Decisive factors affecting plasma resistance and roughness formation in ArF photoresist

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Jinnai, Butsurin; Uesugi, Takuji; Koyama, Koji; Samukawa, Seiji [Institute of Fluid Science, Tohoku University, 2-1-1 Katahira, Aoba-ku, Sendai 980-8577 (Japan); Kato, Keisuke; Yasuda, Atsushi; Maeda, Shinichi [Yokohama Research Laboratories, Mitsubishi Rayon Co., Ltd, 10-1 Daikoku-cho, Tsurumi-ku, Yokohama 230-0053 (Japan); Momose, Hikaru, E-mail: samukawa@ifs.tohoku.ac.j [Corporate Research Laboratories, Mitsubishi Rayon Co., Ltd, 2-1 Miyuki-cho, Otake, Hiroshima 739-0693 (Japan)

2010-10-06

Low plasma resistance and roughness formation in an ArF photoresist are serious issues in plasma processes. To resolve these issues, we investigated several factors that affect the roughness formation and plasma resistance in an ArF photoresist. We used our neutral beam process to categorize the effects of species from the plasma on the ArF photoresist into physical bombardment, chemical reactions and ultraviolet/vacuum ultraviolet (UV/VUV) radiation. The UV/VUV radiation drastically increased the etching rates of the ArF photoresist films, and, in contrast, chemical reactions increased the formation of surface roughness. FTIR analysis indicated that the UV/VUV radiation preferentially dissociates C-H bonds in the ArF photoresist, rather than C=O bonds, because of the dissociation energies of the bonds. This indicated that the etching rates of the ArF photoresist are determined by the UV/VUV radiation because this radiation can break C-H bonds, which account for the majority of structures in the ArF photoresist. In contrast, FTIR analysis showed that chemical species such as radicals and ions were likely to react with C=O bonds, in particular C=O bonds in the lactone groups of the ArF photoresist, due to the structural and electronic effects of the lactone groups. As a result, the etching rates of the ArF photoresist can vary in different bond structures, leading to increased surface roughness in the ArF photoresist.

Over-expression of mango (Mangifera indica L.) MiARF2 inhibits root and hypocotyl growth of Arabidopsis.

Science.gov (United States)

Wu, Bei; Li, Yun-He; Wu, Jian-Yong; Chen, Qi-Zhu; Huang, Xia; Chen, Yun-Feng; Huang, Xue-Lin

2011-06-01

An auxin response factor 2 gene, MiARF2, was cloned in our previous study [1] from the cotyledon section of mango (Mangifera indica L. cv. Zihua) during adventitious root formation, which shares an 84% amino acid sequence similarity to Arabidopsis ARF2. This study was to examine the effects of over-expression of the full-length MiARF2 open reading frame on the root and hypocotyl growth in Arabidopsis. Phenotype analysis showed that the T(3) transgenic lines had about 20-30% reduction in the length of hypocotyls and roots of the seedlings in comparison with the wild-type. The transcription levels of ANT and ARGOS genes which play a role in controlling organ size and cell proliferation in the transgenic seedlings also decreased. Therefore, the inhibited root and hypocotyl growth in the transgenic seedlings may be associated with the down-regulated transcription of ANT and ARGOS by the over-expression of MiARF2. This study also suggests that although MiARF2 only has a single DNA-binding domain (DBD), it can function as other ARF-like proteins containing complete DBD, middle region (MR) and carboxy-terminal dimerization domain (CTD).

Gibberellin Application at Pre-Bloom in Grapevines Down-Regulates the Expressions of VvIAA9 and VvARF7, Negative Regulators of Fruit Set Initiation, during Parthenocarpic Fruit Development

Science.gov (United States)

Jung, Chan Jin; Hur, Youn Young; Yu, Hee-Ju; Noh, Jung-Ho; Park, Kyo-Sun; Lee, Hee Jae

2014-01-01

Fruit set is initiated only after fertilization and is tightly regulated primarily by gibberellins (GAs) and auxins. The application of either of these hormones induces parthenocarpy, fruit set without fertilization, but the molecular mechanism underlying this induction is poorly understood. In the present study, we have shown that the parthenocarpic fruits induced by GA application at pre-bloom result from the interaction of GA with auxin signaling. The transcriptional levels of the putative negative regulators of fruit set initiation, including Vitis auxin/indole-3-acetic acid transcription factor 9 (VvIAA9), Vitis auxin response factor 7 (VvARF7), and VvARF8 were monitored during inflorescence development in seeded diploid ‘Tamnara’ grapevines with or without GA application. Without GA application, VvIAA9, VvARF7, and VvARF8 were expressed at a relatively high level before full bloom, but decreased thereafter following pollination. After GA application at 14 days before full bloom (DBF); however, the expression levels of VvIAA9 and VvARF7 declined at 5 DBF prior to pollination. The effects of GA application on auxin levels or auxin signaling were also analyzed by monitoring the expression patterns of auxin biosynthesis genes and auxin-responsive genes with or without GA application. Transcription levels of the auxin biosynthesis genes Vitis anthranilate synthase β subunit (VvASB1-like), Vitis YUCCA2 (VvYUC2), and VvYUC6 were not significantly changed by GA application. However, the expressions of Vitis Gretchen Hagen3.2 (VvGH3.2) and VvGH3.3, auxin-responsive genes, were up-regulated from 2 DBF to full bloom with GA application. Furthermore, the Vitis GA signaling gene, VvDELLA was up-regulated by GA application during 12 DBF to 7 DBF, prior to down-regulation of VvIAA9 and VvARF7. These results suggest that VvIAA9 and VvARF7 are negative regulators of fruit set initiation in grapevines, and GA signaling is integrated with auxin signaling via VvDELLA during

The transcription factor EGR1 localizes to the nucleolus and is linked to suppression of ribosomal precursor synthesis.

Science.gov (United States)

Ponti, Donatella; Bellenchi, Gian Carlo; Puca, Rosa; Bastianelli, Daniela; Maroder, Marella; Ragona, Giuseppe; Roussel, Pascal; Thiry, Marc; Mercola, Dan; Calogero, Antonella

2014-01-01

EGR1 is an immediate early gene with a wide range of activities as transcription factor, spanning from regulation of cell growth to differentiation. Numerous studies show that EGR1 either promotes the proliferation of stimulated cells or suppresses the tumorigenic growth of transformed cells. Upon interaction with ARF, EGR1 is sumoylated and acquires the ability to bind to specific targets such as PTEN and in turn to regulate cell growth. ARF is mainly localized to the periphery of nucleolus where is able to negatively regulate ribosome biogenesis. Since EGR1 colocalizes with ARF under IGF-1 stimulation we asked the question of whether EGR1 also relocate to the nucleolus to interact with ARF. Here we show that EGR1 colocalizes with nucleolar markers such as fibrillarin and B23 in the presence of ARF. Western analysis of nucleolar extracts from HeLa cells was used to confirm the presence of EGR1 in the nucleolus mainly as the 100 kDa sumoylated form. We also show that the level of the ribosomal RNA precursor 47S is inversely correlated to the level of EGR1 transcripts. The EGR1 iseffective to regulate the synthesis of the 47S rRNA precursor. Then we demonstrated that EGR1 binds to the Upstream Binding Factor (UBF) leading us to hypothesize that the regulating activity of EGR1 is mediated by its interaction within the transcriptional complex of RNA polymerase I. These results confirm the presence of EGR1 in the nucleolus and point to a role for EGR1 in the control of nucleolar metabolism.

The transcription factor EGR1 localizes to the nucleolus and is linked to suppression of ribosomal precursor synthesis.

Directory of Open Access Journals (Sweden)

Donatella Ponti

Full Text Available EGR1 is an immediate early gene with a wide range of activities as transcription factor, spanning from regulation of cell growth to differentiation. Numerous studies show that EGR1 either promotes the proliferation of stimulated cells or suppresses the tumorigenic growth of transformed cells. Upon interaction with ARF, EGR1 is sumoylated and acquires the ability to bind to specific targets such as PTEN and in turn to regulate cell growth. ARF is mainly localized to the periphery of nucleolus where is able to negatively regulate ribosome biogenesis. Since EGR1 colocalizes with ARF under IGF-1 stimulation we asked the question of whether EGR1 also relocate to the nucleolus to interact with ARF. Here we show that EGR1 colocalizes with nucleolar markers such as fibrillarin and B23 in the presence of ARF. Western analysis of nucleolar extracts from HeLa cells was used to confirm the presence of EGR1 in the nucleolus mainly as the 100 kDa sumoylated form. We also show that the level of the ribosomal RNA precursor 47S is inversely correlated to the level of EGR1 transcripts. The EGR1 iseffective to regulate the synthesis of the 47S rRNA precursor. Then we demonstrated that EGR1 binds to the Upstream Binding Factor (UBF leading us to hypothesize that the regulating activity of EGR1 is mediated by its interaction within the transcriptional complex of RNA polymerase I. These results confirm the presence of EGR1 in the nucleolus and point to a role for EGR1 in the control of nucleolar metabolism.

Structural Dynamics Control Allosteric Activation of Cytohesin Family Arf GTPase Exchange Factors

Energy Technology Data Exchange (ETDEWEB)

Malaby, Andrew W.; Das, Sanchaita; Chakravarthy, Srinivas; Irving, Thomas C.; Bilsel, Osman; Lambright, David G.

2018-01-01

Membrane dynamic processes including vesicle biogenesis depend on Arf guanosine triphosphatase (GTPase) activation by guanine nucleotide exchange factors (GEFs) containing a catalytic Sec7 domain and a membrane-targeting module such as a pleckstrin homology (PH) domain. The catalytic output of cytohesin family Arf GEFs is controlled by autoinhibitory interactions that impede accessibility of the exchange site in the Sec7 domain. These restraints can be relieved through activator Arf-GTP binding to an allosteric site comprising the PH domain and proximal autoinhibitory elements (Sec7-PH linker and C-terminal helix). Small-angle X-ray scattering and negative-stain electron microscopy were used to investigate the structural organization and conformational dynamics of cytohesin-3 (Grp1) in autoinhibited and active states. The results support a model in which hinge dynamics in the autoinhibited state expose the activator site for Arf-GTP binding, while subsequent C-terminal helix unlatching and repositioning unleash conformational entropy in the Sec7-PH linker to drive exposure of the exchange site.

Losses of both products of the Cdkn2a/Arf locus contribute to asbestos-induced mesothelioma development and cooperate to accelerate tumorigenesis.

Directory of Open Access Journals (Sweden)

Deborah A Altomare

2011-04-01

Full Text Available The CDKN2A/ARF locus encompasses overlapping tumor suppressor genes p16(INK4A and p14(ARF, which are frequently co-deleted in human malignant mesothelioma (MM. The importance of p16(INK4A loss in human cancer is well established, but the relative significance of p14(ARF loss has been debated. The tumor predisposition of mice singly deficient for either Ink4a or Arf, due to targeting of exons 1α or 1β, respectively, supports the idea that both play significant and nonredundant roles in suppressing spontaneous tumors. To further test this notion, we exposed Ink4a(+/- and Arf(+/- mice to asbestos, the major cause of MM. Asbestos-treated Ink4a(+/- and Arf(+/- mice showed increased incidence and shorter latency of MM relative to wild-type littermates. MMs from Ink4a(+/- mice exhibited biallelic inactivation of Ink4a, loss of Arf or p53 expression and frequent loss of p15(Ink4b. In contrast, MMs from Arf(+/- mice exhibited loss of Arf expression, but did not require loss of Ink4a or Ink4b. Mice doubly deficient for Ink4a and Arf, due to deletion of Cdkn2a/Arf exon 2, showed accelerated asbestos-induced MM formation relative to mice deficient for Ink4a or Arf alone, and MMs exhibited biallelic loss of both tumor suppressor genes. The tumor suppressor function of Arf in MM was p53-independent, since MMs with loss of Arf retained functional p53. Collectively, these in vivo data indicate that both CDKN2A/ARF gene products suppress asbestos carcinogenicity. Furthermore, while inactivation of Arf appears to be crucial for MM pathogenesis, the inactivation of both p16(Ink4a and p19(Arf cooperate to accelerate asbestos-induced tumorigenesis.

Identification of PEG-induced water stress responsive transcripts using co-expression network in Eucalyptus grandis.

Science.gov (United States)

Ghosh Dasgupta, Modhumita; Dharanishanthi, Veeramuthu

2017-09-05

Ecophysiological studies in Eucalyptus have shown that water is the principal factor limiting stem growth. Effect of water deficit conditions on physiological and biochemical parameters has been extensively reported in Eucalyptus. The present study was conducted to identify major polyethylene glycol induced water stress responsive transcripts in Eucalyptus grandis using gene co-expression network. A customized array representing 3359 water stress responsive genes was designed to document their expression in leaves of E. grandis cuttings subjected to -0.225MPa of PEG treatment. The differentially expressed transcripts were documented and significantly co-expressed transcripts were used for construction of network. The co-expression network was constructed with 915 nodes and 3454 edges with degree ranging from 2 to 45. Ninety four GO categories and 117 functional pathways were identified in the network. MCODE analysis generated 27 modules and module 6 with 479 nodes and 1005 edges was identified as the biologically relevant network. The major water responsive transcripts represented in the module included dehydrin, osmotin, LEA protein, expansin, arabinogalactans, heat shock proteins, major facilitator proteins, ARM repeat proteins, raffinose synthase, tonoplast intrinsic protein and transcription factors like DREB2A, ARF9, AGL24, UNE12, WLIM1 and MYB66, MYB70, MYB 55, MYB 16 and MYB 103. The coordinated analysis of gene expression patterns and coexpression networks developed in this study identified an array of transcripts that may regulate PEG induced water stress responses in E. grandis. Copyright © 2017 Elsevier B.V. All rights reserved.

Enhanceosomes as integrators of hypoxia inducible factor (HIF) and other transcription factors in the hypoxic transcriptional response.

Science.gov (United States)

Pawlus, Matthew R; Hu, Cheng-Jun

2013-09-01

Hypoxia is a prevalent attribute of the solid tumor microenvironment that promotes the expression of genes through posttranslational modifications and stabilization of alpha subunits (HIF1α and HIF2α) of hypoxia-inducible factors (HIFs). Despite significant similarities, HIF1 (HIF1α/ARNT) and HIF2 (HIF2α/ARNT) activate common as well as unique target genes and exhibit different functions in cancer biology. More surprisingly, accumulating data indicates that the HIF1- and/or HIF2-mediated hypoxia responses can be oncogenic as well as tumor suppressive. While the role of HIF in the hypoxia response is well established, recent data support the concept that HIF is necessary, but not sufficient for the hypoxic response. Other transcription factors that are activated by hypoxia are also required for the HIF-mediated hypoxia response. HIFs, other transcription factors, co-factors and RNA poll II recruited by HIF and other transcription factors form multifactorial enhanceosome complexes on the promoters of HIF target genes to activate hypoxia inducible genes. Importantly, HIF1 or HIF2 requires distinct partners in activating HIF1 or HIF2 target genes. Because HIF enhanceosome formation is required for the gene activation and distinct functions of HIF1 and HIF2 in tumor biology, disruption of the HIF1 or HIF2 specific enhanceosome complex may prove to be a beneficial strategy in tumor treatment in which tumor growth is specifically dependent upon HIF1 or HIF2 activity. Copyright © 2013 Elsevier Inc. All rights reserved.

Cancer cell specific cytotoxic gene expression mediated by ARF tumor suppressor promoter constructs

International Nuclear Information System (INIS)

Kurayoshi, Kenta; Ozono, Eiko; Iwanaga, Ritsuko; Bradford, Andrew P.; Komori, Hideyuki; Ohtani, Kiyoshi

2014-01-01

Highlights: • ARF promoter showed higher responsiveness to deregulated E2F activity than the E2F1 promoter. • ARF promoter showed higher cancer cell-specificity than E2F1 promoter to drive gene expression. • HSV-TK driven by ARF promoter showed higher cancer cell-specific cytotoxicity than that driven by E2F1 promoter. - Abstract: In current cancer treatment protocols, such as radiation and chemotherapy, side effects on normal cells are major obstacles to radical therapy. To avoid these side effects, a cancer cell-specific approach is needed. One way to specifically target cancer cells is to utilize a cancer specific promoter to express a cytotoxic gene (suicide gene therapy) or a viral gene required for viral replication (oncolytic virotherapy). For this purpose, the selected promoter should have minimal activity in normal cells to avoid side effects, and high activity in a wide variety of cancers to obtain optimal therapeutic efficacy. In contrast to the AFP, CEA and PSA promoters, which have high activity only in a limited spectrum of tumors, the E2F1 promoter exhibits high activity in wide variety of cancers. This is based on the mechanism of carcinogenesis. Defects in the RB pathway and activation of the transcription factor E2F, the main target of the RB pathway, are observed in almost all cancers. Consequently, the E2F1 promoter, which is mainly regulated by E2F, has high activity in wide variety of cancers. However, E2F is also activated by growth stimulation in normal growing cells, suggesting that the E2F1 promoter may also be highly active in normal growing cells. In contrast, we found that the tumor suppressor ARF promoter is activated by deregulated E2F activity, induced by forced inactivation of pRB, but does not respond to physiological E2F activity induced by growth stimulation. We also found that the deregulated E2F activity, which activates the ARF promoter, is detected only in cancer cell lines. These observations suggest that ARF promoter

Cancer cell specific cytotoxic gene expression mediated by ARF tumor suppressor promoter constructs

Energy Technology Data Exchange (ETDEWEB)

Kurayoshi, Kenta [Department of Bioscience, School of Science and Technology, Kwansei Gakuin University, 2-1 Gakuen, Sanda, Hyogo 669-1337 (Japan); Ozono, Eiko [Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary, University of London, John Vane Science Centre, Charterhouse Square, London EC1M 6BQ (United Kingdom); Iwanaga, Ritsuko; Bradford, Andrew P. [Department of Obstetrics and Gynecology, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO 80045 (United States); Komori, Hideyuki [Center for Stem Cell Biology, Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109 (United States); Ohtani, Kiyoshi, E-mail: btm88939@kwansei.ac.jp [Department of Bioscience, School of Science and Technology, Kwansei Gakuin University, 2-1 Gakuen, Sanda, Hyogo 669-1337 (Japan)

2014-07-18

Highlights: • ARF promoter showed higher responsiveness to deregulated E2F activity than the E2F1 promoter. • ARF promoter showed higher cancer cell-specificity than E2F1 promoter to drive gene expression. • HSV-TK driven by ARF promoter showed higher cancer cell-specific cytotoxicity than that driven by E2F1 promoter. - Abstract: In current cancer treatment protocols, such as radiation and chemotherapy, side effects on normal cells are major obstacles to radical therapy. To avoid these side effects, a cancer cell-specific approach is needed. One way to specifically target cancer cells is to utilize a cancer specific promoter to express a cytotoxic gene (suicide gene therapy) or a viral gene required for viral replication (oncolytic virotherapy). For this purpose, the selected promoter should have minimal activity in normal cells to avoid side effects, and high activity in a wide variety of cancers to obtain optimal therapeutic efficacy. In contrast to the AFP, CEA and PSA promoters, which have high activity only in a limited spectrum of tumors, the E2F1 promoter exhibits high activity in wide variety of cancers. This is based on the mechanism of carcinogenesis. Defects in the RB pathway and activation of the transcription factor E2F, the main target of the RB pathway, are observed in almost all cancers. Consequently, the E2F1 promoter, which is mainly regulated by E2F, has high activity in wide variety of cancers. However, E2F is also activated by growth stimulation in normal growing cells, suggesting that the E2F1 promoter may also be highly active in normal growing cells. In contrast, we found that the tumor suppressor ARF promoter is activated by deregulated E2F activity, induced by forced inactivation of pRB, but does not respond to physiological E2F activity induced by growth stimulation. We also found that the deregulated E2F activity, which activates the ARF promoter, is detected only in cancer cell lines. These observations suggest that ARF promoter

[Identification of an auxin response factor-like protein cDNA from mango cotyledon section].

