condition of bone marrow transplantation (BMT). Materials and Methods We performed conventional or competitive BMT with donor hematopoietic cells from Bid−/− or Bid+/+ mice. Flow Cytometry was used for quantification of hematopoietic stem cells (HSCs), hematopoietic progenitor cells (HPCs) and differentiated cells in different lineages (T, B and Myeloid cells). Single cell culture and homing assays were performed to further evaluate HSC functions. HPCs were also measured by the colony-forming cell (CFC) culture. Results Contrary to the widely-recognized role of Bid as a pro-apoptotic protein, the absence of Bid significantly reduced the reconstitution of donor hematopoietic cells in γ-irradiated recipients. Interestingly, however, numbers of HSCs and HPCs, and their functions were not overtly altered. Instead, the regeneration of donor T and B cells was significantly impaired in the absence of Bid. Further analysis indicated an accumulation of the triple negative T cell population in the thymus, and pro-B cells in the bone marrow. Conclusion Our current study demonstrates a positive impact of Bid on hematopoietic regeneration mainly due to its unique effects on donor lymphopoiesis in the transplant recipients. PMID:21703985

  12. A FRET-based study reveals site-specific regulation of spindle position checkpoint proteins at yeast centrosomes

    PubMed Central

    Gryaznova, Yuliya; Caydasi, Ayse Koca; Malengo, Gabriele; Sourjik, Victor; Pereira, Gislene

    2016-01-01

    The spindle position checkpoint (SPOC) is a spindle pole body (SPB, equivalent of mammalian centrosome) associated surveillance mechanism that halts mitotic exit upon spindle mis-orientation. Here, we monitored the interaction between SPB proteins and the SPOC component Bfa1 by FRET microscopy. We show that Bfa1 binds to the scaffold-protein Nud1 and the γ-tubulin receptor Spc72. Spindle misalignment specifically disrupts Bfa1-Spc72 interaction by a mechanism that requires the 14-3-3-family protein Bmh1 and the MARK/PAR-kinase Kin4. Dissociation of Bfa1 from Spc72 prevents the inhibitory phosphorylation of Bfa1 by the polo-like kinase Cdc5. We propose Spc72 as a regulatory hub that coordinates the activity of Kin4 and Cdc5 towards Bfa1. In addition, analysis of spc72∆ cells shows that a mitotic-exit-promoting dominant signal, which is triggered upon elongation of the spindle into the bud, overrides the SPOC. Our data reinforce the importance of daughter-cell-associated factors and centrosome-based regulations in mitotic exit and SPOC control. DOI: http://dx.doi.org/10.7554/eLife.14029.001 PMID:27159239

  13. A FRET-based study reveals site-specific regulation of spindle position checkpoint proteins at yeast centrosomes.

    PubMed

    Gryaznova, Yuliya; Koca Caydasi, Ayse; Malengo, Gabriele; Sourjik, Victor; Pereira, Gislene

    2016-05-09

    The spindle position checkpoint (SPOC) is a spindle pole body (SPB, equivalent of mammalian centrosome) associated surveillance mechanism that halts mitotic exit upon spindle mis-orientation. Here, we monitored the interaction between SPB proteins and the SPOC component Bfa1 by FRET microscopy. We show that Bfa1 binds to the scaffold-protein Nud1 and the γ-tubulin receptor Spc72. Spindle misalignment specifically disrupts Bfa1-Spc72 interaction by a mechanism that requires the 14-3-3-family protein Bmh1 and the MARK/PAR-kinase Kin4. Dissociation of Bfa1 from Spc72 prevents the inhibitory phosphorylation of Bfa1 by the polo-like kinase Cdc5. We propose Spc72 as a regulatory hub that coordinates the activity of Kin4 and Cdc5 towards Bfa1. In addition, analysis of spc72∆ cells shows that a mitotic-exit-promoting dominant signal, which is triggered upon elongation of the spindle into the bud, overrides the SPOC. Our data reinforce the importance of daughter-cell-associated factors and centrosome-based regulations in mitotic exit and SPOC control.

  14. Non-neuronal acetylcholine as an endogenous regulator of proliferation and differentiation of Lgr5-positive stem cells in mice.

    PubMed

    Takahashi, Toshio; Ohnishi, Hiroe; Sugiura, Yuki; Honda, Kurara; Suematsu, Makoto; Kawasaki, Takashi; Deguchi, Tomonori; Fujii, Takeshi; Orihashi, Kaoru; Hippo, Yoshitaka; Watanabe, Takehiro; Yamagaki, Tohru; Yuba, Shunsuke

    2014-10-01

    Non-neuronal acetylcholine (ACh) is predicted to function as a local cell signaling molecule. However, the physiological significance of the synthesis of non-neuronal ACh in the intestine remains unclear. Here, experiments using crypt-villus organoids that lack nerve and immune cells in culture led us to suggest that endogenous ACh is synthesized in the intestinal epithelium to evoke growth and differentiation of the organoids through activation of muscarinic ACh receptors (mAChRs). The extracts of the cultured organoids showed a noticeable capacity for ACh synthesis that was sensitive to a potent inhibitor of choline acetyltransferase. Imaging MS revealed endogenous ACh localized in the epithelial layer in mouse small intestinal epithelium in vivo, suggesting that there are non-neuronal resources of ACh. Treatment of organoids with carbachol downregulated the growth of organoids and the expression of marker genes for epithelial cells. On the other hand, antagonists for mAChRs enhanced the growth and differentiation of organoids, indicating the involvement of mAChRs in regulating the proliferation and differentiation of Lgr5-positive stem cells. Collectively, our data provide evidence that endogenous ACh released from intestinal epithelium maintains homeostasis of intestinal epithelial cell growth and differentiation via mAChRs in mice.

  15. Positive Intervention for Serious Behavior Problems: Best Practices in Implementing the Hughes Bill (A.B. 2586) and the Positive Behavioral Intervention Regulations.

    ERIC Educational Resources Information Center

    Wright, Diana Browning; Gurman, Harvey B.

    This manual provides guidelines to educators attempting to comply with California's Hughes Bill, which is intended to ensure the rights of special education students to have positive behavioral intervention plans designed to bring lasting behavioral changes without interventions that cause pain or trauma. An introductory chapter summarizes the…

  16. The rpoA341 allele of Escherichia coli specifically impairs the transcription of a group of positively-regulated operons.

    PubMed

    Giffard, P M; Booth, I R

    1988-09-01

    The specificity of the transcription defect caused by the rpoA341(phs) allele has been investigated. Three apparently unlinked genetic systems have been found to be impaired in their transcription by this mutant allele of the alpha subunit of RNA polymerase. These three systems, the melAB operon, the cysA locus and the ara regulon, are apparently unrelated other than by their requirement for a regulon-specific positive regulator for the initiation of transcription. Expression of the gene for the positive regulator does not appear to be significantly affected in any of the three systems. However, mutations that render expression of the araBAD operon independent of the regulatory protein also confer insensitivity to the rpoA341 allele. The significance of these observations is discussed in the context of models of positive regulation.

  17. Phosphatidylinositol-4,5-bisphosphate (PIP2) regulation of strong inward rectifier Kir2.1 channels: multilevel positive cooperativity.

    PubMed

    Xie, Lai-Hua; John, Scott A; Ribalet, Bernard; Weiss, James N

    2008-04-01

    Inwardly rectifying potassium (Kir) channels are gated by the interaction of their cytoplasmic regions with membrane-bound phosphatidylinositol-4,5-bisphosphate (PIP(2)). In the present study, we examined how PIP(2) interaction regulates channel availability and channel openings to various subconductance levels (sublevels) as well as the fully open state in the strong inward rectifier Kir2.1 channel. Various Kir2.1 channel constructs were expressed in Xenopus oocytes and single channel or macroscopic currents were recorded from inside-out patches. The wild-type (WT) channel rarely visited the subconductance levels under control conditions. However, upon reducing Kir2.1 channel interaction with PIP(2) by a variety of interventions, including PIP(2) antibodies, screening PIP(2) with neomycin, or mutating PIP(2) binding sites (e.g. K188Q), visitation to the sublevels was markedly increased before channels were converted to an unavailable mode in which they did not open. No channel activity was detected in channels with the double mutation K188A/R189A, a mutant which exhibits extremely weak interaction with PIP(2). By linking subunits together in tandem dimers or tetramers containing mixtures of WT and K188A/R189A subunits, we demonstrate that one functional PIP(2)-interacting WT subunit is sufficient to convert channels from the unavailable to the available mode with a high open probability dominated by the fully open state, with similar kinetics as tetrameric WT channels. Occasional openings to sublevels become progressively less frequent as the number of WT subunits increases. Quantitative analysis reveals that the interaction of PIP(2) with WT subunits exerts strong positive cooperativity in both converting the channels from the unavailable to the available mode, and in promoting the fully open state over sublevels. We conclude that the interaction of PIP(2) with only one Kir2.1 subunit is sufficient for the channel to become available and to open to its full

  18. Fish IRF6 is a positive regulator of IFN expression and involved in both of the MyD88 and TBK1 pathways.

    PubMed

    Li, Shun; Lu, Long-Feng; Wang, Zhao-Xi; Chen, Dan-Dan; Zhang, Yong-An

    2016-10-01

    Interferon (IFN) regulatory factors (IRF) are the crucial transcription factors for IFN expression, leading host cell response to viral infection. In mammals, only IRF6 is unaffected by IFN expression in the IRF family; however, in fish, a lower vertebrate, whether IRF6 is related to IFN regulation is unclear. In this study, we identified that zebrafish IRF6 was a positive regulator of IFN transcription and could be phosphorylated by both MyD88 and TBK1. First, the transcript level of cellular irf6 was upregulated by treatment with poly I:C (a mimic of viral RNAs), indicating IRF6 might be involved in the process of host cell response to viruses. Overexpression of IRF6 could upregulate IFN promoter activity significantly, meaning IRF6 is a positive regulator of IFN transcription. Subsequently, at the protein regulation level and in the interaction relationship, IRF6 was phosphorylated by and associated with both MyD88 and TBK1. In addition, overexpression of IRF6 activated the transcription of isg15, rig-i and mavs of host cells; meanwhile, the transcripts of p, m and n genes of SVCV were significantly declined in IRF6-overexpressing cells. Taken together, our data demonstrate that fish IRF6 is distinguished from the homolog of mammals by being a positive regulator of IFN transcription and phosphorylated by MyD88 and TBK1, suggesting that differences in the IRF6 regulation pattern exist between lower and higher vertebrates.

  19. Positive regulation of the enzymatic activity of gastric H(+),K(+)-ATPase by sialylation of its β-subunit.

    PubMed

    Fujii, Takuto; Watanabe, Midori; Shimizu, Takahiro; Takeshima, Hiroshi; Kushiro, Keiichiro; Takai, Madoka; Sakai, Hideki

    2016-06-01

    The gastric proton pump (H(+),K(+)-ATPase) consists of a catalytic α-subunit (αHK) and a glycosylated β-subunit (βHK). βHK glycosylation is essential for the apical trafficking and stability of αHK in gastric parietal cells. Here, we report the properties of sialic acids at the termini of the oligosaccharide chains of βHK. Sialylation of βHK was found in LLC-PK1 cells stably expressing αHK and βHK by staining of the cells with lectin-tagged fluorescent polymeric nanoparticles. This sialylation was also confirmed by biochemical studies using sialic acid-binding lectin beads and an anti-βHK antibody. The sialic acids of βHK are cleaved enzymatically by neuraminidase (sialidase) and nonenzymatically by an acidic solution (pH5). Interestingly, the enzymatic activity of H(+),K(+)-ATPase was significantly decreased by cleavage of the sialic acids of βHK. In contrast, βHK was not sialylated in the gastric tubulovesicles prepared from the stomach of fed hogs. The H(+),K(+)-ATPase activity in these tubulovesicles was not significantly altered by neuraminidase. Importantly, the sialylation of βHK was observed in the gastric samples prepared from the stomach of famotidine (a histamine H2 receptor antagonist)-treated rats, but not histamine (an acid secretagogue)-treated rats. The enzymatic activity of H(+),K(+)-ATPase in the samples of the famotidine-treated rats was significantly higher than in the histamine-treated rats. The effects of famotidine were weakened by neuraminidase. These results indicate that βHK is sialylated at neutral or weakly acidic pH, but not at acidic pH, suggesting that the sialic acids of βHK positively regulate the enzymatic activity of αHK.

  20. Global Regulator IscR Positively Contributes to Antimonite Resistance and Oxidation in Comamonas testosteroni S44

    PubMed Central

    Liu, Hongliang; Zhuang, Weiping; Zhang, Shengzhe; Rensing, Christopher; Huang, Jun; Li, Jie; Wang, Gejiao

    2015-01-01

    Antimonial compounds can be found as a toxic contaminant in the environment. Knowledge on mechanisms of microbial Sb oxidation and its role in microbial tolerance are limited. Previously, we found that Comamonas testosteroni S44 was resistant to multiple heavy metals and was able to oxidize the toxic antimonite [Sb(III)] to the much less toxic antimonate [Sb(V)]. In this study, transposon mutagenesis was performed in C. testosteroni S44 to isolate genes responsible for Sb(III) resistance and oxidation. An insertion mutation into iscR, which regulates genes involved in the biosynthesis of Fe-S clusters, generated a strain called iscR-280. This mutant strain was complemented with a plasmid carrying iscR to generate strain iscR-280C. Compared to the wild type S44 and iscR-280C, strain iscR-280 showed lower resistance to Sb(III) and a lower Sb(III) oxidation rate. Strain iscR-280 also showed lower resistance to As(III), Cd(II), Cu(II), and H2O2. In addition, intracellular γ-glutamylcysteine ligase (γ-GCL) activity and glutathione (GSH) content were decreased in the mutated strain iscR-280. Real-time RT-PCR and lacZ fusion expression assay indicated that transcription of iscR and iscS was induced by Sb(III). Results of electrophoretic mobility shift assay (EMSA) and bacterial one-hybrid (B1H) system demonstrated a positive interaction between IscR and its promoter region. The diverse defective phenotypes and various expression patterns suggest a role for IscR in contributing to multi-metal(loid)s resistance and Sb(III) oxidation via Fe-S cluster biogenesis and oxidative stress protection. Bacterial Sb(III) oxidation is a detoxification reaction. PMID:26734615

  1. PUMILIO-2 Is Involved in the Positive Regulation of Cellular Proliferation in Human Adipose-Derived Stem Cells

    PubMed Central

    Shigunov, Patrícia; Kuligovski, Crisciele; de Aguiar, Alessandra Melo; Rebelatto, Carmen K.; Moutinho, José A.; Brofman, Paulo S.; Krieger, Marco A.; Goldenberg, Samuel; Munroe, David; Correa, Alejandro

    2012-01-01

    Stem cells can either differentiate into more specialized cells or undergo self-renewal. Several lines of evidence from different organisms suggest that these processes depend on the post-transcriptional regulation of gene expression. The presence of the PUF [Pumilio/FBF (fem-3 binding factor)] domain defines a conserved family of RNA binding proteins involved in repressing gene expression. It has been suggested that a conserved function of PUF proteins is to repress differentiation and sustain the mitotic proliferation of stem cells. In humans, Pumilio-2 (PUM2) is expressed in embryonic stem cells and adult germ cells. Here we show that PUM2 is expressed in a subpopulation of adipose-derived stem cell (ASC) cultures, with a granular pattern of staining in the cytoplasm. Protein levels of PUM2 showed no changes during the differentiation of ASCs into adipocytes. Moreover, RNAi knockdown of pum2 did not alter the rate of adipogenic differentiation compared with wild-type control cells. A ribonomic approach was used to identify PUM2-associated mRNAs. Microarray analysis showed that PUM2-bound mRNAs are part of gene networks involved in cell proliferation and gene expression control. We studied pum2 expression in cell cultures with low or very high levels of proliferation and found that changes in pum2 production were dependent on the proliferation status of the cell. Transient knockdown of pum2 expression by RNAi impaired proliferation of ASCs in vitro. Our results suggest that PUM2 does not repress differentiation of ASCs but rather is involved in the positive control of ASCs division and proliferation. PMID:21649561

  2. AP-1 regulates sphingosine kinase 1 expression in a positive feedback manner in glomerular mesangial cells exposed to high glucose.

    PubMed

    Huang, Kaipeng; Huang, Juan; Chen, Cheng; Hao, Jie; Wang, Shaogui; Huang, Junying; Liu, Peiqing; Huang, Heqing

    2014-03-01

    Our previous studies have confirmed that the sphingosine kinase 1 (SphK1)-sphingosine 1-phosphate (S1P) signaling pathway in the kidney under diabetic conditions is closely correlated with the pathogenesis of diabetic nephropathy (DN). The activation of SphK1-S1P pathway by high glucose (HG) can increase the expression of fibronectin (FN), an important fibrotic component, in glomerular mesangial cells (GMCs) by promoting the DNA-binding activity of transcription factor AP-1. However, the mechanism responsible for the sustained activation of SphK1-S1P pathway remains unclear. Given the binding motifs for AP-1 within the first intron of the SphK1 gene, we speculated that the activated AP-1 in the kidney under HG condition possibly regulates SphK1 expression in a positive feedback manner, thereby promoting the sustained activation of SphK1-S1P pathway and mediating the pathological progression of DN. Here, we observed the effect of AP-1 on SphK1 expression in GMCs and explored the molecular mechanism involved in the sustained activation of SphK1-S1P pathway. We found two consensus binding motifs for AP-1 in the promoter sequences and non-coding region downstream of the transcriptional initiation of the rat SphK1 gene by chromatin immunoprecipitation assay. The treatment of GMCs with both HG and S1P significantly increased the protein expression of c-Jun and c-Fos, and obviously enhanced the phosphorylation of c-Jun at Ser63 and Ser73, and c-Fos at Ser32. Knockdown of c-Jun and c-Fos with siRNAs substantially inhibited the expression of SphK1 and FN, whereas overexpression of c-Jun and c-Fos significantly increased the expression of SphK1 and FN. Curcumin treatment greatly decreased the levels of c-Jun, c-Fos, SphK1, and FN in the kidney tissues of diabetic rats. SiRNAs targeting SphK1 and S1P2 receptor respectively inhibited the phosphorylation of c-Jun (ser63 and ser73) and c-Fos (ser32), as well as FN expression under both normal and HG conditions. Our data

  3. Munc13-4 Is a Rab11-binding Protein That Regulates Rab11-positive Vesicle Trafficking and Docking at the Plasma Membrane*

    PubMed Central

    Johnson, Jennifer L.; He, Jing; Ramadass, Mahalakshmi; Pestonjamasp, Kersi; Kiosses, William B.; Zhang, Jinzhong; Catz, Sergio D.

    2016-01-01

    The small GTPase Rab11 and its effectors control trafficking of recycling endosomes, receptor replenishment and the up-regulation of adhesion and adaptor molecules at the plasma membrane. Despite recent advances in the understanding of Rab11-regulated mechanisms, the final steps mediating docking and fusion of Rab11-positive vesicles at the plasma membrane are not fully understood. Munc13-4 is a docking factor proposed to regulate fusion through interactions with SNAREs. In hematopoietic cells, including neutrophils, Munc13-4 regulates exocytosis in a Rab27a-dependent manner, but its possible regulation of other GTPases has not been explored in detail. Here, we show that Munc13-4 binds to Rab11 and regulates the trafficking of Rab11-containing vesicles. Using a novel Time-resolved Fluorescence Resonance Energy Transfer (TR-FRET) assay, we demonstrate that Munc13-4 binds to Rab11a but not to dominant negative Rab11a. Immunoprecipitation analysis confirmed the specificity of the interaction between Munc13-4 and Rab11, and super-resolution microscopy studies support the interaction of endogenous Munc13-4 with Rab11 at the single molecule level in neutrophils. Vesicular dynamic analysis shows the common spatio-temporal distribution of Munc13-4 and Rab11, while expression of a calcium binding-deficient mutant of Munc13-4 significantly affected Rab11 trafficking. Munc13-4-deficient neutrophils showed normal endocytosis, but the trafficking, up-regulation, and retention of Rab11-positive vesicles at the plasma membrane was significantly impaired. This correlated with deficient NADPH oxidase activation at the plasma membrane in response to Rab11 interference. Our data demonstrate that Munc13-4 is a Rab11-binding partner that regulates the final steps of Rab11-positive vesicle docking at the plasma membrane. PMID:26637356

  4. Munc13-4 Is a Rab11-binding Protein That Regulates Rab11-positive Vesicle Trafficking and Docking at the Plasma Membrane.

    PubMed

    Johnson, Jennifer L; He, Jing; Ramadass, Mahalakshmi; Pestonjamasp, Kersi; Kiosses, William B; Zhang, Jinzhong; Catz, Sergio D

    2016-02-12

    The small GTPase Rab11 and its effectors control trafficking of recycling endosomes, receptor replenishment and the up-regulation of adhesion and adaptor molecules at the plasma membrane. Despite recent advances in the understanding of Rab11-regulated mechanisms, the final steps mediating docking and fusion of Rab11-positive vesicles at the plasma membrane are not fully understood. Munc13-4 is a docking factor proposed to regulate fusion through interactions with SNAREs. In hematopoietic cells, including neutrophils, Munc13-4 regulates exocytosis in a Rab27a-dependent manner, but its possible regulation of other GTPases has not been explored in detail. Here, we show that Munc13-4 binds to Rab11 and regulates the trafficking of Rab11-containing vesicles. Using a novel Time-resolved Fluorescence Resonance Energy Transfer (TR-FRET) assay, we demonstrate that Munc13-4 binds to Rab11a but not to dominant negative Rab11a. Immunoprecipitation analysis confirmed the specificity of the interaction between Munc13-4 and Rab11, and super-resolution microscopy studies support the interaction of endogenous Munc13-4 with Rab11 at the single molecule level in neutrophils. Vesicular dynamic analysis shows the common spatio-temporal distribution of Munc13-4 and Rab11, while expression of a calcium binding-deficient mutant of Munc13-4 significantly affected Rab11 trafficking. Munc13-4-deficient neutrophils showed normal endocytosis, but the trafficking, up-regulation, and retention of Rab11-positive vesicles at the plasma membrane was significantly impaired. This correlated with deficient NADPH oxidase activation at the plasma membrane in response to Rab11 interference. Our data demonstrate that Munc13-4 is a Rab11-binding partner that regulates the final steps of Rab11-positive vesicle docking at the plasma membrane.

  5. Making an Effort to Feel Positive: Insecure Attachment in Infancy Predicts the Neural Underpinnings of Emotion Regulation in Adulthood

    ERIC Educational Resources Information Center

    Moutsiana, Christina; Fearon, Pasco; Murray, Lynne; Cooper, Peter; Goodyer, Ian; Johnstone, Tom; Halligan, Sarah

    2014-01-01

    Background: Animal research indicates that the neural substrates of emotion regulation may be persistently altered by early environmental exposures. If similar processes operate in human development then this is significant, as the capacity to regulate emotional states is fundamental to human adaptation. Methods: We utilised a 22-year longitudinal…

  6. Examining the Effects of Self-Reported Posttraumatic Stress Disorder Symptoms and Positive Relations with Others on Self-Regulated Learning for Student Service Members/Veterans

    ERIC Educational Resources Information Center

    Ness, Bryan M.; Middleton, Michael J.; Hildebrandt, Michael J.

    2015-01-01

    Objectives: To examine the relationships between self-reported posttraumatic stress disorder (PTSD) symptoms, perceived positive relations with others, self-regulation strategy use, and academic motivation among student service members/veterans (SSM/V) enrolled in postsecondary education. Participants: SSM/V (N = 214), defined as veterans, active…

  7. Developmental Trajectories of Intentional Self Regulation in Adolescence: The Role of Parenting and Implications for Positive and Problematic Outcomes among Diverse Youth

    ERIC Educational Resources Information Center

    Bowers, Edmond P.; Gestsdottir, Steinunn; Geldhof, G. John; Nikitin, Jana; von Eye, Alexander; Lerner, Richard M.

    2011-01-01

    This study assessed 1574 Grades 5 to 11 youth (63.6% female) from the 4-H Study of Positive Youth Development (PYD), a longitudinal study involving U.S. adolescents, to assess if patterns of intentional self regulation (ISR) existed; whether these trajectories differed in relation to several Grade 5 parenting characteristics; and whether ISR…

  8. The Effects of College Students' Positive Thinking, Learning Motivation and Self-Regulation through a Self-Reflection Intervention in Taiwan

    ERIC Educational Resources Information Center

    Wang, Hsin-Hui; Chen, Hsiang-Ting; Lin, Huann-Shyang; Hong, Zuway-R

    2017-01-01

    This quasi-experimental study examined the effects of a self-reflection intervention on college (college in this article refers to university-level education) students' positive thinking, learning motivation and self-regulation in Taiwan. One hundred and two college students were selected to participate in an 18-week intervention forming the…

  9. Tomato 14-3-3 protein 7 (TFT7) positively regulates immunity-associated programmed cell death by enhancing accumulation and signaling ability of MAPKKKalpha

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Programmed cell death (PCD) is triggered when Pto, a serine-threonine protein kinase recognizes either the AvrPto or AvrPtoB effector from Pseudomonas syringae pv. tomato. This PCD requires MAPKKKalpha as a positive regulator in tomato and Nicotiana benthamiana. To examine how PCD-eliciting activi...

  10. Transmembrane signalling at the epidermal growth factor receptor. Positive regulation by the C-terminal phosphotyrosine residues.

    PubMed Central

    Magni, M; Pandiella, A; Helin, K; Meldolesi, J; Beguinot, L

    1991-01-01

    Mutant epidermal growth factor (EGF) receptors (obtained by substitution of one, two or three C-terminal autophosphorylable tyrosine residues with phenylalanine residues or by deletion of the C-terminal 19 amino acids, including the distal tyrosine) were expressed in mouse NIH-3T3 fibroblast clones at densities comparable (less than 25% difference) with those in control clones expressing the wild-type receptor. Total EGF-induced phosphorylation of the mutated receptors was not appreciably changed with respect to controls, whereas autophosphorylation at tyrosine residues was decreased, especially in the double and the triple mutants. In the latter mutant, expression of the EGF-receptor-activated lipolytic enzyme phospholipase C gamma was unchanged, whereas its tyrosine phosphorylation induced by the growth factor was lowered to approx. 25% of that in the controls. In all of the cell clones employed, the accumulation of inositol phosphates induced by treatment with fetal calf serum varied only slightly, whereas the same effect induced by EGF was consistently lowered in those lines expressing mutated receptors. This decrease was moderate for those receptors missing only the distal tyrosine (point and deletion mutants), intermediate in the dual mutants and almost complete in the triple mutants. Likewise, increases in intracellular Ca2+ concentrations [( Ca2+]i) induced by fibroblast growth factor were approximately the same in all of the clones, whereas those induced by EGF were decreased in the mutants, again in proportion to the loss of the phosphorylable C-terminal tyrosine residues. The same trend occurred with membrane hyperpolarization, an effect secondary to the increase in [Ca2+]i via the activation of Ca2(+)-dependent K+ channels. We conclude that C-terminal autophosphorylable tyrosine residues play a positive role in the regulation of transmembrane signalling at the EGF receptor. The stepwise decrease in signal generation observed in single, double and triple

  11. The Escherichia coli L-arabinose operon: binding sites of the regulatory proteins and a mechanism of positive and negative regulation.

    PubMed

    Ogden, S; Haggerty, D; Stoner, C M; Kolodrubetz, D; Schleif, R

    1980-06-01

    The locations of DNA binding by the proteins involved with positive and negative regulation of transcription initiation of the L-arabinose operon in Escherichia coli have been determined by the DNase I protection method. Two cyclic AMP receptor protein sites were found, at positions -78 to -107 and -121 to -146, an araC protein--arabinose binding site was found at position -40 to -78, and an araC protein-fucose binding site was found at position -106 to -144. These locations, combined with in vivo data on induction of the two divergently oriented arabinose promoters, suggest the following regulatory mechanism: induction of the araBAD operon occurs when cyclic AMP receptor protein, araC protein, and RNA polymerase are all present and able to bind to DNA. Negative regulation is accomplished by the repressing form of araC protein binding to a site in the regulatory region such that it stimultaneously blocks access of cyclic AMP receptor protein to two sites on the DNA, one site of which serves each of the two promoters. Thus, from a single operator site, the negative regulator represses the two outwardly oriented ara promoters. This regulatory mechanism explains the known positive and negative regulatory properties of the ara promoters.

  12. Human telomerase reverse transcriptase regulates vascular endothelial growth factor expression via human papillomavirus oncogene E7 in HPV-18-positive cervical cancer cells.

    PubMed

    Li, Fang; Cui, Jinquan

    2015-07-01

    Human papillomavirus (HPV) infection induces chronic and precancerous lesions and results in invasive cervical cancer. Human telomerase as well as inflammatory and angiogenic factors such as telomerase reverse transcriptase (hTERT) or vascular endothelial growth factor (VEGF) could play a role in regulating HPV-induced cervical cancer. This study investigated underlying molecular events in HPV-induced HPV-positive cervical cancer through hTERT and VEGF in vitro. Expressions of hTERT, a rate-limiting subunit of telomerase, and VEGF mRNA and proteins were, respectively, assessed by qRT-PCR, ELISA, and TRAP-ELISA in HPV-positive tissue samples and cervical cancer cell lines. To assess hTERT and VEGF secretion, hTERT overexpression and knockdown were conducted in HPV-18-positive Hela cells by hTERT cDNA and shRNA transfection, respectively. Then, the effect of HPV E6 and E7 on VEGF expressions was assessed in HPV-negative cervical cancer cells. Data have shown that VEGF expression levels are associated with hTERT expressions and telomerase activity in HPV-positive cervical cancer tissues and cells. Knockdown of hTERT expression down-regulated VEGF expressions, whereas overexpression of hTERT up-regulated VEGF expressions in HPV-18-positive Hela cells. Furthermore, HPV E7 oncoprotein was necessary for hTERT to up-regulate VEGF expressions in HPV-negative cervical cancer cells. Data from this current study indicate that HPV oncoproteins up-regulated hTERT and telomerase activity and in turn promoted VEGF expressions, which could be a key mechanism for HPV-induced cervical cancer development and progression.

  13. Considering an affect regulation framework for examining the association between body dissatisfaction and positive body image in Black older adolescent females: does body mass index matter?

    PubMed

    Webb, Jennifer B; Butler-Ajibade, Phoebe; Robinson, Seronda A

    2014-09-01

    The present study provided an initial evaluation of an affect regulation model describing the association between body dissatisfaction and two contemporary measures of positive body image among 247 Black college-bound older adolescent females. We further tested whether possessing a higher body mass index (BMI) would strengthen these associations. Self-reported height and weight were used to calculate BMI. Respondents also completed a culturally-sensitive figure rating scale along with assessments of body appreciation and body image flexibility. Results indicated a robust positive association between the two measures of positive body image; BMI was the strongest predictor of both body appreciation and body image flexibility with body size discrepancy (current minus ideal) contributing incremental variance to both models tested. Implications for improving our understanding of the association between positive and negative body image and bolstering positive body image to promote health-protective behaviors among Black young women at this developmental juncture are discussed.