Science.gov (United States)

Xiao, Jie-Ning; Huang, Xue-Lin; Huang, Xia; Li, Xiao-Ju

2004-01-01

Auxin-responsive elements (AuxRE) interact with a new class of plant-specific transcription factors, auxin response factors (ARFs). Some of ARFs have been shown to repress or activate expression of genes with an AuxRE promotor element. In Arabidopsis, ARFs play important roles in early embryo development and vascular strand formation (ARF5), floral patterning (ARF3) and photo- and gravitropic responses (ARF7). Two cut surfaces (distal and proximal) of mango (Mangifera indica L. var. Zi-Hua) cotyledon showed different patterns of adventitious root formation, with only the proximal cut surface, but not the distal one, could be induced to form the roots. Thus, the mango cotyledon is a good system for studying adventitious root formation. A cDNA fragment homologous to the Arabidopsis auxin response factor-like protein and relates to adventitious root formation from the cut sections were isolated using suppressive subtractive hybridization (SSH). Two cDNA clones, designated as MiARF1 (mango auxin response factor 1 gene, GenBank accession number AY255705) and MiARF2 (mango auxin response factor 2 gene, GenBank accession number is AY300808), were identified by 3'RACE. MiARF1, 3 272bp long, contains an open reading frame (ORF) of 2 523bp, 5'UTR of 285bp and 3'UTR of 464bp, MiARF2, 1 474bp long, contains an ORF of 981bp, 5' UTR of 285bp and 3'UTR of 208bp. The deduced MiARF1 and MiARF2 are homologues of auxin response factor (ARF) family of transcriptional regulators, and show high similarity to ARF of Arabidopsis in conserved domains. The motifs of MiARF1 EL-WHACAGPL in DBD (DNA binding domain) and GDDPW in IV domain are identical to that of ARF-like protein of Arabidopsis. MiARF2 is identical to MiARF1 in a large part of DBD, but lacks a carboxyl-terminal domain containing conserved motifs III and IV. Virtual Northern blot showed that the expression of MiARF2 was high in rooting tissue of cultured cotyledon sections but low in non-rooting tissue, and the MiARF1 was

Membrane association of the Arabidopsis ARF exchange factor GNOM involves interaction of conserved domains

DEFF Research Database (Denmark)

Anders, Nadine; Nielsen, Michael M.; Keicher, Jutta

2008-01-01

vesicle formation by activating ARF GTPases on specific membranes in animals, plants, and fungi. However, apart from the catalytic exchange activity of the SEC7 domain, the functional significance of other conserved domains is virtually unknown. Here, we show that a distinct N-terminal domain of GNOM......The GNOM protein plays a fundamental role in Arabidopsis thaliana development by regulating endosome-to-plasma membrane trafficking required for polar localization of the auxin efflux carrier PIN1. GNOM is a family member of large ARF guanine nucleotide exchange factors (ARF-GEFs), which regulate...... mediates dimerization and in addition interacts heterotypically with two other conserved domains in vivo. In contrast with N-terminal dimerization, the heterotypic interaction is essential for GNOM function, as mutations abolishing this interaction inactivate the GNOM protein and compromise its membrane...

The transcription factor MEF2C mediates cardiomyocyte hypertrophy induced by IGF-1 signaling

Energy Technology Data Exchange (ETDEWEB)

Munoz, Juan Pablo; Collao, Andres; Chiong, Mario; Maldonado, Carola; Adasme, Tatiana; Carrasco, Loreto; Ocaranza, Paula; Bravo, Roberto; Gonzalez, Leticia; Diaz-Araya, Guillermo [Centro FONDAP Estudios Moleculares de la Celula, Facultad de Medicina, Universidad de Chile, Santiago 8380492 (Chile); Facultad de Ciencias Quimicas y Farmaceuticas, Facultad de Medicina, Universidad de Chile, Santiago 8380492 (Chile); Hidalgo, Cecilia [Centro FONDAP Estudios Moleculares de la Celula, Facultad de Medicina, Universidad de Chile, Santiago 8380492 (Chile); Instituto de Ciencias Biomedicas, Facultad de Medicina, Universidad de Chile, Santiago 8380492 (Chile); Lavandero, Sergio, E-mail: slavander@uchile.cl [Centro FONDAP Estudios Moleculares de la Celula, Facultad de Medicina, Universidad de Chile, Santiago 8380492 (Chile); Facultad de Ciencias Quimicas y Farmaceuticas, Facultad de Medicina, Universidad de Chile, Santiago 8380492 (Chile); Instituto de Ciencias Biomedicas, Facultad de Medicina, Universidad de Chile, Santiago 8380492 (Chile)

2009-10-09

Myocyte enhancer factor 2C (MEF2C) plays an important role in cardiovascular development and is a key transcription factor for cardiac hypertrophy. Here, we describe MEF2C regulation by insulin-like growth factor-1 (IGF-1) and its role in IGF-1-induced cardiac hypertrophy. We found that IGF-1 addition to cultured rat cardiomyocytes activated MEF2C, as evidenced by its increased nuclear localization and DNA binding activity. IGF-1 stimulated MEF2 dependent-gene transcription in a time-dependent manner, as indicated by increased MEF2 promoter-driven reporter gene activity; IGF-1 also induced p38-MAPK phosphorylation, while an inhibitor of p38-MAPK decreased both effects. Additionally, inhibitors of phosphatidylinositol 3-kinase and calcineurin prevented IGF-1-induced MEF2 transcriptional activity. Via MEF2C-dependent signaling, IGF-1 also stimulated transcription of atrial natriuretic factor and skeletal {alpha}-actin but not of fos-lux reporter genes. These novel data suggest that MEF2C activation by IGF-1 mediates the pro-hypertrophic effects of IGF-1 on cardiac gene expression.

The transcription factor MEF2C mediates cardiomyocyte hypertrophy induced by IGF-1 signaling

International Nuclear Information System (INIS)

Munoz, Juan Pablo; Collao, Andres; Chiong, Mario; Maldonado, Carola; Adasme, Tatiana; Carrasco, Loreto; Ocaranza, Paula; Bravo, Roberto; Gonzalez, Leticia; Diaz-Araya, Guillermo; Hidalgo, Cecilia; Lavandero, Sergio

2009-01-01

Myocyte enhancer factor 2C (MEF2C) plays an important role in cardiovascular development and is a key transcription factor for cardiac hypertrophy. Here, we describe MEF2C regulation by insulin-like growth factor-1 (IGF-1) and its role in IGF-1-induced cardiac hypertrophy. We found that IGF-1 addition to cultured rat cardiomyocytes activated MEF2C, as evidenced by its increased nuclear localization and DNA binding activity. IGF-1 stimulated MEF2 dependent-gene transcription in a time-dependent manner, as indicated by increased MEF2 promoter-driven reporter gene activity; IGF-1 also induced p38-MAPK phosphorylation, while an inhibitor of p38-MAPK decreased both effects. Additionally, inhibitors of phosphatidylinositol 3-kinase and calcineurin prevented IGF-1-induced MEF2 transcriptional activity. Via MEF2C-dependent signaling, IGF-1 also stimulated transcription of atrial natriuretic factor and skeletal α-actin but not of fos-lux reporter genes. These novel data suggest that MEF2C activation by IGF-1 mediates the pro-hypertrophic effects of IGF-1 on cardiac gene expression.

PH Domain-Arf G Protein Interactions Localize the Arf-GEF Steppke for Cleavage Furrow Regulation in Drosophila.

Directory of Open Access Journals (Sweden)

Donghoon M Lee

Full Text Available The recruitment of GDP/GTP exchange factors (GEFs to specific subcellular sites dictates where they activate small G proteins for the regulation of various cellular processes. Cytohesins are a conserved family of plasma membrane GEFs for Arf small G proteins that regulate endocytosis. Analyses of mammalian cytohesins have identified a number of recruitment mechanisms for these multi-domain proteins, but the conservation and developmental roles for these mechanisms are unclear. Here, we report how the pleckstrin homology (PH domain of the Drosophila cytohesin Steppke affects its localization and activity at cleavage furrows of the early embryo. We found that the PH domain is necessary for Steppke furrow localization, and for it to regulate furrow structure. However, the PH domain was not sufficient for the localization. Next, we examined the role of conserved PH domain amino acid residues that are required for mammalian cytohesins to bind PIP3 or GTP-bound Arf G proteins. We confirmed that the Steppke PH domain preferentially binds PIP3 in vitro through a conserved mechanism. However, disruption of residues for PIP3 binding had no apparent effect on GFP-Steppke localization and effects. Rather, residues for binding to GTP-bound Arf G proteins made major contributions to this Steppke localization and activity. By analyzing GFP-tagged Arf and Arf-like small G proteins, we found that Arf1-GFP, Arf6-GFP and Arl4-GFP, but not Arf4-GFP, localized to furrows. However, analyses of embryos depleted of Arf1, Arf6 or Arl4 revealed either earlier defects than occur in embryos depleted of Steppke, or no detectable furrow defects, possibly because of redundancies, and thus it was difficult to assess how individual Arf small G proteins affect Steppke. Nonetheless, our data show that the Steppke PH domain and its conserved residues for binding to GTP-bound Arf G proteins have substantial effects on Steppke localization and activity in early Drosophila embryos.

ADP-ribosylation factor arf6p may function as a molecular switch of new end take off in fission yeast

International Nuclear Information System (INIS)

Fujita, Atsushi

2008-01-01

Small GTPases act as molecular switches in a wide variety of cellular processes. In fission yeast Schizosaccharomyces pombe, the directions of cell growth change from a monopolar manner to a bipolar manner, which is known as 'New End Take Off' (NETO). Here I report the identification of a gene, arf6 + , encoding an ADP-ribosylation factor small GTPase, that may be essential for NETO. arf6Δ cells completely fail to undergo NETO. arf6p localizes at both cell ends and presumptive septa in a cell-cycle dependent manner. And its polarized localization is not dependent on microtubules, actin cytoskeletons and some NETO factors (bud6p, for3p, tea1p, tea3p, and tea4p). Notably, overexpression of a fast GDP/GTP-cycling mutant of arf6p can advance the timing of NETO. These findings suggest that arf6p functions as a molecular switch for the activation of NETO in fission yeast

Expression profiling of AUXIN RESPONSE FACTOR genes during somatic embryogenesis induction in Arabidopsis.

Science.gov (United States)

Wójcikowska, Barbara; Gaj, Małgorzata D

2017-06-01

Extensive modulation of numerous ARF transcripts in the embryogenic culture of Arabidopsis indicates a substantial role of auxin signaling in the mechanism of somatic embryogenesis induction. Somatic embryogenesis (SE) is induced by auxin in plants and auxin signaling is considered to play a key role in the molecular mechanism that controls the embryogenic transition of plant somatic cells. Accordingly, the expression of AUXIN RESPONSE FACTOR (ARF) genes in embryogenic culture of Arabidopsis was analyzed. The study revealed that 14 of the 22 ARFs were transcribed during SE in Arabidopsis. RT-qPCR analysis indicated that the expression of six ARFs (ARF5, ARF6, ARF8, ARF10, ARF16, and ARF17) was significantly up-regulated, whereas five other genes (ARF1, ARF2, ARF3, ARF11, and ARF18) were substantially down-regulated in the SE-induced explants. The activity of ARFs during SE was also monitored with GFP reporter lines and the ARFs that were expressed in areas of the explants engaged in SE induction were detected. A functional test of ARFs transcribed during SE was performed and the embryogenic potential of the arf mutants and overexpressor lines was evaluated. ARFs with a significantly modulated expression during SE coupled with an impaired embryogenic response of the relevant mutant and/or overexpressor line, including ARF1, ARF2, ARF3, ARF5, ARF6, ARF8, and ARF11 were indicated as possibly being involved in SE induction. The study provides evidence that embryogenic induction strongly depends on ARFs, which are key regulators of the auxin signaling. Some clues on the possible functions of the candidate ARFs, especially ARF5, in the mechanism of embryogenic transition are discussed. The results provide guidelines for further research on the auxin-related functional genomics of SE and the developmental plasticity of somatic cells.

The DNA damage checkpoint precedes activation of ARF in response to escalating oncogenic stress during tumorigenesis

DEFF Research Database (Denmark)

Evangelou, K.; Bartkova, J.; Kotsinas, A.

2013-01-01

oncogenes showed that the delayed upregulation of ARF reflected a requirement for a higher, transcriptionally based threshold of oncogenic stress, elicited by at least two oncogenic 'hits', compared with lower activation threshold for DDR. We propose that relative to DDR activation, ARF provides...

Vesicular transport protein Arf6 modulates cytoskeleton dynamics for polar body extrusion in mouse oocyte meiosis.

Science.gov (United States)

Duan, Xing; Zhang, Hao-Lin; Pan, Meng-Hao; Zhang, Yu; Sun, Shao-Chen

2018-02-01

Arf6 (ADP-ribosylation factor 6) is known to play important roles in membrane dynamics through the regulation of actin filament reorganization for multiple cellular processes such as cytokinesis, phagocytosis, cell migration and tumor cell invasion. However, the functions of Arf6 in mammalian oocyte meiosis have not been clarified. In present study we showed that Arf6 expressed in mouse oocytes and was mainly distributed around the spindle during meiosis. Depletion of Arf6 by morpholino microinjection caused oocytes failing to extrude first polar body. Further analysis indicated that Arf6 knock down caused the aberrant actin distribution, which further induced the failure of meiotic spindle movement. And the loss of oocyte polarity also confirmed this. The regulation of Arf6 on actin filaments in mouse oocytes might be due to its effects on the phosphorylation level of cofilin and the expression of Arp2/3 complex. Moreover, we found that the decrease of Arf6 caused the disruption of spindle formation, indicating the multiple roles of Arf6 on cytoskeleton dynamics in meiosis. In summary, our results indicated that Arf6 was involved in mouse oocyte meiosis through its functional roles in actin-mediated spindle movement and spindle organization. Copyright © 2017 Elsevier B.V. All rights reserved.

Role of the Slug Transcription Factor in Chemically-Induced Skin Cancer

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Kristine von Maltzan

2016-02-01

Full Text Available The Slug transcription factor plays an important role in ultraviolet radiation (UVR-induced skin carcinogenesis, particularly in the epithelial-mesenchymal transition (EMT occurring during tumor progression. In the present studies, we investigated the role of Slug in two-stage chemical skin carcinogenesis. Slug and the related transcription factor Snail were expressed at high levels in skin tumors induced by 7,12-dimethylbenz[α]anthracene application followed by 12-O-tetradecanoylphorbol-13-acetate (TPA treatment. TPA-induced transient elevation of Slug and Snail proteins in normal mouse epidermis and studies in Slug transgenic mice indicated that Slug modulates TPA-induced epidermal hyperplasia and cutaneous inflammation. Although Snail family factors have been linked to inflammation via interactions with the cyclooxygenase-2 (COX-2 pathway, a pathway that also plays an important role in skin carcinogenesis, transient TPA induction of Slug and Snail appeared unrelated to COX-2 expression. In cultured human keratinocytes, TPA induced Snail mRNA expression while suppressing Slug expression, and this differential regulation was due specifically to activation of the TPA receptor. These studies show that Slug and Snail exhibit similar patterns of expression during both UVR and chemical skin carcinogenesis, that Slug and Snail can be differentially regulated under some conditions and that in vitro findings may not recapitulate in vivo results.

Hypoxia-Inducible Factor 3 Is an Oxygen-Dependent Transcription Activator and Regulates a Distinct Transcriptional Response to Hypoxia

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Peng Zhang

2014-03-01

Full Text Available Hypoxia-inducible factors (HIFs play key roles in the cellular response to hypoxia. It is widely accepted that whereas HIF-1 and HIF-2 function as transcriptional activators, HIF-3 inhibits HIF-1/2α action. Contrary to this idea, we show that zebrafish Hif-3α has strong transactivation activity. Hif-3α is degraded under normoxia. Mutation of P393, P493, and L503 inhibits this oxygen-dependent degradation. Transcriptomics and chromatin immunoprecipitation analyses identify genes that are regulated by Hif-3α, Hif-1α, or both. Under hypoxia or when overexpressed, Hif-3α binds to its target gene promoters and upregulates their expression. Dominant-negative inhibition and knockdown of Hif-3α abolish hypoxia-induced Hif-3α-promoter binding and gene expression. Hif-3α not only mediates hypoxia-induced growth and developmental retardation but also possesses hypoxia-independent activities. Importantly, transactivation activity is conserved and human HIF-3α upregulates similar genes in human cells. These findings suggest that Hif-3 is an oxygen-dependent transcription factor and activates a distinct transcriptional response to hypoxia.

Heat shock transcription factors regulate heat induced cell death in a ...

Indian Academy of Sciences (India)

2007-03-29

Mar 29, 2007 ... Heat shock transcription factors regulate heat induced cell death in a rat ... the synthesis of heat shock proteins (Hsps) which is strictly regulated by ... The lack of Hsp synthesis in these cells was due to a failure in HSF1 DNA ...

Regulation of endogenous human gene expression by ligand-inducible TALE transcription factors.

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Mercer, Andrew C; Gaj, Thomas; Sirk, Shannon J; Lamb, Brian M; Barbas, Carlos F

2014-10-17

The construction of increasingly sophisticated synthetic biological circuits is dependent on the development of extensible tools capable of providing specific control of gene expression in eukaryotic cells. Here, we describe a new class of synthetic transcription factors that activate gene expression in response to extracellular chemical stimuli. These inducible activators consist of customizable transcription activator-like effector (TALE) proteins combined with steroid hormone receptor ligand-binding domains. We demonstrate that these ligand-responsive TALE transcription factors allow for tunable and conditional control of gene activation and can be used to regulate the expression of endogenous genes in human cells. Since TALEs can be designed to recognize any contiguous DNA sequence, the conditional gene regulatory system described herein will enable the design of advanced synthetic gene networks.

Comprehensive analysis of the renal transcriptional response to acute uranyl nitrate exposure

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Argiles Angel

2006-01-01

Full Text Available Abstract Background Chemical and radiological toxicities related to uranium acute exposure have been widely studied in nuclear fuel workers and military personnel. It is well known that uranyl nitrate induces acute renal failure (ARF. However, the mechanisms of this metal-induced injury are not well defined at the molecular level. Results Renal function and histology were assessed in mice receiving uranyl nitrate (UN(+ and controls (UN(-. To identify the genomic response to uranium exposure, serial analysis gene expression (SAGE of the kidney was performed in both groups. Over 43,000 mRNA SAGE tags were sequenced. A selection of the differentially expressed transcripts was confirmed by real-time quantitative PCR and Western blotting. UN(+ animals developed renal failure and displayed the characteristic histological lesions of UN nephropathy. Of the >14,500 unique tags identified in both libraries, 224 had a modified expression level; they are known to participate in inflammation, ion transport, signal transduction, oxidative stress, apoptosis, metabolism, and catabolism. Several genes that were identified had not previously been evaluated within the context of toxic ARF such as translationally controlled tumor protein, insulin like growth factor binding protein 7 and ribosomal protein S29, all apoptosis related genes. Conclusion We report a comprehensive description of the UN induced modifications in gene expression levels, including the identification of genes previously unrelated to ARF. The study of these genes and the metabolisms they control should improve our understanding of toxic ARF and enlighten on the molecular targets for potential therapeutic interventions.

Bypass of cell cycle arrest induced by transient DNMT1 post-transcriptional silencing triggers aneuploidy in human cells

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Barra Viviana

2012-02-01

Full Text Available Abstract Background Aneuploidy has been acknowledged as a major source of genomic instability in cancer, and it is often considered the result of chromosome segregation errors including those caused by defects in genes controlling the mitotic spindle assembly, centrosome duplication and cell-cycle checkpoints. Aneuploidy and chromosomal instability has been also correlated with epigenetic alteration, however the molecular basis of this correlation is poorly understood. Results To address the functional connection existing between epigenetic changes and aneuploidy, we used RNA-interference to silence the DNMT1 gene, encoding for a highly conserved member of the DNA methyl-transferases. DNMT1 depletion slowed down proliferation of near-diploid human tumor cells (HCT116 and triggered G1 arrest in primary human fibroblasts (IMR90, by inducing p53 stabilization and, in turn, p21waf1 transactivation. Remarkably, p53 increase was not caused by DNA damage and was not observed after p14-ARF post-transcriptional silencing. Interestingly, DNMT1 silenced cells with p53 or p14-ARF depleted did not arrest in G1 but, instead, underwent DNA hypomethylation and became aneuploid. Conclusion Our results suggest that DNMT1 depletion triggers a p14ARF/p53 dependent cell cycle arrest to counteract the aneuploidy induced by changes in DNA methylation.