  14. Considering an Affect Regulation Framework for Examining the Association Between Body Dissatisfaction and Positive Body Image in Black Older Adolescent Females: Does Body Mass Index Matter?

    PubMed Central

    Butler-Ajibade, Phoebe; Robinson, Seronda A.

    2014-01-01

    The present study provided an initial evaluation of an affect regulation model describing the association between body dissatisfaction and two contemporary measures of positive body image among 247 Black college-bound older adolescent females. We further tested whether possessing a higher body mass index (BMI) would strengthen these associations. Self-reported height and weight were used to calculate BMI. Respondents also completed a culturally-sensitive figure rating scale along with assessments of body appreciation and body image flexibility. Results indicated a robust positive association between the two measures of positive body image; BMI was the strongest predictor of both body appreciation and body image flexibility with body size discrepancy (current minus ideal) contributing incremental variance to both models tested. Implications for improving our understanding of the association between positive and negative body image and bolstering positive body image to promote health-protective behaviors among Black young women at this developmental juncture are discussed. PMID:25079011

  15. The E3 deubiquitinase USP17 is a positive regulator of retinoic acid-related orphan nuclear receptor γt (RORγt) in Th17 cells.

    PubMed

    Han, Lei; Yang, Jing; Wang, Xiuwen; Wu, Qingsi; Yin, Shuying; Li, Zhiyuan; Zhang, Jing; Xing, Yue; Chen, Zuojia; Tsun, Andy; Li, Dan; Piccioni, Miranda; Zhang, Yu; Guo, Qiang; Jiang, Lindi; Bao, Liming; Lv, Ling; Li, Bin

    2014-09-12

    Stable retinoic acid-related orphan nuclear receptor γt (RORγt) expression is pivotal for the development and function of Th17 cells. Here we demonstrate that expression of the transcription factor RORγt can be regulated through deubiquitination, which prevents proteasome-mediated degradation. We establish that USP17 stabilizes RORγt protein expression by reducing RORγt polyubiquitination at its Lys-360 residue. In contrast, knockdown of endogenous USP17 in Th17 cells resulted in decreased RORγt protein levels and down-regulation of Th17-related genes. Furthermore, USP17 expression was up-regulated in CD4(+) T cells from systemic lupus erythematosus patients. Our data reveal a molecular mechanism in which RORγt expression in Th17 cells can be positively regulated by USP17, thereby modulating Th17 cell functions.

  16. Induction of GST-P-positive proliferative lesions facilitating lipid peroxidation with possible involvement of transferrin receptor up-regulation and ceruloplasmin down-regulation from the early stage of liver tumor promotion in rats.

    PubMed

    Mizukami, Sayaka; Ichimura, Ryohei; Kemmochi, Sayaka; Taniai, Eriko; Shimamoto, Keisuke; Ohishi, Takumi; Takahashi, Miwa; Mitsumori, Kunitoshi; Shibutani, Makoto

    2010-04-01

    To elucidate the role of metal-related molecules in hepatocarcinogenesis, we examined immunolocalization of transferrin receptor (Tfrc), ceruloplasmin (Cp) and metallothionein (MT)-1/2 in relation to liver cell foci positive for glutathione-S-transferase placental form (GST-P) in the early stage of tumor promotion by fenbendazole (FB), phenobarbital, piperonyl butoxide or thioacetamide in a rat two-stage hepatocarcinogenesis model. To estimate the involvement of oxidative stress responses to the promotion, immunolocalization of 4-hydroxy-2-nonenal, malondialdehyde and acrolein was similarly examined. Our findings showed that MT-1/2 immunoreactivity was not associated with the cellular distribution of GST-P and proliferating cell nuclear antigen, suggesting no role of MT-1/2 in hepatocarcinogenesis. We also found enhanced expression of Tfrc after treatment with strong tumor-promoting chemicals. With regard to Cp, the population showing down-regulation was increased in the GST-P-positive foci in relation to tumor promotion. Up-regulation of Tfrc and down-regulation of Cp was maintained in GST-P-positive neoplastic lesions induced after long-term promotion with FB, suggesting the expression changes occurring downstream of the signaling pathway involved in the formation of GST-P-positive lesions. Furthermore, enhanced accumulation of lipid peroxidation end products was observed in the GST-P-positive foci by promotion. Post-initiation treatment with peroxisome proliferator-activated receptor alpha agonists did not enhance any such distribution changes in GST-P-negative foci. The results thus suggest that facilitation of lipid peroxidation is involved in the induction of GST-P-positive lesions by tumor promotion from an early stage, and up-regulation of Tfrc and down-regulation of Cp may be a signature of enhanced oxidative cellular stress in these lesions.

  17. Reassessment of the Genetic Regulation of Fatty Acid Synthesis in Escherichia coli: Global Positive Control by the Dual Functional Regulator FadR

    PubMed Central

    My, L.; Ghandour Achkar, N.; Viala, J. P.

    2015-01-01

    ABSTRACT In Escherichia coli, the FadR transcriptional regulator represses the expression of fatty acid degradation (fad) genes. However, FadR is also an activator of the expression of fabA and fabB, two genes involved in unsaturated fatty acid synthesis. Therefore, FadR plays an important role in maintaining the balance between saturated and unsaturated fatty acids in the membrane. We recently showed that FadR also activates the promoter upstream of the fabH gene (L. My, B. Rekoske, J. J. Lemke, J. P. Viala, R. L. Gourse, and E. Bouveret, J Bacteriol 195:3784–3795, 2013, doi:10.1128/JB.00384-13). Furthermore, recent transcriptomic and proteomic data suggested that FadR activates the majority of fatty acid (FA) synthesis genes. In the present study, we tested the role of FadR in the expression of all genes involved in FA synthesis. We found that FadR activates the transcription of all tested FA synthesis genes, and we identified the FadR binding site for each of these genes. This necessitated the reassessment of the transcription start sites for accA and accB genes described previously, and we provide evidence for the presence of multiple promoters driving the expression of these genes. We showed further that regulation by FadR impacts the amount of FA synthesis enzymes in the cell. Our results show that FadR is a global regulator of FA metabolism in E. coli, acting both as a repressor of catabolism and an activator of anabolism, two directly opposing pathways. IMPORTANCE In most bacteria, a transcriptional regulator tunes the level of FA synthesis enzymes. Oddly, such a global regulator still was missing for E. coli, which nonetheless is one of the prominent model bacteria used for engineering biofuel production using the FA synthesis pathway. Our work identifies the FadR functional dual regulator as a global activator of almost all FA synthesis genes in E. coli. Because FadR also is the repressor of FA degradation, FadR acts both as a repressor and an activator

  18. Emotion Risk-Factor in Patients With Cardiac Diseases: The Role of Cognitive Emotion Regulation Strategies, Positive Affect and Negative Affect (A Case-Control Study)

    PubMed Central

    Bahremand, Mostafa; Alikhani, Mostafa; Zakiei, Ali; Janjani, Parisa; Aghaei, Abbas

    2016-01-01

    Application of psychological interventions is essential in classic treatments for patient with cardiac diseases. The present study compared cognitive emotion regulation strategies, positive affect, and negative affect for cardiac patients with healthy subjects. This study was a case-control study. Fifty subjects were selected using convenient sampling method from cardiac (coronary artery disease) patients presenting in Imam Ali medical center of Kermanshah, Iran in the spring 2013. Fifty subjects accompanied the patients to the medical center, selected as control group, did not have any history of cardiac diseases. For collecting data, the cognitive emotion regulation questionnaire and positive and negative affect scales were used. For data analysis, multivariate analysis of variance (MANOVA) was applied using the SPSS statistical software (ver. 19.0). In all cognitive emotion regulation strategies, there was a significant difference between the two groups. A significant difference was also detected regarding positive affect between the two groups, but no significant difference was found regarding negative affect. We found as a result that, having poor emotion regulation strategies is a risk factor for developing heart diseases. PMID:26234976

  19. Transgenic analysis reveals LeACS-1 as a positive regulator of ethylene-induced shikonin biosynthesis in Lithospermum erythrorhizon hairy roots.

    PubMed

    Fang, Rongjun; Wu, Fengyao; Zou, Ailan; Zhu, Yu; Zhao, Hua; Zhao, Hu; Liao, Yonghui; Tang, Ren-Jie; Yang, Tongyi; Pang, Yanjun; Wang, Xiaoming; Yang, Rongwu; Qi, Jinliang; Lu, Guihua; Yang, Yonghua

    2016-03-01

    The phytohormone ethylene (ET) is a crucial signaling molecule that induces the biosynthesis of shikonin and its derivatives in Lithospermum erythrorhizon shoot cultures. However, the molecular mechanism and the positive regulators involved in this physiological process are largely unknown. In this study, the function of LeACS-1, a key gene encoding the 1-aminocyclopropane-1-carboxylic acid synthase for ET biosynthesis in L. erythrorhizon hairy roots, was characterized by using overexpression and RNA interference (RNAi) strategies. The results showed that overexpression of LeACS-1 significantly increased endogenous ET concentration and shikonin production, consistent with the up-regulated genes involved in ET biosynthesis and transduction, as well as the genes related to shikonin biosynthesis. Conversely, RNAi of LeACS-1 effectively decreased endogenous ET concentration and shikonin production and down-regulated the expression level of above genes. Correlation analysis showed a significant positive linear relationship between ET concentration and shikonin production. All these results suggest that LeACS-1 acts as a positive regulator of ethylene-induced shikonin biosynthesis in L. erythrorhizon hairy roots. Our work not only gives new insights into the understanding of the relationship between ET and shikonin biosynthesis, but also provides an efficient genetic engineering target gene for secondary metabolite production in non-model plant L. erythrorhizon.

  20. MYB98 Positively Regulates a Battery of Synergid-Expressed Genes Encoding Filiform Apparatus–Localized Proteins[W

    PubMed Central

    Punwani, Jayson A.; Rabiger, David S.; Drews, Gary N.

    2007-01-01

    The synergid cells within the female gametophyte are essential for reproduction in angiosperms. MYB98 encodes an R2R3-MYB protein required for pollen tube guidance and filiform apparatus formation by the synergid cells. To test the predicted function of MYB98 as a transcriptional regulator, we determined its subcellular localization and examined its DNA binding properties. We show that MYB98 binds to a specific DNA sequence (TAAC) and that a MYB98–green fluorescent protein fusion protein localizes to the nucleus, consistent with a role in transcriptional regulation. To identify genes regulated by MYB98, we tested previously identified synergid-expressed genes for reduced expression in myb98 female gametophytes and identified 16 such genes. We dissected the promoter of one of the downstream genes, DD11, and show that it contains a MYB98 binding site required for synergid expression, suggesting that DD11 is regulated directly by MYB98. To gain insight into the functions of the downstream genes, we chose five genes and determined the subcellular localization of the encoded proteins. We show that these five proteins are secreted into the filiform apparatus, suggesting that they play a role in either the formation or the function of this unique structure. Together, these data suggest that MYB98 functions as a transcriptional regulator in the synergid cells and activates the expression of genes required for pollen tube guidance and filiform apparatus formation. PMID:17693534

  1. Pepper CabZIP63 acts as a positive regulator during Ralstonia solanacearum or high temperature-high humidity challenge in a positive feedback loop with CaWRKY40.

    PubMed

    Shen, Lei; Liu, Zhiqin; Yang, Sheng; Yang, Tong; Liang, Jiaqi; Wen, Jiayu; Liu, Yanyan; Li, Jiazhi; Shi, Lanping; Tang, Qian; Shi, Wei; Hu, Jiong; Liu, Cailing; Zhang, Yangwen; Lin, Wei; Wang, Rongzhang; Yu, Huanxin; Mou, Shaoliang; Hussain, Ansar; Cheng, Wei; Cai, Hanyang; He, Li; Guan, Deyi; Wu, Yang; He, Shuilin

    2016-04-01

    CaWRKY40 is known to act as a positive regulator in the response of pepper (Capsicum annuum) to Ralstonia solanacearum inoculation (RSI) or high temperature-high humidity (HTHH), but the underlying mechanism remains elusive. Herein, we report that CabZIP63, a pepper bZIP family member, participates in this process by regulating the expression of CaWRKY40. CabZIP63 was found to localize in the nuclei, be up-regulated by RSI or HTHH, bind to promoters of both CabZIP63(pCabZIP63) and CaWRKY40(pCaWRKY40), and activate pCabZIP63- and pCaWRKY40-driven β-glucuronidase expression in a C- or G-box-dependent manner. Silencing of CabZIP63 by virus-induced gene silencing (VIGS) in pepper plants significantly attenuated their resistance to RSI and tolerance to HTHH, accompanied by down-regulation of immunity- or thermotolerance-associated CaPR1, CaNPR1, CaDEF1, and CaHSP24. Hypersensitive response-mediated cell death and expression of the tested immunity- and thermotolerance-associated marker genes were induced by transient overexpression (TOE) of CabZIP63, but decreased by that of CabZIP63-SRDX. Additionally, binding of CabZIP63 to pCaWRKY40 was up-regulated by RSI or HTHH, and the transcript level of CaWRKY40 and binding of CaWRKY40 to the promoters of CaPR1, CaNPR1, CaDEF1 and CaHSP24 were up-regulated by TOE of CabZIP63. On the other hand, CabZIP63 was also up-regulated transcriptionally by TOE of CaWRKY40. The data suggest collectively that CabZIP63 directly or indirectly regulates the expression of CaWRKY40 at both the transcriptional and post-transcriptional level, forming a positive feedback loop with CaWRKY40 during pepper's response to RSI or HTHH. Altogether, our data will help to elucidate the underlying mechanism of crosstalk between pepper's response to RSI and HTHH.

  2. Pepper CabZIP63 acts as a positive regulator during Ralstonia solanacearum or high temperature–high humidity challenge in a positive feedback loop with CaWRKY40

    PubMed Central

    Shen, Lei; Liu, Zhiqin; Yang, Sheng; Yang, Tong; Liang, Jiaqi; Wen, Jiayu; Liu, Yanyan; Li, Jiazhi; Shi, Lanping; Tang, Qian; Shi, Wei; Hu, Jiong; Liu, Cailing; Zhang, Yangwen; Lin, Wei; Wang, Rongzhang; Yu, Huanxin; Mou, Shaoliang; Hussain, Ansar; Cheng, Wei; Cai, Hanyang; He, Li; Guan, Deyi; Wu, Yang; He, Shuilin

    2016-01-01

    CaWRKY40 is known to act as a positive regulator in the response of pepper (Capsicum annuum) to Ralstonia solanacearum inoculation (RSI) or high temperature–high humidity (HTHH), but the underlying mechanism remains elusive. Herein, we report that CabZIP63, a pepper bZIP family member, participates in this process by regulating the expression of CaWRKY40. CabZIP63 was found to localize in the nuclei, be up-regulated by RSI or HTHH, bind to promoters of both CabZIP63 (pCabZIP63) and CaWRKY40 (pCaWRKY40), and activate pCabZIP63- and pCaWRKY40-driven β-glucuronidase expression in a C- or G-box-dependent manner. Silencing of CabZIP63 by virus-induced gene silencing (VIGS) in pepper plants significantly attenuated their resistance to RSI and tolerance to HTHH, accompanied by down-regulation of immunity- or thermotolerance-associated CaPR1, CaNPR1, CaDEF1, and CaHSP24. Hypersensitive response-mediated cell death and expression of the tested immunity- and thermotolerance-associated marker genes were induced by transient overexpression (TOE) of CabZIP63, but decreased by that of CabZIP63-SRDX. Additionally, binding of CabZIP63 to pCaWRKY40 was up-regulated by RSI or HTHH, and the transcript level of CaWRKY40 and binding of CaWRKY40 to the promoters of CaPR1, CaNPR1, CaDEF1 and CaHSP24 were up-regulated by TOE of CabZIP63. On the other hand, CabZIP63 was also up-regulated transcriptionally by TOE of CaWRKY40. The data suggest collectively that CabZIP63 directly or indirectly regulates the expression of CaWRKY40 at both the transcriptional and post-transcriptional level, forming a positive feedback loop with CaWRKY40 during pepper’s response to RSI or HTHH. Altogether, our data will help to elucidate the underlying mechanism of crosstalk between pepper’s response to RSI and HTHH. PMID:26936828

  3. Analyzing the soybean transcriptome during autoregulation of mycorrhization identifies the transcription factors GmNF-YA1a/b as positive regulators of arbuscular mycorrhization

    PubMed Central

    2013-01-01

    Background Similarly to the legume-rhizobia symbiosis, the arbuscular mycorrhiza interaction is controlled by autoregulation representing a feedback inhibition involving the CLAVATA1-like receptor kinase NARK in shoots. However, little is known about signals and targets down-stream of NARK. To find NARK-related transcriptional changes in mycorrhizal soybean (Glycine max) plants, we analyzed wild-type and two nark mutant lines interacting with the arbuscular mycorrhiza fungus Rhizophagus irregularis. Results Affymetrix GeneChip analysis of non-inoculated and partially inoculated plants in a split-root system identified genes with potential regulation by arbuscular mycorrhiza or NARK. Most transcriptional changes occur locally during arbuscular mycorrhiza symbiosis and independently of NARK. RT-qPCR analysis verified nine genes as NARK-dependently regulated. Most of them have lower expression in roots or shoots of wild type compared to nark mutants, including genes encoding the receptor kinase GmSIK1, proteins with putative function as ornithine acetyl transferase, and a DEAD box RNA helicase. A predicted annexin named GmAnnx1a is differentially regulated by NARK and arbuscular mycorrhiza in distinct plant organs. Two putative CCAAT-binding transcription factor genes named GmNF-YA1a and GmNF-YA1b are down-regulated NARK-dependently in non-infected roots of mycorrhizal wild-type plants and functional gene analysis confirmed a positive role for these genes in the development of an arbuscular mycorrhiza symbiosis. Conclusions Our results indicate GmNF-YA1a/b as positive regulators in arbuscular mycorrhiza establishment, whose expression is down-regulated by NARK in the autoregulated root tissue thereby diminishing subsequent infections. Genes regulated independently of arbuscular mycorrhization by NARK support an additional function of NARK in symbioses-independent mechanisms. PMID:23777981

  4. Calpain-controlled detachment of major glycoproteins from the cytoskeleton regulates adhesive properties of activated phosphatidylserine-positive platelets.

    PubMed

    Artemenko, Elena O; Yakimenko, Alena O; Pichugin, Alexey V; Ataullakhanov, Fazly I; Panteleev, Mikhail A

    2016-02-15

    In resting platelets, adhesive membrane glycoproteins are attached to the cytoskeleton. On strong activation, phosphatidylserine(PS)-positive and -negative platelet subpopulations are formed. Platelet activation is accompanied by cytoskeletal rearrangement, although the glycoprotein attachment status in these two subpopulations is not clear. We developed a new, flow cytometry-based, single-cell approach to investigate attachment of membrane glycoproteins to the cytoskeleton in cell subpopulations. In PS-negative platelets, adhesive glycoproteins integrin αIIbβ3, glycoprotein Ib and, as shown for the first time, P-selectin were associated with the cytoskeleton. In contrast, this attachment was disrupted in PS-positive platelets; it was retained to some extent only in the small convex regions or 'caps'. It correlated with the degradation of talin and filamin observed only in PS-positive platelets. Calpain inhibitors essentially prevented the disruption of membrane glycoprotein attachment in PS-positive platelets, as well as talin and filamin degradation. With the suggestion that detachment of glycoproteins from the cytoskeleton may affect platelet adhesive properties, we investigated the ability of PS-positive platelets to resist shear-induced breakaway from the immobilized fibrinogen. Shear rates of 500/s caused PS-positive platelet breakaway, but their adhesion stability increased more than 10-fold after pretreatment of the platelets with calpain inhibitor. In contrast, the ability of PS-positive platelets to adhere to immobilized von Willebrand's factor at 100/s was low, but this was not affected by the preincubation of platelets with a calpain inhibitor. Our data suggest that calpain-controlled detachment of membrane glycoproteins is a new mechanism that is responsible for the loss of ability of the procoagulant platelets to resist detachment from thrombi by high shear stress.

  5. Characterization of LnmO as a pathway-specific Crp/Fnr-type positive regulator for leinamycin biosynthesis in Streptomyces atroolivaceus and its application for titer improvement.

    PubMed

    Huang, Yong; Yang, Dong; Pan, Guohui; Tang, Gong-Li; Shen, Ben

    2016-12-01

    The cyclic adenosine monophosphate (cAMP) receptor protein/fumarate and nitrate reductase regulatory protein (Crp/Fnr) family of transcriptional regulators are pleiotropic transcriptional regulators that control a broad range of cellular functions. Leinamycin (LNM) is a potent antitumor antibiotic produced by Streptomyces atroolivaceus S-140. We previously cloned and characterized the lnm biosynthetic gene cluster from S. atroolivaceus S-140. We here report inactivation of lnmO in S. atroolivaceus S-140 and overexpression of lnmO in the S. atroolivaceus S-140 wild-type and ∆lnmE mutant SB3033 to investigate its role in LNM biosynthesis. Bioinformatics analysis revealed LnmO as the only regulator within the lnm gene cluster, exhibiting high sequence similarity to known Crp/Fnr family regulators. The inactivation of lnmO in S. atroolivaceus S-140 completely abolished LNM production but caused no apparent morphological changes, supporting that LnmO is indispensable and specific to LNM biosynthesis. Overexpression of lnmO in S. atroolivaceus S-140 and SB3033 resulted in three- and fourfold increase in LNM and LNM E1 production, respectively, supporting that LnmO acts as a positive regulator. While all of the Crp/Fnr family regulators studied to date appeared to be pleiotropic, our results support LnmO as the first Crp/Fnr family regulator that is pathway-specific. LnmO joins the growing list of regulators that could be exploited to improve secondary metabolite production in Streptomyces. Engineered strains overproducing LNM and LNM E1 will facilitate further mechanistic studies and clinical evaluation of LNM and LNM E1 as novel anticancer drugs.

  6. Gene expression profiling reveals novel regulation by bisphenol-A in estrogen receptor-{alpha}-positive human cells

    SciTech Connect

    Singleton, David W.; Feng, Yuxin; Yang, Jun; Puga, Alvaro; Lee, Adrian V.; Khan, Sohaib A. . E-mail: sohaib.khan@uc.edu

    2006-01-15

    Bisphenol-A (BPA) shows proliferative actions in uterus and mammary glands and may influence the development of male and female reproductive tracts in utero or during early postnatal life. Because of its ability to function as an estrogen receptor (ER) agonist, BPA has the potential to disrupt normal endocrine signaling through regulation of ER target genes. Some genes are regulated by both estradiol (E2) and BPA, but those exclusive to either agent have not been described. Using a yeast strain incorporating a vitellogenin A2 ERE-LacZ reporter gene into the genome, we found that BPA induced expression of the reporter in colonies transformed with the ER{alpha} expression plasmid, illustrating BPA-mediated regulation within a chromatin context. Additionally, a reporter gene transiently transfected into the endometrial cancer (Ishikawa) cell line also showed BPA activity, although at 100-fold less potency than E2. To compare global gene expression in response to BPA and E2, we used a variant of the MCF-7 breast cancer cell line stably expressing HA-tagged ER{alpha}. Cultures were treated for 3 h with an ethanol vehicle, E2 (10{sup -8} M), or BPA (10{sup -6} M), followed by isolation of RNA and microarray analysis with the human U95A probe array (Affymetrix, Santa Clara, CA, USA). More than 300 genes were changed 2-fold or more by either or both agents, with roughly half being up-regulated and half down-regulated. A number of growth- and development-related genes, such as HOXC1 and C6, Wnt5A, Frizzled, TGF{beta}-2, and STAT inhibitor 2, were found to be affected exclusively by BPA. We used quantitative real-time PCR to verify regulation of the HOXC6 gene, which showed decreased expression of approximately 2.5-fold by BPA. These results reveal novel effects by BPA and E2, raising interesting possibilities regarding the role of endocrine disruptors in sexual development.

  7. MG428 is a novel positive regulator of recombination that triggers mgpB and mgpC gene variation in Mycoplasma genitalium.

    PubMed

    Burgos, Raul; Totten, Patricia A

    2014-10-01

    The human pathogen Mycoplasma genitalium employs homologous recombination to generate antigenic diversity in the immunodominant MgpB and MgpC proteins. Only recently, some of the molecular factors involved in this process have been characterized, but nothing is known about its regulation. Here, we show that M. genitalium expresses N-terminally truncated RecA isoforms via alternative translation initiation, but only the full-length protein is essential for gene variation. We also demonstrate that overexpression of MG428 positively regulates the expression of recombination genes, including recA, ruvA, ruvB and ORF2, a gene of unknown function co-transcribed with ruvAB. The co-ordinated induction of these genes correlated with an increase of mgpBC gene variation. In contrast, cells lacking MG428 were unable to generate variants despite expressing normal levels of RecA. Similarly, deletion analyses of the recA upstream region defined sequences required for gene variation without abolishing RecA expression. The requirement of these sequences is consistent with the presence of promoter elements associated with MG428-dependent recA induction. Sequences upstream of recA also influence the relative abundance of RecA isoforms, possibly through translational regulation. Overall, these results suggest that MG428 is a positive regulator of recombination and that precise control of recA expression is required to initiate mgpBC variation.

  8. Auxin promotes the transition from chloronema to caulonema in moss protonema by positively regulating PpRSL1and PpRSL2 in Physcomitrella patens.

    PubMed

    Jang, Geupil; Dolan, Liam

    2011-10-01

    Protonemata are multicellular filamentous networks that develop following the germination of a haploid moss spore and comprise two different cell types - chloronema and caulonema. The ROOT HAIR DEFECTIVE SIX-LIKE1 (PpRSL1) and PpRSL2 basic helix-loop-helix transcription factors and auxin promote the development of caulonema in Physcomitrella patens but the mechanism by which these regulators interact during development is unknown. We characterized the role of auxin in regulating the function of PpRSL1 and PpRSL2 in the chloronema-to-caulonema transition during protonema development. Here, we showed that a gradient of cell identity developed along protonemal filaments; cells were chloronemal in proximal regions near the site of spore germination becoming progressively more caulonemal distally as filaments elongated. Auxin controlled this transition by positively regulating the expression of PpRSL1 and PpRSL2 genes. Auxin did not induce caulonemal development in Pprsl1 Pprsl2 double mutants that lack PpRSL1 and PpRSL2 gene activity while constitutive co-expression of PpRSL1 and PpRSL2 in the absence of auxin was sufficient to program constitutive caulonema development. Together, these data indicate that auxin positively regulates PpRSL1 and PpRSL2 whose expression is sufficient to promote caulonema differentiation in moss protonema.

  9. The oncogenic transcription factor IRF4 is regulated by a novel CD30/NF-κB positive feedback loop in peripheral T-cell lymphoma

    PubMed Central

    Boddicker, Rebecca L.; Kip, N. Sertac; Xing, Xiaoming; Zeng, Yu; Yang, Zhi-Zhang; Lee, Jeong-Heon; Almada, Luciana L.; Elsawa, Sherine F.; Knudson, Ryan A.; Law, Mark E.; Ketterling, Rhett P.; Cunningham, Julie M.; Wu, Yanhong; Maurer, Matthew J.; O’Byrne, Megan M.; Cerhan, James R.; Slager, Susan L.; Link, Brian K.; Porcher, Julie C.; Grote, Deanna M.; Jelinek, Diane F.; Dogan, Ahmet; Ansell, Stephen M.; Fernandez-Zapico, Martin E.

    2015-01-01

    Peripheral T-cell lymphomas (PTCLs) are generally aggressive non-Hodgkin lymphomas with poor overall survival rates following standard therapy. One-third of PTCLs express interferon regulatory factor-4 (IRF4), a tightly regulated transcription factor involved in lymphocyte growth and differentiation. IRF4 drives tumor growth in several lymphoid malignancies and has been proposed as a candidate therapeutic target. Because direct IRF4 inhibitors are not clinically available, we sought to characterize the mechanism by which IRF4 expression is regulated in PTCLs. We demonstrated that IRF4 is constitutively expressed in PTCL cells and drives Myc expression and proliferation. Using an inhibitor screen, we identified nuclear factor κB (NF-κB) as a candidate regulator of IRF4 expression and cell proliferation. We then demonstrated that the NF-κB subunits p52 and RelB were transcriptional activators of IRF4. Further analysis showed that activation of CD30 promotes p52 and RelB activity and subsequent IRF4 expression. Finally, we showed that IRF4 transcriptionally regulates CD30 expression. Taken together, these data demonstrate a novel positive feedback loop involving CD30, NF-κB, and IRF4; further evidence for this mechanism was demonstrated in human PTCL tissue samples. Accordingly, NF-κB inhibitors may represent a clinical means to disrupt this feedback loop in IRF4-positive PTCLs. PMID:25833963

  10. MAPK Hog1 closes the S. cerevisiae glycerol channel Fps1 by phosphorylating and displacing its positive regulators

    PubMed Central

    Lee, Jongmin; Reiter, Wolfgang; Dohnal, Ilse; Gregori, Christa; Beese-Sims, Sara; Kuchler, Karl; Ammerer, Gustav; Levin, David E.

    2013-01-01

    The aquaglyceroprin Fps1 is responsible for glycerol transport in yeast in response to changes in extracellular osmolarity. Control of Fps1 channel activity in response to hyperosmotic shock involves a redundant pair of regulators, Rgc1 (regulator of the glycerol channel 1) and Rgc2, and the MAPK Hog1 (high-osmolarity glycerol response 1). However, the mechanism by which these factors influence channel activity is unknown. We show that Rgc2 maintains Fps1 in the open channel state in the absence of osmotic stress by binding to its C-terminal cytoplasmic domain. This interaction involves a tripartite pleckstrin homology (PH) domain within Rgc2 and a partial PH domain within Fps1. Activation of Hog1 in response to hyperosmotic shock induces the rapid eviction of Rgc2 from Fps1 and consequent channel closure. Hog1 was recruited to the N-terminal cytoplasmic domain of Fps1, which it uses as a platform from which to multiply phosphorylate Rgc2. Thus, these results reveal the mechanism by which Hog1 regulates Fps1 in response to hyperosmotic shock. PMID:24298058

  11. EVI1-mediated down regulation of MIR449A is essential for the survival of EVI1 positive leukaemic cells.

    PubMed

    De Weer, An; Van der Meulen, Joni; Rondou, Pieter; Taghon, Tom; Konrad, Torsten A; De Preter, Katleen; Mestdagh, Pieter; Van Maerken, Tom; Van Roy, Nadine; Jeison, Marta; Yaniv, Isaac; Cauwelier, Barbara; Noens, Lucien; Poirel, Hélène-Antoine; Vandenberghe, Peter; Lambert, Frédéric; De Paepe, Anne; Sánchez, Maria García; Odero, Maria; Verhasselt, Bruno; Philippé, Jan; Vandesompele, Joke; Wieser, Rotraud; Dastugue, Nicole; Van Vlierberghe, Pieter; Poppe, Bruce; Speleman, Frank

    2011-08-01

    Chromosomal rearrangements involving the MECOM (MDS1 and EVI1 complex) locus are recurrent genetic events in myeloid leukaemia and are associated with poor prognosis. In this study, we assessed the role of MECOM locus protein EVI1 in the transcriptional regulation of microRNAs (miRNAs) involved in the leukaemic phenotype. For this, we profiled expression of 366 miRNAs in 38 MECOM-rearranged patient samples, normal bone marrow controls and MECOM (EVI1) knock down/re-expression models. Cross-comparison of these miRNA expression profiling data showed that MECOM rearranged leukaemias are characterized by down regulation of MIR449A. Reconstitution of MIR449A expression in MECOM-rearranged cell line models induced apoptosis resulting in a strong decrease in cell viability. These effects might be mediated in part by MIR449A regulation of NOTCH1 and BCL2, which are shown here to be bona fide MIR449A targets. Finally, we confirmed that MIR449A repression is mediated through direct promoter occupation of the EVI1 transcriptional repressor. In conclusion, this study reveals MIR449A as a crucial direct target of the MECOM locus protein EVI1 involved in the pathogenesis of MECOM-rearranged leukaemias and unravels NOTCH1 and BCL2 as important novel targets of MIR449A. This EVI1-MIR449A-NOTCH1/BCL2 regulatory axis might open new possibilities for the development of therapeutic strategies in this poor prognostic leukaemia subgroup.