The Arf GTPase-activating protein family is exploited by Salmonella enterica serovar Typhimurium to invade nonphagocytic host cells.

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Davidson, Anthony C; Humphreys, Daniel; Brooks, Andrew B E; Hume, Peter J; Koronakis, Vassilis

2015-02-10

To establish intracellular infections, Salmonella bacteria trigger host cell membrane ruffling and invasion by subverting cellular Arf guanine nucleotide exchange factors (GEFs) that activate Arf1 and Arf6 GTPases by promoting GTP binding. A family of cellular Arf GTPase-activating proteins (GAPs) can downregulate Arf signaling by stimulating GTP hydrolysis, but whether they do this during infection is unknown. Here, we uncovered a remarkable role for distinct Arf GAP family members in Salmonella invasion. The Arf6 GAPs ACAP1 and ADAP1 and the Arf1 GAP ASAP1 localized at Salmonella-induced ruffles, which was not the case for the plasma membrane-localized Arf6 GAPs ARAP3 and GIT1 or the Golgi-associated Arf1 GAP1. Surprisingly, we found that loss of ACAP1, ADAP1, or ASAP1 impaired Salmonella invasion, revealing that GAPs cannot be considered mere terminators of cytoskeleton remodeling. Salmonella invasion was restored in Arf GAP-depleted cells by expressing fast-cycling Arf derivatives, demonstrating that Arf GTP/GDP cycles facilitate Salmonella invasion. Consistent with this view, both constitutively active and dominant-negative Arf derivatives that cannot undergo GTP/GDP cycles inhibited invasion. Furthermore, we demonstrated that Arf GEFs and GAPs colocalize at invading Salmonella and collaborate to drive Arf1-dependent pathogen invasion. This study revealed that Salmonella bacteria exploit a remarkable interplay between Arf GEFs and GAPs to direct cycles of Arf GTPase activation and inactivation. These cycles drive Salmonella cytoskeleton remodeling and enable intracellular infections. To initiate infections, the Salmonella bacterial pathogen remodels the mammalian actin cytoskeleton and invades host cells by subverting host Arf GEFs that activate Arf1 and Arf6 GTPases. Cellular Arf GAPs deactivate Arf GTPases and negatively regulate cell processes, but whether they target Arfs during infection is unknown. Here, we uncovered an important role for the Arf GAP

Key roles of Arf small G proteins and biosynthetic trafficking for animal development.

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Rodrigues, Francisco F; Harris, Tony J C

2017-04-14

Although biosynthetic trafficking can function constitutively, it also functions specifically for certain developmental processes. These processes require either a large increase to biosynthesis or the biosynthesis and targeted trafficking of specific players. We review the conserved molecular mechanisms that direct biosynthetic trafficking, and discuss how their genetic disruption affects animal development. Specifically, we consider Arf small G proteins, such as Arf1 and Sar1, and their coat effectors, COPI and COPII, and how these proteins promote biosynthetic trafficking for cleavage of the Drosophila embryo, the growth of neuronal dendrites and synapses, extracellular matrix secretion for bone development, lumen development in epithelial tubes, notochord and neural tube development, and ciliogenesis. Specific need for the biosynthetic trafficking system is also evident from conserved CrebA/Creb3-like transcription factors increasing the expression of secretory machinery during several of these developmental processes. Moreover, dysfunctional trafficking leads to a range of developmental syndromes.

ArF Laser -Induced Chemical Vapour Deposition of Polythiene Films from Carbon Disulfide

Czech Academy of Sciences Publication Activity Database

Tomovska, R.; Bastl, Zdeněk; Vorlíček, Vladimír; Vacek, Karel; Šubrt, Jan; Plzák, Zbyněk; Pola, Josef

2003-01-01

Roč. 107, č. 36 (2003), s. 9793-9801 ISSN 1089-5647 R&D Projects: GA MŠk ME 612 Institutional research plan: CEZ:AV0Z4032918; CEZ:AV0Z4040901 Keywords : laser photolysis * ArF * chemical vapour deposition Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.679, year: 2003

ARF1 recruits RAC1 to leading edge in neutrophil chemotaxis.

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Mazaki, Yuichi; Onodera, Yasuhito; Higashi, Tsunehito; Horinouchi, Takahiro; Oikawa, Tsukasa; Sabe, Hisataka

2017-10-02

The small GTPase ARF1 mediates membrane trafficking mostly from the Golgi, and is essential for the G protein-coupled receptor (GPCR)-mediated chemotaxis of neutrophils. In this process, ARF1 is activated by the guanine nucleotide exchanger GBF1, and is inactivated by the GTPase-activating protein GIT2. Neutrophils generate the Gβγ-PAK1-αPIX-GIT2 linear complex during GPCR-induced chemotaxis, in which αPIX activates RAC1/CDC42, which then employs PAK1. However, it has remained unclear as to why GIT2 is included in this complex. We investigated the association between ARF1 and RAC1/CDC42 during the fMLP-stimulated chemotaxis of HL60 cells. We found that the silencing of GBF1 significantly impaired the recruitment of RAC1 to the leading edges, but not PAK1, αPIX, RAC2, or CDC42. A significant population of RAC1 colocalized with ARF1 at the leading edges in stimulated cells, whereas fMLP activated both ARF1 and ARF5. Consistently, the silencing of ARF1, but not ARF5, impaired the recruitment of RAC1, whereas the silencing of RAC1 did not affect the recruitment of ARF1 to the leading edges. Our results indicated that the activation of ARF1 triggers the plasma membrane recruitment of RAC1 in GPCR-mediated chemotaxis, which is essential for cortical actin remodeling. Thus, membrane remodeling at the leading edges appears to precede actin remodeling in chemotaxis. Together with the fact that GIT2, which inactivates ARF1, is an integral component of the machinery activating RAC1, we proposed a model in which the ARF1-RAC1 linkage enables the regulation of ARF1 by repetitive on/off cycles during GPCR-mediated neutrophil chemotaxis.

Radiation activation of transcription factors in mammalian cells

International Nuclear Information System (INIS)

Kraemer, M.; Stein, B.; Mai, S.; Kunz, E.; Koenig, H.; Ponta, H.; Herrlich, P.; Rahmsdorf, H.J.; Loferer, H.; Grunicke, H.H.

1990-01-01

In mammalian cells radiation induces the enhanced transcription of several genes. The cis acting elements in the control region of inducible genes have been delimited by site directed mutagenesis. Several different elements have been found in different genes. They do not only activate gene transcription in response to radiation but also in response to growth factors and to tumor promoter phorbol esters. The transcription factors binding to these elements are present also in non-irradiated cells, but their DNA binding activity and their transactivating capability is increased upon irradiation. The signal chain linking the primary radiation induced signal (damaged DNA) to the activation of transcription factors involves the action of (a) protein kinase(s). (orig.)

Induced Genome-Wide Binding of Three Arabidopsis WRKY Transcription Factors during Early MAMP-Triggered Immunity.

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Birkenbihl, Rainer P; Kracher, Barbara; Somssich, Imre E

2017-01-01

During microbial-associated molecular pattern-triggered immunity (MTI), molecules derived from microbes are perceived by cell surface receptors and upon signaling to the nucleus initiate a massive transcriptional reprogramming critical to mount an appropriate host defense response. WRKY transcription factors play an important role in regulating these transcriptional processes. Here, we determined on a genome-wide scale the flg22-induced in vivo DNA binding dynamics of three of the most prominent WRKY factors, WRKY18, WRKY40, and WRKY33. The three WRKY factors each bound to more than 1000 gene loci predominantly at W-box elements, the known WRKY binding motif. Binding occurred mainly in the 500-bp promoter regions of these genes. Many of the targeted genes are involved in signal perception and transduction not only during MTI but also upon damage-associated molecular pattern-triggered immunity, providing a mechanistic link between these functionally interconnected basal defense pathways. Among the additional targets were genes involved in the production of indolic secondary metabolites and in modulating distinct plant hormone pathways. Importantly, among the targeted genes were numerous transcription factors, encoding predominantly ethylene response factors, active during early MTI, and WRKY factors, supporting the previously hypothesized existence of a WRKY subregulatory network. Transcriptional analysis revealed that WRKY18 and WRKY40 function redundantly as negative regulators of flg22-induced genes often to prevent exaggerated defense responses. © 2016 American Society of Plant Biologists. All rights reserved.

Selective activation of human heat shock gene transcription by nitrosourea antitumor drugs mediated by isocyanate-induced damage and activation of heat shock transcription factor.

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Kroes, R A; Abravaya, K; Seidenfeld, J; Morimoto, R I

1991-01-01

Treatment of cultured human tumor cells with the chloroethylnitrosourea antitumor drug 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) selectively induces transcription and protein synthesis of a subset of the human heat shock or stress-induced genes (HSP90 and HSP70) with little effect on other stress genes or on expression of the c-fos, c-myc, or beta-actin genes. The active component of BCNU and related compounds appears to be the isocyanate moiety that causes carbamoylation of proteins and nucleic acids. Transcriptional activation of the human HSP70 gene by BCNU is dependent on the heat shock element and correlates with the level of heat shock transcription factor and its binding to the heat shock element in vivo. Unlike activation by heat or heavy metals, BCNU-mediated activation is strongly dependent upon new protein synthesis. This suggests that BCNU-induced, isocyanate-mediated damage to newly synthesized protein(s) may be responsible for activation of the heat shock transcription factor and increased transcription of the HSP90 and HSP70 genes. Images PMID:2052560

Hypoxic preconditioning protects photoreceptors against light damage independently of hypoxia inducible transcription factors in rods.

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Kast, Brigitte; Schori, Christian; Grimm, Christian

2016-05-01

Hypoxic preconditioning protects photoreceptors against light-induced degeneration preserving retinal morphology and function. Although hypoxia inducible transcription factors 1 and 2 (HIF1, HIF2) are the main regulators of the hypoxic response, photoreceptor protection does not depend on HIF1 in rods. Here we used rod-specific Hif2a single and Hif1a;Hif2a double knockout mice to investigate the potential involvement of HIF2 in rods for protection after hypoxic preconditioning. To identify potential HIF2 target genes in rods we determined the retinal transcriptome of hypoxic control and rod-specific Hif2a knockouts by RNA sequencing. We show that rods do not need HIF2 for hypoxia-induced increased survival after light exposure. The transcriptomic analysis revealed a number of genes that are potentially regulated by HIF2 in rods; among those were Htra1, Timp3 and Hmox1, candidates that are interesting due to their connection to human degenerative diseases of the retina. We conclude that neither HIF1 nor HIF2 are required in photoreceptors for protection by hypoxic preconditioning. We hypothesize that HIF transcription factors may be needed in other cells to produce protective factors acting in a paracrine fashion on photoreceptor cells. Alternatively, hypoxic preconditioning induces a rod-intrinsic response that is independent of HIF transcription factors. Copyright © 2015 Elsevier Ltd. All rights reserved.

P53- and mevalonate pathway–driven malignancies require Arf6 for metastasis and drug resistance

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Hashimoto, Ari; Oikawa, Tsukasa; Hashimoto, Shigeru; Sugino, Hirokazu; Yoshikawa, Ayumu; Otsuka, Yutaro; Handa, Haruka; Onodera, Yasuhito; Nam, Jin-Min; Oneyama, Chitose; Okada, Masato; Fukuda, Mitsunori

2016-01-01

Drug resistance, metastasis, and a mesenchymal transcriptional program are central features of aggressive breast tumors. The GTPase Arf6, often overexpressed in tumors, is critical to promote epithelial–mesenchymal transition and invasiveness. The metabolic mevalonate pathway (MVP) is associated with tumor invasiveness and known to prenylate proteins, but which prenylated proteins are critical for MVP-driven cancers is unknown. We show here that MVP requires the Arf6-dependent mesenchymal program. The MVP enzyme geranylgeranyl transferase II (GGT-II) and its substrate Rab11b are critical for Arf6 trafficking to the plasma membrane, where it is activated by receptor tyrosine kinases. Consistently, mutant p53, which is known to support tumorigenesis via MVP, promotes Arf6 activation via GGT-II and Rab11b. Inhibition of MVP and GGT-II blocked invasion and metastasis and reduced cancer cell resistance against chemotherapy agents, but only in cells overexpressing Arf6 and components of the mesenchymal program. Overexpression of Arf6 and mesenchymal proteins as well as enhanced MVP activity correlated with poor patient survival. These results provide insights into the molecular basis of MVP-driven malignancy. PMID:27044891

Resveratrol induces growth arrest and apoptosis through activation of FOXO transcription factors in prostate cancer cells.

Directory of Open Access Journals (Sweden)

Qinghe Chen

2010-12-01

Full Text Available Resveratrol, a naturally occurring phytopolyphenol compound, has attracted extensive interest in recent years because of its diverse pharmacological characteristics. Although resveratrol possesses chemopreventive properties against several cancers, the molecular mechanisms by which it inhibits cell growth and induces apoptosis have not been clearly understood. The present study was carried out to examine whether PI3K/AKT/FOXO pathway mediates the biological effects of resveratrol.Resveratrol inhibited the phosphorylation of PI3K, AKT and mTOR. Resveratrol, PI3K inhibitors (LY294002 and Wortmannin and AKT inhibitor alone slightly induced apoptosis in LNCaP cells. These inhibitors further enhanced the apoptosis-inducing potential of resveratrol. Overexpression of wild-type PTEN slightly induced apoptosis. Wild type PTEN and PTEN-G129E enhanced resveratrol-induced apoptosis, whereas PTEN-G129R had no effect on proapoptotic effects of resveratrol. Furthermore, apoptosis-inducing potential of resveratrol was enhanced by dominant negative AKT, and inhibited by wild-type AKT and constitutively active AKT. Resveratrol has no effect on the expression of FKHR, FKHRL1 and AFX genes. The inhibition of FOXO phosphorylation by resveratrol resulted in its nuclear translocation, DNA binding and transcriptional activity. The inhibition of PI3K/AKT pathway induced FOXO transcriptional activity resulting in induction of Bim, TRAIL, p27/KIP1, DR4 and DR5, and inhibition of cyclin D1. Similarly, resveratrol-induced FOXO transcriptional activity was further enhanced when activation of PI3K/AKT pathway was blocked. Over-expression of phosphorylation deficient mutants of FOXO proteins (FOXO1-TM, FOXO3A-TM and FOXO4-TM induced FOXO transcriptional activity, which was further enhanced by resveratrol. Inhibition of FOXO transcription factors by shRNA blocked resveratrol-induced upregulation of Bim, TRAIL, DR4, DR5, p27/KIP1 and apoptosis, and inhibition of cyclin D1 by

NUR TRANSCRIPTION FACTORS IN STRESS AND ADDICTION

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Danae eCampos-Melo

2013-12-01

Full Text Available The Nur transcription factors Nur77 (NGFI-B, NR4A1, Nurr1 (NR4A2 and Nor-1 (NR4A3 are a sub-family of orphan members of the nuclear receptor superfamily. These transcription factors are products of immediate early genes, whose expression is rapidly and transiently induced in the central nervous system by several types of stimuli. Nur factors are present throughout the hypothalamus-pituitary-adrenal axis where are prominently induced in response to stress. Drugs of abuse and stress also induce the expression of Nur factors in nuclei of the motivation/reward circuit of the brain, indicating their participation in the process of drug addiction and in non-hypothalamic responses to stress. Repeated use of addictive drugs and chronic stress induce long-lasting dysregulation of the brain motivation/reward circuit, due to reprogramming of gene expression and enduring alterations in neuronal function. Here, we review the data supporting that Nur transcription factors are key players in the molecular basis of the dysregulation of neuronal circuits involved in chronic stress and addiction.

Identification and Expression Profiling of the Auxin Response Factors in Dendrobium officinale under Abiotic Stresses.

Science.gov (United States)

Chen, Zhehao; Yuan, Ye; Fu, Di; Shen, Chenjia; Yang, Yanjun

2017-05-04

Auxin response factor (ARF) proteins play roles in plant responses to diverse environmental stresses by binding specifically to the auxin response element in the promoters of target genes. Using our latest public Dendrobium transcriptomes, a comprehensive characterization and analysis of 14 DnARF genes were performed. Three selected DnARFs , including DnARF1 , DnARF4 , and DnARF6 , were confirmed to be nuclear proteins according to their transient expression in epidermal cells of Nicotiana benthamiana leaves. Furthermore, the transcription activation abilities of DnARF1 , DnARF4 , and DnARF6 were tested in a yeast system. Our data showed that DnARF6 is a transcriptional activator in Dendrobium officinale . To uncover the basic information of DnARF gene responses to abiotic stresses, we analyzed their expression patterns under various hormones and abiotic treatments. Based on our data, several hormones and significant stress responsive DnARF genes have been identified. Since auxin and ARF genes have been identified in many plant species, our data is imperative to reveal the function of ARF mediated auxin signaling in the adaptation to the challenging Dendrobium environment.

The human ARF tumor suppressor senses blastema activity and suppresses epimorphic tissue regeneration

Science.gov (United States)

Hesse, Robert G; Kouklis, Gayle K; Ahituv, Nadav; Pomerantz, Jason H

2015-01-01

The control of proliferation and differentiation by tumor suppressor genes suggests that evolution of divergent tumor suppressor repertoires could influence species’ regenerative capacity. To directly test that premise, we humanized the zebrafish p53 pathway by introducing regulatory and coding sequences of the human tumor suppressor ARF into the zebrafish genome. ARF was dormant during development, in uninjured adult fins, and during wound healing, but was highly expressed in the blastema during epimorphic fin regeneration after amputation. Regenerative, but not developmental signals resulted in binding of zebrafish E2f to the human ARF promoter and activated conserved ARF-dependent Tp53 functions. The context-dependent activation of ARF did not affect growth and development but inhibited regeneration, an unexpected distinct tumor suppressor response to regenerative versus developmental environments. The antagonistic pleiotropic characteristics of ARF as both tumor and regeneration suppressor imply that inducing epimorphic regeneration clinically would require modulation of ARF –p53 axis activation. DOI: http://dx.doi.org/10.7554/eLife.07702.001 PMID:26575287

A novel proapoptotic gene PANO encodes a post-translational modulator of the tumor suppressor p14ARF

Energy Technology Data Exchange (ETDEWEB)

Watari, Akihiro; Li, Yang; Higashiyama, Shinji; Yutsudo, Masuo, E-mail: yutsudo@biken.osaka-u.ac.jp

2012-02-01

The protein p14ARF is a known tumor suppressor protein controlling cell proliferation and survival, which mainly localizes in nucleoli. However, the regulatory mechanisms that govern its activity or expression remain unclear. Here, we report that a novel proapoptotic nucleolar protein, PANO, modulates the expression and activity of p14ARF in HeLa cells. Overexpression of PANO enhances the stability of p14ARF protein by protecting it from degradation, resulting in an increase in p14ARF expression levels. Overexpression of PANO also induces apoptosis under low serum conditions. This effect is dependent on the nucleolar localization of PANO and inhibited by knocking-down p14ARF. Alternatively, PANO siRNA treated cells exhibit a reduction in p14ARF protein levels. In addition, ectopic expression of PANO suppresses the tumorigenicity of HeLa cells in nude mice. These results indicate that PANO is a new apoptosis-inducing gene by modulating the tumor suppressor protein, p14ARF, and may itself be a new candidate tumor suppressor gene.

Estrogen-induced transcription factor EGR1 regulates c-Kit transcription in the mouse uterus to maintain uterine receptivity for embryo implantation.

Science.gov (United States)

Park, Mira; Kim, Hye-Ryun; Kim, Yeon Sun; Yang, Seung Chel; Yoon, Jung Ah; Lyu, Sang Woo; Lim, Hyunjung Jade; Hong, Seok-Ho; Song, Haengseok

2018-07-15

Early growth response 1 (Egr1) is a key transcription factor that mediates the action of estrogen (E 2 ) to establish uterine receptivity for embryo implantation. However, few direct target genes of EGR1 have been identified in the uterus. Here, we demonstrated that E 2 induced EGR1-regulated transcription of c-Kit, which plays a crucial role in cell fate decisions. Spatiotemporal expression of c-Kit followed that of EGR1 in uteri of ovariectomized mice at various time points after E 2 treatment. E 2 activated ERK1/2 and p38 to induce EGR1, which then activated c-Kit expression in the uterus. EGR1 transfection produced rapid and transient induction of c-KIT in a time- and dose-dependent manner. Furthermore, luciferase assays to measure c-Kit promoter activity confirmed that a functional EGR1 binding site(s) (EBS) was located within -1 kb of the c-Kit promoter. Site-directed mutagenesis and chromatin immunoprecipitation-PCR for three putative EBS within -1 kb demonstrated that the EBS at -818/-805 was critical for EGR1-dependent c-Kit transcription. c-Kit expression was significantly increased in the uterus on day 4 and administration of Masitinib, a c-Kit inhibitor, effectively interfered with embryo implantation. Collectively, our results showed that estrogen induces transcription factor EGR1 to regulate c-Kit transcription for uterine receptivity for embryo implantation in the mouse uterus. Copyright © 2017 Elsevier B.V. All rights reserved.

The structural basis of Arf effector specificity: the crystal structure of ARF6 in a complex with JIP4.

Science.gov (United States)

Isabet, Tatiana; Montagnac, Guillaume; Regazzoni, Karine; Raynal, Bertrand; El Khadali, Fatima; England, Patrick; Franco, Michel; Chavrier, Philippe; Houdusse, Anne; Ménétrey, Julie

2009-09-16

The JNK-interacting proteins, JIP3 and JIP4, are specific effectors of the small GTP-binding protein ARF6. The interaction of ARF6-GTP with the second leucine zipper (LZII) domains of JIP3/JIP4 regulates the binding of JIPs to kinesin-1 and dynactin. Here, we report the crystal structure of ARF6-GTP bound to the JIP4-LZII at 1.9 A resolution. The complex is a heterotetramer with dyad symmetry arranged in an ARF6-(JIP4)(2)-ARF6 configuration. Comparison of the ARF6-JIP4 interface with the equivalent region of ARF1 shows the structural basis of JIP4's specificity for ARF6. Using site-directed mutagenesis and surface plasmon resonance, we further show that non-conserved residues at the switch region borders are the key structural determinants of JIP4 specificity. A structure-derived model of the association of the ARF6-JIP3/JIP4 complex with membranes shows that the JIP4-LZII coiled-coil should lie along the membrane to prevent steric hindrances, resulting in only one ARF6 molecule bound. Such a heterotrimeric complex gives insights to better understand the ARF6-mediated motor switch regulatory function.

Role of Arf GTPases in fungal morphogenesis and virulence.

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Hayet Labbaoui

2017-02-01

Full Text Available Virulence of the human fungal pathogen Candida albicans depends on the switch from budding to filamentous growth, which requires sustained membrane traffic and polarized growth. In many organisms, small GTPases of the Arf (ADP-ribosylation factor family regulate membrane/protein trafficking, yet little is known about their role in fungal filamentous growth. To investigate these GTPases in C. albicans, we generated loss of function mutants in all 3 Arf proteins, Arf1-Arf3, and 2 Arf-like proteins, Arl1 and Arl3. Our results indicate that of these proteins, Arf2 is required for viability and sensitivity to antifungal drugs. Repressible ARF2 expression results in defects in filamentous growth, cell wall integrity and virulence, likely due to alteration of the Golgi. Arl1 is also required for invasive filamentous growth and, although arl1/arl1 cells can initiate hyphal growth, hyphae are substantially shorter than that of the wild-type, due to the inability of this mutant to maintain hyphal growth at a single site. We show that this defect does not result from an alteration of phospholipid distribution and is unlikely to result from the sole Golgin Imh1 mislocalization, as Imh1 is not required for invasive filamentous growth. Rather, our results suggest that the arl1/arl1 hyphal growth defect results from increased secretion in this mutant. Strikingly, the arl1/arl1 mutant is drastically reduced in virulence during oropharyngeal candidiasis. Together, our results highlight the importance of Arl1 and Arf2 as key regulators of hyphal growth and virulence in C. albicans and identify a unique function of Arl1 in secretion.

Transcription Factors Encoded on Core and Accessory Chromosomes of Fusarium oxysporum Induce Expression of Effector Genes

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van der Does, H. Charlotte; Schmidt, Sarah M.; Langereis, Léon; Hughes, Timothy R.

2016-01-01

Proteins secreted by pathogens during host colonization largely determine the outcome of pathogen-host interactions and are commonly called ‘effectors’. In fungal plant pathogens, coordinated transcriptional up-regulation of effector genes is a key feature of pathogenesis and effectors are often encoded in genomic regions with distinct repeat content, histone code and rate of evolution. In the tomato pathogen Fusarium oxysporum f. sp. lycopersici (Fol), effector genes reside on one of four accessory chromosomes, known as the ‘pathogenicity’ chromosome, which can be exchanged between strains through horizontal transfer. The three other accessory chromosomes in the Fol reference strain may also be important for virulence towards tomato. Expression of effector genes in Fol is highly up-regulated upon infection and requires Sge1, a transcription factor encoded on the core genome. Interestingly, the pathogenicity chromosome itself contains 13 predicted transcription factor genes and for all except one, there is a homolog on the core genome. We determined DNA binding specificity for nine transcription factors using oligonucleotide arrays. The binding sites for homologous transcription factors were highly similar, suggesting that extensive neofunctionalization of DNA binding specificity has not occurred. Several DNA binding sites are enriched on accessory chromosomes, and expression of FTF1, its core homolog FTF2 and SGE1 from a constitutive promoter can induce expression of effector genes. The DNA binding sites of only these three transcription factors are enriched among genes up-regulated during infection. We further show that Ftf1, Ftf2 and Sge1 can activate transcription from their binding sites in yeast. RNAseq analysis revealed that in strains with constitutive expression of FTF1, FTF2 or SGE1, expression of a similar set of plant-responsive genes on the pathogenicity chromosome is induced, including most effector genes. We conclude that the Fol

The AP2/EREBP gene PUCHI Co-Acts with LBD16/ASL18 and LBD18/ASL20 downstream of ARF7 and ARF19 to regulate lateral root development in Arabidopsis.

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Kang, Na Young; Lee, Han Woo; Kim, Jungmook

2013-08-01

The developmental process of lateral root formation consists of priming, initiation, primordium development and the emergence of lateral roots from the primary root. Molecular genetic studies with Arabidopsis have revealed several key transcriptional regulators involved in lateral root development. However, their functional interaction has not been fully characterized yet. Here we utilized a genetic approach to understand some of these interactions, revealing that PUCHI functioning in morphogenesis of early lateral root primordium is regulated downstream of ARF7/ARF19 and acts with LBD16(ASL18)/LBD18(ASL20) to regulate lateral root development. We showed that auxin-responsive expression of PUCHI was significantly reduced in arf7 or arf19 single mutants and completely abolished in arf7 arf19 double mutants. Consistent with this, β-glucuronidase (GUS) expression under the PUCHI promoter in arf7 arf19 was greatly reduced in the lateral root primordium compared with that in the wild type and did not respond to exogenous auxin. Results of GUS expression analyses under the PUCHI, LBD16 or LBD18 promoter in lbd16, lbd18 single and double mutants or puchi demonstrated that PUCHI and LBD16 or LBD18 do not regulate each other's expression. Lateral root phenotypes of double and triple mutants of lbd16, lbd18 and puchi showed that the puchi mutation in lbd16 and lbd18 mutants synergistically decreased the number of emerged lateral roots. These analyses also showed that puchi affected lateral root primordium development of lbd16 or lbd18 additively but differentially. Taken together, these results suggest that PUCHI co-acts with LBD16 and LBD18 to control lateral root primordium development and lateral root emergence.

Azidothymidine and cisplatin increase p14ARF expression in OVCAR-3 ovarian cancer cell line

International Nuclear Information System (INIS)

Vaskivuo, Liisa; Rysae, Jaana; Koivuperae, Johanna; Myllynen, Paeivi; Vaskivuo, Tommi; Chvalova, Katerina; Serpi, Raisa; Savolainen, Eeva-Riitta; Puistola, Ulla; Vaehaekangas, Kirsi

2006-01-01

p14 ARF tumor suppressor protein regulates p53 by interfering with mdm2-p53 interaction. p14 ARF is activated in response to oncogenic stimuli but little is known of the responses of endogenous p14 ARF to different types of cellular stress or DNA damage. Azidothymidine (AZT) is being tested in several clinical trials as an enhancer of anticancer chemotherapy. However, the knowledge of the relationship between AZT and cellular pathways, e.g. p53 pathway, is very limited. In this study, we show that AZT, cisplatin (CDDP) and docetaxel (DTX) all induce unique molecular responses in OVCAR-3 ovarian carcinoma cells carrying a mutated p53, while in A2780, ovarian carcinoma and MCF-7 breast carcinoma cells with wild type p53, all of these drugs cause similar p53 responses. We found that endogenous p14 ARF protein in OVCAR-3 cells is down-regulated by DTX but induced by AZT and a short CDDP pulse treatment. In HT-29 colon carcinoma cells with a mutated p53, all treatments down-regulated p14 ARF protein. Both CDDP and AZT increased the expression of p14ARF mRNA in OVCAR-3 cells. Differences in cell death induced by these drugs did not explain the differences in protein and mRNA expressions. No increase in the level of either c-Myc or H-ras oncoproteins was seen in OVCAR-3 cells after AZT or CDDP-treatment. These results suggest that p14 ARF can respond to DNA damage without oncogene activation in cell lines without functional p53

The Role of ARF6 in Biliary Atresia.

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Mylarappa Ningappa

Full Text Available Altered extrahepatic bile ducts, gut, and cardiovascular anomalies constitute the variable phenotype of biliary atresia (BA.To identify potential susceptibility loci, Caucasian children, normal (controls and with BA (cases at two US centers were compared at >550000 SNP loci. Systems biology analysis was carried out on the data. In order to validate a key gene identified in the analysis, biliary morphogenesis was evaluated in 2-5-day post-fertilization zebrafish embryos after morpholino-antisense oligonucleotide knockdown of the candidate gene ADP ribosylation factor-6 (ARF6, Mo-arf6.Among 39 and 24 cases at centers 1 and 2, respectively, and 1907 controls, which clustered together on principal component analysis, the SNPs rs3126184 and rs10140366 in a 3' flanking enhancer region for ARF6 demonstrated higher minor allele frequencies (MAF in each cohort, and 63 combined cases, compared with controls (0.286 vs. 0.131, P = 5.94x10-7, OR 2.66; 0.286 vs. 0.13, P = 5.57x10-7, OR 2.66. Significance was enhanced in 77 total cases, which included 14 additional BA genotyped at rs3126184 only (p = 1.58x10-2, OR = 2.66. Pathway analysis of the 1000 top-ranked SNPs in CHP cases revealed enrichment of genes for EGF regulators (p<1 x10-7, ERK/MAPK and CREB canonical pathways (p<1 x10-34, and functional networks for cellular development and proliferation (p<1 x10-45, further supporting the role of EGFR-ARF6 signaling in BA. In zebrafish embryos, Mo-arf6 injection resulted in a sparse intrahepatic biliary network, several biliary epithelial cell defects, and poor bile excretion to the gall bladder compared with uninjected embryos. Biliary defects were reproduced with the EGFR-blocker AG1478 alone or with Mo-arf6 at lower doses of each agent and rescued with arf6 mRNA.The BA-associated SNPs identify a chromosome 14q21.3 susceptibility locus encompassing the ARF6 gene. arf6 knockdown in zebrafish implicates early biliary dysgenesis as a basis for BA, and also

Wound induced tanscriptional regulation of benzylisoquinoline pathway and characterization of wound inducible PsWRKY transcription factor from Papaver somniferum.

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Sonal Mishra

Full Text Available Wounding is required to be made in the walls of the green seed pod of Opium poppy prior exudation of latex. To withstand this kind of trauma plants regulate expression of some metabolites through an induced transcript level. 167 unique wound-inducible ESTs were identified by a repetitive round of cDNA subtraction after 5 hours of wounding in Papaver somniferum seedlings. Further repetitive reverse northern analysis of these ESTs revealed 80 transcripts showing more than two fold induction, validated through semi-quantitative RT-PCR & real time expression analysis. One of the major classified categories among identified ESTs belonged to benzylisoquinoline transcripts. Tissue specific metabolite analysis of benzylisoquinoline alkaloids (BIAs in response to wounding revealed increased accumulation of narcotine and papaverine. Promoter analysis of seven transcripts of BIAs pathway showed the presence of W-box cis-element with the consensus sequence of TGAC, which is the proposed binding site for WRKY type transcription factors. One of the Wound inducible 'WRKY' EST isolated from our subtracted library was made full-length and named as 'PsWRKY'. Bacterially expressed PsWRKY interacted with the W-box element having consensus sequence TTGACT/C present in the promoter region of BIAs biosynthetic pathway genes. PsWRKY further activated the TYDC promoter in yeast and transiently in tobacco BY2 cells. Preferential expression of PsWRKY in straw and capsule and its interaction with consensus W-box element present in BIAs pathway gene transcripts suggest its possible involvement in the wound induced regulation of BIAs pathway.

The Polycomb group proteins bind throughout the INK4A-ARF locus and are disassociated in senescent cells

DEFF Research Database (Denmark)

Bracken, Adrian P; Kleine-Kohlbrecher, Daniela; Dietrich, Nikolaj

2007-01-01

The p16INK4A and p14ARF proteins, encoded by the INK4A-ARF locus, are key regulators of cellular senescence, yet the mechanisms triggering their up-regulation are not well understood. Here, we show that the ability of the oncogene BMI1 to repress the INK4A-ARF locus requires its direct association...... and is dependent on the continued presence of the EZH2-containing Polycomb-Repressive Complex 2 (PRC2) complex. Significantly, EZH2 is down-regulated in stressed and senescing populations of cells, coinciding with decreased levels of associated H3K27me3, displacement of BMI1, and activation of transcription...

Increased Arf/p53 activity in stem cells, aging and cancer.

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Carrasco-Garcia, Estefania; Moreno, Manuel; Moreno-Cugnon, Leire; Matheu, Ander

2017-04-01

Arf/p53 pathway protects the cells against DNA damage induced by acute stress. This characteristic is the responsible for its tumor suppressor activity. Moreover, it regulates the chronic type of stress associated with aging. This is the basis of its anti-aging activity. Indeed, increased gene dosage of Arf/p53 displays elongated longevity and delayed aging. At a cellular level, it has been recently shown that increased dosage of Arf/p53 delays age-associated stem cell exhaustion and the subsequent decline in tissue homeostasis and regeneration. However, p53 can also promote aging if constitutively activated. In this context, p53 reduces tissue regeneration, which correlates with premature exhaustion of stem cells. We discuss here the current evidence linking the Arf/p53 pathway to the processes of aging and cancer through stem cell regulation. © 2017 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

A-RAF kinase functions in ARF6 regulated endocytic membrane traffic.

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Elena Nekhoroshkova

Full Text Available BACKGROUND: RAF kinases direct ERK MAPK signaling to distinct subcellular compartments in response to growth factor stimulation. METHODOLOGY/PRINCIPAL FINDINGS: Of the three mammalian isoforms A-RAF is special in that one of its two lipid binding domains mediates a unique pattern of membrane localization. Specific membrane binding is retained by an N-terminal fragment (AR149 that corresponds to a naturally occurring splice variant termed DA-RAF2. AR149 colocalizes with ARF6 on tubular endosomes and has a dominant negative effect on endocytic trafficking. Moreover actin polymerization of yeast and mammalian cells is abolished. AR149/DA-RAF2 does not affect the internalization step of endocytosis, but trafficking to the recycling compartment. CONCLUSIONS/SIGNIFICANCE: A-RAF induced ERK activation is required for this step by activating ARF6, as A-RAF depletion or inhibition of the A-RAF controlled MEK-ERK cascade blocks recycling. These data led to a new model for A-RAF function in endocytic trafficking.

Induction of nonapoptotic cell death by activated Ras requires inverse regulation of Rac1 and Arf6.

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Bhanot, Haymanti; Young, Ashley M; Overmeyer, Jean H; Maltese, William A

2010-10-01

Methuosis is a unique form of nonapoptotic cell death triggered by alterations in the trafficking of clathrin-independent endosomes, ultimately leading to extreme vacuolization and rupture of the cell. Methuosis can be induced in glioblastoma cells by expression of constitutively active Ras. This study identifies the small GTPases, Rac1 and Arf6, and the Arf6 GTPase-activating protein, GIT1, as key downstream components of the signaling pathway underlying Ras-induced methuosis. The extent to which graded expression of active H-Ras(G12V) triggers cytoplasmic vacuolization correlates with the amount of endogenous Rac1 in the active GTP state. Blocking Rac1 activation with the specific Rac inhibitor, EHT 1864, or coexpression of dominant-negative Rac1(T17N), prevents the accumulation of vacuoles induced by H-Ras(G12V). Coincident with Rac1 activation, H-Ras(G12V) causes a decrease in the amount of active Arf6, a GTPase that functions in the recycling of clathrin-independent endosomes. The effect of H-Ras(G12V) on Arf6 is blocked by EHT 1864, indicating that the decrease in Arf6-GTP is directly linked to the activation of Rac1. Constitutively active Rac1(G12V) interacts with GIT1 in immunoprecipitation assays. Ablation of GIT1 by short hairpin RNA prevents the decrease in active Arf6, inhibits vacuolization, and prevents loss of cell viability in cells expressing Rac1(G12V). Together, the results suggest that perturbations of endosome morphology associated with Ras-induced methuosis are due to downstream activation of Rac1 combined with reciprocal inactivation of Arf6. The latter seems to be mediated through Rac1 stimulation of GIT1. Further insights into this pathway could suggest opportunities for the induction of methuosis in cancers that are resistant to apoptotic cell death.

The domain architecture of large guanine nucleotide exchange factors for the small GTP-binding protein Arf

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Geldner Niko

2005-02-01

Full Text Available Abstract Background Small G proteins, which are essential regulators of multiple cellular functions, are activated by guanine nucleotide exchange factors (GEFs that stimulate the exchange of the tightly bound GDP nucleotide by GTP. The catalytic domain responsible for nucleotide exchange is in general associated with non-catalytic domains that define the spatio-temporal conditions of activation. In the case of small G proteins of the Arf subfamily, which are major regulators of membrane trafficking, GEFs form a heterogeneous family whose only common characteristic is the well-characterized Sec7 catalytic domain. In contrast, the function of non-catalytic domains and how they regulate/cooperate with the catalytic domain is essentially unknown. Results Based on Sec7-containing sequences from fully-annotated eukaryotic genomes, including our annotation of these sequences from Paramecium, we have investigated the domain architecture of large ArfGEFs of the BIG and GBF subfamilies, which are involved in Golgi traffic. Multiple sequence alignments combined with the analysis of predicted secondary structures, non-structured regions and splicing patterns, identifies five novel non-catalytic structural domains which are common to both subfamilies, revealing that they share a conserved modular organization. We also report a novel ArfGEF subfamily with a domain organization so far unique to alveolates, which we name TBS (TBC-Sec7. Conclusion Our analysis unifies the BIG and GBF subfamilies into a higher order subfamily, which, together with their being the only subfamilies common to all eukaryotes, suggests that they descend from a common ancestor from which species-specific ArfGEFs have subsequently evolved. Our identification of a conserved modular architecture provides a background for future functional investigation of non-catalytic domains.