  12. Glyceollin, a novel regulator of mTOR/p70S6 in estrogen receptor positive breast cancer

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An estimated 70% of breast cancer tumors utilize estrogen receptor (ER) signaling to maintain tumorigenesis, and targeting of the estrogen receptor is a common method of treatment for these tumor types. However, ER-positive (+) breast cancers often acquire drug resistant or altered ER activity in r...

  13. Youth Development Program Participation and Intentional Self-Regulation Skills: Contextual and Individual Bases of Pathways to Positive Youth Development

    ERIC Educational Resources Information Center

    Mueller, Megan Kiely; Phelps, Erin; Bowers, Edmond P.; Agans, Jennifer P.; Urban, Jennifer Brown; Lerner, Richard M.

    2011-01-01

    The present research used data from Grades 8, 9, and 10 of the 4-H Study of Positive Youth Development, a longitudinal study involving U.S. adolescents, in order to better elucidate the process through which the strengths of youth and the ecological resources promoting healthy development (such as out-of-school-time programs) may contribute to…

  14. Oestrogen receptor-α contributes to the regulation of the hedgehog signalling pathway in ERα-positive gastric cancer

    PubMed Central

    Kameda, C; Nakamura, M; Tanaka, H; Yamasaki, A; Kubo, M; Tanaka, M; Onishi, H; Katano, M

    2010-01-01

    Background: Oestrogen receptor-alpha (ERα) is highly expressed in diffuse-type gastric cancer and oestrogen increases the proliferation of ERα-positive gastric cancer. However, a detailed mechanism by which oestrogen increases the proliferation of these cells is still unclear. Methods: We used 17-β-oestradiol (E2) as a stimulator against the ERα pathway. Pure anti-oestrogen drug ICI 182 780 (ICI) and small interfering RNA against ERα (ERα siRNA) were used as inhibitors. Cyclopamine (Cyc) was used as the hedgehog (Hh) pathway inhibitor. Two human ERα-positive gastric cancer cells were used as target cells. Effects of the stimulator and inhibitor on E2-induced cell proliferation were also examined. Results: In ERα-positive cells, E2 increased not only cell proliferation but also one of the ligands of the Hh pathway, Shh expression. 17-β-Oestradiol-induced cell proliferation was suppressed by ICI, ERα siRNA or Cyc. The increased expression of Shh induced by E2 was suppressed by ICI and ERα siRNA but not by Cyc. Furthermore, recombinant Shh activated the Hh pathway and increased cell proliferation, whereas anti-Shh antibody suppressed E2-induced cell proliferation. When a relationship between ERα and Shh expressions was analysed using surgically resected gastric cancer specimens, a positive correlation was found, suggesting a linkage between the ERα and Hh pathways. Conclusion: Our data indicate that activation of the ERα pathway promotes cell proliferation by activating the Hh pathway in a ligand-dependent manner through Shh induction of ERα-positive gastric cancer. PMID:20087349

  15. Restoration of microRNA-214 expression reduces growth of myeloma cells through positive regulation of P53 and inhibition of DNA replication

    PubMed Central

    Misiewicz-Krzeminska, Irena; Sarasquete, María E.; Quwaider, Dalia; Krzeminski, Patryk; Ticona, Fany V.; Paíno, Teresa; Delgado, Manuel; Aires, Andreia; Ocio, Enrique M.; García-Sanz, Ramón; San Miguel, Jesús F.; Gutiérrez, Norma C.

    2013-01-01

    MicroRNA have been demonstrated to be deregulated in multiple myeloma. We have previously reported that miR-214 is down-regulated in multiple myeloma compared to in normal plasma cells. The functional role of miR-214 in myeloma pathogenesis was explored by transfecting myeloma cell lines with synthetic microRNA followed by gene expression profiling. Putative miR-214 targets were validated by luciferase reporter assay. Ectopic expression of miR-214 reduced cell growth and induced apoptosis of myeloma cells. In order to identify the potential direct target genes of miR-214 which could be involved in the biological pathways regulated by this microRNA, gene expression profiling of the H929 myeloma cell line transfected with precursor miR-214 was carried out. Functional analysis revealed significant enrichment for DNA replication, cell cycle phase and DNA binding. miR-214 directly down-regulated the expression of PSMD10, which encodes the oncoprotein gankyrin, and ASF1B, a histone chaperone required for DNA replication, by binding to their 3'-untranslated regions. In addition, gankyrin inhibition induced an increase of P53 mRNA levels and subsequent up-regulation of CDKN1A (p21Waf1/Cip1) and BAX transcripts, which are direct transcriptional targets of p53. In conclusion, MiR-214 functions as a tumor suppressor in myeloma by positive regulation of p53 and inhibition of DNA replication. PMID:23100276

  16. Expression and clinical significance of estrogen-regulated long non-coding RNAs in estrogen receptor α-positive ovarian cancer progression.

    PubMed

    Qiu, Jun-Jun; Ye, Le-Chi; Ding, Jing-Xin; Feng, Wei-Wei; Jin, Hong-Yan; Zhang, Ying; Li, Qing; Hua, Ke-Qin

    2014-04-01

    Estrogen (E2) has long been implicated in epithelial ovarian cancer (EOC) progression. The effects of E2 on cancer progression can be mediated by numerous target genes, including coding RNAs and, more recently, non-coding RNAs (ncRNAs). Among the ncRNAs, long ncRNAs (lncRNAs) have emerged as new regulators in cancer progression; therefore, our aim was to determine whether the expression of any lncRNAs is regulated by E2 and, if so, whether a subset of these lncRNAs have some clinical significance in EOC progression. A microarray was performed to identify E2-regulated lncRNAs in E2 receptor (ER) α-positive EOC cells. Bioinformatics analyses of lncRNAs were conducted, focusing on gene ontology and pathway analyses. Quantitative real-time polymerase chain reactions were performed to confirm the expression of certain lncRNAs in ERα-positive EOC tissues. The correlation between certain lncRNA expression and clinicopathological factors as well as prognosis in ERα-positive EOC patients was then analyzed. We showed that 115 lncRNAs exhibited significant changes in E2-treated SKOV3 cells compared with untreated controls. Most of these lncRNAs were predicated to have potential to contribute to cancer progression. Notably, three candidates (TC0100223, TC0101686 and TC0101441) were aberrantly expressed in ERα-positive compared to ERα-negative EOC tissues, showing correlations with some malignant cancer phenotypes such as advanced FIGO stage and/or high histological grade. Furthermore, multivariate analysis indicated that TC0101441 was an independent prognostic factor for overall survival. Taken together, these results indicate for the first time that E2 can modulate lncRNA expression in ERα-positive EOC cells and that certain lncRNAs are correlated with advanced cancer progression and suggestive of a prognostic indicator in ERα-positive EOC patients. Knowledge of these E2-regulated lncRNAs could aid in the future understanding of the estrogenic effect on EOC progression

  17. FGF/FGFR2 signaling regulates the generation and correct positioning of Bergmann glia cells in the developing mouse cerebellum.

    PubMed

    Meier, Florian; Giesert, Florian; Delic, Sabit; Faus-Kessler, Theresa; Matheus, Friederike; Simeone, Antonio; Hölter, Sabine M; Kühn, Ralf; Weisenhorn, Daniela M Vogt; Wurst, Wolfgang; Prakash, Nilima

    2014-01-01

    The normal cellular organization and layering of the vertebrate cerebellum is established during embryonic and early postnatal development by the interplay of a complex array of genetic and signaling pathways. Disruption of these processes and of the proper layering of the cerebellum usually leads to ataxic behaviors. Here, we analyzed the relative contribution of Fibroblast growth factor receptor 2 (FGFR2)-mediated signaling to cerebellar development in conditional Fgfr2 single mutant mice. We show that during embryonic mouse development, Fgfr2 expression is higher in the anterior cerebellar primordium and excluded from the proliferative ventricular neuroepithelium. Consistent with this finding, conditional Fgfr2 single mutant mice display the most prominent defects in the anterior lobules of the adult cerebellum. In this context, FGFR2-mediated signaling is required for the proper generation of Bergmann glia cells and the correct positioning of these cells within the Purkinje cell layer, and for cell survival in the developing cerebellar primordium. Using cerebellar microexplant cultures treated with an FGFR agonist (FGF9) or antagonist (SU5402), we also show that FGF9/FGFR-mediated signaling inhibits the outward migration of radial glia and Bergmann glia precursors and cells, and might thus act as a positioning cue for these cells. Altogether, our findings reveal the specific functions of the FGFR2-mediated signaling pathway in the generation and positioning of Bergmann glia cells during cerebellar development in the mouse.

  18. The inflammation-related gene S100A12 is positively regulated by C/EBPβ and AP-1 in pigs.

    PubMed

    Li, Xinyun; Tang, Juan; Xu, Jing; Zhu, Mengjin; Cao, Jianhua; Liu, Ying; Yu, Mei; Zhao, Shuhong

    2014-08-08

    S100A12 is involved in the inflammatory response and is considered an important marker for many inflammatory diseases in humans. Our previous studies indicated that the S100A12 gene was abundant in the immune tissues of pigs and was significantly upregulated during infection with Haemophilus parasuis (HPS) or porcine circovirus type 2 (PCV2). In this study, the mechanism of transcriptional regulation of S100A12 was investigated in pigs. Our results showed that S100A12, CCAAT/enhancer-binding protein beta (C/EBPβ) and activator protein-1 (AP-1) genes were up-regulated in PK-15 (ATCC, CCL-33) cells when treated with LPS or Poly I: C. Additionally, the promoter activity and expression level of the S100A12 gene were significantly upregulated when C/EBPβ or AP-1 were overexpressed. We utilized electromobility shift assays (EMSA) to confirm that C/EBPβ and AP-1 could directly bind the S100A12 gene promoter. We also found that the transcriptional activity and expression levels of C/EBPβ and AP-1 could positively regulate each other. Furthermore, the promoter activity of the S100A12 gene was higher when C/EBPβ and AP-1 were cotransfected than when they were transfected individually. We concluded that the S100A12 gene was cooperatively and positively regulated by C/EBPβ and AP-1 in pigs. Our study offers new insight into the transcriptional regulation of the S100A12 gene.

  19. Gene Positioning

    PubMed Central

    Ferrai, Carmelo; de Castro, Inês Jesus; Lavitas, Liron; Chotalia, Mita; Pombo, Ana

    2010-01-01

    Eukaryotic gene expression is an intricate multistep process, regulated within the cell nucleus through the activation or repression of RNA synthesis, processing, cytoplasmic export, and translation into protein. The major regulators of gene expression are chromatin remodeling and transcription machineries that are locally recruited to genes. However, enzymatic activities that act on genes are not ubiquitously distributed throughout the nucleoplasm, but limited to specific and spatially defined foci that promote preferred higher-order chromatin arrangements. The positioning of genes within the nuclear landscape relative to specific functional landmarks plays an important role in gene regulation and disease. PMID:20484389

  20. TaTypA, a Ribosome-Binding GTPase Protein, Positively Regulates Wheat Resistance to the Stripe Rust Fungus

    PubMed Central

    Liu, Peng; Myo, Thwin; Ma, Wei; Lan, Dingyun; Qi, Tuo; Guo, Jia; Song, Ping; Guo, Jun; Kang, Zhensheng

    2016-01-01

    Tyrosine phosphorylation protein A (TypA/BipA) belongs to the ribosome-binding GTPase superfamily. In many bacterial species, TypA acts as a global stress and virulence regulator and also mediates resistance to the antimicrobial peptide bactericidal permeability-increasing protein. However, the function of TypA in plants under biotic stresses is not known. In this study, we isolated and functionally characterized a stress-responsive TypA gene (TaTypA) from wheat, with three copies located on chromosomes 6A, 6B, and 6D, respectively. Transient expression assays indicated chloroplast localization of TaTypA. The transcript levels of TaTypA were up-regulated in response to treatment with methyl viologen, which induces reactive oxygen species (ROS) in chloroplasts through photoreaction, cold stress, and infection by an avirulent strain of the stripe rust pathogen. Knock down of the expression of TaTypA through virus-induced gene silencing decreased the resistance of wheat to stripe rust accompanied by weakened ROS accumulation and hypersensitive response, an increase in TaCAT and TaSOD expression, and an increase in pathogen hyphal growth and branching. Our findings suggest that TaTypA contributes to resistance in an ROS-dependent manner. PMID:27446108

  1. Dietary fish oil positively regulates plasma leptin and adiponectin levels in sucrose-fed, insulin-resistant rats.

    PubMed

    Rossi, Andrea S; Lombardo, Yolanda B; Lacorte, Jean-Marc; Chicco, Adriana G; Rouault, Christine; Slama, Gérard; Rizkalla, Salwa W

    2005-08-01

    Insulin resistance and adiposity induced by a long-term sucrose-rich diet (SRD) in rats could be reversed by fish oil (FO). Regulation of plasma leptin and adiponectin levels, as well as their gene expression, by FO might be implicated in these findings. This study was designed to evaluate the long-term regulation of leptin and adiponectin by dietary FO in a dietary model of insulin resistance induced by long-term SRD in rats and to determine their impact on adiposity and insulin sensitivity. Rats were randomized to consume a control diet (CD; n = 25) or an SRD (n = 50) for 7 mo. Subsequently, the SRD-fed rats were randomized to consume SRD+FO or to continue on SRD for an additional 2 mo. Long-term SRD induced overweight and decreased both plasma leptin and adiponectin levels without change in gene expression. Dyslipidemia, adiposity, and insulin resistance accompanied these modifications. Shifting the source of fat to FO for 2 mo increased plasma levels of both adipokines, reversed insulin resistance and dyslipidemia, and improved adiposity. These results were not associated with modifications in gene expression. These results suggest that increasing both adipokines by dietary FO might play an essential role in the normalization of insulin resistance and adiposity in dietary-induced, insulin-resistant models.

  2. The size and internal structure of a heterochromatic block determine its ability to induce position effect variegation in Drosophila melanogaster.

    PubMed Central

    Tolchkov, E V; Rasheva, V I; Bonaccorsi, S; Westphal, T; Gvozdev, V A

    2000-01-01

    In the In(1LR)pn2a rearrangement, the 1A-2E euchromatic segment is transposed to the vicinity of X heterochromatin (Xh), resulting in position effect variegation (PEV) of the genes in the 2BE region. Practically the whole X-linked heterochromatin is situated adjacent to variegated euchromatic genes. Secondary rearrangements showing weakening or reversion of PEV were obtained by irradiation of the In(1LR)pn2a. These rearrangements demonstrate a positive correlation between the strength of PEV of the wapl locus and the sizes of the adjacent heterochromatic blocks carrying the centromere. The smallest PEV-inducing fragment consists of a block corresponding to approximately 10% of Xh and containing the entire XR, the centromere, and a very proximal portion of XL heterochromatin. Heterochromatic blocks retaining the entire XR near the 2E region, but lacking the centromere, show no PEV. Reversion of PEV was also observed as a result of an internal rearrangement of the Xh blocks where the centromere is moved away from the eu-heterochromatin boundary but the amount of X heterochromatin remaining adjacent to 2E is unchanged. We propose a primary role of the X pericentromeric region in PEV induction and an enhancing effect of the other blocks, positively correlated with their size. PMID:10747057

  3. The heat-inducible essential response regulator WalR positively regulates transcription of sigI, mreBH and lytE in Bacillus subtilis under heat stress.

    PubMed

    Huang, Wan-Zhen; Wang, Jyun-Jhih; Chen, Hui-Ju; Chen, Jung-Tze; Shaw, Gwo-Chyuan

    2013-12-01

    The actin homolog MreBH governs cell morphogenesis of Bacillus subtilis through localization of the cell wall hydrolase LytE. The alternative sigma factor SigI of B. subtilis coordinately regulates transcription of mreBH and lytE. Transcription of sigI, mreBH and lytE is heat-inducible. The essential response regulator WalR (YycF) plays a key role in coordinating cell wall metabolism with cell proliferation. We now demonstrate that mreBH is a new member of the WalR regulon. We also found that WalR can positively and directly regulate sigI transcription under heat stress through a binding site located upstream of the σ(I) promoter of sigI. In addition, we found that a WalR binding site located upstream of the SigI binding site in the regulatory region of lytE is important for lytE expression under heat stress. Moreover, we found that walR is a new member of the heat shock stimulon of B. subtilis. WalR appears to coordinately and positively regulate transcription of sigI, mreBH and lytE under heat stress. Finally, our work shows for the first time that WalR can stimulate activities of σ(I) promoters under heat stress.

  4. The Zebrafish Period2 Protein Positively Regulates the Circadian Clock through Mediation of Retinoic Acid Receptor (RAR)-related Orphan Receptor α (Rorα)*

    PubMed Central

    Wang, Mingyong; Zhong, Zhaomin; Zhong, Yingbin; Zhang, Wei; Wang, Han

    2015-01-01

    We report the characterization of a null mutant for zebrafish circadian clock gene period2 (per2) generated by transcription activator-like effector nuclease and a positive role of PER2 in vertebrate circadian regulation. Locomotor experiments showed that per2 mutant zebrafish display reduced activities under light-dark and 2-h phase delay under constant darkness, and quantitative real time PCR analyses showed up-regulation of cry1aa, cry1ba, cry1bb, and aanat2 but down-regulation of per1b, per3, and bmal1b in per2 mutant zebrafish, suggesting that Per2 is essential for the zebrafish circadian clock. Luciferase reporter assays demonstrated that Per2 represses aanat2 expression through E-box and enhances bmal1b expression through the Ror/Rev-erb response element, implicating that Per2 plays dual roles in the zebrafish circadian clock. Cell transfection and co-immunoprecipitation assays revealed that Per2 enhances bmal1b expression through binding to orphan nuclear receptor Rorα. The enhancing effect of mouse PER2 on Bmal1 transcription is also mediated by RORα even though it binds to REV-ERBα. Moreover, zebrafish Per2 also appears to have tissue-specific regulatory roles in numerous peripheral organs. These findings help define the essential functions of Per2 in the zebrafish circadian clock and in particular provide strong evidence for a positive role of PER2 in the vertebrate circadian system. PMID:25544291

  5. The environmental endocrine disruptor p-nitrophenol interacts with FKBP51, a positive regulator of androgen receptor and inhibits androgen receptor signaling in human cells.

    PubMed

    Wu, Dan; Tao, Xuanyu; Chen, Zhi-Peng; Han, Jian-Ting; Jia, Wen-Juan; Zhu, Ning; Li, Xiangkai; Wang, Zhiping; He, Yong-Xing

    2016-04-15

    The compound p-nitrophenol, which shows the anti-androgenic activity, can easily become anthropogenic pollutants and pose a threat to the environment and human health. Previous work indicates that the anti-androgenic mechanism of p-nitrophenol is complex and may involve several components in the AR signaling pathway, but the molecular details of how p-nitrophenol inhibits AR signaling are still not quite clear. Here, we characterized p-nitrophenol binds to the FK1 domain of an AR positive regulator FKBP51 with micromolar affinity and structural analysis of FK1 domain in complex with p-nitrophenol revealed that p-nitrophenol occupies a hydrophobic FK1 pocket that is vital for AR activity enhancement. Molecular dynamics simulation indicated that p-nitrophenol is stably bound to the FK1 pocket and the hotspot residues that involved p-nitrophenol binding are mainly hydrophobic and overlap with the AR interaction site. Furthermore, we showed that p-nitrophenol inhibits the androgen-dependent growth of human prostate cancer cells, possibly through down-regulating the expression levels of AR activated downstream genes. Taken together, our data suggests that p-nitrophenol suppresses the AR signaling pathway at least in part by blocking the interaction between AR and its positive regulator FKBP51. We believe that our findings could provide new guidelines for assessing the potential health effects of p-nitrophenol.

  6. The use of nanometer tetrabasic lead sulfate as positive active material additive for valve regulated lead-acid battery

    NASA Astrophysics Data System (ADS)

    Lang, Xiaoshi; Wang, Dianlong; Hu, Chiyu; Tang, Shenzhi; Zhu, Junsheng; Guo, Chenfeng

    2014-12-01

    Conventional tetrabasic lead sulfate used as positive active material additive shows the results of the low effective lead dioxide conversion rate due to the large grain size and crossed the crystal structure. In this paper, we study on a type of nanometer tetrabasic lead sulfate. Through the XRD and SEM test and Material Studio software calculation, the purity of tetrabasic lead sulfate is very high, the grain size of the nanometer 4BS is almost unanimous, and can be controlled below 200 nm. When charged and discharged in 1.75 V-2.42 V with the current density of 40 mA g-1, 80 mA g-1 and 160 mA g-1, the effective lead dioxide conversion rate of nanometer 4BS after formation can achieve to 83.48%, 71.42%, and 66.96%. Subsequently, the nanometer 4BS as additive is added to positive paste of lead-acid battery. When the batteries are tested galvanostatically between 1.75 V and 2.42 V at 0.25 C charge and 0.5 C discharge rates at room temperature. The ratio of adding nanometer 4BS is 0%, 1% and 4% and the initial discharge specific capacities are 60 mAh g-1, 65 mAh g-1 and 68 mAh g-1. After 80 cycles, the initial discharge capacity of positive active material with 1% nanometer 4BS decreased less than 10%, while adding 4% nanometer 4BS, the initial discharge capacity doesn't decrease obviously.

  7. The Highly Conserved Proline at Position 438 in Pseudorabies Virus gH Is Important for Regulation of Membrane Fusion

    PubMed Central

    Schröter, Christina; Klupp, Barbara G.; Fuchs, Walter; Gerhard, Marika; Backovic, Marija; Rey, Felix A.

    2014-01-01

    ABSTRACT Membrane fusion in herpesviruses requires viral glycoproteins (g) gB and gH/gL. While gB is considered the actual fusion protein but is nonfusogenic per se, the function of gH/gL remains enigmatic. Crystal structures for different gH homologs are strikingly similar despite only moderate amino acid sequence conservation. A highly conserved sequence motif comprises the residues serine-proline-cysteine corresponding to positions 437 to 439 in pseudorabies virus (PrV) gH. The PrV-gH structure shows that proline438 induces bending at the end of an alpha-helix, thereby placing cysteine404 and cysteine439 in juxtaposition to allow formation of a strictly conserved disulfide bond. However, PrV vaccine strain Bartha unexpectedly carries a serine at this conserved position. To test the influence of this substitution, we constructed different gH chimeras carrying proline or serine at position 438 in gH derived from either PrV strain Kaplan or strain Bartha. Mutants expressing gH with serine438 showed reduced fusion activity in transient-fusion assays and during infection, with delayed penetration kinetics and a small-plaque phenotype which indicates that proline438 is important for efficient fusion. A more drastic effect was observed when disulfide bond formation was completely blocked by mutation of cysteine404 to serine. Although PrV expressing gHC404S was viable, plaque size and penetration kinetics were drastically reduced. Alteration of serine438 to proline in gH of strain Bartha enhanced cell-to-cell spread and penetration kinetics, but restoration of full activity required additional alteration of aspartic acid to valine at position 59. IMPORTANCE The role of the gH/gL complex in herpesvirus membrane fusion is still unclear. Structural studies predicted a critical role for proline438 in PrV gH to allow the formation of a conserved disulfide bond and correct protein folding. Functional analyses within this study corroborated these structural predictions

  8. Delays Induce Different Switch in a Stochastic Single Genetic Regulation System with a Positive Autoregulatory Feedback Loop

    NASA Astrophysics Data System (ADS)

    Wang, Can-Jun

    2013-04-01

    The steady properties of a stochastic single genetic regulation system with the different time delays, which appear in the deterministic and fluctuating forces, are investigated based on the small delay time approximation method. Using the approximation probability density approach, the delayed Fokker-Planck equation is obtained. The effects of two different time delays on the stationary probability distribution and the mean value are discussed. It is found that with the time delay τ1 in the deterministic force increasing, the TF-A monomer concentration shifts from "off" state to "on" state. However, with the time delay τ2 in the fluctuating force increasing, the TF-A monomer concentration shifts from "on" state to "off" state. In the switch process, two kinds of time delays play an opposite role. The theoretical predictions are found to be in good agreement with numerical results.

  9. PTP1D is a positive regulator of the prolactin signal leading to beta-casein promoter activation.

    PubMed Central

    Ali, S; Chen, Z; Lebrun, J J; Vogel, W; Kharitonenkov, A; Kelly, P A; Ullrich, A

    1996-01-01

    Stimulation of the prolactin receptor (PRLR), a member of the cytokine/growth hormone receptor family, results in activation of the associated Jak2 tyrosine kinase and downstream signaling pathways. We report that PTP1D, a cytoplasmic protein tyrosine phosphatase containing two Src homology 2 (SH2) domains, physically associates with the PRLR-Jak2 complex and is tyrosine-phosphorylated upon stimulation with prolactin. The formation of the trimeric PRLR-Jak2-PTP1D complex is critical for transmission of a lactogenic signal, while PTP1D phosphorylation is necessary, but not sufficient. The dominant negative inhibitory effect of a phosphatase-deficient mutant on expression of a beta-casein promoter-controlled reporter gene is evidence for an essential role of fully functional PTP1D in the regulation of milk protein gene transcription. Images PMID:8598196

  10. Death associated protein 1 (DAP 1) positively regulates virus replication and apoptosis of hemocytes in shrimp Marsupenaeus japonicus.

    PubMed

    Xia, Wen-Li; Kang, Li-Hua; Liu, Chang-Bin; Kang, Cui-Jie

    2017-04-01

    Death-associated protein 1 (DAP1) is a small proline-rich cytoplasmic protein that functions both in the apoptosis and autophage process of mammalian and in the clinical cancer of human. However, little knowledge is known about the homologue gene of DAP1 and its roles in the physiological process of invertebrates. In this paper, we report a novel function of DAP1 in the antivirus immunity of shrimp. A homologue gene of DAP1 was cloned from Marsupenaeus japonicus and named as Mjdap-1. The full-length of Mjdap-1 was 1761 bp with a 309 bp open reading frame that encoded 102 amino acids. Reverse transcription-PCR results showed that Mjdap-1 was expressed in all tested tissues, including hemocytes, gills, intestines, stomach, heart, hepatopancreas, testes, and ovaries. In shrimp, Mjdap-1 transcripts were up-regulated by white spot syndrome virus (WSSV) infection; Mjdap-1 knockdown decreased the virus copy in vivo and the mortality of M. japonicus to WSSV challenge. Conversely, injecting the purified recombinant MjDAP1 protein promoted the amplification of virus in shrimp. Flow cytometric assay showed, the virus infection-induced apoptosis of hemocytes was enhanced by MjDAP1 protein injection and inhibited in MjDAP1 knockdown shrimp. Furthermore, the expression of apoptosis-inducing factor (AIF) was regulated by Mjdap-1, but the caspase transcripts were not affected. Our results suggested that MjDAP1 facilitated the amplification of virus in shrimp, which may be attributed to the promotion of hemocyte apoptosis in an AIF-dependent manner. These results provided a new insight into the function of this protein that may be used for virus disease control.

  11. RNAi screen in Drosophila larvae identifies histone deacetylase 3 as a positive regulator of the hsp70 heat shock gene expression during heat shock

    PubMed Central

    Achary, Bhavana G.; Campbell, Katie M.; Co, Ivy S.; Gilmour, David S.

    2014-01-01

    Transcription regulation of the Drosophila hsp70 gene is a complex process that involves regulation of multiple steps including establishment of paused Pol II and release of Pol II into elongation upon heat shock activation. While the major players involved in regulation of gene expression have been studied in detail, additional factors involved in this process continue to be discovered. To identify factors involved in hsp70 expression, we developed a screen that capitalizes on a visual assessment of heat shock activation using a hsp70-beta galactosidase reporter and publicly available RNAi fly lines to deplete candidate proteins. We validated the screen by showing that depletion of HSF, CycT, Cdk9, Nurf 301, or ELL prevented full induction of hsp70 by heat shock. Our screen also identified the histone deacetylase HDAC3 and its associated protein SMRTER as positive regulators of hsp70 activation. Additionally we show that HDAC3 and SMRTER contribute to hsp70 gene expression at a step subsequent to HSF-mediated activation and release of the paused Pol II that resides at the promoter prior to heat shock induction. PMID:24607507

  12. RNAi screen in Drosophila larvae identifies histone deacetylase 3 as a positive regulator of the hsp70 heat shock gene expression during heat shock.

    PubMed

    Achary, Bhavana G; Campbell, Katie M; Co, Ivy S; Gilmour, David S

    2014-05-01

    The transcription regulation of the Drosophila hsp70 gene is a complex process that involves the regulation of multiple steps, including the establishment of paused Pol II and release of Pol II into elongation upon heat shock activation. While the major players involved in the regulation of gene expression have been studied in detail, additional factors involved in this process continue to be discovered. To identify factors involved in hsp70 expression, we developed a screen that capitalizes on a visual assessment of heat shock activation using a hsp70-beta galactosidase reporter and publicly available RNAi fly lines to deplete candidate proteins. We validated the screen by showing that the depletion of HSF, CycT, Cdk9, Nurf 301, or ELL prevented the full induction of hsp70 by heat shock. Our screen also identified the histone deacetylase HDAC3 and its associated protein SMRTER as positive regulators of hsp70 activation. Additionally, we show that HDAC3 and SMRTER contribute to hsp70 gene expression at a step subsequent to HSF-mediated activation and release of the paused Pol II that resides at the promoter prior to heat shock induction.

  13. MILI, a PIWI-interacting RNA-binding protein, is required for germ line stem cell self-renewal and appears to positively regulate translation.

    PubMed

    Unhavaithaya, Yingdee; Hao, Yi; Beyret, Ergin; Yin, Hang; Kuramochi-Miyagawa, Satomi; Nakano, Toru; Lin, Haifan

    2009-03-06

    The Argonaute/PIWI protein family consists of Argonaute and PIWI subfamilies. Argonautes function in RNA interference and micro-RNA pathways; whereas PIWIs bind to PIWI-interacting RNAs and regulate germ line development, stem cell maintenance, epigenetic regulation, and transposition. However, the role of PIWIs in mammalian stem cells has not been demonstrated, and molecular mechanisms mediated by PIWIs remain elusive. Here we show that MILI, a murine PIWI protein, is expressed in the cytoplasm of testicular germ line stem cells, spermatogonia, and early spermatocytes, where it is enriched in chromatoid bodies. MILI is essential for the self-renewing division and differentiation of germ line stem cells but does not affect initial establishment of the germ line stem cell population at 7 days postpartum. Furthermore, MILI forms a stable RNA-independent complex with eIF3a and associates with the eIF4E- and eIF4G-containing m7G cap-binding complex. In isolated 7 days postpartum seminiferous tubules containing mostly germ line stem cells, the mili mutation has no effect on the cellular mRNA level yet significantly reduces the rate of protein synthesis. These observations indicate that MILI may positively regulate translation and that such regulation is required for germ line stem cell self-renewal.