TBC1D24 regulates neuronal migration and maturation through modulation of the ARF6-dependent pathway

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Falace, Antonio; Buhler, Emmanuelle; Fadda, Manuela; Watrin, Françoise; Lippiello, Pellegrino; Pallesi-Pocachard, Emilie; Baldelli, Pietro; Benfenati, Fabio; Zara, Federico; Represa, Alfonso; Fassio, Anna; Cardoso, Carlos

2014-01-01

Alterations in the formation of brain networks are associated with several neurodevelopmental disorders. Mutations in TBC1 domain family member 24 (TBC1D24) are responsible for syndromes that combine cortical malformations, intellectual disability, and epilepsy, but the function of TBC1D24 in the brain remains unknown. We report here that in utero TBC1D24 knockdown in the rat developing neocortex affects the multipolar-bipolar transition of neurons leading to delayed radial migration. Furthermore, we find that TBC1D24-knockdown neurons display an abnormal maturation and retain immature morphofunctional properties. TBC1D24 interacts with ADP ribosylation factor (ARF)6, a small GTPase crucial for membrane trafficking. We show that in vivo, overexpression of the dominant-negative form of ARF6 rescues the neuronal migration and dendritic outgrowth defects induced by TBC1D24 knockdown, suggesting that TBC1D24 prevents ARF6 activation. Overall, our findings demonstrate an essential role of TBC1D24 in neuronal migration and maturation and highlight the physiological relevance of the ARF6-dependent membrane-trafficking pathway in brain development. PMID:24469796

Estrogen induced concentration dependent differential gene expression in human breast cancer (MCF7) cells: Role of transcription factors

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Chandrasekharan, Sabarinath; Kandasamy, Krishna Kumar; Dayalan, Pavithra; Ramamurthy, Viraragavan

2013-01-01

Highlights: •Estradiol (E2) at low dose induced cell proliferation in breast cancer cells. •E2 at high concentration induced cell stress in breast cancer cells. •Estrogen receptor physically interacts only with a few transcription factors. •Differential expression of genes with Oct-1 binding sites increased under stress. •Transcription factor binding sites showed distinct spatial distribution on genes. -- Abstract: Background: Breast cancer cells respond to estrogen in a concentration dependent fashion, resulting in proliferation or apoptosis. The mechanism of this concentration dependent differential outcome is not well understood yet. Methodology: Meta-analysis of the expression data of MCF7 cells treated with low (1 nM) or high (100 nM) dose of estradiol (E2) was performed. We identified genes differentially expressed at the low or the high dose, and examined the nature of regulatory elements in the vicinity of these genes. Specifically, we looked for the difference in the presence, abundance and spatial distribution of binding sites for estrogen receptor (ER) and selected transcription factors (TFs) in the genomic region up to 25 kb upstream and downstream from the transcription start site (TSS) of these genes. Results: It was observed that at high dose E2 induced the expression of stress responsive genes, while at low dose, genes involved in cell cycle were induced. We found that the occurrence of transcription factor binding regions (TFBRs) for certain factors such as Sp1 and SREBP1 were higher on regulatory regions of genes expressed at low dose. At high concentration of E2, genes with a higher frequency of Oct-1 binding regions were predominantly involved. In addition, there were differences in the spatial distribution pattern of the TFBRs in the genomic regions among the two sets of genes. Discussion: E2 induced predominantly proliferative/metabolic response at low concentrations; but at high concentration, stress–rescue responses were induced

Estrogen induced concentration dependent differential gene expression in human breast cancer (MCF7) cells: Role of transcription factors

Energy Technology Data Exchange (ETDEWEB)

Chandrasekharan, Sabarinath, E-mail: csab@bio.psgtech.ac.in [Department of Biotechnology, PSG College of Technology, Coimbatore 641004 (India); Kandasamy, Krishna Kumar [Max Planck Institute for Biology of Ageing, Cologne (Germany); Dayalan, Pavithra; Ramamurthy, Viraragavan [Department of Biotechnology, PSG College of Technology, Coimbatore 641004 (India)

2013-08-02

Highlights: •Estradiol (E2) at low dose induced cell proliferation in breast cancer cells. •E2 at high concentration induced cell stress in breast cancer cells. •Estrogen receptor physically interacts only with a few transcription factors. •Differential expression of genes with Oct-1 binding sites increased under stress. •Transcription factor binding sites showed distinct spatial distribution on genes. -- Abstract: Background: Breast cancer cells respond to estrogen in a concentration dependent fashion, resulting in proliferation or apoptosis. The mechanism of this concentration dependent differential outcome is not well understood yet. Methodology: Meta-analysis of the expression data of MCF7 cells treated with low (1 nM) or high (100 nM) dose of estradiol (E2) was performed. We identified genes differentially expressed at the low or the high dose, and examined the nature of regulatory elements in the vicinity of these genes. Specifically, we looked for the difference in the presence, abundance and spatial distribution of binding sites for estrogen receptor (ER) and selected transcription factors (TFs) in the genomic region up to 25 kb upstream and downstream from the transcription start site (TSS) of these genes. Results: It was observed that at high dose E2 induced the expression of stress responsive genes, while at low dose, genes involved in cell cycle were induced. We found that the occurrence of transcription factor binding regions (TFBRs) for certain factors such as Sp1 and SREBP1 were higher on regulatory regions of genes expressed at low dose. At high concentration of E2, genes with a higher frequency of Oct-1 binding regions were predominantly involved. In addition, there were differences in the spatial distribution pattern of the TFBRs in the genomic regions among the two sets of genes. Discussion: E2 induced predominantly proliferative/metabolic response at low concentrations; but at high concentration, stress–rescue responses were induced

ARF tumor suppression in the nucleolus.

Science.gov (United States)

Maggi, Leonard B; Winkeler, Crystal L; Miceli, Alexander P; Apicelli, Anthony J; Brady, Suzanne N; Kuchenreuther, Michael J; Weber, Jason D

2014-06-01

Since its discovery close to twenty years ago, the ARF tumor suppressor has played a pivotal role in the field of cancer biology. Elucidating ARF's basal physiological function in the cell has been the focal interest of numerous laboratories throughout the world for many years. Our current understanding of ARF is constantly evolving to include novel frameworks for conceptualizing the regulation of this critical tumor suppressor. As a result of this complexity, there is great need to broaden our understanding of the intricacies governing the biology of the ARF tumor suppressor. The ARF tumor suppressor is a key sensor of signals that instruct a cell to grow and proliferate and is appropriately localized in nucleoli to limit these processes. This article is part of a Special Issue entitled: Role of the Nucleolus in Human Disease. Copyright © 2014 Elsevier B.V. All rights reserved.

The transcription factor LEF-1 induces an epithelial–mesenchymal transition in MDCK cells independent of β-catenin

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Kobayashi, Wakako; Ozawa, Masayuki, E-mail: mozawa@m.kufm.kagoshima-u.ac.jp

2013-12-06

Highlights: •The transcription factor LEF-1 induces an EMT in MDCK cells. •A mutant LEF-1 that cannot interact with β-catenin retained the ability. •The nuclear function of β-catenin was not necessary for the LEF-1-induced EMT. •The mRNA levels of Slug, ZEB1, and ZEB2 increased significantly in these cells. -- Abstract: The epithelial–mesenchymal transition (EMT), a key process in the tumor metastatic cascade, is characterized by the loss of cell–cell junctions and cell polarity, as well as the acquisition of migratory and invasive properties. LEF-1 is a member of the lymphoid enhancer-binding factor/T-cell factor (LEF/TCF) family of DNA-binding transcription factors, which interact with nuclear β-catenin and act as central transcriptional mediators of Wnt signaling. To investigate the role of LEF-1 in EMT, we generated stable LEF-1 transfectants using MDCK cells. The transfectants had a spindle-shaped mesenchymal morphology, and enhanced migration and invasiveness relative to control cells. These EMT changes were accompanied by the downregulation of an epithelial marker protein, E-cadherin, and the upregulation of mesenchymal marker proteins, vimentin and N-cadherin. Consistent with these observations, the mRNA levels of Slug, ZEB1, and ZEB2—EMT-related transcription factors—increased significantly. Although the N-terminally deleted mutant LEF-1 cannot interact with β-catenin, it retained the ability to induce EMT. Consistent with these observations, neither the expression of a dominant negative β-catenin/engrailed chimera, nor the expression of a cytoplasmic domain of E-cadherin that sequesters β-catenin from binding to LEF/TCF, reversed LEF-1-induced EMT. Together, these data indicated that the nuclear function of β-catenin was not necessary for the induction of Slug, ZEB1, and ZEB2 expression leading to EMT.

Phorbol-ester-induced activation of the NF-κB transcription factor involves dissociation of an apparently cytoplasmic NF-κB/inhibitor complex

International Nuclear Information System (INIS)

Baeuerle, P.A.; Lenardo, M.; Pierce, J.W.; Baltimore, D.

1988-01-01

There is increasing evidence that inducible transcription of genes is mediated through the induction of the activity of trans-acting protein factors. The NF-κB transcription factor provides a model system to study the posttranslational activation of a phorbol-ester-inducible transcription factor. The finding that NF-κB activity is undectable in subcellular fractions from unstimulated cells suggests that NF-κB exists as an inactive precursor. The authors showed that NF-κB is detectable in two different forms. After selective removal of endogenous NF-κB, they demonstrate the existence of a protein inhibitor in cytosolic fractions of unstimulated cells that is able in vitro to convert NF-κB into an inactive desoxycholate-dependent form. The data are consistent with a molecular mechanism of inducible gene expression by which an apparently cytoplasmic transcription factor-inhibitor complex is dissociated by the action of TPA-activated protein kinase C

Induction of Non-Apoptotic Cell Death by Activated Ras Requires Inverse Regulation of Rac1 and Arf6

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Bhanot, Haymanti; Young, Ashley M.; Overmeyer, Jean H.; Maltese, William A.

2010-01-01

Methuosis is a unique form of non-apoptotic cell death triggered by alterations in the trafficking of clathrin-independent endosomes, ultimately leading to extreme vacuolization and rupture of the cell. Methuosis can be induced in glioblastoma cells by expression of constitutively active Ras. This study identifies the small GTPases, Rac1 and Arf6, and the Arf6 GTPase-activating-protein, GIT1, as key downstream components of the signaling pathway underlying Ras-induced methuosis. The extent to which graded expression of active H-Ras(G12V) triggers cytoplasmic vacuolization correlates with the amount of endogenous Rac1 in the active GTP state. Blocking Rac1 activation with the specific Rac inhibitor, EHT 1864, or co-expression of dominant-negative Rac1(T17N), prevents the accumulation of vacuoles induced by H-Ras(G12V). Coincident with Rac1 activation, H-Ras(G12V) causes a decrease in the amount of active Arf6, a GTPase that functions in recycling of clathrin-independent endosomes. The effect of H-Ras(G12V) on Arf6 is blocked by EHT 1864, indicating that the decrease in Arf6-GTP is directly linked to activation of Rac1. Constitutively active Rac1(G12V) interacts with GIT1 in immunoprecipitation assays. Ablation of GIT1 by shRNA prevents the decrease in active Arf6, inhibits vacuolization, and prevents loss of cell viability in cells expressing Rac1(G12V). Together the results suggest that perturbations of endosome morphology associated with Ras-induced methuosis are due to downstream activation of Rac1, combined with reciprocal inactivation of Arf6. The latter appears to be mediated through Rac1 stimulation of GIT1. Further insights into this pathway could suggest opportunities for induction of methuosis in cancers that are resistant to apoptotic cell death. PMID:20713492

Expression of transcription factors Slug in the lens epithelial cells undergoing epithelial-mesenchymal transition induced by connective tissue growth factor

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Ying-Na Wang

2015-10-01

Full Text Available AIM:To investigate the expression of transcription factors Slug in human lens epithelial cells (HLECs undergoing epithelial-mesenchymal transition (EMT induced by connective tissue growth factor (CTGF.METHODS: HLECs were treated with CTGF of different concentrations (20, 50 and 100 ng/mL or without CTGF (control for 24h. The morphological changes of HLECs were analysed by microscopy. The expression and cellular localization of Slug was evaluated by immumo-fluorescence. Expressions of Slug, E-cadherin and alpha smooth muscle actin (α-SMA were further determined by Western blot analysis. RESULTS: HLECs showed spidle fibrolasts-like characteristics and loosely connected each other after CTGF treatment. The immuno-fluorescence staining indicated that Slug was localized in the nuclei and its expression was induced by CTGF. The relative expressions of Slug protein were 1.64±0.11, 1.96 ±0.03, 3.12 ±0.10, and 4.08±0.14, respectively, in response to control group and treatment with CTGF of 20, 50 and 100 ng/mL (F=443.86, PCONCLUSION: Transcription factor Slug may be involved in EMT of HLECs induced by CTGF in vitro.

A Taiwanese Propolis Derivative Induces Apoptosis through Inducing Endoplasmic Reticular Stress and Activating Transcription Factor-3 in Human Hepatoma Cells

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Fat-Moon Suk

2013-01-01

Full Text Available Activating transcription factor-(ATF- 3, a stress-inducible transcription factor, is rapidly upregulated under various stress conditions and plays an important role in inducing cancer cell apoptosis. NBM-TP-007-GS-002 (GS-002 is a Taiwanese propolin G (PPG derivative. In this study, we examined the antitumor effects of GS-002 in human hepatoma Hep3B and HepG2 cells in vitro. First, we found that GS-002 significantly inhibited cell proliferation and induced cell apoptosis in dose-dependent manners. Several main apoptotic indicators were found in GS-002-treated cells, such as the cleaved forms of caspase-3, caspase-9, and poly(ADP-ribose polymerase (PARP. GS-002 also induced endoplasmic reticular (ER stress as evidenced by increases in ER stress-responsive proteins including glucose-regulated protein 78 (GRP78, growth arrest- and DNA damage-inducible gene 153 (GADD153, phosphorylated eukaryotic initiation factor 2α (eIF2α, phosphorylated protein endoplasmic-reticular-resident kinase (PERK, and ATF-3. The induction of ATF-3 expression was mediated by mitogen-activated protein kinase (MAPK signaling pathways in GS-002-treated cells. Furthermore, we found that GS-002 induced more cell apoptosis in ATF-3-overexpressing cells. These results suggest that the induction of apoptosis by the propolis derivative, GS-002, is partially mediated through ER stress and ATF-3-dependent pathways, and GS-002 has the potential for development as an antitumor drug.

Manipulation of Auxin Response Factor 19 affects seed size in the woody perennial Jatropha curcas

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Sun, Yanwei; Wang, Chunming; Wang, Ning; Jiang, Xiyuan; Mao, Huizhu; Zhu, Changxiang; Wen, Fujiang; Wang, Xianghua; Lu, Zhijun; Yue, Genhua; Xu, Zengfu; Ye, Jian

2017-01-01

Seed size is a major determinant of seed yield but few is known about the genetics controlling of seed size in plants. Phytohormones cytokinin and brassinosteroid were known to be involved in the regulation of herbaceous plant seed development. Here we identified a homolog of Auxin Response Factor 19 (JcARF19) from a woody plant Jatropha curcas and genetically demonstrated its functions in controlling seed size and seed yield. Through Virus Induced Gene Silencing (VIGS), we found that JcARF19 was a positive upstream modulator in auxin signaling and may control plant organ size in J. curcas. Importantly, transgenic overexpression of JcARF19 significantly increased seed size and seed yield in plants Arabidopsis thaliana and J. curcas, indicating the importance of auxin pathway in seed yield controlling in dicot plants. Transcripts analysis indicated that ectopic expression of JcARF19 in J. curcas upregulated auxin responsive genes encoding essential regulators in cell differentiation and cytoskeletal dynamics of seed development. Our data suggested the potential of improving seed traits by precisely engineering auxin signaling in woody perennial plants. PMID:28102350

The Drosophila Arf GEF Steppke controls MAPK activation in EGFR signaling.

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Hahn, Ines; Fuss, Bernhard; Peters, Annika; Werner, Tamara; Sieberg, Andrea; Gosejacob, Dominic; Hoch, Michael

2013-06-01

Guanine nucleotide exchange factors (GEFs) of the cytohesin protein family are regulators of GDP/GTP exchange for members of the ADP ribosylation factor (Arf) of small GTPases. They have been identified as modulators of various receptor tyrosine kinase signaling pathways including the insulin, the vascular epidermal growth factor (VEGF) and the epidermal growth factor (EGF) pathways. These pathways control many cellular functions, including cell proliferation and differentiation, and their misregulation is often associated with cancerogenesis. In vivo studies on cytohesins using genetic loss of function alleles are lacking, however, since knockout mouse models are not available yet. We have recently identified mutants for the single cytohesin Steppke (Step) in Drosophila and we could demonstrate an essential role of Step in the insulin signaling cascade. In the present study, we provide in vivo evidence for a role of Step in EGFR signaling during wing and eye development. By analyzing step mutants, transgenic RNA interference (RNAi) and overexpression lines for tissue specific as well as clonal analysis, we found that Step acts downstream of the EGFR and is required for the activation of mitogen-activated protein kinase (MAPK) and the induction of EGFR target genes. We further demonstrate that step transcription is induced by EGFR signaling whereas it is negatively regulated by insulin signaling. Furthermore, genetic studies and biochemical analysis show that Step interacts with the Connector Enhancer of KSR (CNK). We propose that Step may be part of a larger signaling scaffold coordinating receptor tyrosine kinase-dependent MAPK activation.

Blue light alters miR167 expression and microRNA-targeted auxin response factor genes in Arabidopsis thaliana plants.

Science.gov (United States)

Pashkovskiy, Pavel P; Kartashov, Alexander V; Zlobin, Ilya E; Pogosyan, Sergei I; Kuznetsov, Vladimir V

2016-07-01

The effect of blue LED (450 nm) on the photomorphogenesis of Arabidopsis thaliana Col-0 plants and the transcript levels of several genes, including miRNAs, photoreceptors and auxin response factors (ARF) was investigated. It was observed that blue light accelerated the generative development, reduced the rosette leaf number, significantly reduced the leaf area, dry biomass and led to the disruption of conductive tissue formation. The blue LED differentially influenced the transcript levels of several phytochromes (PHY a, b, c, d, and e), cryptochromes (CRY 1 and 2) and phototropins (PHOT 1 and 2). At the same time, the blue LED significantly increased miR167 expression compared to a fluorescent lamp or white LEDs. This increase likely resulted in the enhanced transcription of the auxin response factor genes ARF4 and ARF8, which are regulated by this miRNA. These findings support the hypothesis that the effects of blue light on A. thaliana are mediated by auxin signalling pathway involving miRNA-dependent regulation of ARF gene expression. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

UV-induced transcription from the human immunodeficiency virus type 1 (HIV-1) long terminal repeat and UV-induced secretion of an extracellular factor that induces HIV-1 transcription in nonirradiated cells

International Nuclear Information System (INIS)

Stein, B.; Kraemer, M.R.; Rahmsdorf, H.J.; Ponta, H.; Herrlich, P.

1989-01-01

UV irradiation, but not visible sunlight, induces the transcription of human immunodeficiency virus type 1 (HIV-1). Chimeric constructs carrying all or parts of the HIV-1 long terminal repeat linked to an indicator gene were transfected into HeLa cells or murine and human T-cell lines, and their response to irradiation was tested. The cis-acting element conferring UV responsiveness is identical to the sequence binding transcription factor NF kappa B. UV irradiation enhances NF kappa B binding activity as assayed by gel retardation experiments. Interestingly, the requirement for UV irradiation can be replaced by cocultivation of transfected cells with UV-irradiated nontransfected (HIV-1-negative) cells. A UV-induced extracellular protein factor is detected in the culture medium conditioned by UV-treated cells. The factor is produced upon UV irradiation by several murine and human cell lines, including HeLa, Molt-4, and Jurkat, and acts on several cells. These data suggest that the UV response of keratinocytes in human skin can be magnified and spread to deeper layers that are more shielded, including the Langerhans cells, and that this indirect UV response may contribute to the activation of HIV-1 in humans

Potential Therapeutic Uses of p19ARF Mimics in Mammary Tumorigenesis

National Research Council Canada - National Science Library

Hann, Stephen R

2005-01-01

Since many breast tumors have deregulated c-Myc we hypothesize that an ARF mimic would be a valuable therapeutic agent for breast cancer to inhibit c-Myc-induced transformation/tumorigenesis without...

The Role of the E2F Transcription Factor Family in UV-Induced Apoptosis

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Orla Gannon

2011-12-01

Full Text Available The E2F transcription factor family is traditionally associated with cell cycle control. However, recent data has shown that activating E2Fs (E2F1-3a are potent activators of apoptosis. In contrast, the recently cloned inhibitory E2Fs (E2F7 and 8 appear to antagonize E2F-induced cell death. In this review we will discuss (i the potential role of E2Fs in UV-induced cell death and (ii the implications of this to the development of UV-induced cutaneous malignancies.