  14. Epigenetic mechanisms and associated brain circuits in the regulation of positive emotions: A role for transposable elements.

    PubMed

    Gaudi, Simona; Guffanti, Guia; Fallon, James; Macciardi, Fabio

    2016-10-15

    Epigenetic programming and reprogramming are at the heart of cellular differentiation and represent developmental and evolutionary mechanisms in both germline and somatic cell lines. Only about 2% of our genome is composed of protein-coding genes, while the remaining 98%, once considered "junk" DNA, codes for regulatory/epigenetic elements that control how genes are expressed in different tissues and across time from conception to death. While we already know that epigenetic mechanisms are at play in cancer development and in regulating metabolism (cellular and whole body), the role of epigenetics in the developing prenatal and postnatal brain, and in maintaining a proper brain activity throughout the various stages of life, in addition to having played a critical role in human evolution, is a relatively new domain of knowledge. Here we present the current state-of-the-art techniques and results of these studies within the domain of emotions, and then speculate on how genomic and epigenetic mechanisms can modify and potentially alter our emotional (limbic) brain and affect our social interactions. J. Comp. Neurol. 524:2944-2954, 2016. © 2016 Wiley Periodicals, Inc.

  15. Positive regulation of the Shewanella oneidensis OmpS38, a major porin facilitating anaerobic respiration, by Crp and Fur

    PubMed Central

    Gao, Tong; Ju, Lili; Yin, Jianhua; Gao, Haichun

    2015-01-01

    Major porins are among the most abundant proteins embedded in the outer membrane (OM) of Gram-negative bacteria, playing crucial roles in maintenance of membrane structural integrity and OM permeability. Although many OM proteins (especially c-type cytochromes) in Shewanella oneidensis, a research model for respiratory versatility, have been extensively studied, physiological significance of major porins remains largely unexplored. In this study, we show that OmpS38 and OmpA are two major porins, neither of which is responsive to changes in osmolarity or contributes to the intrinsic resistance to β-lactam antibiotics. However, OmpS38 but not OmpA is largely involved in respiration of non-oxygen electron acceptors. We then provide evidence that expression of ompS38 is transcribed from two promoters, the major of which is favored under anaerobic conditions while the other appears constitutive. The major promoter is under the direct control of Crp, the master regulator dictating respiration. As a result, the increase in the level of OmpS38 correlates with an elevated activity in Crp under anaerobic conditions. In addition, we show that the activity of the major promoter is also affected by Fur, presumably indirectly, the transcription factor for iron-dependent gene expression. PMID:26381456

  16. A constitutive active MAPK/ERK pathway due to BRAFV600E positively regulates AHR pathway in PTC

    PubMed Central

    Regazzo, Daniela; Bertazza, Loris; Galuppini, Francesca; Guzzardo, Vincenza; Jaffrain-Rea, Marie Lise; Vianello, Federica; Ciato, Denis; Ceccato, Filippo; Watutantrige-Fernando, Sara; Bisognin, Andrea; Bortoluzzi, Stefania; Pennelli, Gianmaria; Boscaro, Marco; Scaroni, Carla; Mian, Caterina

    2015-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor mediating the toxicity and tumor-promoting properties of dioxin. AHR has been reported to be overexpressed and constitutively active in a variety of solid tumors, but few data are currently available concerning its role in thyroid cancer. In this study we quantitatively explored a series of 51 paired-normal and papillary thyroid carcinoma (PTC) tissues for AHR-related genes. We identified an increased AHR expression/activity in PTC, independently from its nuclear dimerization partner and repressor but strictly related to a constitutive active MAPK/ERK pathway. The AHR up-regulation followed by an increased expression of AHR target genes was confirmed by a meta-analysis of published microarray data, suggesting a ligand-independent active AHR pathway in PTC. In-vitro studies using a PTC-derived cell line (BCPAP) and HEK293 cells showed that BRAFV600E may directly modulate AHR localization, induce AHR expression and activity in an exogenous ligand-independent manner. The AHR pathway might represent a potential novel therapeutic target for PTC in the clinical practice. PMID:26392334

  17. Glycoproteomic Approach Identifies KRAS as a Positive Regulator of CREG1 in Non-small Cell Lung Cancer Cells

    PubMed Central

    Clark, David J.; Mei, Yuping; Sun, Shisheng; Zhang, Hui; Yang, Austin J.; Mao, Li

    2016-01-01

    Protein glycosylation plays a fundamental role in a multitude of biological processes, and the associated aberrant expression of glycoproteins in cancer has made them attractive biomarkers and therapeutic targets. In this study, we examined differentially expressed glycoproteins in cell lines derived from three different states of lung tumorigenesis: an immortalized bronchial epithelial cell (HBE) line, a non-small cell lung cancer (NSCLC) cell line harboring a Kirsten rat sarcoma viral oncogene homolog (KRAS) activation mutation and a NSCLC cell line harboring an epidermal growth factor receptor (EGFR) activation deletion. Using a Triple SILAC proteomic quantification strategy paired with hydrazide chemistry N-linked glycopeptide enrichment, we quantified 118 glycopeptides in the three cell lines derived from 82 glycoproteins. Proteomic profiling revealed 27 glycopeptides overexpressed in both NSCLC cell lines, 6 glycopeptides overexpressed only in the EGFR mutant cells and 19 glycopeptides overexpressed only in the KRAS mutant cells. Further investigation of a panel of NSCLC cell lines found that Cellular repressor of E1A-stimulated genes (CREG1) overexpression was closely correlated with KRAS mutation status in NSCLC cells and could be down-regulated by inhibition of KRAS expression. Our results indicate that CREG1 is a down-stream effector of KRAS in a sub-type of NSCLC cells and a novel candidate biomarker or therapeutic target for KRAS mutant NSCLC. PMID:26722374

  18. [Remarks about the position of the medico-legal expert in imperative regulations in the Penal and Civil Codes].

    PubMed

    Chowaniec, Czesław; Nowak, Agnieszka; Chowaniec, Małgorzata; Kobek, Mariusz

    2005-01-01

    In the monograph 'Medico-legal opinions--essays on theory', prof. K. Jaegermann wrote that 'the use of an expert requires theoretical or fairly clear knowledge about the mutual relationship between judge and expert'. In his opinion knowledge of this kind plays a significant role in estimating the usefulness of so-called expert evidence. Practical knowledge about the relationship between the judge and expert is necessary but not a decisive condition not only for a lawyer to be a judge but also for a physician to be a medico-legal expert. An expert can be not only a person appointed by the court but must also possess proper knowledge in a particular field, namely, the required professional and specialist qualifications and must also considered to be impartial. On the basis of the analysis of law in force and imperative regulations in Penal and Civil Codes, the authors have presented remarks relating to the expert's status as well as the lack of judicial control over the activity of experts appointed by court. Verification of professional qualifications in court experts and a reduction of those appointed 'ad hoc' are suggested. In the authors opinion co-operation between lawyers and experts should be improved. It is also essential to introduce statutory legal protection of court experts as well as to undertake activities leading to equaling the status of Polish court experts to that of other European countries.

  19. Distinct forms of synaptic inhibition and neuromodulation regulate calretinin-positive neuron excitability in the spinal cord dorsal horn.

    PubMed

    Smith, K M; Boyle, K A; Mustapa, M; Jobling, P; Callister, R J; Hughes, D I; Graham, B A

    2016-06-21

    The dorsal horn (DH) of the spinal cord contains a heterogenous population of neurons that process incoming sensory signals before information ascends to the brain. We have recently characterized calretinin-expressing (CR+) neurons in the DH and shown that they can be divided into excitatory and inhibitory subpopulations. The excitatory population receives high-frequency excitatory synaptic input and expresses delayed firing action potential discharge, whereas the inhibitory population receives weak excitatory drive and exhibits tonic or initial bursting discharge. Here, we characterize inhibitory synaptic input and neuromodulation in the two CR+ populations, in order to determine how each is regulated. We show that excitatory CR+ neurons receive mixed inhibition from GABAergic and glycinergic sources, whereas inhibitory CR+ neurons receive inhibition, which is dominated by glycine. Noradrenaline and serotonin produced robust outward currents in excitatory CR+ neurons, predicting an inhibitory action on these neurons, but neither neuromodulator produced a response in CR+ inhibitory neurons. In contrast, enkephalin (along with selective mu and delta opioid receptor agonists) produced outward currents in inhibitory CR+ neurons, consistent with an inhibitory action but did not affect the excitatory CR+ population. Our findings show that the pharmacology of inhibitory inputs and neuromodulator actions on CR+ cells, along with their excitatory inputs can define these two subpopulations further, and this could be exploited to modulate discrete aspects of sensory processing selectively in the DH.

  20. Positive regulation of the Shewanella oneidensis OmpS38, a major porin facilitating anaerobic respiration, by Crp and Fur.

    PubMed

    Gao, Tong; Ju, Lili; Yin, Jianhua; Gao, Haichun

    2015-09-18

    Major porins are among the most abundant proteins embedded in the outer membrane (OM) of Gram-negative bacteria, playing crucial roles in maintenance of membrane structural integrity and OM permeability. Although many OM proteins (especially c-type cytochromes) in Shewanella oneidensis, a research model for respiratory versatility, have been extensively studied, physiological significance of major porins remains largely unexplored. In this study, we show that OmpS38 and OmpA are two major porins, neither of which is responsive to changes in osmolarity or contributes to the intrinsic resistance to β-lactam antibiotics. However, OmpS38 but not OmpA is largely involved in respiration of non-oxygen electron acceptors. We then provide evidence that expression of ompS38 is transcribed from two promoters, the major of which is favored under anaerobic conditions while the other appears constitutive. The major promoter is under the direct control of Crp, the master regulator dictating respiration. As a result, the increase in the level of OmpS38 correlates with an elevated activity in Crp under anaerobic conditions. In addition, we show that the activity of the major promoter is also affected by Fur, presumably indirectly, the transcription factor for iron-dependent gene expression.

  1. Long Noncoding RNA DANCR Is a Positive Regulator of Proliferation and Chondrogenic Differentiation in Human Synovium-Derived Stem Cells.

    PubMed

    Zhang, Lei; Yang, Chao; Chen, Shuo; Wang, Guihua; Shi, Ben; Tao, Xin; Zhou, Liwu; Zhao, Jianning

    2017-02-01

    Cartilage tissues have limited capacity for repair after damage and then cause osteoarthritis, so finding alternative treatment is ongoing. Mesenchymal stem cells (MSCs) have become a promising therapy for cartilage damage and diseases due to the advantages of easy separation, high proliferative potentiality, and genetic stability. Synovium-derived MSCs (SMSCs) have been recognized as an ideal source for cartilage repair. In our previous study, we found that Sox4 promoted proliferation and chondrogenesis of SMSCs through upregulation of long noncoding RNA (lncRNA) DANCR. However, the exact molecular mechanism by which DANCR promotes proliferation and chondrogenesis of SMSCs remains unknown. In the present study, we investigated the effect of lncRNA DANCR on the proliferation and chondrogenesis of SMSCs. We found that overexpression of DANCR could promote proliferation and chondrogenesis of SMSCs, while knockdown of DANCR had the opposite effect. Moreover, our data demonstrated that DANCR directly interacted with myc, Smad3, and STAT3 mRNA to regulate their stability. Finally, we found that the promotion of SMSC proliferation induced by DANCR depended on myc. Also, DANCR activated chondrogenesis of SMSCs via upregulation of Smad3 and STAT3 expression. Our growing knowledge of the role of DANCR is pointing toward its potential use as a novel therapeutic approach for cartilage damage and diseases.

  2. A constitutive active MAPK/ERK pathway due to BRAFV600E positively regulates AHR pathway in PTC.

    PubMed

    Occhi, Gianluca; Barollo, Susi; Regazzo, Daniela; Bertazza, Loris; Galuppini, Francesca; Guzzardo, Vincenza; Jaffrain-Rea, Marie Lise; Vianello, Federica; Ciato, Denis; Ceccato, Filippo; Watutantrige-Fernando, Sara; Bisognin, Andrea; Bortoluzzi, Stefania; Pennelli, Gianmaria; Boscaro, Marco; Scaroni, Carla; Mian, Caterina

    2015-10-13

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor mediating the toxicity and tumor-promoting properties of dioxin. AHR has been reported to be overexpressed and constitutively active in a variety of solid tumors, but few data are currently available concerning its role in thyroid cancer. In this study we quantitatively explored a series of 51 paired-normal and papillary thyroid carcinoma (PTC) tissues for AHR-related genes. We identified an increased AHR expression/activity in PTC, independently from its nuclear dimerization partner and repressor but strictly related to a constitutive active MAPK/ERK pathway. The AHR up-regulation followed by an increased expression of AHR target genes was confirmed by a meta-analysis of published microarray data, suggesting a ligand-independent active AHR pathway in PTC. In-vitro studies using a PTC-derived cell line (BCPAP) and HEK293 cells showed that BRAFV600E may directly modulate AHR localization, induce AHR expression and activity in an exogenous ligand-independent manner. The AHR pathway might represent a potential novel therapeutic target for PTC in the clinical practice.

  3. Type I and II positive allosteric modulators differentially modulate agonist-induced up-regulation of α7 nicotinic acetylcholine receptors.

    PubMed

    Thomsen, Morten S; Mikkelsen, Jens D

    2012-10-01

    Long-term treatment with nicotine or selective α7 nicotinic acetylcholine receptor (nAChR) agonists increases the number of α7 nAChRs and this up-regulation may be involved in the mechanism underlying the sustained procognitive effect of these compounds. Here, we investigate the influence of type I and II α7 nAChR positive allosteric modulators (PAMs) on agonist-induced α7 nAChR up-regulation. We show that the type II PAMs, PNU-120596 (10 μM) or TQS (1 and 10 μM), inhibit up-regulation, as measured by protein levels, induced by the α7 nAChR agonist A-582941 (10 nM or 10 μM), in SH-EP1 cells stably expressing human α7 nAChR, whereas the type I PAMs AVL-3288 or NS1738 do not. Contrarily, neither type I nor II PAMs affect 10 μM nicotine-induced receptor up-regulation, suggesting that nicotine and A-582941 induce up-regulation through different mechanisms. We further show in vivo that 3 mg/kg PNU-120596 inhibits up-regulation of the α7 nAChR induced by 10 mg/kg A-582941, as measured by [(125)I]-bungarotoxin autoradiography, whereas 1 mg/kg AVL-3288 does not. Given that type II PAMs decrease desensitization of the receptor, whereas type I PAMs do not, these results suggest that receptor desensitization is involved in A-582941-induced up-regulation. Our results are the first to show an in vivo difference between type I and II α7 nAChR PAMs, and demonstrate an agonist-dependent effect of type II PAMs occurring on a much longer time scale than previously appreciated. Furthermore, our data suggest that nicotine and A-582941 induce up-regulation through different mechanisms, and that this confers differential sensitivity to the effects of α7 nAChR PAMs. These results may have implications for the clinical development of α7 nAChR PAMs.

  4. Signal-regulator interactions. Genetic analysis of the effector binding site of xylS, the benzoate-activated positive regulator of Pseudomonas TOL plasmid meta-cleavage pathway operon.

    PubMed

    Ramos, J L; Michan, C; Rojo, F; Dwyer, D; Timmis, K

    1990-01-20

    This study reports a genetic analysis of the interactions between a positive regulator of gene expression and its effector molecules. Transcription of the TOL plasmid meta-cleavage pathway operon is specifically stimulated by the XylS protein positive regulator either through activation of this regulator by benzoate effectors or through its hyperproduction. One xylS mutant that exhibits constitutive expression of the operon promoter has been characterized, together with six mutants encoding altered XylS proteins that recognize as effectors benzoate analogues that are non-effectors for the XylS wild-type protein. The changes in two mutant regulators are located at the N-terminal end of the protein, within a putative beta-pleated domain. These mutant proteins exhibit a markedly increased affinity for normal benzoate effectors, with K's values fivefold to 60-fold lower than those of the wild-type XylS protein. They are additionally activated by new effectors having certain substituents at position 2, 3 and 4 of the aromatic ring. Two other mutant proteins recognize new effectors having substituents at position 4 and 5 of the aromatic ring, and contain mutations at their C-terminal end within a putative alpha-helix-rich domain. Three other mutations, one of which leads to constitutive expression from Pm, each result in an amino acid change in the central region of the regulator. These findings suggest but do not prove that the effector binding pocket of the XylS protein may be composed of two or more non-contiguous segments of its primary structure. The XylS protein exhibits homology with the AraC protein of Escherichia coli, a protein that stimulates transcription from ara promoters when it is activated by arabinose or benzoate. Mutations influencing effector activation of the XylS protein characterized in this study are all located in regions exhibiting a high degree of homology with the corresponding aligned sequence of AraC protein.

  5. Evolutionary divergence of intrinsic and trans-regulated nucleosome positioning sequences reveals plastic rules for chromatin organization

    PubMed Central

    Tsankov, Alex; Yanagisawa, Yoshimi; Rhind, Nicholas; Regev, Aviv; Rando, Oliver J.

    2011-01-01

    The packaging of eukaryotic genomes into nuclesomes plays critical roles in chromatin organization and gene regulation. Studies in Saccharomyces cerevisiae indicate that nucleosome occupancy is partially encoded by intrinsic antinucleosomal DNA sequences, such as poly(A) sequences, as well as by binding sites for trans-acting factors that can evict nucleosomes, such as Reb1 and the Rsc3/30 complex. Here, we use genome-wide nucleosome occupancy maps in 13 Ascomycota fungi to discover large-scale evolutionary reprogramming of both intrinsic and trans determinants of chromatin structure. We find that poly(G)s act as intrinsic antinucleosomal sequences, comparable to the known function of poly(A)s, but that the abundance of poly(G)s has diverged greatly between species, obscuring their antinucleosomal effect in low-poly(G) species such as S. cerevisiae. We also develop a computational method that uses nucleosome occupancy maps for discovering trans-acting general regulatory factor (GRF) binding sites. Our approach reveals that the specific sequences bound by GRFs have diverged substantially across evolution, corresponding to a number of major evolutionary transitions in the repertoire of GRFs. We experimentally validate a proposed evolutionary transition from Cbf1 as a major GRF in pre-whole-genome duplication (WGD) yeasts to Reb1 in post-WGD yeasts. We further show that the mating type switch-activating protein Sap1 is a GRF in S. pombe, demonstrating the general applicability of our approach. Our results reveal that the underlying mechanisms that determine in vivo chromatin organization have diverged and that comparative genomics can help discover new determinants of chromatin organization. PMID:21914852

  6. Berberine regulates peroxisome proliferator-activated receptors and positive transcription elongation factor b expression in diabetic adipocytes.

    PubMed

    Zhou, Jiyin; Zhou, Shiwen

    2010-12-15

    Berberine has hypoglycemic and hypolipidemic effects on diabetic rats. This study investigated the relationship between hypoglycemic and hypolipidemic effects of berberine and peroxisome proliferator-activated receptors (PPARs) and positive transcription elongation factor b (P-TEFb) (including cyclin-dependent kinase 9 (CDK9) and cyclin T1) in white adipose tissue of diabetic rats and RNA interference-treated 3T3-L1 cells. Berberine promoted differentiation and inhibited lipid accumulation of 3T3-L1 cells, further decreased PPARα/δ/γ, CDK9 and cyclin T1 mRNA and protein expression and decreased tumor necrosis factor α content in supernatants of both control and RNA interference-treated 3T3-L1 cells. After a 16-week induction with 35 mg/kg streptozotocin (i.p.) and high-carbohydrate/high-fat diet, diabetic rats were treated with 75, 150 and 300 mg/kg berberine and 100 mg/kg fenofibrate or 4 mg/kg rosiglitazone for another 16 weeks. Berberine decreased white adipose tissue to body weight ratio and adipocyte size and increased adipocyte number. Berberine upregulated PPARα/δ/γ, CDK9 and cyclin T1 mRNA and protein expression in adipose tissue, decreased tumor necrosis factor α and free fatty acid content and increased lipoprotein lipase activity in serum and adipose tissue. Berberine modulated metabolic related PPARs expression and differentiation related P-TEFb expression in adipocytes, which are associated with its hypoglycemic and hypolipidemic effects.

  7. Mammalian Atg18 (WIPI2) localizes to omegasome-anchored phagophores and positively regulates LC3 lipidation.

    PubMed

    Polson, Hannah E J; de Lartigue, Jane; Rigden, Daniel J; Reedijk, Marco; Urbé, Sylvie; Clague, Michael J; Tooze, Sharon A

    2010-05-01

    Autophagosome formation is a complex process that begins with the nucleation of a pre-autophagosomal structure (PAS) that expands into a phagophore or isolation membrane, the precursor of the autophagosome. A key event in the formation of the phagophore is the production of PtdIns3P by the phosphatidylinsitol kinase Vps34. In yeast the two closely related proteins, Atg18 and Atg21, are the only known effectors of PtdIns3P that act in the autophagy pathway. The recruitment of Atg18 or Atg21 to the PAS is an essential step in the formation of the phagophore. Our bioinformatic analysis of the Atg18 and Atg21 orthologues in all eukaryotes shows that WIPI1 and WIPI2 are both mammalian orthologues of Atg18. We show that WIPI2 is a mammalian effector of PtdIns3P and is ubiquitously expressed in a variety of cell lines. WIPI2 is recruited to early autophagosomal structures along with Atg16L and ULK1 and is required for the formation of LC3-positive autophagosomes. Furthermore, when WIPI2 is depleted, we observe a remarkable accumulation of omegasomes, ER-localized PtdIns3P-containing structures labeled by DFCP1 (double FYVE domain-containing protein 1), which are thought to act as platforms for autophagosome formation. In view of our data we propose a role for WIPI2 in the progression of omegasomes into autophagosomes.

  8. Identification of OsbZIP72 as a positive regulator of ABA response and drought tolerance in rice.

    PubMed

    Lu, Guojun; Gao, Chenxi; Zheng, Xingnan; Han, Bin

    2009-02-01

    Abscisic Acid (ABA) is an important phytohormone involved in abiotic stress resistance in plants. A group of bZIP transcription factors play important roles in the ABA signaling pathway in Arabidopsis. However, little is known about the function of their orthologs in rice, where they may hold a great potential for developing drought resistant food crops. In this study, our phylogenetic analysis showed that this group of bZIPs was evolutionarily conserved between Arabidopsis and rice, which implies that they may share similar functions. We demonstrated with quantitative RT-PCR that the expressions of most of these OsbZIPs were significantly induced by ABA, ACC, and abiotic stresses. OsbZIP72, a member of this group, was proved to be an ABRE binding factor in rice using the yeast hybrid systems. We showed that it could bind to ABRE and transactivate the downstream reporter genes in yeast, and the transactivity was depending on its N-terminal region. Transgenic rice overexpressing OsbZIP72 showed a hypersensitivity to ABA, elevated levels of expression of ABA response gene such as LEAs, and an enhanced ability of drought tolerance. These results suggest that OsbZIP72 plays a positive role in drought resistance through ABA signaling, and is potential useful for engineering drought tolerant rice.

  9. ZIC1 LEVELS REGULATE MOSSY FIBER NEURON POSITION AND AXON LATERALITY CHOICE IN THE VENTRAL BRAIN STEM

    PubMed Central

    DiPietrantonio, Heather J.; Dymecki, Susan M.

    2010-01-01

    Pontine gray neurons of the brain stem are a major source of mossy fiber (MF) afferents to granule cells of the cerebellum. Achieving this connectivity involves an early regionalization of pontine gray neuron cell bodies within the brainstem pontine nuclei, as well as establishing the proper ratio of crossed versus uncrossed MF projections to contralateral versus ipsilateral cerebellar territories. Here, we report expression of the transcription factor Zic1 in newly postmitotic pontine gray neurons and present functional experiments in embryonic and postnatal mice that implicate Zic1 levels as a key determinant of pontine neuron cell body position within the pons and axon laterality. Reducing Zic1 levels embryonically via in utero electroporation of short hairpin RNA interference (shRNAi) vectors shifted the postnatal distribution of pontine neurons from caudolateral to rostromedial territories; by contrast, increasing Zic1 levels resulted in the reciprocal shift, with electroporated cells redistributing caudolaterally. Associated with the latter was a change in axon laterality, with a greater proportion of marked projections now targeting the ipsilateral instead of contralateral cerebellum. Zic1 levels in pontine gray neurons, therefore, play an important role in the development of pontocerebellar circuitry. PMID:19303920

  10. Interlocked positive and negative feedback network motifs regulate β-catenin activity in the adherens junction pathway

    PubMed Central

    Klinke, David J.; Horvath, Nicholas; Cuppett, Vanessa; Wu, Yueting; Deng, Wentao; Kanj, Rania

    2015-01-01

    The integrity of epithelial tissue architecture is maintained through adherens junctions that are created through extracellular homotypic protein–protein interactions between cadherin molecules. Cadherins also provide an intracellular scaffold for the formation of a multiprotein complex that contains signaling proteins, including β-catenin. Environmental factors and controlled tissue reorganization disrupt adherens junctions by cleaving the extracellular binding domain and initiating a series of transcriptional events that aim to restore tissue homeostasis. However, it remains unclear how alterations in cell adhesion coordinate transcriptional events, including those mediated by β-catenin in this pathway. Here were used quantitative single-cell and population-level in vitro assays to quantify the endogenous pathway dynamics after the proteolytic disruption of the adherens junctions. Using prior knowledge of isolated elements of the overall network, we interpreted these data using in silico model-based inference to identify the topology of the regulatory network. Collectively the data suggest that the regulatory network contains interlocked network motifs consisting of a positive feedback loop, which is used to restore the integrity of adherens junctions, and a negative feedback loop, which is used to limit β-catenin–induced gene expression. PMID:26224311

  11. Regulation of the Alkane Hydroxylase CYP153 Gene in a Gram-Positive Alkane-Degrading Bacterium, Dietzia sp. Strain DQ12-45-1b

    PubMed Central

    Liang, Jie-Liang; JiangYang, Jing-Hong

    2015-01-01

    CYP153, one of the most common medium-chain n-alkane hydroxylases belonging to the cytochrome P450 superfamily, is widely expressed in n-alkane-degrading bacteria. CYP153 is also thought to cooperate with AlkB in degrading various n-alkanes. However, the mechanisms regulating the expression of the protein remain largely unknown. In this paper, we studied CYP153 gene transcription regulation by the potential AraC family regulator (CypR) located upstream of the CYP153 gene cluster in a broad-spectrum n-alkane-degrading Gram-positive bacterium, Dietzia sp. strain DQ12-45-1b. We first identified the transcriptional start site and the promoter of the CYP153 gene cluster. Sequence alignment of upstream regions of CYP153 gene clusters revealed high conservation in the −10 and −35 regions in Actinobacteria. Further analysis of the β-galactosidase activity in the CYP153 gene promoter-lacZ fusion cell indicated that the CYP153 gene promoter was induced by n-alkanes comprised of 8 to 14 carbon atoms, but not by derived decanol and decanic acid. Moreover, we constructed a cypR mutant strain and found that the CYP153 gene promoter activities and CYP153 gene transcriptional levels in the mutant strain were depressed compared with those in the wild-type strain in the presence of n-alkanes, suggesting that CypR served as an activator for the CYP153 gene promoter. By comparing CYP153 gene arrangements in Actinobacteria and Proteobacteria, we found that the AraC family regulator is ubiquitously located upstream of the CYP153 gene, suggesting its universal regulatory role in CYP153 gene transcription. We further hypothesize that the observed mode of CYP153 gene regulation is shared by many Actinobacteria. PMID:26567302

  12. Regulation of the Alkane Hydroxylase CYP153 Gene in a Gram-Positive Alkane-Degrading Bacterium, Dietzia sp. Strain DQ12-45-1b.

    PubMed

    Liang, Jie-Liang; JiangYang, Jing-Hong; Nie, Yong; Wu, Xiao-Lei

    2015-11-13

    CYP153, one of the most common medium-chain n-alkane hydroxylases belonging to the cytochrome P450 superfamily, is widely expressed in n-alkane-degrading bacteria. CYP153 is also thought to cooperate with AlkB in degrading various n-alkanes. However, the mechanisms regulating the expression of the protein remain largely unknown. In this paper, we studied CYP153 gene transcription regulation by the potential AraC family regulator (CypR) located upstream of the CYP153 gene cluster in a broad-spectrum n-alkane-degrading Gram-positive bacterium, Dietzia sp. strain DQ12-45-1b. We first identified the transcriptional start site and the promoter of the CYP153 gene cluster. Sequence alignment of upstream regions of CYP153 gene clusters revealed high conservation in the -10 and -35 regions in Actinobacteria. Further analysis of the β-galactosidase activity in the CYP153 gene promoter-lacZ fusion cell indicated that the CYP153 gene promoter was induced by n-alkanes comprised of 8 to 14 carbon atoms, but not by derived decanol and decanic acid. Moreover, we constructed a cypR mutant strain and found that the CYP153 gene promoter activities and CYP153 gene transcriptional levels in the mutant strain were depressed compared with those in the wild-type strain in the presence of n-alkanes, suggesting that CypR served as an activator for the CYP153 gene promoter. By comparing CYP153 gene arrangements in Actinobacteria and Proteobacteria, we found that the AraC family regulator is ubiquitously located upstream of the CYP153 gene, suggesting its universal regulatory role in CYP153 gene transcription. We further hypothesize that the observed mode of CYP153 gene regulation is shared by many Actinobacteria.

  13. Inhibition of miR-146b expression increases radioiodine-sensitivity in poorly differential thyroid carcinoma via positively regulating NIS expression

    SciTech Connect

    Li, Luchuan; Lv, Bin; Chen, Bo; Guan, Ming; Sun, Yongfeng; Li, Haipeng; Zhang, Binbin; Ding, Changyuan; He, Shan; Zeng, Qingdong

    2015-07-10

    Dedifferentiated thyroid carcinoma (DTC) with the loss of radioiodine uptake (RAIU) is often observed in clinical practice under radioiodine therapy, indicating the challenge for poor prognosis. MicroRNA (miRNA) has emerged as a promising therapeutic target in many diseases; yet, the role of miRNAs in RAIU has not been generally investigated. Based on recent studies about miRNA expression in papillary or follicular thyroid carcinomas, the expression profiles of several thyroid relative miRNAs were investigated in one DTC cell line, derived from normal DTC cells by radioiodine treatment. The top candidate miR-146b, with the most significant overexpression profiles in dedifferentiated cells, was picked up. Further research found that miR-146b could be negatively regulated by histone deacetylase 3 (HDAC3) in normal cells, indicating the correlation between miR-146b and Na{sup +}/I{sup −} symporter (NIS)-mediated RAIU. Fortunately, it was confirmed that miR-146b could regulate NIS expression/activity; what is more important, miR-146b interference would contribute to the recovery of radioiodine-sensitivity in dedifferentiated cells via positively regulating NIS. In the present study, it was concluded that NIS-mediated RAIU could be modulated by miR-146b; accordingly, miR-146b might serve as one of targets to enhance efficacy of radioactive therapy against poorly differential thyroid carcinoma (PDTC). - Highlights: • Significant upregulated miR-146b was picked up from thyroid relative miRNAs in DTC. • MiR-146b was negatively regulated by HDAC3 in normal thyroid carcinoma cells. • NIS activity and expression could be regulated by miR-146b in thyroid carcinoma. • MiR-146b inhibition could recover the decreased radioiodine-sensitivity of DTC cells.

  14. DC-SCRIPT is a novel regulator of the tumor suppressor gene CDKN2B and induces cell cycle arrest in ERα-positive breast cancer cells.

    PubMed

    Ansems, Marleen; Søndergaard, Jonas Nørskov; Sieuwerts, Anieta M; Looman, Maaike W G; Smid, Marcel; de Graaf, Annemarie M A; de Weerd, Vanja; Zuidscherwoude, Malou; Foekens, John A; Martens, John W M; Adema, Gosse J

    2015-02-01

    Breast cancer is one of the most common causes of cancer-related deaths in women. The estrogen receptor (ERα) is well known for having growth promoting effects in breast cancer. Recently, we have identified DC-SCRIPT (ZNF366) as a co-suppressor of ERα and as a strong and independent prognostic marker in ESR1 (ERα gene)-positive breast cancer patients. In this study, we further investigated the molecular mechanism on how DC-SCRIPT inhibits breast cancer cell growth. DC-SCRIPT mRNA levels from 190 primary ESR1-positive breast tumors were related to global gene expression, followed by gene ontology and pathway analysis. The effect of DC-SCRIPT on breast cancer cell growth and cell cycle arrest was investigated using novel DC-SCRIPT-inducible MCF7 breast cancer cell lines. Genome-wide expression profiling of DC-SCRIPT-expressing MCF7 cells was performed to investigate the effect of DC-SCRIPT on cell cycle-related gene expression. Findings were validated by real-time PCR in a cohort of 1,132 ESR1-positive breast cancer patients. In the primary ESR1-positive breast tumors, DC-SCRIPT expression negatively correlated with several cell cycle gene ontologies and pathways. DC-SCRIPT expression strongly reduced breast cancer cell growth in vitro, breast tumor growth in vivo, and induced cell cycle arrest. In addition, in the presence of DC-SCRIPT, multiple cell cycles related genes were differentially expressed including the tumor suppressor gene CDKN2B. Moreover, in 1,132 primary ESR1-positive breast tumors, DC-SCRIPT expression also correlated with CDKN2B expression. Collectively, these data show that DC-SCRIPT acts as a novel regulator of CDKN2B and induces cell cycle arrest in ESR1-positive breast cancer cells.

  15. Chromatin-associated RNA interference components contribute to transcriptional regulation in Drosophila.

    PubMed

    Cernilogar, Filippo M; Onorati, Maria Cristina; Kothe, Greg O; Burroughs, A Maxwell; Parsi, Krishna Mohan; Breiling, Achim; Lo Sardo, Federica; Saxena, Alka; Miyoshi, Keita; Siomi, Haruhiko; Siomi, Mikiko C; Carninci, Piero; Gilmour, David S; Corona, Davide F V; Orlando, Valerio

    2011-11-06

    RNA interference (RNAi) pathways have evolved as important modulators of gene expression that operate in the cytoplasm by degrading RNA target molecules through the activity of short (21-30 nucleotide) RNAs. RNAi components have been reported to have a role in the nucleus, as they are involved in epigenetic regulation and heterochromatin formation. However, although RNAi-mediated post-transcriptional gene silencing is well documented, the mechanisms of RNAi-mediated transcriptional gene silencing and, in particular, the role of RNAi components in chromatin dynamics, especially in animal multicellular organisms, are elusive. Here we show that the key RNAi components Dicer 2 (DCR2) and Argonaute 2 (AGO2) associate with chromatin (with a strong preference for euchromatic, transcriptionally active, loci) and interact with the core transcription machinery. Notably, loss of function of DCR2 or AGO2 showed that transcriptional defects are accompanied by the perturbation of RNA polymerase II positioning on promoters. Furthermore, after heat shock, both Dcr2 and Ago2 null mutations, as well as missense mutations that compromise the RNAi activity, impaired the global dynamics of RNA polymerase II. Finally, the deep sequencing of the AGO2-associated small RNAs (AGO2 RIP-seq) revealed that AGO2 is strongly enriched in small RNAs that encompass the promoter regions and other regions of heat-shock and other genetic loci on both the sense and antisense DNA strands, but with a strong bias for the antisense strand, particularly after heat shock. Taken together, our results show that DCR2 and AGO2 are globally associated with transcriptionally active loci and may have a pivotal role in shaping the transcriptome by controlling the processivity of RNA polymerase II.

  16. Regulation and Gene Expression Profiling of NKG2D Positive Human Cytomegalovirus-Primed CD4+ T-Cells

    PubMed Central

    Jensen, Helle; Folkersen, Lasse; Skov, Søren

    2012-01-01

    NKG2D is a stimulatory receptor expressed by natural killer (NK) cells, CD8+ T-cells, and γδ T-cells. NKG2D expression is normally absent from CD4+ T-cells, however recently a subset of NKG2D+ CD4+ T-cells has been found, which is specific for human cytomegalovirus (HCMV). This particular subset of HCMV-specific NKG2D+ CD4+ T-cells possesses effector-like functions, thus resembling the subsets of NKG2D+ CD4+ T-cells found in other chronic inflammations. However, the precise mechanism leading to NKG2D expression on HCMV-specific CD4+ T-cells is currently not known. In this study we used genome-wide analysis of individual genes and gene set enrichment analysis (GSEA) to investigate the gene expression profile of NKG2D+ CD4+ T-cells, generated from HCMV-primed CD4+ T-cells. We show that the HCMV-primed NKG2D+ CD4+ T-cells possess a higher differentiated phenotype than the NKG2D– CD4+ T-cells, both at the gene expression profile and cytokine profile. The ability to express NKG2D at the cell surface was primarily determined by the activation or differentiation status of the CD4+ T-cells and not by the antigen presenting cells. We observed a correlation between CD94 and NKG2D expression in the CD4+ T-cells following HCMV stimulation. However, knock-down of CD94 did not affect NKG2D cell surface expression or signaling. In addition, we show that NKG2D is recycled at the cell surface of activated CD4+ T-cells, whereas it is produced de novo in resting CD4+ T-cells. These findings provide novel information about the gene expression profile of HCMV-primed NKG2D+ CD4+ T-cells, as well as the mechanisms regulating NKG2D cell surface expression. PMID:22870231

  17. RLN2 Is a Positive Regulator of AKT-2-Induced Gene Expression Required for Osteosarcoma Cells Invasion and Chemoresistance

    PubMed Central

    Ma, Jinfeng; Huang, Hai; Han, Zenggang; Zhu, Changzheng; Yue, Bin

    2015-01-01

    The aim of the study was to determine the effect of H2 relaxin (RLN2) on invasion, migration, and chemosensitivity to cisplatin in human osteosarcoma U2-OS and MG-63 cells and then to investigate the effect of RLN2 on the AKT/NF-κB signaling pathway. The expression of RLN2, p-AKT (Ser473), and p-ERK1/2 (Phospho-Thr202/Tyr204) proteins was detected by western blot in OS tissues from 21 patients with pulmonary metastatic disease, and the correlation between RLN2 and p-AKT or RLN2 and p-ERK1/2 expression was investigated. RLN2 expression was inhibited by RLN2 siRNA transfection in the MG-63 cells. RLN2 was overexpressed in the U2-OS cells by treatment with recombinant relaxin. The results showed that positive relation was found between RLN2 and p-AKT expression in tissues of OS. Silencing RLN2 inhibited cell migratory and invasive ability and angiogenesis formation and increased the chemosensitivity to cisplatin in MG-63 cells. RLN2 overexpression promoted migratory and invasive ability and angiogenesis and increased the chemoresistance to cisplatin in U2-OS cells. Silencing RLN2 inhibited the activity of AKT/NF-κB signaling pathway in MG-63 cells, and vice versa. Blockage of both pathways by specific inhibitors abrogated RLN2-induced survival and invasion of OS cells, and vice versa. Our results indicated RLN2 confers to migratory and invasive ability, angiogenesis, and chemoresistance to cisplatin via modulating the AKT/NF-κB signaling pathway in vitro. PMID:26229955

  18. A novel hypoxia-induced miR-147a regulates cell proliferation through a positive feedback loop of stabilizing HIF-1α

    PubMed Central

    Wang, Fan; Zhang, Haoxiang; Xu, Naihan; Huang, Nunu; Tian, Caiming; Ye, Anlin; Hu, Guangnan; He, Jie; Zhang, Yaou

    2016-01-01

    ABSTRACT Hypoxia is a general event in solid tumor growth. Therefore, induced cellular responses by hypoxia are important for tumorigenesis and tumor growth. MicroRNAs (miRNAs) have recently emerged as important regulators of hypoxia induced cellular responses. Here we report that miR-147a is a novel and crucial hypoxia induced miRNA. HIF-1α up-regulates the expression of miR-147a, and miR-147a in turn stabilizes and accumulates HIF-1α protein via directly targeting HIF-3α, a dominant negative regulator of HIF-1α. Subsequent studies in xenograft mouse model reveal that miR-147a is capable of inhibiting tumor growth. Collectively, these data demonstrate a positive feedback loop between HIF-1α, miR-147a and HIF-3α, which provide a new insight into the mechanism of miR-147a induced cell proliferation arrest under hypoxia. PMID:27260617

  19. A calmodulin like EF hand protein positively regulates oxalate decarboxylase expression by interacting with E-box elements of the promoter

    PubMed Central

    Kamthan, Ayushi; Kamthan, Mohan; Kumar, Avinash; Sharma, Pratima; Ansari, Sekhu; Thakur, Sarjeet Singh; Chaudhuri, Abira; Datta, Asis

    2015-01-01

    Oxalate decarboxylase (OXDC) enzyme has immense biotechnological applications due to its ability to decompose anti-nutrient oxalic acid. Flammulina velutipes, an edible wood rotting fungus responds to oxalic acid by induction of OXDC to maintain steady levels of pH and oxalate anions outside the fungal hyphae. Here, we report that upon oxalic acid induction, a calmodulin (CaM) like protein-FvCaMLP, interacts with the OXDC promoter to regulate its expression. Electrophoretic mobility shift assay showed that FvCamlp specifically binds to two non-canonical E-box elements (AACGTG) in the OXDC promoter. Moreover, substitutions of amino acids in the EF hand motifs resulted in loss of DNA binding ability of FvCamlp. F. velutipes mycelia treated with synthetic siRNAs designed against FvCaMLP showed significant reduction in FvCaMLP as well as OXDC transcript pointing towards positive nature of the regulation. FvCaMLP is different from other known EF hand proteins. It shows sequence similarity to both CaMs and myosin regulatory light chain (Cdc4), but has properties typical of a calmodulin, like binding of 45Ca2+, heat stability and Ca2+ dependent electrophoretic shift. Hence, FvCaMLP can be considered a new addition to the category of unconventional Ca2+ binding transcriptional regulators. PMID:26455820

  20. Epstein-Barr Virus nuclear antigen 1 (EBNA1) confers resistance to apoptosis in EBV-positive B-lymphoma cells through up-regulation of survivin

    SciTech Connect

    Lu Jie; Murakami, Masanao; Verma, Subhash C.; Cai Qiliang; Haldar, Sabyasachi; Kaul, Rajeev; Wasik, Mariusz A.; Middeldorp, Jaap; Robertson, Erle S.

    2011-02-05

    Resistance to apoptosis is an important component of the overall mechanism which drives the tumorigenic process. EBV is a ubiquitous human gamma-herpesvirus which preferentially establishes latent infection in viral infected B-lymphocytes. EBNA1 is typically expressed in most forms of EBV-positive malignancies and is important for replication of the latent episome in concert with replication of the host cells. Here, we investigate the effects of EBNA1 on survivin up-regulation in EBV-infected human B-lymphoma cells. We present evidence which demonstrates that EBNA1 forms a complex with Sp1 or Sp1-like proteins bound to their cis-element at the survivin promoter. This enhances the activity of the complex and up-regulates survivin. Knockdown of survivin and EBNA1 showed enhanced apoptosis in infected cells and thus supports a role for EBNA1 in suppressing apoptosis in EBV-infected cells. Here, we suggest that EBV encoded EBNA1 can contribute to the oncogenic process by up-regulating the apoptosis suppressor protein, survivin in EBV-associated B-lymphoma cells.

  1. A calmodulin like EF hand protein positively regulates oxalate decarboxylase expression by interacting with E-box elements of the promoter.

    PubMed

    Kamthan, Ayushi; Kamthan, Mohan; Kumar, Avinash; Sharma, Pratima; Ansari, Sekhu; Thakur, Sarjeet Singh; Chaudhuri, Abira; Datta, Asis

    2015-10-12

    Oxalate decarboxylase (OXDC) enzyme has immense biotechnological applications due to its ability to decompose anti-nutrient oxalic acid. Flammulina velutipes, an edible wood rotting fungus responds to oxalic acid by induction of OXDC to maintain steady levels of pH and oxalate anions outside the fungal hyphae. Here, we report that upon oxalic acid induction, a calmodulin (CaM) like protein-FvCaMLP, interacts with the OXDC promoter to regulate its expression. Electrophoretic mobility shift assay showed that FvCamlp specifically binds to two non-canonical E-box elements (AACGTG) in the OXDC promoter. Moreover, substitutions of amino acids in the EF hand motifs resulted in loss of DNA binding ability of FvCamlp. F. velutipes mycelia treated with synthetic siRNAs designed against FvCaMLP showed significant reduction in FvCaMLP as well as OXDC transcript pointing towards positive nature of the regulation. FvCaMLP is different from other known EF hand proteins. It shows sequence similarity to both CaMs and myosin regulatory light chain (Cdc4), but has properties typical of a calmodulin, like binding of (45)Ca(2+), heat stability and Ca(2+) dependent electrophoretic shift. Hence, FvCaMLP can be considered a new addition to the category of unconventional Ca(2+) binding transcriptional regulators.

  2. Apoptosis signal-regulating kinase 1 and cyclin D1 compose a positive feedback loop contributing to tumor growth in gastric cancer

    PubMed Central

    Hayakawa, Yoku; Hirata, Yoshihiro; Nakagawa, Hayato; Sakamoto, Kei; Hikiba, Yohko; Kinoshita, Hiroto; Nakata, Wachiko; Takahashi, Ryota; Tateishi, Keisuke; Tada, Motohisa; Akanuma, Masao; Yoshida, Haruhiko; Takeda, Kohsuke; Ichijo, Hidenori; Omata, Masao; Maeda, Shin; Koike, Kazuhiko

    2011-01-01

    Mitogen-activated protein kinase (MAPK) pathways regulate multiple cellular functions and are highly active in many types of human cancers. Apoptosis signal-regulating kinase 1 (ASK1) is an upstream MAPK involved in apoptosis, inflammation, and carcinogenesis. This study investigated the role of ASK1 in the development of gastric cancer. In human gastric cancer specimens, we observed increased ASK1 expression, compared to nontumor epithelium. Using a chemically induced murine gastric tumorigenesis model, we observed increased tumor ASK1 expression, and ASK1 knockout mice had both fewer and smaller tumors than wild-type (WT) mice. ASK1 siRNA inhibited cell proliferation through the accumulation of cells in G1 phase of the cell cycle, and reduced cyclin D1 expression in gastric cancer cells, whereas these effects were uncommon in other cancer cells. ASK1 overexpression induced the transcription of cyclin D1, through AP-1 activation, and ASK1 levels were regulated by cyclin D1, via the Rb–E2F pathway. Exogenous ASK1 induced cyclin D1 expression, followed by elevated expression of endogenous ASK1. These results indicate an autoregulatory mechanism of ASK1 in the development of gastric cancer. Targeting this positive feedback loop, ASK1 may present a potential therapeutic target for the treatment of advanced gastric cancer. PMID:21187402

  3. Zinc-Finger Transcription Factor ZAT6 Positively Regulates Cadmium Tolerance through the Glutathione-Dependent Pathway in Arabidopsis1[OPEN

    PubMed Central

    Chen, Jian; Yan, Xingxing; Liu, Yunlei; Wang, Ren; Fan, Tingting; Ren, Yongbing; Tang, Xiaofeng; Xiao, Fangming

    2016-01-01

    Cadmium (Cd) is an environmental pollutant with high toxicity to animals and plants. It has been established that the glutathione (GSH)-dependent phytochelatin (PC) synthesis pathway is one of the most important mechanisms contributing to Cd accumulation and tolerance in plants. However, the transcription factors involved in regulating GSH-dependent PC synthesis pathway remain largely unknown. Here, we identified an Arabidopsis (Arabidopsis thaliana) Cd-resistant mutant xcd2-D (XVE system-induced cadmium-tolerance2) using a forward genetics approach. The mutant gene underlying xcd2-D mutation was revealed to encode a known zinc-finger transcription factor, ZAT6. Transgenic plants overexpressing ZAT6 showed significant increase of Cd tolerance, whereas loss of function of ZAT6 led to decreased Cd tolerance. Increased Cd accumulation and tolerance in ZAT6-overexpressing lines was GSH dependent and associated with Cd-activated synthesis of PC, which was correlated with coordinated activation of PC-synthesis related gene expression. By contrast, loss of function of ZAT6 reduced Cd accumulation and tolerance, which was accompanied by abolished PC synthesis and gene expression. Further analysis revealed that ZAT6 positively regulates the transcription of GSH1, GSH2, PCS1, and PCS2, but ZAT6 is capable of specifically binding to GSH1 promoter in vivo. Consistently, overexpression of GSH1 has been shown to restore Cd sensitivity in the zat6-1 mutant, suggesting that GSH1 is a key target of ZAT6. Taken together, our data provide evidence that ZAT6 coordinately activates PC synthesis-related gene expression and directly targets GSH1 to positively regulate Cd accumulation and tolerance in Arabidopsis. PMID:26983992

  4. The cotton MYB108 forms a positive feedback regulation loop with CML11 and participates in the defense response against Verticillium dahliae infection

    PubMed Central

    Cheng, Huan-Qing; Han, Li-Bo; Yang, Chun-Lin; Wu, Xiao-Min; Zhong, Nai-Qin; Wu, Jia-He; Wang, Fu-Xin; Xia, Gui-Xian

    2016-01-01

    Accumulating evidence indicates that plant MYB transcription factors participate in defense against pathogen attack, but their regulatory targets and related signaling processes remain largely unknown. Here, we identified a defense-related MYB gene (GhMYB108) from upland cotton (Gossypium hirsutum) and characterized its functional mechanism. Expression of GhMYB108 in cotton plants was induced by Verticillium dahliae infection and responded to the application of defense signaling molecules, including salicylic acid, jasmonic acid, and ethylene. Knockdown of GhMYB108 expression led to increased susceptibility of cotton plants to V. dahliae, while ecotopic overexpression of GhMYB108 in Arabidopsis thaliana conferred enhanced tolerance to the pathogen. Further analysis demonstrated that GhMYB108 interacted with the calmodulin-like protein GhCML11, and the two proteins form a positive feedback loop to enhance the transcription of GhCML11 in a calcium-dependent manner. Verticillium dahliae infection stimulated Ca2+ influx into the cytosol in cotton root cells, but this response was disrupted in both GhCML11-silenced plants and GhMYB108-silenced plants in which expression of several calcium signaling-related genes was down-regulated. Taken together, these results indicate that GhMYB108 acts as a positive regulator in defense against V. dahliae infection by interacting with GhCML11. Furthermore, the data also revealed the important roles and synergetic regulation of MYB transcription factor, Ca2+, and calmodulin in plant immune responses. PMID:26873979

  5. The curli biosynthesis regulator CsgD co-ordinates the expression of both positive and negative determinants for biofilm formation in Escherichia coli.

    PubMed

    Brombacher, Eva; Dorel, Corinne; Zehnder, Alexander J B; Landini, Paolo

    2003-10-01

    Production of curli, extracellular structures important for biofilm formation, is positively regulated by OmpR, which constitutes with the EnvZ protein an osmolarity-sensing two-component regulatory system. The expression of curli is cryptic in most Escherichia coli laboratory strains such as MG1655, due to the lack of csgD expression. The csgD gene encodes a transcription activator of the curli-subunit-encoding csgBA operon. The ompR234 up-mutation can restore csgD expression, resulting in curli production and increased biofilm formation. In this report, it is shown that ompR234-dependent csgD expression, in addition to csgBA activation during stationary phase of growth, stimulates expression of the yaiC gene and negatively regulates at least two other genes, pepD and yagS. The promoter regions of these four genes share a conserved 11 bp sequence (CGGGKGAKNKA), necessary for csgBA and yaiC regulation by CsgD. While at both the csgBA and yaiC promoters the sequence is located upstream of the promoter elements, in both yagS and pepD it overlaps either the putative -10 sequence or the transcription start point, suggesting that CsgD can function as both an activator and a repressor. Adhesion experiments show that csgD-independent expression of both yagS and pepD from a multicopy plasmid negatively affects biofilm formation, which, in contrast, is stimulated by yaiC expression. Thus it is proposed that CsgD stimulates biofilm formation in E. coli by contemporary activation of adhesion positive determinants (the curli-encoding csg operons and the product of the yaiC gene) and repression of negative effectors such as yagS and pepD.

  6. A Critical Role of Lyst-Interacting Protein5, a Positive Regulator of Multivesicular Body Biogenesis, in Plant Responses to Heat and Salt Stresses1

    PubMed Central

    Wang, Fei; Yang, Yan; Wang, Zhe; Zhou, Jie; Fan, Baofang; Chen, Zhixiang

    2015-01-01

    Multivesicular bodies (MVBs) are unique endosomes containing vesicles in the lumen and play critical roles in many cellular processes. We have recently shown that Arabidopsis (Arabidopsis thaliana) Lyst-Interacting Protein5 (LIP5), a positive regulator of the Suppressor of K+ Transport Growth Defect1 (SKD1) AAA ATPase in MVB biogenesis, is a critical target of the mitogen-activated protein kinases MPK3 and MPK6 and plays an important role in the plant immune system. In this study, we report that the LIP5-regulated MVB pathway also plays a critical role in plant responses to abiotic stresses. Disruption of LIP5 causes compromised tolerance to both heat and salt stresses. The critical role of LIP5 in plant tolerance to abiotic stresses is dependent on its ability to interact with Suppressor of K+ Transport Growth Defect1. When compared with wild-type plants, lip5 mutants accumulate increased levels of ubiquitinated protein aggregates and NaCl under heat and salt stresses, respectively. Further analysis using fluorescent dye and MVB markers reveals that abiotic stress increases the formation of endocytic vesicles and MVBs in a largely LIP5-dependent manner. LIP5 is also required for the salt-induced increase of intracellular reactive oxygen species, which have been implicated in signaling of salt stress responses. Basal levels of LIP5 phosphorylation by MPKs and the stability of LIP5 are elevated by salt stress, and mutation of MPK phosphorylation sites in LIP5 reduces the stability and compromises the ability to complement the lip5 salt-sensitive mutant phenotype. These results collectively indicate that the MVB pathway is positively regulated by pathogen/stress-responsive MPK3/6 through LIP5 phosphorylation and plays a critical role in broad plant responses to biotic and abiotic stresses. PMID:26229051

  7. The protein phosphatase 2C clade A protein OsPP2C51 positively regulates seed germination by directly inactivating OsbZIP10.

    PubMed

    Bhatnagar, Nikita; Min, Myung-Ki; Choi, Eun-Hye; Kim, Namhyo; Moon, Seok-Jun; Yoon, Insun; Kwon, Taekryoun; Jung, Ki-Hong; Kim, Beom-Gi

    2017-03-01

    Protein phosphatase 2C clade A members are major signaling components in the ABA-dependent signaling cascade that regulates seed germination. To elucidate the role of PP2CA genes in germination of rice seed, we selected OsPP2C51, which shows highly specific expression in the embryo compared with other protein phosphatases based on microarray data. GUS histochemical assay confirmed that OsPP2C51 is expressed in the seed embryo and that this expression pattern is unique compared with those of other OsPP2CA genes. Data obtained from germination assays and alpha-amylase assays of OsPP2C51 knockout and overexpression lines suggest that OsPP2C51 positively regulates seed germination in rice. The expression of alpha-amylase synthesizing genes was high in OsPP2C51 overexpressing plants, suggesting that elevated levels of OsPP2C51 might enhance gene expression related to higher rates of seed germination. Analysis of protein interactions between ABA signaling components showed that OsPP2C51 interacts with OsPYL/RCAR5 in an ABA-dependent manner. Furthermore, interactions were observed between OsPP2C51 and SAPK2, and between OsPP2C51 and OsbZIP10 and we found out that OsPP2C51 can dephosphorylates OsbZIP10. These findings suggest the existence of a new branch in ABA signaling pathway consisting of OsPYL/RCAR-OsPP2C-bZIP apart from the previously reported OsPYL/RCAR-OsPP2C-SAPK-bZIP. Overall, our result suggests that OsPP2C51 is a positive regulator of seed germination by directly suppressing active phosphorylated OsbZIP10.

  8. Triptolide inhibits proliferation of Epstein–Barr virus-positive B lymphocytes by down-regulating expression of a viral protein LMP1

    SciTech Connect

    Zhou, Heng; Guo, Wei; Long, Cong; Wang, Huan; Wang, Jingchao; Sun, Xiaoping

    2015-01-16

    Highlights: • Triptolide inhibits proliferation of EBV-positive lymphoma cells in vitro and in vivo. • Triptolide reduces expression of LMP1 by decreasing its transcription level. • Triptolide inhibits ED-L1 promoter activity. - Abstract: Epstein–Barr virus (EBV) infects various types of cells and mainly establishes latent infection in B lymphocytes. The viral latent membrane protein 1 (LMP1) plays important roles in transformation and proliferation of B lymphocytes infected with EBV. Triptolide is a compound of Tripterygium extracts, showing anti-inflammatory, immunosuppressive, and anti-cancer activities. In this study, it is determined whether triptolide inhibits proliferation of Epstein–Barr virus-positive B lymphocytes. The CCK-8 assays were performed to examine cell viabilities of EBV-positive B95-8 and P3HR-1 cells treated by triptolide. The mRNA and protein levels of LMP1 were examined by real time-PCR and Western blotting, respectively. The activities of two LMP1 promoters (ED-L1 and TR-L1) were determined by Dual luciferase reportor assay. The results showed that triptolide inhibited the cell viability of EBV-positive B lymphocytes, and the over-expression of LMP1 attenuated this inhibitory effect. Triptolide decreased the LMP1 expression and transcriptional levels in EBV-positive B cells. The activity of LMP1 promoter ED-L1 in type III latent infection was strongly suppressed by triptolide treatment. In addition, triptolide strongly reduced growth of B95-8 induced B lymphoma in BALB/c nude mice. These results suggest that triptolide decreases proliferation of EBV-induced B lymphocytes possibly by a mechanism related to down-regulation of the LMP1 expression.

  9. Contribution of position alpha4S336 on functional expression and up-regulation of alpha4beta2 neuronal nicotinic receptors.

    PubMed

    López-Hernández, Gretchen Y; Biaggi-Labiosa, Nilza M; Torres-Cintrón, Alexis; Ortiz-Acevedo, Alejandro; Lasalde-Dominicci, José A

    2009-02-01

    Phosphorylation of the nicotinic acetylcholine receptor (nAChR) is believed to play a critical role in its nicotine-induced desensitization and up-regulation. We examined the contribution of a consensus PKC site in the alpha4 M3/M4 intracellular loop (alpha4S336) on the desensitization and up-regulation of alpha4beta2 nAChRs expressed in oocytes. Position alpha4S336 was replaced with either alanine to abolish potential phosphorylation at this site or with aspartic acid to mimic phosphorylation at this same site. Mutations alpha4S336A and alpha4S336D displayed a threefold increase in the ACh-induced response and an increase in ACh EC(50). Epibatidine binding revealed a three and sevenfold increase in surface expression for the alpha4S336A and alpha4S336D mutations, respectively, relative to wild-type, therefore, both mutations enhanced expression of the alpha4beta2 nAChR. Interestingly, the EC(50)'s and peak currents for nicotine activation remained unaffected in both mutants. Both mutations abolished the nicotine-induced up-regulation that is normally observed in the wild-type. The present data suggest that adding or removing a negative charge at this phosphorylation site cannot be explained by a simple straightforward on-and-off mechanism; rather a more complex mechanism(s) may govern the functional expression of the alpha4beta2 nAChR. Along the same line, our data support the idea that phosphorylation at multiple consensus sites in the alpha4 subunit could play a remarkable role on the regulation of the functional expression of the alpha4beta2 nAChR.

  10. Major latex protein-like protein 43 (MLP43) functions as a positive regulator during abscisic acid responses and confers drought tolerance in Arabidopsis thaliana

    PubMed Central

    Wang, Yanping; Yang, Li; Chen, Xi; Ye, Tiantian; Zhong, Bao; Liu, Ruijie; Wu, Yan; Chan, Zhulong

    2016-01-01

    Drought stress is one of the disadvantageous environmental conditions for plant growth and reproduction. Given the importance of abscisic acid (ABA) to plant growth and abiotic stress responses, identification of novel components involved in ABA signalling transduction is critical. In this study, we screened numerous Arabidopsis thaliana mutants by seed germination assay and identified a mutant mlp43 (major latex protein-like 43) with decreased ABA sensitivity in seed germination. The mlp43 mutant was sensitive to drought stress while the MLP43-overexpressed transgenic plants were drought tolerant. The tissue-specific expression pattern analysis showed that MLP43 was predominantly expressed in cotyledons, primary roots and apical meristems, and a subcellular localization study indicated that MLP43 was localized in the nucleus and cytoplasm. Physiological and biochemical analyses indicated that MLP43 functioned as a positive regulator in ABA- and drought-stress responses in Arabidopsis through regulating water loss efficiency, electrolyte leakage, ROS levels, and as well as ABA-responsive gene expression. Moreover, metabolite profiling analysis indicated that MLP43 could modulate the production of primary metabolites under drought stress conditions. Reconstitution of ABA signalling components in Arabidopsis protoplasts indicated that MLP43 was involved in ABA signalling transduction and acted upstream of SnRK2s by directly interacting with SnRK2.6 and ABF1 in a yeast two-hybrid assay. Moreover, ABA and drought stress down-regulated MLP43 expression as a negative feedback loop regulation to the performance of MLP43 in ABA and drought stress responses. Therefore, this study provided new insights for interpretation of physiological and molecular mechanisms of Arabidopsis MLP43 mediating ABA signalling transduction and drought stress responses. PMID:26512059

  11. Major latex protein-like protein 43 (MLP43) functions as a positive regulator during abscisic acid responses and confers drought tolerance in Arabidopsis thaliana.