Elicitor-induced transcription factors for metabolic reprogramming of secondary metabolism in Medicago truncatula

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Dixon Richard A

2008-12-01

Full Text Available Abstract Background Exposure of Medicago truncatula cell suspension cultures to pathogen or wound signals leads to accumulation of various classes of flavonoid and/or triterpene defense molecules, orchestrated via a complex signalling network in which transcription factors (TFs are essential components. Results In this study, we analyzed TFs responding to yeast elicitor (YE or methyl jasmonate (MJ. From 502 differentially expressed TFs, WRKY and AP2/EREBP gene families were over-represented among YE-induced genes whereas Basic Helix-Loop-Helix (bHLH family members were more over-represented among the MJ-induced genes. Jasmonate ZIM-domain (JAZ transcriptional regulators were highly induced by MJ treatment. To investigate potential involvement of WRKY TFs in signalling, we expressed four Medicago WRKY genes in tobacco. Levels of soluble and wall bound phenolic compounds and lignin were increased in all cases. WRKY W109669 also induced tobacco endo-1,3-β-glucanase (NtPR2 and enhanced the systemic defense response to tobacco mosaic virus in transgenic tobacco plants. Conclusion These results confirm that Medicago WRKY TFs have broad roles in orchestrating metabolic responses to biotic stress, and that they also represent potentially valuable reagents for engineering metabolic changes that impact pathogen resistance.

Hypoxia, hypoxia-inducible transcription factor, and macrophages in human atherosclerotic plaques are correlated with intraplaque angiogenesis

NARCIS (Netherlands)

Sluimer, Judith C.; Gasc, Jean-Marie; van Wanroij, Job L.; Kisters, Natasja; Groeneweg, Mathijs; Sollewijn Gelpke, Maarten D.; Cleutjens, Jack P.; van den Akker, Luc H.; Corvol, Pierre; Wouters, Bradly G.; Daemen, Mat J.; Bijnens, Ann-Pascale J.

2008-01-01

We sought to examine the presence of hypoxia in human carotid atherosclerosis and its association with hypoxia-inducible transcription factor (HIF) and intraplaque angiogenesis. Atherosclerotic plaques develop intraplaque angiogenesis, which is a typical feature of hypoxic tissue and expression of

Brassinosteroid-Induced Transcriptional Repression and Dephosphorylation-Dependent Protein Degradation Negatively Regulate BIN2-Interacting AIF2 (a BR Signaling-Negative Regulator) bHLH Transcription Factor.

Science.gov (United States)

Kim, Yoon; Song, Ji-Hye; Park, Seon-U; Jeong, You-Seung; Kim, Soo-Hwan

2017-02-01

Brassinosteroids (BRs) are plant polyhydroxy-steroids that play important roles in plant growth and development via extensive signal integration through direct interactions between regulatory components of different signaling pathways. Recent studies have shown that diverse helix-loop-helix/basic helix-loop-helix (HLH/bHLH) family proteins are actively involved in control of BR signaling pathways and interact with other signaling pathways. In this study, we show that ATBS1-INTERACTING FACTOR 2 (AIF2), a nuclear-localized atypical bHLH transcription factor, specifically interacts with BRASSINOSTEROID-INSENSITIVE 2 (BIN2) among other BR signaling molecules. Overexpression of AIF2 down-regulated transcript expression of growth-promoting genes, thus resulting in retardation of growth. AIF2 renders plants hyposensitive to BR-induced root growth inhibition, but shows little effects on BR-promoted hypocotyl elongation. Notably, AIF2 was dephosphorylated by BR, and the dephosphorylated AIF2 was subject to proteasome-mediated degradation. AIF2 degradation was greatly induced by BR and ABA, but relatively slightly by other hormones such as auxin, gibberellin, cytokinin and ethylene. Moreover, AIF2 transcription was significantly suppressed by a BRI1/BZR1-mediated BR signaling pathway through a direct binding of BRASSINAZOLE RESISTANT 1 (BZR1) to the BR response element (BRRE) region of the AIF2 promoter. In conclusion, our study suggests that BIN2-driven AIF2 phosphorylation could augment the BIN2/AIF2-mediated negative circuit of BR signaling pathways, and the BR-induced transcriptional repression and protein degradation negatively regulate AIF2 transcription factor, reinforcing the BZR1/BES1-mediated positive BR signaling pathway. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

ArF Excimer Laser-induced Deposition of Ag/C Nanocomposite Thin Films in the Presence of n-Hexane

Czech Academy of Sciences Publication Activity Database

Gondal, M.A.; Fajgar, Radek; Chang, X.; Shen, K.; Xu, Q.

2014-01-01

Roč. 311, AUG 30 (2014), s. 95-100 ISSN 0169-4332 Grant - others:NNSFCH(CN) 51172044; NSFJP(CN) BK2011617; KFUPM(CN) RG 1311-1 Institutional support: RVO:67985858 Keywords : ArF excimer laser * nanocomposite * laser deposition Subject RIV: CH - Nuclear ; Quantum Chemistry Impact factor: 2.711, year: 2014

Flavonoids as Putative Inducers of the Transcription Factors Nrf2, FoxO, and PPARγ

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Kathrin Pallauf

2017-01-01

Full Text Available Dietary flavonoids have been shown to extend the lifespan of some model organisms and may delay the onset of chronic ageing-related diseases. Mechanistically, the effects could be explained by the compounds scavenging free radicals or modulating signalling pathways. Transcription factors Nrf2, FoxO, and PPARγ possibly affect ageing by regulating stress response, adipogenesis, and insulin sensitivity. Using Hek-293 cells transfected with luciferase reporter constructs, we tested the potency of flavonoids from different subclasses (flavonols, flavones, flavanols, and isoflavones to activate these transcription factors. Under cell-free conditions (ABTS and FRAP assays, we tested their free radical scavenging activities and used α-tocopherol and ascorbic acid as positive controls. Most of the tested flavonoids, but not the antioxidant vitamins, stimulated Nrf2-, FoxO-, and PPARγ-dependent promoter activities. Flavonoids activating Nrf2 also tended to induce a FoxO and PPARγ response. Interestingly, activation patterns of cellular stress response by flavonoids were not mirrored by their activities in ABTS and FRAP assays, which depended mostly on hydroxylation in the flavonoid B ring and, in some cases, extended that of the vitamins. In conclusion, the free radical scavenging properties of flavonoids do not predict whether these molecules can stimulate a cellular response linked to activation of longevity-associated transcription factors.

Flavonoids as Putative Inducers of the Transcription Factors Nrf2, FoxO, and PPARγ.

Science.gov (United States)

Pallauf, Kathrin; Duckstein, Nils; Hasler, Mario; Klotz, Lars-Oliver; Rimbach, Gerald

2017-01-01

Dietary flavonoids have been shown to extend the lifespan of some model organisms and may delay the onset of chronic ageing-related diseases. Mechanistically, the effects could be explained by the compounds scavenging free radicals or modulating signalling pathways. Transcription factors Nrf2, FoxO, and PPAR γ possibly affect ageing by regulating stress response, adipogenesis, and insulin sensitivity. Using Hek-293 cells transfected with luciferase reporter constructs, we tested the potency of flavonoids from different subclasses (flavonols, flavones, flavanols, and isoflavones) to activate these transcription factors. Under cell-free conditions (ABTS and FRAP assays), we tested their free radical scavenging activities and used α -tocopherol and ascorbic acid as positive controls. Most of the tested flavonoids, but not the antioxidant vitamins, stimulated Nrf2-, FoxO-, and PPAR γ -dependent promoter activities. Flavonoids activating Nrf2 also tended to induce a FoxO and PPAR γ response. Interestingly, activation patterns of cellular stress response by flavonoids were not mirrored by their activities in ABTS and FRAP assays, which depended mostly on hydroxylation in the flavonoid B ring and, in some cases, extended that of the vitamins. In conclusion, the free radical scavenging properties of flavonoids do not predict whether these molecules can stimulate a cellular response linked to activation of longevity-associated transcription factors.

The calcium-sensing receptor changes cell shape via a beta-arrestin-1 ARNO ARF6 ELMO protein network.

Science.gov (United States)

Bouschet, Tristan; Martin, Stéphane; Kanamarlapudi, Venkateswarlu; Mundell, Stuart; Henley, Jeremy M

2007-08-01

G-protein-coupled receptors (GPCRs) transduce the binding of extracellular stimuli into intracellular signalling cascades that can lead to morphological changes. Here, we demonstrate that stimulation of the calcium-sensing receptor (CaSR), a GPCR that promotes chemotaxis by detecting increases in extracellular calcium, triggers plasma membrane (PM) ruffling via a pathway that involves beta-arrestin 1, Arf nucleotide binding site opener (ARNO), ADP-ribosylating factor 6 (ARF6) and engulfment and cell motility protein (ELMO). Expression of dominant negative beta-arrestin 1 or its knockdown with siRNA impaired the CaSR-induced PM ruffling response. Expression of a catalytically inactive ARNO also reduced CaSR-induced PM ruffling. Furthermore, beta-arrestin 1 co-immunoprecipitated with the CaSR and ARNO under resting conditions. Agonist treatment did not markedly alter beta-arrestin 1 binding to the CaSR or to ARNO but it did elicit the translocation and colocalisation of the CaSR, beta-arrestin 1 and ARNO to membrane protrusions. Furthermore, ARF6 and ELMO, two proteins known to couple ARNO to the cytoskeleton, were required for CaSR-dependent morphological changes and translocated to the PM ruffles. These data suggest that cells ruffle upon CaSR stimulation via a mechanism that involves translocation of beta-arrestin 1 pre-assembled with the CaSR or ARNO, and that ELMO plays an essential role in this CaSR-signalling-induced cytoskeletal reorganisation.

[The role of Smads and related transcription factors in the signal transduction of bone morphogenetic protein inducing bone formation].

Science.gov (United States)

Xu, Xiao-liang; Dai, Ke-rong; Tang, Ting-ting

2003-09-01

To clarify the mechanisms of the signal transduction of bone morphogenetic proteins (BMPs) inducing bone formation and to provide theoretical basis for basic and applying research of BMPs. We looked up the literature of the role of Smads and related transcription factors in the signal transduction of BMPs inducing bone formation. The signal transduction processes of BMPs included: 1. BMPs combined with type II and type I receptors; 2. the type I receptor phosphorylated Smads; and 3. Smads entered the cell nucleus, interacted with transcription factors and influenced the transcription of related proteins. Smads could be divided into receptor-regulated Smads (R-Smads: Smad1, Smad2, Smad3, Smad5, Smad8 and Smad9), common-mediator Smad (co-Smad: Smad4), and inhibitory Smads (I-Smads: Smad6 and Smad7). Smad1, Smad5, Smad8, and probable Smad9 were involved in the signal transduction of BMPs. Multiple kinases, such as focal adhesion kinase (FAK), Ras-extracellular signal-regulated kinase (ERK), phosphatidylinositol 3-kinase (PI3K), and Akt serine/threonine kinase were related to Smads signal transduction. Smad1 and Smad5 related with transcription factors included core binding factor A1 (CBFA1), smad-interacting protein 1 (SIP1), ornithine decarboxylase antizyme (OAZ), activating protein-1 (AP-1), xenopus ventralizing homeobox protein-2 (Xvent-2), sandostatin (Ski), antiproliferative proteins (Tob), and homeodomain-containing transcriptian factor-8 (Hoxc-8), et al. CBFA1 could interact with Smad1, Smad2, Smad3, and Smad5, so it was involved in TGF-beta and BMP-2 signal transduction, and played an important role in the bone formation. Cleidocranial dysplasia (CCD) was thought to be caused by heterozygous mutations in CBFA1. The CBFA1 knockout mice showed no osteogenesis and had maturational disturbance of chondrocytes. Smads and related transcription factors, especially Smad1, Smad5, Smad8 and CBFA1, play an important role in the signal transduction of BMPs inducing bone

HIV-1 Nef hijacks clathrin coats by stabilizing AP-1:Arf1 polygons.

Science.gov (United States)

Shen, Qing-Tao; Ren, Xuefeng; Zhang, Rui; Lee, Il-Hyung; Hurley, James H

2015-10-23

The lentiviruses HIV and simian immunodeficiency virus (SIV) subvert intracellular membrane traffic as part of their replication cycle. The lentiviral Nef protein helps viruses evade innate and adaptive immune defenses by hijacking the adaptor protein 1 (AP-1) and AP-2 clathrin adaptors. We found that HIV-1 Nef and the guanosine triphosphatase Arf1 induced trimerization and activation of AP-1. Here we report the cryo-electron microscopy structures of the Nef- and Arf1-bound AP-1 trimer in the active and inactive states. A central nucleus of three Arf1 molecules organizes the trimers. We combined the open trimer with a known dimer structure and thus predicted a hexagonal assembly with inner and outer faces that bind the membranes and clathrin, respectively. Hexagons were directly visualized and the model validated by reconstituting clathrin cage assembly. Arf1 and Nef thus play interconnected roles in allosteric activation, cargo recruitment, and coat assembly, revealing an unexpectedly intricate organization of the inner AP-1 layer of the clathrin coat. Copyright © 2015, American Association for the Advancement of Science.

Which Way In? The RalF Arf-GEF Orchestrates Rickettsia Host Cell Invasion

Science.gov (United States)

Rennoll-Bankert, Kristen E.; Rahman, M. Sayeedur; Gillespie, Joseph J.; Guillotte, Mark L.; Kaur, Simran J.; Lehman, Stephanie S.; Beier-Sexton, Magda; Azad, Abdu F.

2015-01-01

Bacterial Sec7-domain-containing proteins (RalF) are known only from species of Legionella and Rickettsia, which have facultative and obligate intracellular lifestyles, respectively. L. pneumophila RalF, a type IV secretion system (T4SS) effector, is a guanine nucleotide exchange factor (GEF) of ADP-ribosylation factors (Arfs), activating and recruiting host Arf1 to the Legionella-containing vacuole. In contrast, previous in vitro studies showed R. prowazekii (Typhus Group) RalF is a functional Arf-GEF that localizes to the host plasma membrane and interacts with the actin cytoskeleton via a unique C-terminal domain. As RalF is differentially encoded across Rickettsia species (e.g., pseudogenized in all Spotted Fever Group species), it may function in lineage-specific biology and pathogenicity. Herein, we demonstrate RalF of R. typhi (Typhus Group) interacts with the Rickettsia T4SS coupling protein (RvhD4) via its proximal C-terminal sequence. RalF is expressed early during infection, with its inactivation via antibody blocking significantly reducing R. typhi host cell invasion. For R. typhi and R. felis (Transitional Group), RalF ectopic expression revealed subcellular localization with the host plasma membrane and actin cytoskeleton. Remarkably, R. bellii (Ancestral Group) RalF showed perinuclear localization reminiscent of ectopically expressed Legionella RalF, for which it shares several structural features. For R. typhi, RalF co-localization with Arf6 and PI(4,5)P2 at entry foci on the host plasma membrane was determined to be critical for invasion. Thus, we propose recruitment of PI(4,5)P2 at entry foci, mediated by RalF activation of Arf6, initiates actin remodeling and ultimately facilitates bacterial invasion. Collectively, our characterization of RalF as an invasin suggests that, despite carrying a similar Arf-GEF unknown from other bacteria, different intracellular lifestyles across Rickettsia and Legionella species have driven divergent roles for Ral

Developmental Expression and Hypoxic Induction of Hypoxia Inducible Transcription Factors in the Zebrafish.

Science.gov (United States)

Köblitz, Louise; Fiechtner, Birgit; Baus, Katharina; Lussnig, Rebecca; Pelster, Bernd

2015-01-01

The hypoxia inducible transcription factor (HIF) has been shown to coordinate the hypoxic response of vertebrates and is expressed in three different isoforms, HIF-1α, HIF-2α and HIF-3α. Knock down of either Hif-1α or Hif-2α in mice results in lethality in embryonic or perinatal stages, suggesting that this transcription factor is not only controlling the hypoxic response, but is also involved in developmental phenomena. In the translucent zebrafish embryo the performance of the cardiovascular system is not essential for early development, therefore this study was designed to analyze the expression of the three Hif-isoforms during zebrafish development and to test the hypoxic inducibility of these transcription factors. To complement the existing zfHif-1α antibody we expressed the whole zfHif-2α protein and used it for immunization and antibody generation. Similarly, fragments of the zfHif-3α protein were used for immunization and generation of a zfHif-3α specific antibody. To demonstrate presence of the Hif-isoforms during development [between 1 day post fertilization (1 dpf) and 9 dpf] affinity-purified antibodies were used. Hif-1α protein was present under normoxic conditions in all developmental stages, but no significant differences between the different developmental stages could be detected. Hif-2α was also present from 1 dpf onwards, but in post hatching stages (between 5 and 9 dpf) the expression level was significantly higher than prior to hatching. Similarly, Hif-3α was expressed from 1 dpf onwards, and the expression level significantly increased until 5 dpf, suggesting that Hif-2α and Hif-3α play a particular role in early development. Hypoxic exposure (oxygen partial pressure = 5 kPa) in turn caused a significant increase in the level of Hif-1α protein even at 1 dpf and in later stages, while neither Hif-2α nor Hif-3α protein level were affected. In these early developmental stages Hif-1α therefore appears to be more important for

Induction of Non-Apoptotic Cell Death by Activated Ras Requires Inverse Regulation of Rac1 and Arf6

OpenAIRE

Bhanot, Haymanti; Young, Ashley M.; Overmeyer, Jean H.; Maltese, William A.

2010-01-01

Methuosis is a unique form of non-apoptotic cell death triggered by alterations in the trafficking of clathrin-independent endosomes, ultimately leading to extreme vacuolization and rupture of the cell. Methuosis can be induced in glioblastoma cells by expression of constitutively active Ras. This study identifies the small GTPases, Rac1 and Arf6, and the Arf6 GTPase-activating-protein, GIT1, as key downstream components of the signaling pathway underlying Ras-induced methuosis. The extent to...

A Bivalent Securinine Compound SN3-L6 Induces Neuronal Differentiation via Translational Upregulation of Neurogenic Transcription Factors

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Yumei Liao

2018-04-01

Full Text Available Developing therapeutic approaches that target neuronal differentiation will be greatly beneficial for the regeneration of neurons and synaptic networks in neurological diseases. Protein synthesis (mRNA translation has recently been shown to regulate neurogenesis of neural stem/progenitor cells (NSPCs. However, it has remained unknown whether engineering translational machinery is a valid approach for manipulating neuronal differentiation. The present study identifies that a bivalent securinine compound SN3-L6, previously designed and synthesized by our group, induces potent neuronal differentiation through a novel translation-dependent mechanism. An isobaric tag for relative and absolute quantitation (iTRAQ-based proteomic analysis in Neuro-2a progenitor cells revealed that SN3-L6 upregulated a group of neurogenic transcription regulators, and also upregulated proteins involved in RNA processing, translation, and protein metabolism. Notably, puromycylation and metabolic labeling of newly synthesized proteins demonstrated that SN3-L6 induced rapid and robust activation of general mRNA translation. Importantly, mRNAs of the proneural transcription factors Foxp1, Foxp4, Hsf1, and Erf were among the targets that were translationally upregulated by SN3-L6. Either inhibition of translation or knockdown of these transcription factors blocked SN3-L6 activity. We finally confirmed that protein synthesis of a same set of transcription factors was upregulated in primary cortical NPCs. These findings together identify a new compound for translational activation and neuronal differentiation, and provide compelling evidence that reprogramming transcriptional regulation network at translational levels is a promising strategy for engineering NSPCs.

Molecular dynamics simulations of membrane deformation induced by amphiphilic helices of Epsin, Sar1p, and Arf1

Science.gov (United States)

Li, Zhen-Lu

2018-03-01

The N-terminal amphiphilic helices of proteins Epsin, Sar1p, and Arf1 play a critical role in initiating membrane deformation. The interactions of these amphiphilic helices with the lipid membranes are investigated in this study by combining the all-atom and coarse-grained simulations. In the all-atom simulations, the amphiphilic helices of Epsin and Sar1p are found to have a shallower insertion depth into the membrane than the amphiphilic helix of Arf1, but remarkably, the amphiphilic helices of Epsin and Sar1p induce higher asymmetry in the lipid packing between the two monolayers of the membrane. The insertion depth of amphiphilic helix into the membrane is determined not only by the overall hydrophobicity but also by the specific distributions of polar and non-polar residues along the helix. To directly compare their ability to deform the membrane, the coarse-grained simulations are performed to investigate the membrane deformation under the insertion of multiple helices. Project supported by the National Natural Science Foundation of China (Grant Nos. 91427302 and 11474155).