    PubMed

    Wang, Yanping; Yang, Li; Chen, Xi; Ye, Tiantian; Zhong, Bao; Liu, Ruijie; Wu, Yan; Chan, Zhulong

    2016-01-01

    Drought stress is one of the disadvantageous environmental conditions for plant growth and reproduction. Given the importance of abscisic acid (ABA) to plant growth and abiotic stress responses, identification of novel components involved in ABA signalling transduction is critical. In this study, we screened numerous Arabidopsis thaliana mutants by seed germination assay and identified a mutant mlp43 (major latex protein-like 43) with decreased ABA sensitivity in seed germination. The mlp43 mutant was sensitive to drought stress while the MLP43-overexpressed transgenic plants were drought tolerant. The tissue-specific expression pattern analysis showed that MLP43 was predominantly expressed in cotyledons, primary roots and apical meristems, and a subcellular localization study indicated that MLP43 was localized in the nucleus and cytoplasm. Physiological and biochemical analyses indicated that MLP43 functioned as a positive regulator in ABA- and drought-stress responses in Arabidopsis through regulating water loss efficiency, electrolyte leakage, ROS levels, and as well as ABA-responsive gene expression. Moreover, metabolite profiling analysis indicated that MLP43 could modulate the production of primary metabolites under drought stress conditions. Reconstitution of ABA signalling components in Arabidopsis protoplasts indicated that MLP43 was involved in ABA signalling transduction and acted upstream of SnRK2s by directly interacting with SnRK2.6 and ABF1 in a yeast two-hybrid assay. Moreover, ABA and drought stress down-regulated MLP43 expression as a negative feedback loop regulation to the performance of MLP43 in ABA and drought stress responses. Therefore, this study provided new insights for interpretation of physiological and molecular mechanisms of Arabidopsis MLP43 mediating ABA signalling transduction and drought stress responses.

  12. Local transformations of androgens into estradiol by aromatase P450 is involved in the regulation of prolactin and the proliferation of pituitary prolactin-positive cells.

    PubMed

    García Barrado, María José; Blanco, Enrique J; Carretero Hernández, Marta; Iglesias Osma, María Carmen; Carretero, Manuel; Herrero, Julio J; Burks, Deborah Jane; Carretero, José

    2014-01-01

    In previous studies we demonstrated the immunohistochemical expression of aromatase in pituitary cells. In order to determine whether pituitary aromatase is involved in the paracrine regulation of prolactin-producing pituitary cells and the physiological relevance of pituitary aromatase in the control of these cells, an in vivo and in vitro immunocytochemical and morphometric study of prolactin-positive pituitary cells was carried out on the pituitary glands of adult male rats treated with the aromatase antagonist fadrozole. Moreover, we analyzed the expression of mRNA for the enzyme in pituitary cells of male adult rats by in situ hybridization. The aromatase-mRNA was seen to be located in the cytoplasm of 41% of pituitary cells and was well correlated with the immunocytochemical staining. After in vivo treatment with fadrozole, the size (cellular and nuclear areas) of prolactin cells, as well as the percentage of prolactin-positive cells and the percentage of proliferating-prolactin cells, was significantly decreased. Moreover, fadrozole decreased serum prolactin levels. In vitro, treatment with fadrozole plus testosterone induced similar effects on prolactin-positive cells, inhibiting their cellular proliferation. Our results suggest that under physiological conditions aromatase P450 exerts a relevant control over male pituitary prolactin-cells, probably transforming testosterone to estradiol in the pituitary gland.

  13. DBZ regulates cortical cell positioning and neurite development by sustaining the anterograde transport of Lis1 and DISC1 through control of Ndel1 dual-phosphorylation.

    PubMed

    Okamoto, Masayuki; Iguchi, Tokuichi; Hattori, Tsuyoshi; Matsuzaki, Shinsuke; Koyama, Yoshihisa; Taniguchi, Manabu; Komada, Munekazu; Xie, Min-Jue; Yagi, Hideshi; Shimizu, Shoko; Konishi, Yoshiyuki; Omi, Minoru; Yoshimi, Tomohiko; Tachibana, Taro; Fujieda, Shigeharu; Katayama, Taiichi; Ito, Akira; Hirotsune, Shinji; Tohyama, Masaya; Sato, Makoto

    2015-02-18

    Cell positioning and neuronal network formation are crucial for proper brain function. Disrupted-in-Schizophrenia 1 (DISC1) is anterogradely transported to the neurite tips, together with Lis1, and functions in neurite extension via suppression of GSK3β activity. Then, transported Lis1 is retrogradely transported and functions in cell migration. Here, we show that DISC1-binding zinc finger protein (DBZ), together with DISC1, regulates mouse cortical cell positioning and neurite development in vivo. DBZ hindered Ndel1 phosphorylation at threonine 219 and serine 251. DBZ depletion or expression of a double-phosphorylated mimetic form of Ndel1 impaired the transport of Lis1 and DISC1 to the neurite tips and hampered microtubule elongation. Moreover, application of DISC1 or a GSK3β inhibitor rescued the impairments caused by DBZ insufficiency or double-phosphorylated Ndel1 expression. We concluded that DBZ controls cell positioning and neurite development by interfering with Ndel1 from disproportionate phosphorylation, which is critical for appropriate anterograde transport of the DISC1-complex.

  14. AtPP2CG1, a protein phosphatase 2C, positively regulates salt tolerance of Arabidopsis in abscisic acid-dependent manner

    SciTech Connect

    Liu, Xin; Zhu, Yanming; Zhai, Hong; Cai, Hua; Ji, Wei; Luo, Xiao; Li, Jing; Bai, Xi

    2012-06-15

    Highlights: Black-Right-Pointing-Pointer AtPP2CG1 positively regulates salt tolerance in ABA-dependent manner. Black-Right-Pointing-Pointer AtPP2CG1 up-regulates the expression of marker genes in different pathways. Black-Right-Pointing-Pointer AtPP2CG1 expresses in the vascular system and trichomes of Arabidopsis. -- Abstract: AtPP2CG1 (Arabidopsis thaliana protein phosphatase 2C G Group 1) was predicted as an abiotic stress candidate gene by bioinformatic analysis in our previous study. The gene encodes a putative protein phosphatase 2C that belongs to Group G of PP2C. There is no report of Group G genes involved in abiotic stress so far. Real-time RT-PCR analysis showed that AtPP2CG1 expression was induced by salt, drought, and abscisic acid (ABA) treatment. The expression levels of AtPP2CG1 in the ABA synthesis-deficient mutant abi2-3 were much lower than that in WT plants under salt stress suggesting that the expression of AtPP2CG1 acts in an ABA-dependent manner. Over-expression of AtPP2CG1 led to enhanced salt tolerance, whereas its loss of function caused decreased salt tolerance. These results indicate that AtPP2CG1 positively regulates salt stress in an ABA-dependent manner. Under salt treatment, AtPP2CG1 up-regulated the expression levels of stress-responsive genes, including RD29A, RD29B, DREB2A and KIN1. GUS activity was detected in roots, leaves, stems, flower, and trichomes of AtPP2CG1 promoter-GUS transgenic plants. AtPP2CG1 protein was localized in nucleus and cytoplasm via AtPP2CG1:eGFP and YFP:AtPP2CG1 fusion approaches.

  15. Transcriptional regulation of genes encoding the selenium-free [NiFe]-hydrogenases in the archaeon Methanococcus voltae involves positive and negative control elements.

    PubMed Central

    Noll, I; Müller, S; Klein, A

    1999-01-01

    Methanococcus voltae harbors genetic information for two pairs of homologous [NiFe]-hydrogenases. Two of the enzymes contain selenocysteine, while the other two gene groups encode apparent isoenzymes that carry cysteinyl residues in the homologous positions. The genes coding for the selenium-free enzymes, frc and vhc, are expressed only under selenium limitation. They are transcribed out of a common intergenic region. A series of deletions made in the intergenic region localized a common negative regulatory element for the vhc and frc promoters as well as two activator elements that are specific for each of the two transcription units. Repeated sequences, partially overlapping the frc promoter, were also detected. Mutations in these repeated heptanucleotide sequences led to a weak induction of a reporter gene under the control of the frc promoters in the presence of selenium. This result suggests that the heptamer repeats contribute to the negative regulation of the frc transcription unit. PMID:10430564

  16. Brachypodium distachyon BdPP2CA6 Interacts with BdPYLs and BdSnRK2 and Positively Regulates Salt Tolerance in Transgenic Arabidopsis

    PubMed Central

    Zhang, Fan; Wei, Qiuhui; Shi, Jiaochun; Jin, Xia; He, Yuan; Zhang, Yang; Luo, Qingchen; Wang, Yuesheng; Chang, Junli; Yang, Guangxiao; He, Guangyuan

    2017-01-01

    The phytohormone abscisic acid (ABA) is essential in plant responding to biotic and abiotic stresses. Although ABA signaling model is well established in Arabidopsis, ABA receptor PYL family and clade A PP2C subfamily are not yet characterized in monocot model plant Brachypodium distachyon. In this study, we identified 12 PYLs and 8 clade A PP2Cs from B. distachyon genome and successfully cloned 12 PYLs and 7 clade A PP2Cs. Bioinformatic and expression analyses showed that most of the identified genes respond to several signal molecules and abiotic stresses. Protein–protein interaction analysis revealed that many BdPYLs and BdPP2CAs participate in the classic ABA-PYL-PP2C-SnRK2 signaling pathway. A clade A PP2C, designated BdPP2CA6, interacted with BdPYL11 in the absence of ABA and localized in nucleus. Most clade A PP2C members from Arabidopsis showed negatively regulation in ABA signaling pathway, whereas BdPP2CA6-overexpression transgenic Arabidopsis showed ABA hypersensitive phenotype, resulting in enhanced stomatal closure and salinity tolerance. Our results indicate that BdPP2CA6 positively regulates ABA and stress signal pathway in transgenic Arabidopsis plant seedlings. PMID:28293246

  17. Staphylococcal superantigen-like genes, ssl5 and ssl8, are positively regulated by Sae and negatively by Agr in the Newman strain.

    PubMed

    Pantrangi, Madhulatha; Singh, Vineet K; Wolz, Christiane; Shukla, Sanjay K

    2010-07-01

    Some of the staphylococcal superantigen-like (SSL) proteins SSL5, SSL7, SSL9, and SSL11 act as immunomodulatory proteins in Staphylococcus aureus. However, little is known about their regulatory mechanisms. We determined the expression levels of ssl5 and ssl8 in seven clinically important S. aureus strains and their regulatory mechanisms in the Newman strain, which had the highest ssl5 and ssl8 expression. Independent comparisons of ssl5 or ssl8 coding and upstream sequences in these strains identified multiple haplotypes that did not correlate with the differential expression of ssl5 and ssl8, suggesting the role of additional regulatory elements. Using knockout mutant strains of known S. aureus global regulators such as Agr, Sae, and SigB in the Newman strain, we showed that both ssl5 and ssl8 were induced by Sae and repressed by Agr, suggesting that Sae and Agr are the positive and the negative regulators, respectively, of these two ssl genes. Moreover, we observed upregulation of sae in the agr mutant and upregulation of agr in the sae mutant compared with the isogenic Newman strain, suggesting that the Agr and Sae may be inhibiting each other. The SigB mutation did not affect ssl5 and ssl8 expression, but they were downregulated in the agr/sigB double mutant, indicating that SigB probably acts synergistically with Agr in their upregulation.

  18. Genomics of a metamorphic timing QTL: met1 maps to a unique genomic position and regulates morph and species-specific patterns of brain transcription.

    PubMed

    Page, Robert B; Boley, Meredith A; Kump, David K; Voss, Stephen R

    2013-01-01

    Very little is known about genetic factors that regulate life history transitions during ontogeny. Closely related tiger salamanders (Ambystoma species complex) show extreme variation in metamorphic timing, with some species foregoing metamorphosis altogether, an adaptive trait called paedomorphosis. Previous studies identified a major effect quantitative trait locus (met1) for metamorphic timing and expression of paedomorphosis in hybrid crosses between the biphasic Eastern tiger salamander (Ambystoma tigrinum tigrinum) and the paedomorphic Mexican axolotl (Ambystoma mexicanum). We used existing hybrid mapping panels and a newly created hybrid cross to map the met1 genomic region and determine the effect of met1 on larval growth, metamorphic timing, and gene expression in the brain. We show that met1 maps to the position of a urodele-specific chromosome rearrangement on linkage group 2 that uniquely brought functionally associated genes into linkage. Furthermore, we found that more than 200 genes were differentially expressed during larval development as a function of met1 genotype. This list of differentially expressed genes is enriched for proteins that function in the mitochondria, providing evidence of a link between met1, thyroid hormone signaling, and mitochondrial energetics associated with metamorphosis. Finally, we found that met1 significantly affected metamorphic timing in hybrids, but not early larval growth rate. Collectively, our results show that met1 regulates species and morph-specific patterns of brain transcription and life history variation.

  19. Nucleotide sequencing and characterization of Pseudomonas putida catR: a positive regulator of the catBC operon is a member of the LysR family.

    PubMed Central

    Rothmel, R K; Aldrich, T L; Houghton, J E; Coco, W M; Ornston, L N; Chakrabarty, A M

    1990-01-01

    Pseudomonas putida utilizes the catBC operon for growth on benzoate as a sole carbon source. This operon is positively regulated by the CatR protein, which is encoded from a gene divergently oriented from the catBC operon. The catR gene encodes a 32.2-kilodalton polypeptide that binds to the catBC promoter region in the presence or absence of the inducer cis-cis-muconate, as shown by gel retardation studies. However, the inducer is required for transcriptional activation of the catBC operon. The catR promoter has been localized to a 385-base-pair fragment by using the broad-host-range promoter-probe vector pKT240. This fragment also contains the catBC promoter whose -35 site is separated by only 36 nucleotides from the predicted CatR translational start. Dot blot analysis suggests that CatR binding to this dual promoter-control region, in addition to inducing the catBC operon, may also regulate its own expression. Data from a computer homology search using the predicted amino acid sequence of CatR, deduced from the DNA sequence, showed CatR to be a member of a large class of procaryotic regulatory proteins designated the LysR family. Striking homology was seen between CatR and a putative regulatory protein, TfdS. Images FIG. 4 FIG. 5 FIG. 6 PMID:1688844

  20. Positive regulation of Fas gene expression by MSSP and abrogation of Fas-mediated apoptosis induction in MSSP-deficient mice.

    PubMed

    Nomura, Jun; Matsumoto, Ken-ichi; Iguchi-Ariga, Sanae M M; Ariga, Hiroyoshi

    2005-05-01

    MSSP has been identified as a transcription factor that regulates the c-myc gene. MSSP was later found to positively or negatively regulate a variety of genes, including alpha-smooth actin, MHC class I, MHC class 2 and the thyrotropin receptor. The knockout mice for the Mssp gene developed by us revealed that these mice became partially embryonic lethal due to a low concentration of progesterone at E2.5. In this study, we further analyzed Mssp-knockout mice and found that the expression of the Fas gene was repressed, resulting in abrogation of Fas-mediated induction of apoptosis both in Mssp-knockout mice and primary thymocytes. MSSP was then found to stimulate promoter activity of the Fas gene by binding to a region spanning -1035 to -635 in chromatin immunoprecipitation assays. Binding of MSSP in the MSSP-binding sequence, TCTAAT, located in this region was confirmed by mobility shift assays, and deletion of this sequence from the Fas promoter was found to result in loss of MSSP-dependent stimulating activity. The results suggest that MSSP is an important mediator for Fas-induced apoptosis in vivo and in vitro.

  1. Brachypodium distachyon BdPP2CA6 Interacts with BdPYLs and BdSnRK2 and Positively Regulates Salt Tolerance in Transgenic Arabidopsis.

    PubMed

    Zhang, Fan; Wei, Qiuhui; Shi, Jiaochun; Jin, Xia; He, Yuan; Zhang, Yang; Luo, Qingchen; Wang, Yuesheng; Chang, Junli; Yang, Guangxiao; He, Guangyuan

    2017-01-01

    The phytohormone abscisic acid (ABA) is essential in plant responding to biotic and abiotic stresses. Although ABA signaling model is well established in Arabidopsis, ABA receptor PYL family and clade A PP2C subfamily are not yet characterized in monocot model plant Brachypodium distachyon. In this study, we identified 12 PYLs and 8 clade A PP2Cs from B. distachyon genome and successfully cloned 12 PYLs and 7 clade A PP2Cs. Bioinformatic and expression analyses showed that most of the identified genes respond to several signal molecules and abiotic stresses. Protein-protein interaction analysis revealed that many BdPYLs and BdPP2CAs participate in the classic ABA-PYL-PP2C-SnRK2 signaling pathway. A clade A PP2C, designated BdPP2CA6, interacted with BdPYL11 in the absence of ABA and localized in nucleus. Most clade A PP2C members from Arabidopsis showed negatively regulation in ABA signaling pathway, whereas BdPP2CA6-overexpression transgenic Arabidopsis showed ABA hypersensitive phenotype, resulting in enhanced stomatal closure and salinity tolerance. Our results indicate that BdPP2CA6 positively regulates ABA and stress signal pathway in transgenic Arabidopsis plant seedlings.

  2. Characterization of the Pathway-Specific Positive Transcriptional Regulator for Actinorhodin Biosynthesis in Streptomyces coelicolor A3(2) as a DNA-Binding Protein

    PubMed Central

    Arias, Paloma; Fernández-Moreno, Miguel A.; Malpartida, Francisco

    1999-01-01

    The ActII-ORF4 protein has been characterized as a DNA-binding protein that positively regulates the transcription of the actinorhodin biosynthetic genes. The target regions for the ActII-ORF4 protein were located within the act cluster. These regions, at high copy number, generate a nonproducer strain by in vivo titration of the regulator. The mutant phenotype could be made to revert with extra copies of the wild-type actII-ORF4 gene but not with the actII-ORF4-177 mutant. His-tagged recombinant wild-type ActII-ORF4 and mutant ActII-ORF4-177 proteins were purified from Escherichia coli cultures; both showed specific DNA-binding activity for the actVI-ORF1–ORFA and actIII-actI intergenic regions. DNase I footprinting assays clearly located the DNA-binding sites within the −35 regions of the corresponding promoters, showing the consensus sequence 5′-TCGAG-3′. Although both gene products (wild-type and mutant ActII-ORF4) showed DNA-binding activity, only the wild-type gene was capable of activating transcription of the act genes; thus, two basic functions can be differentiated within the regulatory protein: a specific DNA-binding activity and a transcriptional activation of the act biosynthetic genes. PMID:10559161

  3. Genomics of a Metamorphic Timing QTL: met1 Maps to a Unique Genomic Position and Regulates Morph and Species-Specific Patterns of Brain Transcription

    PubMed Central

    Page, Robert B.; Boley, Meredith A.; Kump, David K.; Voss, Stephen R.

    2013-01-01

    Very little is known about genetic factors that regulate life history transitions during ontogeny. Closely related tiger salamanders (Ambystoma species complex) show extreme variation in metamorphic timing, with some species foregoing metamorphosis altogether, an adaptive trait called paedomorphosis. Previous studies identified a major effect quantitative trait locus (met1) for metamorphic timing and expression of paedomorphosis in hybrid crosses between the biphasic Eastern tiger salamander (Ambystoma tigrinum tigrinum) and the paedomorphic Mexican axolotl (Ambystoma mexicanum). We used existing hybrid mapping panels and a newly created hybrid cross to map the met1 genomic region and determine the effect of met1 on larval growth, metamorphic timing, and gene expression in the brain. We show that met1 maps to the position of a urodele-specific chromosome rearrangement on linkage group 2 that uniquely brought functionally associated genes into linkage. Furthermore, we found that more than 200 genes were differentially expressed during larval development as a function of met1 genotype. This list of differentially expressed genes is enriched for proteins that function in the mitochondria, providing evidence of a link between met1, thyroid hormone signaling, and mitochondrial energetics associated with metamorphosis. Finally, we found that met1 significantly affected metamorphic timing in hybrids, but not early larval growth rate. Collectively, our results show that met1 regulates species and morph-specific patterns of brain transcription and life history variation. PMID:23946331

  4. The RING Finger E3 Ligase SpRing is a Positive Regulator of Salt Stress Signaling in Salt-Tolerant Wild Tomato Species.

    PubMed

    Qi, Shilian; Lin, Qingfang; Zhu, Huishan; Gao, Fenghua; Zhang, Wenhao; Hua, Xuejun

    2016-03-01

    Protein ubiquitination in plants plays critical roles in many biological processes, including adaptation to abiotic stresses. Previously, RING finger E3 ligase has been characterized during salt stress response in several plant species, but little is known about its function in tomato. Here, we report that SpRing, a stress-inducible gene, is involved in salt stress signaling in wild tomato species Solanum pimpinellifolium 'PI365967'. In vitro ubiquitination assay revealed that SpRing is an E3 ubiquitin ligase and the RING finger conserved region is required for its activity. SpRing is expressed in all tissues of wild tomato and up-regulated by salt, drought and osmotic stresses, but repressed by low temperature. Green fluorescent protein (GFP) fusion analysis showed that SpRing is localized at the endoplasmic reticulum. Silencing of SpRing through a virus-induced gene silencing approach led to increased sensitivity to salt stress in wild tomato. Overexpression of SpRing in Arabidopsis thaliana resulted in enhanced salt tolerance during seed germination and early seedling development. The expression levels of certain key stress-related genes are altered both in SpRing-overexpressing Arabidopsis plants and virus-induced gene silenced tomato seedlings. Taken together, our results indicate that SpRing is involved in salt stress and functions as a positive regulator of salt tolerance.

  5. The Yeast Hrs1 Gene Is Involved in Positive and Negative Regulation of Transcription and Shows Genetic Characteristics Similar to Sin4 and Gal11

    PubMed Central

    Piruat, J. I.; Chavez, S.; Aguilera, A.

    1997-01-01

    We provide genetic evidence that HRS1/PGD1, a yeast gene previously identified as a suppressor of the hyper-recombination phenotype of hpr1, has positive and negative roles in transcriptional regulation. We have analyzed three differently regulated promoters, GAL1, PHO5 and HSP26, by β-galactosidase assays of lacZ-fused promoters and by Northern analysis of the endogenous genes. Transcription of these promoters was derepressed in hrs1δ mutants under conditions in which it is normally repressed in wild type. Under induced conditions it was either strongly reduced or significantly enhanced depending on the promoter system analyzed. Constitutive transcription was not affected, as determined in ADH1 and TEF2. In addition, Hrs1p was required for mating-factor expression, telomere-linked DNA silencing and DNA supercoiling of plasmids. Furthermore, hrs1δ suppressed Ty-insertion mutations and conferred a Gal(-) phenotype. Many of these phenotypes also result from mutations in GAL11, SIN4 or RGR1, which encode proteins of the RNA polII mediator. We also show that gal11δ and sin4δ partially suppress the hyper-rec phenotype of hpr1 mutants, although to a lesser extent than hrs1δ. Our results provide new evidence for the connection between hpr1δ-induced deletions and transcription. We discuss the possibility that Hrs1p might be a component of the RNA polII transcription machinery. PMID:9409823

  6. Mry, a trans-acting positive regulator of the M protein gene of Streptococcus pyogenes with similarity to the receptor proteins of two-component regulatory systems.

    PubMed Central

    Perez-Casal, J; Caparon, M G; Scott, J R

    1991-01-01

    In the Streptococcus pyogenes M6 strain D471, an insertion of the conjugative transposon Tn916 into a region 2 kb upstream of the promoter of emm6 (the structural gene for the M protein) rendered the strain M negative (M. G. Caparon and J. R. Scott, Proc. Natl. Acad. Sci. USA 84:8677-8681, 1987). In the present work, we show that this insertion mutation, mry-1, is 244 bp upstream of an open reading frame encoding a protein we call Mry. This protein is visible on a gel after transcription and translation in vitro. We have developed a technique for complementation analysis in S. pyogenes and have used it to show that the wild-type mry gene is dominant to two mutant alleles. This dominance indicates that Mry acts in trans as a positive regulator of the emm6 gene. The translated DNA sequence of mry has two regions of similarity to the motif common to the receptor protein of two-component regulatory systems. In addition, the N terminus of Mry has two regions resembling a helix-turn-helix motif. Mry does not appear to be a global regulator of virulence determinants in the group A streptococcus because there is no effect of the mry-1 mutation on production of the hyaluronic acid capsule or streptokinase. Images PMID:1849511

  7. TetR is a positive regulator of the tetanus toxin gene in Clostridium tetani and is homologous to botR.

    PubMed

    Marvaud, J C; Eisel, U; Binz, T; Niemann, H; Popoff, M R

    1998-12-01

    The TetR gene immediately upstream from the tetanus toxin (TeTx) gene was characterized. It encodes a 21,562-Da protein which is related (50 to 65% identity) to the equivalent genes (botR) in Clostridium botulinum. TetR has the feature of a DNA binding protein with a basic pI (9.53). It contains a helix-turn-helix motif and shows 29% identity with other putative regulatory genes in Clostridium, i.e., uviA from C. perfringens and txeR from C. difficile. We report for the first time the transformation of C. tetani by electroporation, which permitted us to investigate the function of tetR. Overexpression of tetR in C. tetani induced an increase in TeTx production and in the level of the corresponding mRNA. This indicates that TetR is a transcriptional activator of the TeTx gene. Overexpression of botR/A (60% identity with TetR at the amino acid level) in C. tetani induced an increase in TeTx production comparable to that for overexpression of tetR. However, botR/C (50% identity with TetR at the amino acid level) was less efficient. This supports that TetR positively regulates the TeTx gene in C. tetani and that a conserved mechanism of regulation of the neurotoxin genes is involved in C. tetani and C. botulinum.

  8. Capsicum annuum transcription factor WRKYa positively regulates defense response upon TMV infection and is a substrate of CaMK1 and CaMK2.

    PubMed

    Huh, Sung Un; Lee, Gil-Je; Jung, Ji Hoon; Kim, Yunsik; Kim, Young Jin; Paek, Kyung-Hee

    2015-01-23

    Plants are constantly exposed to pathogens and environmental stresses. To minimize damage caused by these potentially harmful factors, plants respond by massive transcriptional reprogramming of various stress-related genes via major transcription factor families. One of the transcription factor families, WRKY, plays an important role in diverse stress response of plants and is often useful to generate genetically engineered crop plants. In this study, we carried out functional characterization of CaWRKYa encoding group I WRKY member, which is induced during hypersensitive response (HR) in hot pepper (Capsicum annuum) upon Tobacco mosaic virus (TMV) infection. CaWRKYa was involved in L-mediated resistance via transcriptional reprogramming of pathogenesis-related (PR) gene expression and affected HR upon TMV-P0 infection. CaWRKYa acts as a positive regulator of this defense system and could bind to the W-box of diverse PR genes promoters. Furthermore, we found Capsicum annuum mitogen-activated protein kinase 1 (CaMK1) and 2 (CaMK2) interacted with CaWRKYa and phosphorylated the SP clusters but not the MAPK docking (D)-domain of CaWRKYa. Thus, these results demonstrated that CaWRKYa was regulated by CaMK1 and CaMK2 at the posttranslational level in hot pepper.

  9. Positive regulation of meiotic DNA double-strand break formation by activation of the DNA damage checkpoint kinase Mec1(ATR).

    PubMed

    Gray, Stephen; Allison, Rachal M; Garcia, Valerie; Goldman, Alastair S H; Neale, Matthew J

    2013-07-31

    During meiosis, formation and repair of programmed DNA double-strand breaks (DSBs) create genetic exchange between homologous chromosomes-a process that is critical for reductional meiotic chromosome segregation and the production of genetically diverse sexually reproducing populations. Meiotic DSB formation is a complex process, requiring numerous proteins, of which Spo11 is the evolutionarily conserved catalytic subunit. Precisely how Spo11 and its accessory proteins function or are regulated is unclear. Here, we use Saccharomyces cerevisiae to reveal that meiotic DSB formation is modulated by the Mec1(ATR) branch of the DNA damage signalling cascade, promoting DSB formation when Spo11-mediated catalysis is compromised. Activation of the positive feedback pathway correlates with the formation of single-stranded DNA (ssDNA) recombination intermediates and activation of the downstream kinase, Mek1. We show that the requirement for checkpoint activation can be rescued by prolonging meiotic prophase by deleting the NDT80 transcription factor, and that even transient prophase arrest caused by Ndt80 depletion is sufficient to restore meiotic spore viability in checkpoint mutants. Our observations are unexpected given recent reports that the complementary kinase pathway Tel1(ATM) acts to inhibit DSB formation. We propose that such antagonistic regulation of DSB formation by Mec1 and Tel1 creates a regulatory mechanism, where the absolute frequency of DSBs is maintained at a level optimal for genetic exchange and efficient chromosome segregation.

  10. SALT TOLERANCE HOMOLOG2, a B-box protein in Arabidopsis that activates transcription and positively regulates light-mediated development.

    PubMed

    Datta, Sourav; Hettiarachchi, Chamari; Johansson, Henrik; Holm, Magnus

    2007-10-01

    CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1) and ELONGATED HYPOCOTYL5 (HY5) are two major regulators of light signaling in plants. Here, we identify SALT TOLERANCE HOMOLOG2 (STH2) as a gene that interacts genetically with both of these key regulators. STH2 encodes a B-box-containing protein that interacts physically with HY5 in yeast and in plant cells. Whereas STH2 is uniformly nuclear by itself, it shows a COP1-dependent localization to speckles when coexpressed with COP1. We identified two independent T-DNA insertion lines in STH2. Both alleles are hyposensitive to blue, red, and far-red light. The sth2 mutant, like hy5, shows an enhanced number of lateral roots and accumulates less anthocyanin. Analysis of double mutants between sth2 and hy5 indicates that STH2 has both HY5-dependent and -independent functions. Furthermore, besides partially suppressing the hypocotyl phenotype of dark-grown cop1 alleles, sth2 also suppresses the reduced number of lateral roots and high anthocyanin levels in light-grown cop1 alleles. Interestingly, we found that STH2 can activate transcription. Transient transfection assays in protoplasts using a LUC reporter driven by the chalcone isomerase promoter show that the B-boxes in STH2 and a functional G-box element in the promoter are required for this activity. In conclusion, we have identified STH2, a B-box protein in Arabidopsis thaliana, as a positive regulator of photomorphogenesis and report that the B-box domain plays a direct role in activating transcription in plants.

  11. Expression of SOD and APX genes positively regulates secondary cell wall biosynthesis and promotes plant growth and yield in Arabidopsis under salt stress.

    PubMed

    Shafi, Amrina; Chauhan, Rohit; Gill, Tejpal; Swarnkar, Mohit K; Sreenivasulu, Yelam; Kumar, Sanjay; Kumar, Neeraj; Shankar, Ravi; Ahuja, Paramvir Singh; Singh, Anil Kumar

    2015-04-01

    Abiotic stresses cause accumulation of reactive oxygen species (ROS), such as hydrogen peroxide (H2O2) in plants. Sophisticated mechanisms are required to maintain optimum level of H2O2 that acts as signalling molecule regulating adaptive response to salt stress. CuZn-superoxide dismutase (CuZn-SOD) and ascorbate peroxidase (APX) constitute first line of defence against oxidative stress. In the present study, PaSOD and RaAPX genes from Potentilla atrosanguinea and Rheum australe, respectively were overexpressed individually as well as in combination in Arabidopsis thaliana. Interestingly, PaSOD and dual transgenic lines exhibit enhanced lignin deposition in their vascular bundles with altered S:G ratio under salt stress. RNA-seq analysis revealed that expression of PaSOD gene in single and dual transgenics positively regulates expression of lignin biosynthesis genes and transcription factors (NACs, MYBs, C3Hs and WRKY), leading to enhanced and ectopic deposition of lignin in vascular tissues with larger xylem fibres and alters S:G ratio, as well. In addition, transgenic plants exhibit growth promotion, higher biomass production and increased yield under salt stress as compared to wild type plants. Our results suggest that in dual transgenics, ROS generated during salt stress gets converted into H2O2 by SOD and its optimum level was maintained by APX. This basal level of H2O2 acts as messenger for transcriptional activation of lignin biosynthesis in vascular tissue, which provides mechanical strength to plants. These findings reveal an important role of PaSOD and RaAPX in enhancing salt tolerance of transgenic Arabidopsis via increased accumulation of compatible solutes and by regulating lignin biosynthesis.

  12. Positive feedback regulation between gamma-aminobutyric acid type A (GABA(A)) receptor signaling and brain-derived neurotrophic factor (BDNF) release in developing neurons.

    PubMed

    Porcher, Christophe; Hatchett, Caroline; Longbottom, Rebecca E; McAinch, Kristina; Sihra, Talvinder S; Moss, Stephen J; Thomson, Alex M; Jovanovic, Jasmina N

    2011-06-17

    During the early development of the nervous system, γ-aminobutyric acid (GABA) type A receptor (GABA(A)R)-mediated signaling parallels the neurotrophin/tropomyosin-related kinase (Trk)-dependent signaling in controlling a number of processes from cell proliferation and migration, via dendritic and axonal outgrowth, to synapse formation and plasticity. Here we present the first evidence that these two signaling systems regulate each other through a complex positive feedback mechanism. We first demonstrate that GABA(A)R activation leads to an increase in the cell surface expression of these receptors in cultured embryonic cerebrocortical neurons, specifically at the stage when this activity causes depolarization of the plasma membrane and Ca(2+) influx through L-type voltage-gated Ca(2+) channels. We further demonstrate that GABA(A)R activity triggers release of the brain-derived neurotrophic factor (BDNF), which, in turn by activating TrkB receptors, mediates the observed increase in cell surface expression of GABA(A)Rs. This BDNF/TrkB-dependent increase in surface levels of GABA(A)Rs requires the activity of phosphoinositide 3-kinase (PI3K) and protein kinase C (PKC) and does not involve the extracellular signal-regulated kinase (ERK) 1/2 activity. The increase in GABA(A)R surface levels occurs due to an inhibition of the receptor endocytosis by BDNF, whereas the receptor reinsertion into the plasma membrane remains unaltered. Thus, GABA(A)R activity is a potent regulator of the BDNF release during neuronal development, and at the same time, it is strongly enhanced by the activity of the BDNF/TrkB/PI3K/PKC signaling pathway.

  13. The small GTPase HRas shapes local PI3K signals through positive feedback and regulates persistent membrane extension in migrating fibroblasts

    PubMed Central

    Thevathasan, Jervis Vermal; Tan, Elisabeth; Zheng, Hui; Lin, Yu-Chun; Li, Yang; Inoue, Takanari; Fivaz, Marc

    2013-01-01

    Self-amplification of phosphoinositide 3-kinase (PI3K) signaling is believed to regulate asymmetric membrane extension and cell migration, but the molecular organization of the underlying feedback circuit is elusive. Here we use an inducible approach to synthetically activate PI3K and interrogate the feedback circuitry governing self-enhancement of 3′-phosphoinositide (3-PI) signals in NIH3T3 fibroblasts. Synthetic activation of PI3K initially leads to uniform production of 3-PIs at the plasma membrane, followed by the appearance of asymmetric and highly amplified 3-PI signals. A detailed spatiotemporal analysis shows that local self-amplifying 3-PI signals drive rapid membrane extension with remarkable directional persistence and initiate a robust migratory response. This positive feedback loop is critically dependent on the small GTPase HRas. Silencing of HRas abrogates local amplification of 3-PI signals upon synthetic PI3K activation and results in short-lived protrusion events that do not support cell migration. Finally, our data indicate that this feedback circuit is likely to operate during platelet-derived growth factor–induced random cell migration. We conclude that positive feedback between PI3K and HRas is essential for fibroblasts to spontaneously self-organize and generate a productive migratory response in the absence of spatial cues. PMID:23676667

  14. A functional C-terminal TRAF3-binding site in MAVS participates in positive and negative regulation of the IFN antiviral response.

    PubMed

    Paz, Suzanne; Vilasco, Myriam; Werden, Steven J; Arguello, Meztli; Joseph-Pillai, Deshanthe; Zhao, Tiejun; Nguyen, Thi Lien-Anh; Sun, Qiang; Meurs, Eliane F; Lin, Rongtuan; Hiscott, John

    2011-06-01

    Recognition of viral RNA structures by the cytosolic sensor retinoic acid-inducible gene-I (RIG-I) results in the activation of signaling cascades that culminate with the generation of the type I interferon (IFN) antiviral response. Onset of antiviral and inflammatory responses to viral pathogens necessitates the regulated spatiotemporal recruitment of signaling adapters, kinases and transcriptional proteins to the mitochondrial antiviral signaling protein (MAVS). We previously demonstrated that the serine/threonine kinase IKKε is recruited to the C-terminal region of MAVS following Sendai or vesicular stomatitis virus (VSV) infection, mediated by Lys63-linked polyubiquitination of MAVS at Lys500, resulting in inhibition of downstream IFN signaling (Paz et al, Mol Cell Biol, 2009). In this study, we demonstrate that C-terminus of MAVS harbors a novel TRAF3-binding site in the aa450-468 region of MAVS. A consensus TRAF-interacting motif (TIM), 455-PEENEY-460, within this site is required for TRAF3 binding and activation of IFN antiviral response genes, whereas mutation of the TIM eliminates TRAF3 binding and the downstream IFN response. Reconstitution of MAVS(-/-) mouse embryo fibroblasts with a construct expressing a TIM-mutated version of MAVS failed to restore the antiviral response or block VSV replication, whereas wild-type MAVS reconstituted antiviral inhibition of VSV replication. Furthermore, recruitment of IKKε to an adjacent C-terminal site (aa 468-540) in MAVS via Lys500 ubiquitination decreased TRAF3 binding and protein stability, thus contributing to IKKε-mediated shutdown of the IFN response. This study demonstrates that MAVS harbors a functional C-terminal TRAF3-binding site that participates in positive and negative regulation of the IFN antiviral response.

  15. Tryptophan-2,3-dioxygenase is regulated by prostaglandin E2 in malignant glioma via a positive signaling loop involving prostaglandin E receptor-4.

    PubMed

    Ochs, Katharina; Ott, Martina; Rauschenbach, Katharina J; Deumelandt, Katrin; Sahm, Felix; Opitz, Christiane A; von Deimling, Andreas; Wick, Wolfgang; Platten, Michael

    2015-12-27

    Malignant gliomas and other types of tumors generate a local immunosuppressive microenvironment, which prohibits an effective anti-tumor immune response and promotes tumor growth. Along with others, we have recently demonstrated that catabolism of the essential amino acid tryptophan via tryptophan-2,3-dioxygenase (TDO) is an important mechanism mediating tumor-associated immunosuppression particularly in gliomas. The pathways regulating TDO in tumors, however, are poorly understood. Here we show that prostaglandins enhance TDO expression and enzymatic activity in malignant gliomas via activation of prostaglandin E receptor-4 (EP4). Stimulation with prostaglandin E2 (PGE2 ) up-regulated TDO-mediated kynurenine release in human glioma cell lines while knockdown of the PGE2 receptor EP4 inhibited TDO expression and activity. In human malignant glioma tissue expression of the PGE2 -producing enzyme cyclooxygenase-2 (COX2) and its receptor EP4 were associated with TDO expression both on transcript and protein level. High expression of EP4 correlated with poor survival in malignant glioma patients WHO III-IV. Importantly, treatment of glioma cells with an EP4 inhibitor decreased TDO expression and activity. Moreover, TDO-over-expressing murine gliomas showed increased COX2 and EP4 expression suggesting a positive feedback mechanism in vivo. In summary, targeting EP4 may inhibit - in addition to immunosuppressive COX2 signaling - tryptophan degradation as another important immunosuppressive pathway and thus, could provide a dual clinically relevant immunotherapeutic avenue for the treatment of malignant gliomas. This article is protected by copyright. All rights reserved.

  16. FOXO3 is a glucocorticoid receptor target and regulates LKB1 and its own expression based on cellular AMP levels via a positive autoregulatory loop.

    PubMed

    Lützner, Nicolas; Kalbacher, Hubert; Krones-Herzig, Anja; Rösl, Frank

    2012-01-01

    FOXO3 is a transcription factor involved in the regulation of multiple physiological processes including cell cycle arrest, apoptosis, oxidative stress-response and energy metabolism. Although much is known about its post-translational modification, the transcriptional regulation of FOXO3, as well as the cross-talk between transcription and post-translational events, is still poorly understood. In the present study, we show that FOXO3 is an immediate early glucocorticoid receptor (GR) target, whose transcription is even further enhanced by conditions that mimic metabolic stress. Induction of FOXO3 transcription by GR-binding steroids was reversed by concomitant treatment with the GR antagonist RU-486, but further enhanced by stimuli that activate the AMP-activated protein kinase (AMPK). Analysis of genomic DNA and chromatin immunoprecipitation, as well as luciferase reporter assays, revealed two functional glucocorticoid responsive elements within the FOXO3 promoter. Furthermore, we provide functional evidence for a phosphorylation switch that explains how glucocorticoids induce transcriptional activation of the gene but subsequently inactivate the corresponding protein by site-specific phosphorylation. Only when AMPK is stimulated, pre-existing FOXO3 becomes reverted toward an active form. Energy deprived conditions thus activate FOXO3 on two different levels, namely transcriptional and post-translational. In that way, FOXO3 acts as a metabolic stress sensor that coordinates expression of LKB1, the master upstream kinase involved in metabolic sensing, depending on the energy status of the cell. Additionally, we show that FOXO3 binds and activates its own promoter via a positive autoregulatory feedback loop. In conclusion, our data explain how catabolic glucocorticoid hormones and high intracellular AMP levels cooperate in inducing FOXO3 transcription and in activating the corresponding protein.

  17. Negative regulation of p120GAP GTPase promoting activity by p210bcr/abl: implication for RAS-dependent Philadelphia chromosome positive cell growth

    PubMed Central

    1994-01-01

    The p210bcr/abl tyrosine kinase appears to be responsible for initiating and maintaining the leukemic phenotype in chronic myelogenous leukemia (CML) patients. p21ras-p120GAP interactions play a central role in transducing mitogenic signals. Therefore, we investigated whether p21ras and p120GAP are regulated by p210bcr/abl, and whether this activation is functionally significant for CML cell proliferation. We report that transient expression of p210bcr/abl in fibroblast-like cells induces simultaneous activation of p21ras and inhibition of GTPase-promoting activity of p120GAP, and confirm these data showing that downregulation of p210bcr/abl expression in CML cells with bcr/abl antisense oligodeoxynucleotides induces both inhibition of p21ras activation and stimulation of GTPase-promoting activity of p120GAP. Tyrosine phosphorylation of two p120GAP-associated proteins, p190 and p62, which may affect p120GAP activity, also depends on p210bcr/abl tyrosine kinase expression. Direct dependence of these effects on the kinase activity is proven in experiments in which expression of c-MYB protein in fibroblast-like cells or downregulation of c-MYB expression resulting in analogous inhibition of CML cell proliferation does not result in the same changes. Use of specific antisense oligodeoxynucleotides to downregulate p21ras expression revealed a requirement for functional p21ras in the proliferation of Philadelphia chromosome-positive CML primary cells. Thus, the p210bcr/abl-dependent regulation of p120GAP activity is responsible, in part, for the maintenance of p21ras in the active GTP-bound form, a crucial requirement for CML cell proliferation. PMID:8195713

  18. GsCBRLK, a calcium/calmodulin-binding receptor-like kinase, is a positive regulator of plant tolerance to salt and ABA stress.

    PubMed

    Yang, Liang; Ji, Wei; Zhu, Yanming; Gao, Peng; Li, Yong; Cai, Hua; Bai, Xi; Guo, Dianjing

    2010-05-01

    Calcium/calmodulin-dependent kinases play vital roles in protein phosphorylation in eukaryotes, yet little is known about the phosphorylation process of calcium/calmodulin-dependent protein kinase and its role in stress signal transduction in plants. A novel plant-specific calcium-dependent calmodulin-binding receptor-like kinase (GsCBRLK) has been isolated from Glycine soja. A subcellular localization study using GFP fusion protein indicated that GsCBRLK is localized in the plasma membrane. Binding assays demonstrated that calmodulin binds to GsCBRLK with an affinity of 25.9 nM in a calcium-dependent manner and the binding motif lies between amino acids 147 to169 within subdomain II of the kinase domain. GsCBRLK undergoes autophosphorylation and Myelin Basis Protein phosphorylation in the presence of calcium. It was also found that calcium/calmodulin positively regulates GsCBRLK kinase activity through direct interaction between the calmodulin-binding domain and calmodulin. So, it is likely that GsCBRLK responds to an environmental stimulus in two ways: by increasing the protein expression level and by regulating its kinase activity through the calcium/calmodulin complex. Furthermore, cold, salinity, drought, and ABA stress induce GsCBRLK gene transcripts. Over-expression of GsCBRLK in transgenic Arabidopsis resulted in enhanced plant tolerance to high salinity and ABA and increased the expression pattern of a number of stress gene markers in response to ABA and high salt. These results identify GsCBRLK as a molecular link between the stress- and ABA-induced calcium/calmodulin signal and gene expression in plant cells.

  19. Transcription enhancer factor 1 interacts with a basic helix-loop-helix zipper protein, Max, for positive regulation of cardiac alpha-myosin heavy-chain gene expression.

    PubMed Central

    Gupta, M P; Amin, C S; Gupta, M; Hay, N; Zak, R

    1997-01-01

    The M-CAT binding factor transcription enhancer factor 1 (TEF-1) has been implicated in the regulation of several cardiac and skeletal muscle genes. Previously, we identified an E-box-M-CAT hybrid (EM) motif that is responsible for the basal and cyclic AMP-inducible expression of the rat cardiac alpha-myosin heavy chain (alpha-MHC) gene in cardiac myocytes. In this study, we report that two factors, TEF-1 and a basic helix-loop-helix leucine zipper protein, Max, bind to the alpha-MHC EM motif. We also found that Max was a part of the cardiac troponin T M-CAT-TEF-1 complex even when the DNA template did not contain an apparent E-box binding site. In the protein-protein interaction assay, a stable association of Max with TEF-1 was observed when glutathione S-transferase (GST)-TEF-1 or GST-Max was used to pull down in vitro-translated Max or TEF-1, respectively. In addition, Max was coimmunoprecipitated with TEF-1, thus documenting an in vivo TEF-1-Max interaction. In the transient transcription assay, overexpression of either Max or TEF-1 resulted a mild activation of the alpha-MHC-chloramphenicol acetyltransferase (CAT) reporter gene at lower concentrations and repression of this gene at higher concentrations. However, when Max and TEF-1 expression plasmids were transfected together, the repression mediated by a single expression plasmid was alleviated and a three- to fourfold transactivation of the alpha-MHC-CAT reporter gene was observed. This effect was abolished once the EM motif in the promoter-reporter construct was mutated, thus suggesting that the synergistic transactivation function of the TEF-1-Max heterotypic complex is mediated through binding of the complex to the EM motif. These results demonstrate a novel association between Max and TEF-1 and indicate a positive cooperation between these two factors in alpha-MHC gene regulation. PMID:9199327

  20. A Positive Feedback Loop between Glial Cells Missing 1 and Human Chorionic Gonadotropin (hCG) Regulates Placental hCGβ Expression and Cell Differentiation.

    PubMed

    Cheong, Mei-Leng; Wang, Liang-Jie; Chuang, Pei-Yun; Chang, Ching-Wen; Lee, Yun-Shien; Lo, Hsiao-Fan; Tsai, Ming-Song; Chen, Hungwen

    2016-01-01

    Human chorionic gonadotropin (hCG) is composed of a common α subunit and a placenta-specific β subunit. Importantly, hCG is highly expressed in the differentiated and multinucleated syncytiotrophoblast, which is formed via trophoblast cell fusion and stimulated by cyclic AMP (cAMP). Although the ubiquitous activating protein 2 (AP2) transcription factors TFAP2A and TFAP2C may regulate hCGβ expression, it remains unclear how cAMP stimulates placenta-specific hCGβ gene expression and trophoblastic differentiation. Here we demonstrated that the placental transcription factor glial cells missing 1 (GCM1) binds to a highly conserved promoter region in all six hCGβ paralogues by chromatin immunoprecipitation-on-chip (ChIP-chip) analyses. We further showed that cAMP stimulates GCM1 and the CBP coactivator to activate the hCGβ promoter through a GCM1-binding site (GBS1), which also constitutes a previously identified AP2 site. Given that TFAP2C may compete with GCM1 for GBS1, cAMP enhances the association between the hCGβ promoter and GCM1 but not TFAP2C. Indeed, the hCG-cAMP-protein kinase A (PKA) signaling pathway also stimulates Ser269 and Ser275 phosphorylation of GCM1, which recruits CBP to mediate GCM1 acetylation and stabilization. Consequently, hCG stimulates the expression of GCM1 target genes, including the fusogenic protein syncytin-1, to promote placental cell fusion. Our study reveals a positive feedback loop between GCM1 and hCG regulating placental hCGβ expression and cell differentiation.

  1. A Positive Feedback Loop Links Opposing Functions of P-TEFb/Cdk9 and Histone H2B Ubiquitylation to Regulate Transcript Elongation in Fission Yeast

    PubMed Central

    Jacques, Pierre-Étienne; Pagé, Viviane; Nagy, Stephen; Racine, Ariane; St. Amour, Courtney V.; Zhang, Chao; Shokat, Kevan M.; Schwer, Beate; Robert, François; Fisher, Robert P.; Tanny, Jason C.

    2012-01-01

    Transcript elongation by RNA polymerase II (RNAPII) is accompanied by conserved patterns of histone modification. Whereas histone modifications have established roles in transcription initiation, their functions during elongation are not understood. Mono-ubiquitylation of histone H2B (H2Bub1) plays a key role in coordinating co-transcriptional histone modification by promoting site-specific methylation of histone H3. H2Bub1 also regulates gene expression through an unidentified, methylation-independent mechanism. Here we reveal bidirectional communication between H2Bub1 and Cdk9, the ortholog of metazoan positive transcription elongation factor b (P-TEFb), in the fission yeast Schizosaccharomyces pombe. Chemical and classical genetic analyses indicate that lowering Cdk9 activity or preventing phosphorylation of its substrate, the transcription processivity factor Spt5, reduces H2Bub1 in vivo. Conversely, mutations in the H2Bub1 pathway impair Cdk9 recruitment to chromatin and decrease Spt5 phosphorylation. Moreover, an Spt5 phosphorylation-site mutation, combined with deletion of the histone H3 Lys4 methyltransferase Set1, phenocopies morphologic and growth defects due to H2Bub1 loss, suggesting independent, partially redundant roles for Cdk9 and Set1 downstream of H2Bub1. Surprisingly, mutation of the histone H2B ubiquitin-acceptor residue relaxes the Cdk9 activity requirement in vivo, and cdk9 mutations suppress cell-morphology defects in H2Bub1-deficient strains. Genome-wide analyses by chromatin immunoprecipitation also demonstrate opposing effects of Cdk9 and H2Bub1 on distribution of transcribing RNAPII. Therefore, whereas mutual dependence of H2Bub1 and Spt5 phosphorylation indicates positive feedback, mutual suppression by cdk9 and H2Bub1-pathway mutations suggests antagonistic functions that must be kept in balance to regulate elongation. Loss of H2Bub1 disrupts that balance and leads to deranged gene expression and aberrant cell morphologies, revealing a

  2. Protein kinase A and regulation of neonatal Nav1.5 expression in human breast cancer cells: activity-dependent positive feedback and cellular migration.

    PubMed

    Chioni, Athina-Myrto; Shao, Dongmin; Grose, Richard; Djamgoz, Mustafa B A

    2010-02-01

    Voltage-gated Na(+) channels (VGSCs) are expressed in excitable cells (e.g. neurons and muscles), as well as in some classically 'non-excitable' cells (e.g. fibroblasts), and in carcinomas. In general, functional expression of VGSCs in plasma membrane (PM) is hierarchical and dynamic. Previously, we have shown that an activity-dependent positive feedback mechanism involving cAMP-dependent protein kinase A (PKA) plays a significant role in upregulation of VGSCs in strongly metastatic rat prostate cancer Mat-LyLu cells expressing Nav1.7. Here, we investigated the possible role of PKA in VGSC regulation and its functional consequences in strongly metastatic human breast cancer (BCa) MDA-MB-231 cells, where the neonatal splice form of Nav1.5 (nNav1.5) is the predominant VGSC present. Treatment with the PKA activator forskolin for 24h increased mRNA and PM protein levels of nNav1.5, without changing the total VGSC protein level. Opposite effects were obtained by application of the PKA inhibitor KT5720 or the highly specific VGSC blocker tetrodotoxin (TTX), the latter implying activity-dependent upregulation. We tested the possibility, therefore, that the activity dependence of VGSC (nNav1.5) expression involved PKA. Indeed, TTX pretreatment reduced the level of phosphorylated PKA and eliminated basal and PKA-stimulated cellular migration. These data suggested that activity-dependent positive feedback mediated by PKA plays an important role in the functional expression of nNav1.5 in BCa, and in turn, this enhances the cells' metastatic potential.

  3. Cross-Species Antiviral Activity of Goose Interferons against Duck Plague Virus Is Related to Its Positive Self-Feedback Regulation and Subsequent Interferon Stimulated Genes Induction

    PubMed Central

    Zhou, Hao; Chen, Shun; Zhou, Qin; Wei, Yunan; Wang, Mingshu; Jia, Renyong; Zhu, Dekang; Liu, Mafeng; Liu, Fei; Yang, Qiao; Wu, Ying; Sun, Kunfeng; Chen, Xiaoyue; Cheng, Anchun

    2016-01-01

    Interferons are a group of antiviral cytokines acting as the first line of defense in the antiviral immunity. Here, we describe the antiviral activity of goose type I interferon (IFNα) and type II interferon (IFNγ) against duck plague virus (DPV). Recombinant goose IFNα and IFNγ proteins of approximately 20 kDa and 18 kDa, respectively, were expressed. Following DPV-enhanced green fluorescent protein (EGFP) infection of duck embryo fibroblast cells (DEFs) with IFNα and IFNγ pre-treatment, the number of viral gene copies decreased more than 100-fold, with viral titers dropping approximately 100-fold. Compared to the control, DPV-EGFP cell positivity was decreased by goose IFNα and IFNγ at 36 hpi (3.89%; 0.79%) and 48 hpi (17.05%; 5.58%). In accordance with interferon-stimulated genes being the “workhorse” of IFN activity, the expression of duck myxovirus resistance (Mx) and oligoadenylate synthetases-like (OASL) was significantly upregulated (p < 0.001) by IFN treatment for 24 h. Interestingly, duck cells and goose cells showed a similar trend of increased ISG expression after goose IFNα and IFNγ pretreatment. Another interesting observation is that the positive feedback regulation of type I IFN and type II IFN by goose IFNα and IFNγ was confirmed in waterfowl for the first time. These results suggest that the antiviral activities of goose IFNα and IFNγ can likely be attributed to the potency with which downstream genes are induced by interferon. These findings will contribute to our understanding of the functional significance of the interferon antiviral system in aquatic birds and to the development of interferon-based prophylactic and therapeutic approaches against viral disease. PMID:27438848

  4. Positive and Negative Transcriptional Regulation of the Foxp3 gene is Mediated by TGF-β Signal Transducer Smad3 Access and Binding to Enhancer I

    PubMed Central

    Xu, Lili; Kitani, Atsushi; Stuelten, Christina; McGrady, George; Fuss, Ivan; Strober, Warren

    2010-01-01

    The molecular mechanisms underlying retinoic acid (RA) augmentation of T cell receptor (TCR) and transforming growth factor-β (TGF-β)-induced Foxp3 transcription and inhibition of the latter by cytokines such as IL-27 were here shown to be related processes involving modifications of baseline (TGF-β-induced) phosphorylated Smad3 (pSmad3) binding to a conserved enhancer region (enhancer I). RA augmentation involved the binding of retinoic acid receptor (RAR) and retinoid X receptor (RXR) to a dominant site in enhancer I and a subordinate site in the promoter. This led to increased histone acetylation in the region of the Smad3 binding site and increased binding of pSmad3. Cytokine (IL-27) inhibition involved binding of pStat3 to a gene silencer in a second conserved enhancer region (enhancer II) downstream from enhancer I; this led to loss of pSmad3 binding to enhancer I. Thus, control of accessibility and binding of pSmad3 provides a common framework for positive and negative regulation of TGF-β-induced Foxp3 transcription. PMID:20870174

  5. Nrf2 protects the lung against inflammation induced by titanium dioxide nanoparticles: A positive regulator role of Nrf2 on cytokine release.

    PubMed

    Delgado-Buenrostro, Norma L; Medina-Reyes, Estefany I; Lastres-Becker, Isabel; Freyre-Fonseca, Verónica; Ji, Zhaoxia; Hernández-Pando, Rogelio; Marquina, Brenda; Pedraza-Chaverri, José; Espada, Sandra; Cuadrado, Antonio; Chirino, Yolanda I

    2015-07-01

    Titanium dioxide nanoparticles (TiO2 NPs) have been classified as possibly carcinogenic to humans and they are an important nanomaterial widely used in pharmaceutical and paint industries. Inhalation is one of the most important routes of exposure in occupational settings. Several experimental models have shown that oxidative stress and inflammation are key mediators of cell damage. In this regard, Nrf2 modulates cytoprotection against oxidative stress and inflammation, however, its role in inflammation induced by TiO2 NPs exposure has been less investigated. The aim of this work was to investigate the role of Nrf2 in the cytokines produced after 4 weeks of TiO2 NPs exposure (5 mg/kg/2 days/week) using wild-type and Nrf2 knockout C57bl6 mice. Results showed that Nrf2 protects against inflammation and oxidative damage induced by TiO2 NPs exposure, however, Nrf2 is a positive mediator in the expression of IFN-γ, TNF-α, and TGF-β in bronchial epithelium and alveolar space after 4 weeks of exposure. These results suggest that Nrf2 has a central role in up-regulation of cytokines released during inflammation induced by TiO2 NPs and those cytokines are needed to cope with histological alterations in lung tissue.

  6. A putative positive feedback regulation mechanism in CsACS2 expression suggests a modified model for sex determination in cucumber (Cucumis sativus L.)

    PubMed Central

    Wang, Shu; Tao, Qianyi; Pan, Junsong; Si, Longting; Gong, Zhenhui; Cai, Run

    2012-01-01

    It is well established that the plant hormone ethylene plays a key role in cucumber sex determination. Since the unisexual control gene M was cloned and shown to encode an ethylene synthase, instead of an ethylene receptor, the ‘one-hormone hypothesis’, which was used to explain the cucumber sex phenotype, has been challenged. Here, the physiological function of CsACS2 (the gene encoded by the M locus) was studied using the transgenic tobacco system. The results indicated that overexpression of CsACS2 increased ethylene production in the tobacco plant, and the native cucumber promoter had no activity in transgenic tobacco (PM). However, when PM plants were treated with exogenous ethylene, CsACS2 expression could be detected. In cucumber, ethylene treatment could also induce transcription of CsACS2, while inhibition of ethylene action reduced the expression level. These findings suggest a positive feedback regulation mechanism for CsACS2, and a modified ‘one-hormone hypothesis’ for sex determination in cucumber is proposed. PMID:22577183

  7. AtrbohD and AtrbohF positively regulate abscisic acid-inhibited primary root growth by affecting Ca2+ signalling and auxin response of roots in Arabidopsis.

    PubMed

    Jiao, Yiheng; Sun, Lirong; Song, Yalin; Wang, Limin; Liu, Liping; Zhang, Liyue; Liu, Bo; Li, Ning; Miao, Chen; Hao, Fushun

    2013-11-01

    Reactive oxygen species (ROS) originating from the NADPH oxidases AtrbohD and AtrbohF play an important role in abscisic acid (ABA)-inhibited primary root growth in Arabidopsis. However, the mechanisms underlying this process remain elusive. In this study, the double mutant atrbohD1/F1 and atrbohD2/F2, in which both AtrbohD and AtrbohF were disrupted, were less sensitive to ABA suppression of root cell elongation than wild-type (WT) plants. Furthermore, the double mutants showed impaired ABA responses in roots, including ROS generation, cytosolic Ca(2+) increases, and activation of plasma membrane Ca(2+)-permeable channels compared with WT. Exogenous H2O2 can activate the Ca(2+) currents in roots of atrbohD1/F1. In addition, exogenous application of the auxin transport inhibitor naphthylphthalamic acid effectively promoted ABA inhibition of root growth of the mutants relative to that of WT. The ABA-induced decreases in auxin sensitivity of the root tips were more pronounced in WT than in atrbohD1/F1. These findings suggest that both AtrbohD and AtrbohF are essential for ABA-promoted ROS production in roots. ROS activate Ca(2+) signalling and reduce auxin sensitivity of roots, thus positively regulating ABA-inhibited primary root growth in Arabidopsis.

  8. A putative positive feedback regulation mechanism in CsACS2 expression suggests a modified model for sex determination in cucumber (Cucumis sativus L.).

    PubMed

    Li, Zheng; Wang, Shu; Tao, Qianyi; Pan, Junsong; Si, Longting; Gong, Zhenhui; Cai, Run

    2012-07-01

    It is well established that the plant hormone ethylene plays a key role in cucumber sex determination. Since the unisexual control gene M was cloned and shown to encode an ethylene synthase, instead of an ethylene receptor, the 'one-hormone hypothesis', which was used to explain the cucumber sex phenotype, has been challenged. Here, the physiological function of CsACS2 (the gene encoded by the M locus) was studied using the transgenic tobacco system. The results indicated that overexpression of CsACS2 increased ethylene production in the tobacco plant, and the native cucumber promoter had no activity in transgenic tobacco (PM). However, when PM plants were treated with exogenous ethylene, CsACS2 expression could be detected. In cucumber, ethylene treatment could also induce transcription of CsACS2, while inhibition of ethylene action reduced the expression level. These findings suggest a positive feedback regulation mechanism for CsACS2, and a modified 'one-hormone hypothesis' for sex determination in cucumber is proposed.

  9. PacMYBA, a sweet cherry R2R3-MYB transcription factor, is a positive regulator of salt stress tolerance and pathogen resistance.

    PubMed

    Shen, Xinjie; Guo, Xinwei; Guo, Xiao; Zhao, Di; Zhao, Wei; Chen, Jingsheng; Li, Tianhong

    2017-03-01

    Plant R2R3-MYB transcription factors play crucial roles in stress responses. We previously isolated a R2R3-MYB homolog from sweet cherry cv. Hong Deng, designated PacMYBA (GenBank accession No. KF974774). To explore the role of PacMYBA in the plant stress response, we heterologously expressed PacMYBA in transgenic Arabidopsis thaliana plants. In a previous study, we demonstrated that PacMYBA is mainly localized to the nucleus and could be induced by abscisic acid (ABA). Analysis of the promoter sequence of PacMYBA revealed that it contains several stress-related cis-elements. QPCR results showed that PacMYBA is induced by salt, salicylic (SA), and jasmonic acid (JA) in sweet cherry leaves. Transgenic Arabidopsis plants heterologously expressing PacMYBA exhibited enhanced salt-tolerance and increased resistance to Pseudomonas syringe pv. tomato (Pst) DC3000 infection. Overexpression of PacMYBA decreased the osmotic potential (OP), increased the free proline content, and increased the peroxidase content in transgenic Arabidopsis plants. Furthermore, overexpression of PacMYBA also affected the expression levels of salt stress- and pathogen defense-related genes in the transgenic plants. These results indicate that PacMYBA is a positive regulator of salt stress tolerance and pathogen resistance.

  10. The Arabidopsis mediator complex subunit16 positively regulates salicylate-mediated systemic acquired resistance and jasmonate/ethylene-induced defense pathways.

    PubMed

    Zhang, Xudong; Wang, Chenggang; Zhang, Yanping; Sun, Yijun; Mou, Zhonglin

    2012-10-01

    Systemic acquired resistance (SAR) is a long-lasting plant immunity against a broad spectrum of pathogens. Biological induction of SAR requires the signal molecule salicylic acid (SA) and involves profound transcriptional changes that are largely controlled by the transcription coactivator nonexpressor of pathogenesis-related genes1 (NPR1). However, it is unclear how SAR signals are transduced from the NPR1 signaling node to the general transcription machinery. Here, we report that the Arabidopsis thaliana Mediator subunit16 (MED16) is an essential positive regulator of SAR. Mutations in MED16 reduced NPR1 protein levels and completely compromised biological induction of SAR. These mutations also significantly suppressed SA-induced defense responses, altered the transcriptional changes induced by the avirulent bacterial pathogen Pseudomonas syringae pv tomato (Pst) DC3000/avrRpt2, and rendered plants susceptible to both Pst DC3000/avrRpt2 and Pst DC3000. In addition, mutations in MED16 blocked the induction of several jasmonic acid (JA)/ethylene (ET)-responsive genes and compromised resistance to the necrotrophic fungal pathogens Botrytis cinerea and Alternaria brassicicola. The Mediator complex acts as a bridge between specific transcriptional activators and the RNA polymerase II transcription machinery; therefore, our data suggest that MED16 may be a signaling component in the gap between the NPR1 signaling node and the general transcription machinery and may relay signals from both the SA and the JA/ET pathways.

  11. BnaC9.SMG7b Functions as a Positive Regulator of the Number of Seeds per Silique in Brassica napus by Regulating the Formation of Functional Female Gametophytes.

    PubMed

    Li, Shipeng; Chen, Lei; Zhang, Liwu; Li, Xi; Liu, Ying; Wu, Zhikun; Dong, Faming; Wan, Lili; Liu, Kede; Hong, Dengfeng; Yang, Guangsheng

    2015-12-01

    Number of seeds per silique (NSS) is an important determinant of seed yield potential in Brassicaceae crops, and it is controlled by naturally occurring quantitative trait loci. We previously mapped a major quantitative trait locus, qSS.C9, on the C9 chromosome that controls NSS in Brassica napus. To gain a better understanding of how qSS.C9 controls NSS in B. napus, we isolated this locus through a map-based cloning strategy. qSS.C9 encodes a predicted small protein with 119 amino acids, designated as BnaC9.SMG7b, that shows homology with the Ever ShorterTelomere1 tertratricopeptide repeats and Ever Shorter Telomere central domains of Arabidopsis (Arabidopsis thaliana) SUPPRESSOR WITH MORPHOGENETIC EFFECTS ON GENITALIA7 (SMG7). BnaC9.SMG7b plays a role in regulating the formation of functional female gametophyte, thus determining the formation of functional megaspores and then mature ovules. Natural loss or artificial knockdown of BnaC9.SMG7b significantly reduces the number of functional ovules per silique and thus, results in decreased seed number, indicating that qSS.C9 is a positive regulator of NSS in B. napus. Sequence and function analyses show that BnaC9.SMG7b experiences a subfunctionalization process that causes loss of function in nonsense-mediated mRNA decay, such as in Arabidopsis SMG7. Haplotype analysis in 84 accessions showed that the favorable BnaC9.SMG7b alleles are prevalent in modern B. napus germplasms, suggesting that this locus has been a major selection target of B. napus improvement. Our results represent the first step toward unraveling the molecular mechanism that controls the natural variation of NSS in B. napus.

  12. BnaC9.SMG7b Functions as a Positive Regulator of the Number of Seeds per Silique in Brassica napus by Regulating the Formation of Functional Female Gametophytes1

    PubMed Central

    Li, Shipeng; Chen, Lei; Zhang, Liwu; Li, Xi; Liu, Ying; Wu, Zhikun; Dong, Faming; Wan, Lili; Liu, Kede; Yang, Guangsheng

    2015-01-01

    Number of seeds per silique (NSS) is an important determinant of seed yield potential in Brassicaceae crops, and it is controlled by naturally occurring quantitative trait loci. We previously mapped a major quantitative trait locus, qSS.C9, on the C9 chromosome that controls NSS in Brassica napus. To gain a better understanding of how qSS.C9 controls NSS in B. napus, we isolated this locus through a map-based cloning strategy. qSS.C9 encodes a predicted small protein with 119 amino acids, designated as BnaC9.SMG7b, that shows homology with the Ever ShorterTelomere1 tertratricopeptide repeats and Ever Shorter Telomere central domains of Arabidopsis (Arabidopsis thaliana) SUPPRESSOR WITH MORPHOGENETIC EFFECTS ON GENITALIA7 (SMG7). BnaC9.SMG7b plays a role in regulating the formation of functional female gametophyte, thus determining the formation of functional megaspores and then mature ovules. Natural loss or artificial knockdown of BnaC9.SMG7b significantly reduces the number of functional ovules per silique and thus, results in decreased seed number, indicating that qSS.C9 is a positive regulator of NSS in B. napus. Sequence and function analyses show that BnaC9.SMG7b experiences a subfunctionalization process that causes loss of function in nonsense-mediated mRNA decay, such as in Arabidopsis SMG7. Haplotype analysis in 84 accessions showed that the favorable BnaC9.SMG7b alleles are prevalent in modern B. napus germplasms, suggesting that this locus has been a major selection target of B. napus improvement. Our results represent the first step toward unraveling the molecular mechanism that controls the natural variation of NSS in B. napus. PMID:26494121

  13. VOLTAGE REGULATOR

    DOEpatents

    Von Eschen, R.L.; Scheele, P.F.

    1962-04-24

    A transistorized voltage regulator which provides very close voitage regulation up to about 180 deg F is described. A diode in the positive line provides a constant voltage drop from the input to a regulating transistor emitter. An amplifier is coupled to the positive line through a resistor and is connected between a difference circuit and the regulating transistor base which is negative due to the difference in voltage drop across thc diode and the resistor so that a change in the regulator output causes the amplifier to increase or decrease the base voltage and current and incrcase or decrease the transistor impedance to return the regulator output to normal. (AEC)

  14. CDC-25.1 controls the rate of germline mitotic cell cycle by counteracting WEE-1.3 and by positively regulating CDK-1 in Caenorhabditis elegans.

    PubMed

    Yoon, Sunghee; Kawasaki, Ichiro; Shim, Yhong-Hee

    2012-04-01

    In Caenorhabditis elegans, cdc-25.1 loss-of-function mutants display a lack of germline proliferation. We found that the proliferation defect of cdc-25.1 mutants was suppressed by wee-1.3 RNAi. Further, among the seven cdk and seven cyclin homologs examined, cdk-1 and cyb-3 RNAi treatment caused the most severe germline proliferation defects in an rrf-1 mutant background, which were similar to those of the cdc-25.1 mutants. In addition, while RNAi of cyd-1 and cye-1 caused significant germline proliferation defects, RNAi of cdk-2 and cdk-4 did not. Compared with the number of germ nuclei in wee-1.3(RNAi) worms, the number in wee-1.3(RNAi);cdk-1(RNAi) and wee-1.3(RNAi);cyb-3(RNAi) worms further decreased to the level of cdk-1(RNAi) and cyb-3(RNAi) worms, respectively, indicating that cdk-1 and cyb-3 are epistatic and function downstream of cdc-25.1 and wee-1.3 in the control of the cell cycle. BrdU labeling of adult worms showed that, while 100% of the wild-type germ nuclei in the mitotic region incorporated BrdU when labeled for more than 12 h at 20°C, a small fraction of the cdc-25.1 mutant germ nuclei failed to incorporate BrdU even when labeled for 68 h. These results indicate that CDC-25.1 is required for maintaining proper rate of germline mitotic cell cycle. We propose that CDC-25.1 regulates the rate of germline mitotic cell cycle by counteracting WEE-1.3 and by positively controlling CDK-1, which forms a complex primarily with CYB-3, but also possibly with CYD-1 and CYE-1.

  15. HvDep1 Is a Positive Regulator of Culm Elongation and Grain Size in Barley and Impacts Yield in an Environment-Dependent Manner

    PubMed Central

    Wendt, Toni; Holme, Inger; Dockter, Christoph; Preuß, Aileen; Thomas, William; Waugh, Robbie; Braumann, Ilka

    2016-01-01

    Heterotrimeric G proteins are intracellular membrane-attached signal transducers involved in various cellular processes in both plants and animals. They consist of three subunits denoted as α, β and γ. The γ-subunits of the so-called AGG3 type, which comprise a transmembrane domain, are exclusively found in plants. In model species, these proteins have been shown to participate in the control of plant height, branching and seed size and could therefore impact the harvestable yield of various crop plants. Whether AGG3-type γ-subunits influence yield in temperate cereals like barley and wheat remains unknown. Using a transgenic complementation approach, we show here that the Scottish malting barley cultivar (cv.) Golden Promise carries a loss-of-function mutation in HvDep1, an AGG3-type subunit encoding gene that positively regulates culm elongation and seed size in barley. Somewhat intriguingly, agronomic field data collected over a 12-year period reveals that the HvDep1 loss-of-function mutation in cv. Golden Promise has the potential to confer either a significant increase or decrease in harvestable yield depending on the environment. Our results confirm the role of AGG3-type subunit-encoding genes in shaping plant architecture, but interestingly also indicate that the impact HvDep1 has on yield in barley is both genotypically and environmentally sensitive. This may explain why widespread exploitation of variation in AGG3-type subunit-encoding genes has not occurred in temperate cereals while in rice the DEP1 locus is widely exploited to improve harvestable yield. PMID:28005988

  16. The Arabidopsis AAA ATPase SKD1 is involved in multivesicular endosome function and interacts with its positive regulator LYST-INTERACTING PROTEIN5.

    PubMed

    Haas, Thomas J; Sliwinski, Marek K; Martínez, Dana E; Preuss, Mary; Ebine, Kazuo; Ueda, Takashi; Nielsen, Erik; Odorizzi, Greg; Otegui, Marisa S

    2007-04-01

    In yeast and mammals, the AAA ATPase Vps4p/SKD1 (for Vacuolar protein sorting 4/SUPPRESSOR OF K(+) TRANSPORT GROWTH DEFECT1) is required for the endosomal sorting of secretory and endocytic cargo. We identified a VPS4/SKD1 homolog in Arabidopsis thaliana, which localizes to the cytoplasm and to multivesicular endosomes. In addition, green fluorescent protein-SKD1 colocalizes on multivesicular bodies with fluorescent fusion protein endosomal Rab GTPases, such as ARA6/RabF1, RHA1/RabF2a, and ARA7/RabF2b, and with the endocytic marker FM4-64. The expression of SKD1(E232Q), an ATPase-deficient version of SKD1, induces alterations in the endosomal system of tobacco (Nicotiana tabacum) Bright Yellow 2 cells and ultimately leads to cell death. The inducible expression of SKD1(E232Q) in Arabidopsis resulted in enlarged endosomes with a reduced number of internal vesicles. In a yeast two-hybrid screen using Arabidopsis SKD1 as bait, we isolated a putative homolog of mammalian LYST-INTERACTING PROTEIN5 (LIP5)/SKD1 BINDING PROTEIN1 and yeast Vta1p (for Vps twenty associated 1 protein). Arabidopsis LIP5 acts as a positive regulator of SKD1 by increasing fourfold to fivefold its in vitro ATPase activity. We isolated a knockout homozygous Arabidopsis mutant line with a T-DNA insertion in LIP5. lip5 plants are viable and show no phenotypic alterations under normal growth conditions, suggesting that basal SKD1 ATPase activity is sufficient for plant development and growth.

  17. The Lectin Receptor Kinase-VI.2 Is Required for Priming and Positively Regulates Arabidopsis Pattern-Triggered Immunity[C][W

    PubMed Central

    Singh, Prashant; Kuo, Yi-Chun; Mishra, Swati; Tsai, Chia-Hong; Chien, Chih-Cheng; Chen, Ching-Wei; Desclos-Theveniau, Marie; Chu, Po-Wei; Schulze, Birgit; Chinchilla, Delphine; Boller, Thomas; Zimmerli, Laurent

    2012-01-01

    Plant cells can be sensitized toward a subsequent pathogen attack by avirulent pathogens or by chemicals such as β-aminobutyric acid (BABA). This process is called priming. Using a reverse genetic approach in Arabidopsis thaliana, we demonstrate that the BABA-responsive L-type lectin receptor kinase-VI.2 (LecRK-VI.2) contributes to disease resistance against the hemibiotrophic Pseudomonas syringae and the necrotrophic Pectobacterium carotovorum bacteria. Accordingly, LecRK-VI.2 mRNA levels increased after bacterial inoculation or treatments with microbe-associated molecular patterns (MAMPs). We also show that LecRK-VI.2 is required for full activation of pattern-triggered immunity (PTI); notably, lecrk-VI.2-1 mutants show reduced upregulation of PTI marker genes, impaired callose deposition, and defective stomatal closure. Overexpression studies combined with genome-wide microarray analyses indicate that LecRK-VI.2 positively regulates the PTI response. LecRK-VI.2 is demonstrated to act upstream of mitogen-activated protein kinase signaling, but independently of reactive oxygen production and BOTRYTIS-INDUCED KINASE1 phosphorylation. In addition, complex formation between the MAMP receptor FLAGELLIN SENSING2 and its signaling partner BRASSINOSTEROID INSENSITIVE1-ASSOCIATED KINASE1 is observed in flg22-treated lecrk-VI.2-1 mutants. LecRK-VI.2 is also required for full BABA-induced resistance and priming of PTI. Our work identifies LecRK-VI.2 as a novel mediator of the Arabidopsis PTI response and provides insight into molecular mechanisms governing priming. PMID:22427336

  18. OsCCD1, a novel small calcium-binding protein with one EF-hand motif, positively regulates osmotic and salt tolerance in rice.

    PubMed

    Jing, Pei; Zou, Juanzi; Kong, Lin; Hu, Shiqi; Wang, Biying; Yang, Jun; Xie, Guosheng

    2016-06-01

    Calcium-binding proteins play key roles in the signal transduction in the growth and stress response in eukaryotes. However, a subfamily of proteins with one EF-hand motif has not been fully studied in higher plants. Here, a novel small calcium-binding protein with a C-terminal centrin-like domain (CCD1) in rice, OsCCD1, was characterized to show high similarity with a TaCCD1 in wheat. As a result, OsCCD1 can bind Ca(2+) in the in vitro EMSA and the fluorescence staining calcium-binding assays. Transient expression of green fluorescent protein (GFP)-tagged OsCCD1 in rice protoplasts showed that OsCCD1 was localized in the nucleus and cytosol of rice cells. OsCCD1 transcript levels were transiently induced by osmotic stress and salt stress through the calcium-mediated ABA signal. The rice seedlings of T-DNA mutant lines showed significantly less tolerance to osmotic and salt stresses than wild type plants (p<0.01). Conversely, its overexpressors can significantly enhance the tolerance to osmotic and salt stresses than wild type plants (p<0.05). Semi-quantitative RT-PCR analysis revealed that, OsDREB2B, OsAPX1 and OsP5CS genes are involved in the rice tolerance to osmotic and salt stresses. In sum, OsCCD1 gene probably affects the DREB2B and its downstream genes to positively regulate osmotic and salt tolerance in rice seedlings.

  19. Overexpression of CaWRKY27, a subgroup IIe WRKY transcription factor of Capsicum annuum, positively regulates tobacco resistance to Ralstonia solanacearum infection.

    PubMed

    Dang, Fengfeng; Wang, Yuna; She, Jianju; Lei, Yufen; Liu, Zhiqin; Eulgem, Thomas; Lai, Yan; Lin, Jing; Yu, Lu; Lei, Dan; Guan, Deyi; Li, Xia; Yuan, Qian; He, Shuilin

    2014-03-01

    WRKY proteins are encoded by a large gene family and are linked to many biological processes across a range of plant species. The functions and underlying mechanisms of WRKY proteins have been investigated primarily in model plants such as Arabidopsis and rice. The roles of these transcription factors in non-model plants, including pepper and other Solanaceae, are poorly understood. Here, we characterize the expression and function of a subgroup IIe WRKY protein from pepper (Capsicum annuum), denoted as CaWRKY27. The protein localized to nuclei and activated the transcription of a reporter GUS gene construct driven by the 35S promoter that contained two copies of the W-box in its proximal upstream region. Inoculation of pepper cultivars with Ralstonia solanacearum induced the expression of CaWRKY27 transcript in 76a, a bacterial wilt-resistant pepper cultivar, whereas it downregulated the expression of CaWRKY27 transcript in Gui-1-3, a bacterial wilt-susceptible pepper cultivar. CaWRKY27 transcript levels were also increased by treatments with salicylic acid (SA), methyl jasmonate (MeJA) and ethephon (ETH). Transgenic tobacco plants overexpressing CaWRKY27 exhibited resistance to R. solanacearum infection compared to that of wild-type plants. This resistance was coupled with increased transcript levels in a number of marker genes, including hypersensitive response genes, and SA-, JA- and ET-associated genes. By contrast, virus-induced gene silencing (VIGS) of CaWRKY27 increased the susceptibility of pepper plants to R. solanacearum infection. These results suggest that CaWRKY27 acts as a positive regulator in tobacco resistance responses to R. solanacearum infection through modulation of SA-, JA- and ET-mediated signaling pathways.

  20. Positive Regulation by γ-Aminobutyric Acid B Receptor Subunit-1 of Chondrogenesis through Acceleration of Nuclear Translocation of Activating Transcription Factor-4*

    PubMed Central

    Takahata, Yoshifumi; Hinoi, Eiichi; Takarada, Takeshi; Nakamura, Yukari; Ogawa, Shinya; Yoneda, Yukio

    2012-01-01

    A view that signaling machineries for the neurotransmitter γ-aminobutyric acid (GABA) are functionally expressed by cells outside the central nervous system is now prevailing. In this study, we attempted to demonstrate functional expression of GABAergic signaling molecules by chondrocytes. In cultured murine costal chondrocytes, mRNA was constitutively expressed for metabotropic GABAB receptor subunit-1 (GABABR1), but not for GABABR2. Immunohistochemical analysis revealed the predominant expression of GABABR1 by prehypertrophic to hypertrophic chondrocytes in tibial sections of newborn mice. The GABABR agonist baclofen failed to significantly affect chondrocytic differentiation determined by Alcian blue staining and alkaline phosphatase activity in cultured chondrocytes, whereas newborn mice knocked out of GABABR1 (KO) showed a decreased body size and delayed calcification in hyoid bone and forelimb and hindlimb digits. Delayed calcification was also seen in cultured metatarsals from KO mice with a marked reduction of Indian hedgehog gene (Ihh) expression. Introduction of GABABR1 led to synergistic promotion of the transcriptional activity of activating transcription factor-4 (ATF4) essential for normal chondrogenesis, in addition to facilitating ATF4-dependent Ihh promoter activation. Although immunoreactive ATF4 was negligibly detected in the nucleus of chondrocytes from KO mice, ATF4 expression was again seen in the nucleus and cytoplasm after the retroviral introduction of GABABR1 into cultured chondrocytes from KO mice. In nuclear extracts of KO chondrocytes, a marked decrease was seen in ATF4 DNA binding. These results suggest that GABABR1 positively regulates chondrogenesis through a mechanism relevant to the acceleration of nuclear translocation of ATF4 for Ihh expression in chondrocytes. PMID:22879594

  1. Simulated Microgravity Regulates Gene Transcript Profiles of 2T3 Preosteoblasts: Comparison of the Random Positioning Machine and the Rotating Wall Vessel Bioreactor

    NASA Technical Reports Server (NTRS)

    Patel, Mamta J.; Liu, Wenbin; Sykes, Michelle C.; Ward, Nancy E.; Risin, Semyon A.; Risin, Diana; Hanjoong, Jo

    2007-01-01

    Microgravity of spaceflight induces bone loss due in part to decreased bone formation by osteoblasts. We have previously examined the microgravity-induced changes in gene expression profiles in 2T3 preosteoblasts using the Random Positioning Machine (RPM) to simulate microgravity conditions. Here, we hypothesized that exposure of preosteoblasts to an independent microgravity simulator, the Rotating Wall Vessel (RWV), induces similar changes in differentiation and gene transcript profiles, resulting in a more confined list of gravi-sensitive genes that may play a role in bone formation. In comparison to static 1g controls, exposure of 2T3 cells to RWV for 3 days inhibited alkaline phosphatase activity, a marker of differentiation, and downregulated 61 genes and upregulated 45 genes by more than two-fold as shown by microarray analysis. The microarray results were confirmed with real time PCR for downregulated genes osteomodulin, bone morphogenic protein 4 (BMP4), runx2, and parathyroid hormone receptor 1. Western blot analysis validated the expression of three downregulated genes, BMP4, peroxiredoxin IV, and osteoglycin, and one upregulated gene peroxiredoxin I. Comparison of the microarrays from the RPM and the RWV studies identified 14 gravi-sensitive genes that changed in the same direction in both systems. Further comparison of our results to a published database showing gene transcript profiles of mechanically loaded mouse tibiae revealed 16 genes upregulated by the loading that were shown to be downregulated by RWV and RPM. These mechanosensitive genes identified by the comparative studies may provide novel insights into understanding the mechanisms regulating bone formation and potential targets of countermeasure against decreased bone formation both in astronauts and in general patients with musculoskeletal disorders.

  2. Multiplex ligation dependent probe amplification (MLPA) for rapid distinction between unique sequence positive and negative marker chromosomes in prenatal diagnosis

    PubMed Central

    2011-01-01

    Background Small supernumerary marker chromosomes (sSMC) are extra structurally abnormal chromosomes that cannot be unambiguously identified with conventional chromosome banding techniques. These marker chromosomes may cause an abnormal phenotype or be harmless depending on different factors such as genetic content, chromosomal origin and level of mosaicism. When a sSMC is found during prenatal diagnosis, the main question is whether the sSMC contains euchromatin since in most cases this will lead to phenotypic abnormalities. We present the use of Multiplex Ligation Dependent probe Amplification (MLPA) for rapid distinction between non-euchromatic and euchromatic sSMC. Results 29 well-defined sSMC found during prenatal diagnosis were retrospectively investigated with MLPA with the SALSA MLPA centromere kits P181 and P182 as well as with the SALSA MLPA telomere kits P036B and P070 (MRC Holland BV, Amsterdam, The Netherlands). All unique-sequence positive sSMC were correctly identified with MLPA, whereas the unique-sequence negative sSMC had normal MLPA results. Conclusions Although different techniques exist for identification of sSMC, we show that MLPA is a valuable adjunctive tool for rapidly distinguishing between unique-sequence positive and negative sSMC. In case of positive MLPA results, genetic microarray analysis or, if not available, targeted FISH can be applied for further identification and determination of the exact breakpoints, which is important for prediction of the fetal phenotype. In case of a negative MLPA result, which means that the sSMC most probably does not contain genes, the parents can already be reassured and parental karyotyping can be initiated to assess the heritability. In the mean time, FISH techniques are needed for determination of the chromosomal origin. PMID:21235775

  3. The qTSN Positive Effect on Panicle and Flag Leaf Size of Rice is Associated with an Early Down-Regulation of Tillering

    PubMed Central

    Adriani, Dewi E.; Lafarge, Tanguy; Dardou, Audrey; Fabro, Aubrey; Clément-Vidal, Anne; Yahya, Sudirman; Dingkuhn, Michael; Luquet, Delphine

    2016-01-01

    The qTSN4 was identified as rice QTL (Quantitative Traits Locus) increasing total spikelet number per panicle and flag leaf area but potentially reducing panicle number depending on the environment. So far, this trade-off was mainly observed at grain maturity and not specifically studied in details, limiting the apprehension of the agronomic interest of qTSN4. This study aimed to understand the effect of qTSN4 and of the environment on panicle sizing, its trade-off with panicle number, and finally plant grain production. It compared two high yielding genotypes to their Near Isogenic Lines (NIL) carrying either QTL qTSN4 or qTSN12, two distinct QTLs contributing to the enlarged panicle size, thereafter designated as qTSN. Traits describing C sink (organ appearance rate, size, biomass) and source (leaf area, photosynthesis, sugar availability) were dynamically characterized along plant and/or panicle development within two trials (greenhouse, field), each comparing two treatments contrasting for plant access to light (with or without shading, high or low planting densities). The positive effect of qTSN on panicle size and flag leaf area of the main tiller was confirmed. More precisely, it could be shown that qTSN increased leaf area and internode cross-section, and in some cases of the photosynthetic rate and starch reserves, of the top 3–4 phytomers of the main tiller. This was accompanied by an earlier tillering cessation, that coincided with the initiation of these phytomers, and an enhanced panicle size on the main tiller. Plant leaf area at flowering was not affected by qTSN but fertile tiller number was reduced to an extent that depended on the environment. Accordingly, plant grain production was enhanced by qTSN only under shading in the greenhouse experiment, where panicle number was not affected and photosynthesis and starch storage in internodes was enhanced. The effect of qTSN on rice phenotype was thus expressed before panicle initiation (PI). Whether

  4. Homemade-device-induced negative pressure promotes wound healing more efficiently than VSD-induced positive pressure by regulating inflammation, proliferation and remodeling

    PubMed Central

    Liu, Jinyan; Hu, Feng; Tang, Jintian; Tang, Shijie; Xia, Kun; Wu, Song; Yin, Chaoqi; Wang, Shaohua; He, Quanyong; Xie, Huiqing; Zhou, Jianda

    2017-01-01

    Vacuum sealing drainage (VSD) is an effective technique used to promote wound healing. However, recent studies have shown that it exerts positive pressure (PP) rather than negative pressure (NP) on skin. In this study, we created a homemade device that could maintain NP on the wound, and compared the therapeutic effects of VSD-induced PP to those of our home-made device which induced NP on wound healing. The NP induced by our device required less time for wound healing and decreased the wound area more efficiently than the PP induced by VSD. NP and PP both promoted the inflammatory response by upregulating neutrophil infiltration and interleukin (IL)-1β expression, and downregulating IL-10 expression. Higher levels of epidermal growth factor (EGF), transforming growth factor (TGF)-β and platelet-derived growth factor (PDGF), and lower levels of basic fibroblast growth factor (bFGF) were observed in the wound tissue treated with NP compared to the wound tissue exposed to PP. Proliferation in the wound tissue exposed to NP on day 10 was significantly higher than that in wound tissue exposed to PP. NP generated more fibroblasts, keratinized stratified epithelium, and less epithelia with stemness than PP. The levels of ccollagen I and III were both decreased in both the NP and PP groups. NP induced a statistically significant increase in the expression of fibronectin (FN) on days 3 and 10 compared to PP. Furthermore, the level of matrix metalloproteinase (MMP)-13 increased in the NP group, but decreased in the PP group on day 3. NP also induced a decrease in the levels of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2 during the early stages of wound healing, which was significantly different from the increasing effect of PP on TIMP-1 and TIMP-2 levels at the corresponding time points. On the whole, our data indicate that our homemade device which induced NP, was more efficient than VSD-induced PP on wound healing by regulating inflammation, secretion

  5. Keratinocyte-derived IL-24 plays a role in the positive feedback regulation of epidermal inflammation in response to environmental and endogenous toxic stressors

    SciTech Connect

    Jin, Sun Hee; Choi, Dalwoong; Chun, Young-Jin; Noh, Minsoo

    2014-10-15

    Keratinocytes are the major cellular components of human epidermis and play a key role in the modulating cutaneous inflammation and toxic responses. In human chronic skin diseases, the common skin inflammatory phenotypes like skin barrier disruption and epidermal hyperplasia are manifested in epidermal keratinocytes by interactions with T helper (Th) cells. To find a common gene expression signature of human keratinocytes in chronic skin diseases, we performed a whole genome microarray analysis on normal human epidermal keratinocytes (NHKs) treated with IFNγ, IL-4, IL-17A or IL-22, major cytokines from Th1, Th2, Th17 or Th22 cells, respectively. The microarray results showed that the four genes, IL-24, PDZK1IP1, H19 and filaggrin, had common expression profiles in NHKs exposed to Th cell cytokines. In addition, the acute phase pro-inflammatory cytokines, IL-1β, IL-6 and TNFα, also change the gene transcriptional profile of IL-24, PDZK1IP1, H19, and filaggrin in NHKs as those of Th cytokines. Therefore, the signature gene set, consisting of IL-24, PDZK1IP1, H19, and filaggrin, provides essential insights for understanding the process of cutaneous inflammation and toxic responses. We demonstrate that environmental toxic stressors, such as chemical irritants and ultraviolet irradiation stimulate the production of IL-24 in NHKs. IL-24 stimulates the JAK1-STAT3 and MAPK pathways in NHKs, and promotes the secretion of pro-inflammatory mediators IL-8, PGE2, and MMP-1. These results suggest that keratinocyte-derived IL-24 participates in the positive feedback regulation of epidermal inflammation in response to both endogenous and environmental toxic stressors. - Highlights: • Cutaneous inflammatory gene signature consists of PDZK1IP1, IL-24, H19 and filaggrin. • Pro-inflammatory cytokines increase IL-24 production in human keratinocytes. • Environmental toxic stressors increase IL-24 production in human keratinocytes. • IL-24 stimulates human keratinocytes to

  6. Positive regulation of the beta-galactosidase gene from Kluyveromyces lactis is mediated by an upstream activation site that shows homology to the GAL upstream activation site of Saccharomyces cerevisiae.

    PubMed Central

    Ruzzi, M; Breunig, K D; Ficca, A G; Hollenberg, C P

    1987-01-01

    In contrast to the Escherichia coli lac operon, the yeast beta-galactosidase gene is positively regulated. In the 5'-noncoding region of the Kluyveromyces lactis LAC4 gene, we mapped an upstream activation site (UAS) that is re