Genome-Wide Identification, Phylogeny, and Expression Analysis of ARF Genes Involved in Vegetative Organs Development in Switchgrass

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Jianli Wang

2018-01-01

Full Text Available Auxin response factors (ARFs have been reported to play vital roles during plant growth and development. In order to reveal specific functions related to vegetative organs in grasses, an in-depth study of the ARF gene family was carried out in switchgrass (Panicum virgatum L., a warm-season C4 perennial grass that is mostly used as bioenergy and animal feedstock. A total of 47 putative ARF genes (PvARFs were identified in the switchgrass genome (2n = 4x = 36, 42 of which were anchored to the seven pairs of chromosomes and found to be unevenly distributed. Sixteen PvARFs were predicted to be potential targets of small RNAs (microRNA160 and 167. Phylogenetically speaking, PvARFs were divided into seven distinct subgroups based on the phylogeny, exon/intron arrangement, and conserved motif distribution. Moreover, 15 pairs of PvARFs have different temporal-spatial expression profiles in vegetative organs (2nd, 3rd, and 4th internode and leaves, which implies that different PvARFs have specific functions in switchgrass growth and development. In addition, at least 14 pairs of PvARFs respond to naphthylacetic acid (NAA treatment, which might be helpful for us to study on auxin response in switchgrass. The comprehensive analysis, described here, will facilitate the future functional analysis of ARF genes in grasses.

Repression of meiotic genes by antisense transcription and by Fkh2 transcription factor in Schizosaccharomyces pombe.

Science.gov (United States)

Chen, Huei-Mei; Rosebrock, Adam P; Khan, Sohail R; Futcher, Bruce; Leatherwood, Janet K

2012-01-01

In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s) of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the "unspliced" signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression.

Ultraviolet Radiation and the Slug Transcription Factor Induce Proinflammatory and Immunomodulatory Mediator Expression in Melanocytes

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Stephanie H. Shirley

2012-01-01

Full Text Available Despite extensive investigation, the precise contribution of the ultraviolet radiation (UVR component of sunlight to melanoma etiology remains unclear. UVR induces keratinocytes to secrete proinflammatory and immunomodulatory mediators that promote inflammation and skin tumor development; expression of the slug transcription factor in keratinocytes is required for maximal production of these mediators. In the present studies we examined the possibility that UVR-exposed melanocytes also produce proinflammatory mediators and that Slug is important in this process. Microarray studies revealed that both UVR exposure and Slug overexpression altered transcription of a variety of proinflammatory mediators by normal human melanocytes; some of these mediators are also known to stimulate melanocyte growth and migration. There was little overlap in the spectra of cytokines produced by the two stimuli. However IL-20 was similarly induced by both stimuli and the NFκB pathway appeared to be important in both circumstances. Further exploration of UVR-induced and Slug-dependent pathways of cytokine induction in melanocytes may reveal novel targets for melanoma therapy.

Manganese Superoxide Dismutase Gene Expression Is Induced by Nanog and Oct4, Essential Pluripotent Stem Cells’ Transcription Factors

Science.gov (United States)

Solari, Claudia; Vázquez Echegaray, Camila; Cosentino, María Soledad; Petrone, María Victoria; Waisman, Ariel; Luzzani, Carlos; Francia, Marcos; Villodre, Emilly; Lenz, Guido; Miriuka, Santiago; Barañao, Lino; Guberman, Alejandra

2015-01-01

Pluripotent stem cells possess complex systems that protect them from oxidative stress and ensure genomic stability, vital for their role in development. Even though it has been reported that antioxidant activity diminishes along stem cell differentiation, little is known about the transcriptional regulation of the involved genes. The reported modulation of some of these genes led us to hypothesize that some of them could be regulated by the transcription factors critical for self-renewal and pluripotency in embryonic stem cells (ESCs) and in induced pluripotent stem cells (iPSCs). In this work, we studied the expression profile of multiple genes involved in antioxidant defense systems in both ESCs and iPSCs. We found that Manganese superoxide dismutase gene (Mn-Sod/Sod2) was repressed during diverse differentiation protocols showing an expression pattern similar to Nanog gene. Moreover, Sod2 promoter activity was induced by Oct4 and Nanog when we performed a transactivation assay using two different reporter constructions. Finally, we studied Sod2 gene regulation by modulating the expression of Oct4 and Nanog in ESCs by shRNAs and found that downregulation of any of them reduced Sod2 expression. Our results indicate that pluripotency transcription factors positively modulate Sod2 gene transcription. PMID:26642061

Hydrophilicity and morphological investigation of polycarbonate irradiated by ArF excimer laser

Energy Technology Data Exchange (ETDEWEB)

Jaleh, B. [Bu-Ali-Sina University, Physics Department, Postal Code 65174, Hamedan (Iran, Islamic Republic of)], E-mail: jaleh@basu.ac.ir; Parvin, P. [Amir Kabir University of Technology, Physics Department, P.O. Box: 15875-4413, Tehran (Iran, Islamic Republic of); Laser Research Center, AEOI, P.O. Box: 11365-8486, Tehran (Iran, Islamic Republic of); Sheikh, N. [Nuclear Science and Technology Research Institute, Radiation Applications Research School, Tehran (Iran, Islamic Republic of); Zamanipour, Z. [Laser Research Center, AEOI, P.O. Box: 11365-8486, Tehran (Iran, Islamic Republic of); Sajad, B. [Azzahra University, Physics Department, Tehran (Iran, Islamic Republic of)

2007-12-15

Lasers are used to modify polymeric materials. In this work, a number of polycarbonate (PC) pieces were exposed by ArF excimer laser, 193 nm, at various UV doses from 10 to 100 J/cm{sup 2} with 50-500 mJ/pulse at 10 Hz pulse repetition rate. Morphology of PC has been investigated by scanning electron microscope (SEM) at three regimes pre-ablation, slow and fast ablation. SEM identifies that the conical defects are created on the polymer surface to grow opposite to the direction of laser irradiation. It increases the superficial absorptivity of the material dependent on the ArF laser induced conical microstructure geometry. The contact angle measurement was performed here, in order to determine the hydrophilicity of the irradiated polymer at various coherent doses. It is shown that the contact angle of PC samples which are exposed to the ArF laser significantly alters with UV dose below 7 J/cm{sup 2}.

Inhibitory effects of curcumin and capsaicin on phorbol ester-induced activation of eukaryotic transcription factors, NF-kappaB and AP-1.

Science.gov (United States)

Surh, Y J; Han, S S; Keum, Y S; Seo, H J; Lee, S S

2000-01-01

Recently, considerable attention has been focused on identifying dietary and medicinal phytochemicals that can inhibit, retard or reverse the multi-stage carcinogenesis. Spices and herbs contain phenolic substances with potent antioxidative and chemopreventive properties. Curcumin, a yellow colouring agent from turmeric and capsaicin, a pungent principle of red pepper exhibit profound anticarcinogenic and antimutagenic activities. Two well-defined eukaryotic transcription factors, nuclear factor-kappa B (NF-kappaB) and activator protein 1 (AP-1) have been implicated in pathogenesis of many human diseases including cancer. These transcription factors are known to be activated by a wide array of external stimuli, such as tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA), tumor necrosis factor, reactive oxygen species, bacterial lipopolysaccharide, and ultraviolet. In the present study, we found that topical application of TPA onto dorsal skin of female ICR mice resulted in marked activation of epidermal NF-kappaB and AP-1. Curcumin and capsaicin, when topically applied prior to TPA, significantly attenuated TPA-induced activation of each transcription factor in mouse skin. Likewise, both compounds inhibited NF-kappaB and AP-1 activation in cultured human promyelocytic leukemia (HL-60) cells stimulated with TPA. Based on these findings, it is likely that curcumin and capsaicin exert anti-tumor promotional effects through suppression of the tumor promoter-induced activation of transcription factors, NF-kappaB and AP-1.

Repression of Meiotic Genes by Antisense Transcription and by Fkh2 Transcription Factor in Schizosaccharomyces pombe

Science.gov (United States)

Chen, Huei-Mei; Rosebrock, Adam P.; Khan, Sohail R.; Futcher, Bruce; Leatherwood, Janet K.

2012-01-01

In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s) of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the “unspliced” signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression. PMID:22238674

Repression of meiotic genes by antisense transcription and by Fkh2 transcription factor in Schizosaccharomyces pombe.

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Huei-Mei Chen

Full Text Available In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the "unspliced" signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression.

Curcumin and synthetic analogs induce reactive oxygen species and decreases specificity protein (Sp) transcription factors by targeting microRNAs

International Nuclear Information System (INIS)

Gandhy, Shruti U; Kim, KyoungHyun; Larsen, Lesley; Rosengren, Rhonda J; Safe, Stephen

2012-01-01

Curcumin inhibits growth of several cancer cell lines, and studies in this laboratory in bladder and pancreatic cancer cells show that curcumin downregulates specificity protein (Sp) transcription factors Sp1, Sp3 and Sp4 and pro-oncogenic Sp-regulated genes. In this study, we investigated the anticancer activity of curcumin and several synthetic cyclohexanone and piperidine analogs in colon cancer cells. The effects of curcumin and synthetic analogs on colon cancer cell proliferation and apoptosis were determined using standardized assays. The changes in Sp proteins and Sp-regulated gene products were analysed by western blots, and real time PCR was used to determine microRNA-27a (miR-27a), miR-20a, miR-17-5p and ZBTB10 and ZBTB4 mRNA expression. The IC 50 (half-maximal) values for growth inhibition (24 hr) of colon cancer cells by curcumin and synthetic cyclohexanone and piperidine analogs of curcumin varied from 10 μM for curcumin to 0.7 μM for the most active synthetic piperidine analog RL197, which was used along with curcumin as model agents in this study. Curcumin and RL197 inhibited RKO and SW480 colon cancer cell growth and induced apoptosis, and this was accompanied by downregulation of specificity protein (Sp) transcription factors Sp1, Sp3 and Sp4 and Sp-regulated genes including the epidermal growth factor receptor (EGFR), hepatocyte growth factor receptor (c-MET), survivin, bcl-2, cyclin D1 and NFκB (p65 and p50). Curcumin and RL197 also induced reactive oxygen species (ROS), and cotreatment with the antioxidant glutathione significantly attenuated curcumin- and RL197-induced growth inhibition and downregulation of Sp1, Sp3, Sp4 and Sp-regulated genes. The mechanism of curcumin-/RL197-induced repression of Sp transcription factors was ROS-dependent and due to induction of the Sp repressors ZBTB10 and ZBTB4 and downregulation of microRNAs (miR)-27a, miR-20a and miR-17-5p that regulate these repressors. These results identify a new and highly potent

Curcumin and synthetic analogs induce reactive oxygen species and decreases specificity protein (Sp transcription factors by targeting microRNAs

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Gandhy Shruti U

2012-11-01

Full Text Available Abstract Background Curcumin inhibits growth of several cancer cell lines, and studies in this laboratory in bladder and pancreatic cancer cells show that curcumin downregulates specificity protein (Sp transcription factors Sp1, Sp3 and Sp4 and pro-oncogenic Sp-regulated genes. In this study, we investigated the anticancer activity of curcumin and several synthetic cyclohexanone and piperidine analogs in colon cancer cells. Methods The effects of curcumin and synthetic analogs on colon cancer cell proliferation and apoptosis were determined using standardized assays. The changes in Sp proteins and Sp-regulated gene products were analysed by western blots, and real time PCR was used to determine microRNA-27a (miR-27a, miR-20a, miR-17-5p and ZBTB10 and ZBTB4 mRNA expression. Results The IC50 (half-maximal values for growth inhibition (24 hr of colon cancer cells by curcumin and synthetic cyclohexanone and piperidine analogs of curcumin varied from 10 μM for curcumin to 0.7 μM for the most active synthetic piperidine analog RL197, which was used along with curcumin as model agents in this study. Curcumin and RL197 inhibited RKO and SW480 colon cancer cell growth and induced apoptosis, and this was accompanied by downregulation of specificity protein (Sp transcription factors Sp1, Sp3 and Sp4 and Sp-regulated genes including the epidermal growth factor receptor (EGFR, hepatocyte growth factor receptor (c-MET, survivin, bcl-2, cyclin D1 and NFκB (p65 and p50. Curcumin and RL197 also induced reactive oxygen species (ROS, and cotreatment with the antioxidant glutathione significantly attenuated curcumin- and RL197-induced growth inhibition and downregulation of Sp1, Sp3, Sp4 and Sp-regulated genes. The mechanism of curcumin-/RL197-induced repression of Sp transcription factors was ROS-dependent and due to induction of the Sp repressors ZBTB10 and ZBTB4 and downregulation of microRNAs (miR-27a, miR-20a and miR-17-5p that regulate these repressors

Improving plasma resistance and lowering roughness in an ArF photoresist by adding a chemical reaction inhibitor

International Nuclear Information System (INIS)

Jinnai, Butsurin; Uesugi, Takuji; Koyama, Koji; Samukawa, Seiji; Kato, Keisuke; Yasuda, Atsushi; Maeda, Shinichi; Momose, Hikaru

2010-01-01

Major challenges associated with 193 nm lithography using an ArF photoresist are low plasma resistance and roughness formation in the ArF photoresist during plasma processes. We have previously found decisive factors affecting the plasma resistance and roughness formation in an ArF photoresist: plasma resistance is determined by UV/VUV radiation, and roughness formation is dominated by chemical reactions. In this study, based on our findings on the interaction between plasma radiation species and ArF photoresist polymers, we proposed an ArF photoresist with a chemical reaction inhibitor, which can trap reactive species from the plasma, and characterized the performances of the resultant ArF photoresist through neutral beam experiments. Hindered amine light stabilizers, i.e. 4-hydroxy-2,2,6,6-tetramethyl-1-piperidinyloxy (HO-TEMPO), were used as the chemical reaction inhibitor. Etching rates of the ArF photoresist films were not dependent on the HO-TEMPO content in the irradiations without chemical reactions or under UV/VUV radiation. However, in the irradiation with chemical reactions, the etching rates of the ArF photoresist films decreased as the HO-TEMPO content increased. In addition, the surface roughness decreased with the increase in the additive amount of chemical reaction inhibitor. According to FTIR analysis, a chemical reaction inhibitor can inhibit the chemical reactions in ArF photoresist films through plasma radicals. These results indicate that a chemical reaction inhibitor is effective against chemical reactions, resulting in improved plasma resistance and less roughness in an ArF photoresist. These results also support our suggested mechanism of plasma resistance and roughness formation in an ArF photoresist.

Hypoxia induces cyclophilin B through the activation of transcription factor 6 in gastric adenocarcinoma cells.

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Jeong, Kwon; Kim, Kiyoon; Kim, Hunsung; Oh, Yoojung; Kim, Seong-Jin; Jo, Yunhee; Choe, Wonchae

2015-06-01

Hypoxia is an important form of physiological stress that induces cell death, due to the resulting endoplasmic reticulum (ER) stress, particularly in solid tumors. Although previous studies have indicated that cyclophilin B (CypB) plays a role in ER stress, there is currently no direct information supporting the mechanism of CypB involvement under hypoxic conditions. However, it has previously been demonstrated that ER stress positively regulates the expression of CypB. In the present study, it was demonstrated that CypB is transcriptionally regulated by hypoxia-mediated activation of transcription factor 6 (ATF6), an ER stress transcription factor. Subsequently, the effects of ATF6 on CypB promoter activity were investigated and an ATF6-responsive region in the promoter was identified. Hypoxia and ATF6 expression each increased CypB promoter activity. Collectively, these results demonstrate that ATF6 positively regulates the expression of CypB by binding to an ATF6-responsive region in the promoter, which may play an important role in the attenuation of apoptosis in the adaption to hypoxia. These results suggest that CypB may be a key molecule in the adaptation of cells to hypoxic conditions.

The cellular transcription factor CREB corresponds to activating transcription factor 47 (ATF-47) and forms complexes with a group of polypeptides related to ATF-43.

OpenAIRE

Hurst, H C; Masson, N; Jones, N C; Lee, K A

1990-01-01

Promoter elements containing the sequence motif CGTCA are important for a variety of inducible responses at the transcriptional level. Multiple cellular factors specifically bind to these elements and are encoded by a multigene family. Among these factors, polypeptides termed activating transcription factor 43 (ATF-43) and ATF-47 have been purified from HeLa cells and a factor referred to as cyclic AMP response element-binding protein (CREB) has been isolated from PC12 cells and rat brain. We...

High miR156 Expression Is Required for Auxin-Induced Adventitious Root Formation via MxSPL26 Independent of PINs and ARFs in Malus xiaojinensis

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Xiaozhao Xu

2017-06-01

Full Text Available Adventitious root formation is essential for the vegetative propagation of perennial woody plants. During the juvenile-to-adult phase change mediated by the microRNA156 (miR156, the adventitious rooting ability decreases dramatically in many species, including apple rootstocks. However, the mechanism underlying how miR156 affects adventitious root formation is unclear. In the present study, we showed that in the presence of the synthetic auxin indole-3-butyric acid (IBA, semi-lignified leafy cuttings from juvenile phase (Mx-J and rejuvenated (Mx-R Malus xiaojinensis trees exhibited significantly higher expression of miR156, PIN-FORMED1 (PIN1, PIN10, and rootless concerning crown and seminal roots-like (RTCS-like genes, thus resulting in higher adventitious rooting ability than those from adult phase (Mx-A trees. However, the expression of SQUAMOSA-PROMOTER BINDING PROTEIN-LIKE26 (SPL26 and some auxin response factor (ARF gene family members were substantially higher in Mx-A than in Mx-R cuttings. The expression of NbRTCS-like but not NbPINs and NbARFs varied with miR156 expression in tobacco (Nicotiana benthamiana plants transformed with 35S:MdMIR156a6 or 35S:MIM156 constructs. Overexpressing the miR156-resistant MxrSPL genes in tobacco confirmed the involvement of MxSPL20, MxSPL21&22, and MxSPL26 in adventitious root formation. Together, high expression of miR156 was necessary for auxin-induced adventitious root formation via MxSPL26, but independent of MxPINs and MxARFs expression in M. xiaojinensis leafy cuttings.

WRKY transcription factors

Science.gov (United States)

Bakshi, Madhunita; Oelmüller, Ralf

2014-01-01

WRKY transcription factors are one of the largest families of transcriptional regulators found exclusively in plants. They have diverse biological functions in plant disease resistance, abiotic stress responses, nutrient deprivation, senescence, seed and trichome development, embryogenesis, as well as additional developmental and hormone-controlled processes. WRKYs can act as transcriptional activators or repressors, in various homo- and heterodimer combinations. Here we review recent progress on the function of WRKY transcription factors in Arabidopsis and other plant species such as rice, potato, and parsley, with a special focus on abiotic, developmental, and hormone-regulated processes. PMID:24492469

Ultraviolet Radiation and the Slug Transcription Factor Induce Pro inflammatory and Immunomodulatory Mediator Expression in Melanocytes

International Nuclear Information System (INIS)

Shirley, S. H.; Kusewitt, D. F.; Grimm, E. A.

2012-01-01

Despite extensive investigation, the precise contribution of the ultraviolet radiation (UVR) component of sunlight to melanoma etiology remains unclear. UVR induces keratinocytes to secrete pro inflammatory and immunomodulatory mediators that promote inflammation and skin tumor development; expression of the slug transcription factor in keratinocytes is required for maximal production of these mediators. In the present studies we examined the possibility that UVR-exposed melanocytes also produce pro inflammatory mediators and that Slug is important in this process. Micro array studies revealed that both UVR exposure and Slug overexpression altered transcription of a variety of pro inflammatory mediators by normal human melanocytes; some of these mediators are also known to stimulate melanocyte growth and migration. There was little overlap in the spectra of cytokines produced by the two stimuli. However IL-20 was similarly induced by both stimuli and the NFκB pathway appeared to be important in both circumstances. Further exploration of UVR-induced and Slug-dependent pathways of cytokine induction in melanocytes may reveal novel targets for melanoma therapy.

Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors.

Science.gov (United States)

Perdigão, Pedro; Gaj, Thomas; Santa-Marta, Mariana; Barbas, Carlos F; Goncalves, Joao

2016-01-01

The presence of replication-competent HIV-1 -which resides mainly in resting CD4+ T cells--is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression. Here we show that engineered transcriptional modulators based on customizable transcription activator-like effector (TALE) proteins can induce gene expression from the HIV-1 long terminal repeat promoter, and that combinations of TALE transcription factors can synergistically reactivate latent viral expression in cell line models of HIV-1 latency. We further show that complementing TALE transcription factors with Vorinostat, a histone deacetylase inhibitor, enhances HIV-1 expression in latency models. Collectively, these findings demonstrate that TALE transcription factors are a potentially effective alternative to current pharmacological routes for reactivating latent virus and that combining synthetic transcriptional activators with histone deacetylase inhibitors could lead to the development of improved therapies for latent HIV-1 infection.

Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors.

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Pedro Perdigão

Full Text Available The presence of replication-competent HIV-1 -which resides mainly in resting CD4+ T cells--is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression. Here we show that engineered transcriptional modulators based on customizable transcription activator-like effector (TALE proteins can induce gene expression from the HIV-1 long terminal repeat promoter, and that combinations of TALE transcription factors can synergistically reactivate latent viral expression in cell line models of HIV-1 latency. We further show that complementing TALE transcription factors with Vorinostat, a histone deacetylase inhibitor, enhances HIV-1 expression in latency models. Collectively, these findings demonstrate that TALE transcription factors are a potentially effective alternative to current pharmacological routes for reactivating latent virus and that combining synthetic transcriptional activators with histone deacetylase inhibitors could lead to the development of improved therapies for latent HIV-1 infection.

ARF and ATM/ATR cooperate in p53-mediated apoptosis upon oncogenic stress

International Nuclear Information System (INIS)

Pauklin, Siim; Kristjuhan, Arnold; Maimets, Toivo; Jaks, Viljar

2005-01-01

Induction of apoptosis is pivotal for eliminating cells with damaged DNA or deregulated proliferation. We show that tumor suppressor ARF and ATM/ATR kinase pathways cooperate in the induction of apoptosis in response to elevated expression of c-myc, β-catenin or human papilloma virus E7 oncogenes. Overexpression of oncogenes leads to the formation of phosphorylated H2AX foci, induction of Rad51 protein levels and ATM/ATR-dependent phosphorylation of p53. Inhibition of ATM/ATR kinases abolishes both induction of Rad51 and phosphorylation of p53, and remarkably reduces the level of apoptosis induced by co-expression of oncogenes and ARF. However, the induction of apoptosis is downregulated in p53-/- cells and does not depend on activities of ATM/ATR kinases, indicating that efficient induction of apoptosis by oncogene activation depends on coordinated action of ARF and ATM/ATR pathways in the regulation of p53

Identification and network-enabled characterization of auxin response factor genes in Medicago truncatula

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David J. Burks

2016-12-01

Full Text Available The Auxin Response Factor (ARF family of transcription factors is an important regulator of environmental response and symbiotic nodulation in the legume Medicago truncatula. While previous studies have identified members of this family, a recent spurt in gene expression data coupled with genome update and reannotation calls for a reassessment of the prevalence of ARF genes and their interaction networks in M. truncatula. We performed a comprehensive analysis of the M. truncatula genome and transcriptome that entailed search for novel ARF genes and the co-expression networks. Our investigation revealed 8 novel M. truncatula ARF (MtARF genes, of the total 22 identified, and uncovered novel gene co-expression networks as well. Furthermore, the topological clustering and single enrichment analysis of several network models revealed the roles of individual members of the MtARF family in nitrogen regulation, nodule initiation, and post-embryonic development through a specialized protein packaging and secretory pathway. In summary, this study not just shines new light on an important gene family, but also provides a guideline for identification of new members of gene families and their functional characterization through network analyses.

The Arabidopsis transcription factor ABIG1 relays ABA signaled growth inhibition and drought induced senescence.

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Liu, Tie; Longhurst, Adam D; Talavera-Rauh, Franklin; Hokin, Samuel A; Barton, M Kathryn

2016-10-04

Drought inhibits plant growth and can also induce premature senescence. Here we identify a transcription factor, ABA INSENSITIVE GROWTH 1 (ABIG1) required for abscisic acid (ABA) mediated growth inhibition, but not for stomatal closure. ABIG1 mRNA levels are increased both in response to drought and in response to ABA treatment. When treated with ABA, abig1 mutants remain greener and produce more leaves than comparable wild-type plants. When challenged with drought, abig1 mutants have fewer yellow, senesced leaves than wild-type. Induction of ABIG1 transcription mimics ABA treatment and regulates a set of genes implicated in stress responses. We propose a model in which drought acts through ABA to increase ABIG1 transcription which in turn restricts new shoot growth and promotes leaf senescence. The results have implications for plant breeding: the existence of a mutant that is both ABA resistant and drought resistant points to new strategies for isolating drought resistant genetic varieties.

The transcription factor KLF2 restrains CD4⁺ T follicular helper cell differentiation.

Science.gov (United States)

Lee, June-Yong; Skon, Cara N; Lee, You Jeong; Oh, Soohwan; Taylor, Justin J; Malhotra, Deepali; Jenkins, Marc K; Rosenfeld, M Geoffrey; Hogquist, Kristin A; Jameson, Stephen C

2015-02-17

T follicular helper (Tfh) cells are essential for efficient B cell responses, yet the factors that regulate differentiation of this CD4(+) T cell subset are incompletely understood. Here we found that the KLF2 transcription factor serves to restrain Tfh cell generation. Induced KLF2 deficiency in activated CD4(+) T cells led to increased Tfh cell generation and B cell priming, whereas KLF2 overexpression prevented Tfh cell production. KLF2 promotes expression of the trafficking receptor S1PR1, and S1PR1 downregulation is essential for efficient Tfh cell production. However, KLF2 also induced expression of the transcription factor Blimp-1, which repressed transcription factor Bcl-6 and thereby impaired Tfh cell differentiation. Furthermore, KLF2 induced expression of the transcription factors T-bet and GATA3 and enhanced Th1 differentiation. Hence, our data indicate KLF2 is pivotal for coordinating CD4(+) T cell differentiation through two distinct and complementary mechanisms: via control of T cell localization and by regulation of lineage-defining transcription factors. Copyright © 2015 Elsevier Inc. All rights reserved.

Secretion of soluble vascular endothelial growth factor receptor 1 (sVEGFR1/sFlt1 requires Arf1, Arf6, and Rab11 GTPases.

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Jae-Joon Jung

Full Text Available The soluble form of vascular endothelial growth factor receptor 1 (sVEGFR-1/sFlt1 is generated by alternative splicing of the FLT1 gene. Secretion of sFlt1 from endothelial cells plays an important role in blood vessel sprouting and morphogenesis. However, excess sFlt1 secretion is associated with diseases such as preeclampsia and chronic kidney disease. To date, the secretory transport process involved in the secretion of sFlt1 is poorly understood. In the present study, we investigated the itinerary of sFlt1 trafficking along the secretory pathway. To understand the timecourse of sFlt1 secretion, endothelial cells stably expressing sFlt1 were metabolically radiolabeled with [(35S]-methionine and cysteine. Our results indicate that after initial synthesis the levels of secreted [(35S]-sFlt1 in the extracellular medium peaks at 8 hours. Treatment with brefeldin A (BFA, a drug which blocks trafficking between the endoplasmic reticulum (ER and the Golgi complex, inhibited extracellular release of sFlt1 suggesting that ER to Golgi and intra-Golgi trafficking of sFlt1 are essential for its secretion. Furthermore, we show that ectopic expression of dominant-negative mutant forms of Arf1, Arf6, and Rab11 as well as siRNA-mediated knockdown of these GTPases block secretion of sFlt1 during normoxic and hypoxic conditions suggesting role for these small GTPases. This work is the first to report role of regulatory proteins involved in sFlt1 trafficking along the secretory pathway and may provide insights and new molecular targets for the modulation of sFlt-1 release during physiological and pathological conditions.

C/EBPβ represses p53 to promote cell survival downstream of DNA damage independent of oncogenic Ras and p19Arf

Science.gov (United States)

Ewing, SJ; Zhu, S; Zhu, F; House, JS; Smart, RC

2013-01-01

CCAAT/enhancer-binding protein-β (C/EBPβ) is a mediator of cell survival and tumorigenesis. When C/EBPβ−/− mice are treated with carcinogens that produce oncogenic Ras mutations in keratinocytes, they respond with abnormally elevated keratinocyte apoptosis and a block in skin tumorigenesis. Although this aberrant carcinogen-induced apoptosis results from abnormal upregulation of p53, it is not known whether upregulated p53 results from oncogenic Ras and its ability to induce p19Arf and/or activate DNA-damage response pathways or from direct carcinogen-induced DNA damage. We report that p19Arf is dramatically elevated in C/EBPβ−/− epidermis and that C/EBPβ represses a p19Arf promoter reporter. To determine whether p19Arf is responsible for the proapoptotic phenotype in C/EBPβ−/− mice, C/EBPβ−/−;p19Arf−/− mice were generated. C/EBPβ−/−;p19Arf−/− mice responded to carcinogen treatment with increased p53 and apoptosis, indicating p19Arf is not essential. To ascertain whether oncogenic Ras activation induces aberrant p53 and apoptosis in C/EBPβ−/− epidermis, we generated K14-ER:Ras; C/EBPβ−/− mice. Oncogenic Ras activation induced by 4-hydroxytamoxifen did not produce increased p53 or apoptosis. Finally, when C/EBPβ−/− mice were treated with differing types of DNA-damaging agents, including alkylating chemotherapeutic agents, they displayed aberrant levels of p53 and apoptosis. These results indicate that C/EBPβ represses p53 to promote cell survival downstream of DNA damage and suggest that inhibition of C/EBPβ may be a target for cancer cotherapy to increase the efficacy of alkylating chemotherapeutic agents. PMID:18636078

Interconversion of two GDP-bound conformations and their selection in an Arf-family small G protein.

Science.gov (United States)

Okamura, Hideyasu; Nishikiori, Masaki; Xiang, Hongyu; Ishikawa, Masayuki; Katoh, Etsuko

2011-07-13

ADP-ribosylation factor (Arf) and other Arf-family small G proteins participate in many cellular functions via their characteristic GTP/GDP conformational cycles, during which a nucleotide(∗)Mg(2+)-binding site communicates with a remote N-terminal helix. However, the conformational interplay between the nucleotides, the helix, the protein core, and Mg(2+) has not been fully delineated. Herein, we report a study of the dynamics of an Arf-family protein, Arl8, under various conditions by means of NMR relaxation spectroscopy. The data indicated that, when GDP is bound, the protein core, which does not include the N-terminal helix, reversibly transition between an Arf-family GDP form and another conformation that resembles the Arf-family GTP form. Additionally, we found that the N-terminal helix and Mg(2+), respectively, stabilize the aforementioned former and latter conformations in a population-shift manner. Given the dynamics of the conformational changes, we can describe the Arl8 GTP/GDP cycle in terms of an energy diagram. Copyright © 2011 Elsevier Ltd. All rights reserved.

Transcription factor Sox4 is required for PUMA-mediated apoptosis induced by histone deacetylase inhibitor, TSA.

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Jang, Sang-Min; Kang, Eun-Jin; Kim, Jung-Woong; Kim, Chul-Hong; An, Joo-Hee; Choi, Kyung-Hee

2013-08-23

PUMA is a crucial regulator of apoptotic cell death mediated by p53-dependent and p53-independent mechanisms. In many cancer cells, PUMA expression is induced in response to DNA-damaging reagent in a p53-dependent manner. However, few studies have investigated transcription factors that lead to the induction of PUMA expression via p53-independent apoptotic signaling. In this study, we found that the transcription factor Sox4 increased PUMA expression in response to trichostatin A (TSA), a histone deacetylase inhibitor in the p53-null human lung cancer cell line H1299. Ectopic expression of Sox4 led to the induction of PUMA expression at the mRNA and protein levels, and TSA-mediated up-regulation of PUMA transcription was repressed by the knockdown of Sox4. Using luciferase assays and chromatin immunoprecipitation, we also determined that Sox4 recruits p300 on the PUMA promoter region and increases PUMA gene expression in response to TSA treatment. Taken together, these results suggest that Sox4 is required for p53-independent apoptotic cell death mediated by PUMA induction via TSA treatment. Crown Copyright © 2013. Published by Elsevier Inc. All rights reserved.

Mapping temperature-induced conformational changes in the Escherichia coli heat shock transcription factor sigma 32 by amide hydrogen exchange

DEFF Research Database (Denmark)

Rist, Wolfgang; Jørgensen, Thomas J D; Roepstorff, Peter

2003-01-01

Stress conditions such as heat shock alter the transcriptional profile in all organisms. In Escherichia coli the heat shock transcription factor, sigma 32, out-competes upon temperature up-shift the housekeeping sigma-factor, sigma 70, for binding to core RNA polymerase and initiates heat shock...... gene transcription. To investigate possible heat-induced conformational changes in sigma 32 we performed amide hydrogen (H/D) exchange experiments under optimal growth and heat shock conditions combined with mass spectrometry. We found a rapid exchange of around 220 of the 294 amide hydrogens at 37...... degrees C, indicating that sigma 32 adopts a highly flexible structure. At 42 degrees C we observed a slow correlated exchange of 30 additional amide hydrogens and localized it to a helix-loop-helix motif within domain sigma 2 that is responsible for the recognition of the -10 region in heat shock...

ARF6, PI3-kinase and host cell actin cytoskeleton in Toxoplasma gondii cell invasion

International Nuclear Information System (INIS)

Vieira da Silva, Claudio; Alves da Silva, Erika; Costa Cruz, Mario; Chavrier, Philippe; Arruda Mortara, Renato

2009-01-01

Toxoplasma gondii infects a variety of different cell types in a range of different hosts. Host cell invasion by T. gondii occurs by active penetration of the host cell, a process previously described as independent of host actin polymerization. Also, the parasitophorous vacuole has been shown to resist fusion with endocytic and exocytic pathways of the host cell. ADP-ribosylation factor-6 (ARF6) belongs to the ARF family of small GTP-binding proteins. ARF6 regulates membrane trafficking and actin cytoskeleton rearrangements at the plasma membrane. Here, we have observed that ARF6 is recruited to the parasitophorous vacuole of tachyzoites of T. gondii RH strain and it also plays an important role in the parasite cell invasion with activation of PI3-kinase and recruitment of PIP 2 and PIP 3 to the parasitophorous vacuole of invading parasites. Moreover, it was verified that maintenance of host cell actin cytoskeleton integrity is important to parasite invasion.

Nucleocytoplasmic shuttling of transcription factors

DEFF Research Database (Denmark)

Cartwright, P; Helin, K

2000-01-01

To elicit the transcriptional response following intra- or extracellular stimuli, the signals need to be transmitted to their site of action within the nucleus. The nucleocytoplasmic shuttling of transcription factors is a mechanism mediating this process. The activation and inactivation...... of the transcriptional response is essential for cells to progress through the cell cycle in a normal manner. The involvement of cytoplasmic and nuclear accessory molecules, and the general nuclear membrane transport components, are essential for this process. Although nuclear import and export for different...... transcription factor families are regulated by similar mechanisms, there are several differences that allow for the specific activation of each transcription factor. This review discusses the general import and export pathways found to be common amongst many different transcription factors, and highlights...

Mitochondrial transcription factor A protects human retinal ...

African Journals Online (AJOL)

Purpose: To investigate the impact of mitochondrial transcription factor A (TFAM), as a modulator of NF-κB, on proliferation of hypoxia-induced human retinal endothelial cell (HREC), and the probable mechanism. Methods: After exposure to hypoxia (1 % O2) for 5 days, cell proliferation and cell cycle of HREC were ...

A novel zinc finger protein Zfp277 mediates transcriptional repression of the Ink4a/arf locus through polycomb repressive complex 1

DEFF Research Database (Denmark)

Negishi, Masamitsu; Saraya, Atsunori; Mochizuki, Shinobu

2010-01-01

. METHODOLOGY/PRINCIPAL FINDINGS: We examined the function of Zinc finger domain-containing protein 277 (Zfp277), a novel zinc finger protein that interacts with the PcG protein Bmi1. Zfp277 binds to the Ink4a/Arf locus in a Bmi1-independent manner and interacts with polycomb repressor complex (PRC) 1 through...... is essential for the recruitment of PRC1 to the Ink4a/Arf locus. Our findings also highlight dynamic regulation of both Zfp277 and PcG proteins by the oxidative stress pathways....

Natural antioxidants exhibit chemopreventive characteristics through the regulation of CNC b-Zip transcription factors in estrogen-induced breast carcinogenesis.

Science.gov (United States)

Chatterjee, Anwesha; Ronghe, Amruta; Singh, Bhupendra; Bhat, Nimee K; Chen, Jie; Bhat, Hari K

2014-12-01

The objective of the present study was to characterize the role of resveratrol (Res) and vitamin C (VC) in prevention of estrogen-induced breast cancer through regulation of cap "n"collar (CNC) b-zip transcription factors. Human breast epithelial cell line MCF-10A was treated with 17β-estradiol (E2) and VC or Res with or without E2. mRNA and protein expression levels of CNC b-zip transcription factors nuclear factor erythroid 2-related factor 1 (Nrf1), nuclear factor erythroid 2 related factor 2 (Nrf2), nuclear factor erythroid 2 related factor 3 (Nrf3), and Nrf2-regulated antioxidant enzymes superoxide dismutase 3 (SOD3) and quinone oxidoreductase 1 (NQO1) were quantified. The treatment with E2 suppressed, whereas VC and Res prevented E2-mediated decrease in the expression levels of SOD3, NQO1, Nrf2 mRNA, and protein in MCF-10A cells. The treatment with E2, Res, or VC significantly increased mRNA and protein expression levels of Nrf1. 17β-Estradiol treatment significantly increased but VC or Res decreased Nrf3 mRNA and protein expression levels. Our studies demonstrate that estrogen-induced breast cancer might be prevented through upregulation of antioxidant enzymes via Nrf-dependent pathways. © 2014 Wiley Periodicals, Inc.

Transcriptional regulation by competing transcription factor modules.

Directory of Open Access Journals (Sweden)

Rutger Hermsen

2006-12-01

Full Text Available Gene regulatory networks lie at the heart of cellular computation. In these networks, intracellular and extracellular signals are integrated by transcription factors, which control the expression of transcription units by binding to cis-regulatory regions on the DNA. The designs of both eukaryotic and prokaryotic cis-regulatory regions are usually highly complex. They frequently consist of both repetitive and overlapping transcription factor binding sites. To unravel the design principles of these promoter architectures, we have designed in silico prokaryotic transcriptional logic gates with predefined input-output relations using an evolutionary algorithm. The resulting cis-regulatory designs are often composed of modules that consist of tandem arrays of binding sites to which the transcription factors bind cooperatively. Moreover, these modules often overlap with each other, leading to competition between them. Our analysis thus identifies a new signal integration motif that is based upon the interplay between intramodular cooperativity and intermodular competition. We show that this signal integration mechanism drastically enhances the capacity of cis-regulatory domains to integrate signals. Our results provide a possible explanation for the complexity of promoter architectures and could be used for the rational design of synthetic gene circuits.

Activation of nuclear transcription factor-kappaB in mouse brain induced by a simulated microgravity environment

Science.gov (United States)

Wise, Kimberly C.; Manna, Sunil K.; Yamauchi, Keiko; Ramesh, Vani; Wilson, Bobby L.; Thomas, Renard L.; Sarkar, Shubhashish; Kulkarni, Anil D.; Pellis, Neil R.; Ramesh, Govindarajan T.

2005-01-01

Microgravity induces inflammatory responses and modulates immune functions that may increase oxidative stress. Exposure to a microgravity environment induces adverse neurological effects; however, there is little research exploring the etiology of these effects resulting from exposure to such an environment. It is also known that spaceflight is associated with increase in oxidative stress; however, this phenomenon has not been reproduced in land-based simulated microgravity models. In this study, an attempt has been made to show the induction of reactive oxygen species (ROS) in mice brain, using ground-based microgravity simulator. Increased ROS was observed in brain stem and frontal cortex with concomitant decrease in glutathione, on exposing mice to simulated microgravity for 7 d. Oxidative stress-induced activation of nuclear factor-kappaB was observed in all the regions of the brain. Moreover, mitogen-activated protein kinase kinase was phosphorylated equally in all regions of the brain exposed to simulated microgravity. These results suggest that exposure of brain to simulated microgravity can induce expression of certain transcription factors, and these have been earlier argued to be oxidative stress dependent.

WAVE regulatory complex activation by cooperating GTPases Arf and Rac1

DEFF Research Database (Denmark)

Koronakis, Vassilis; Hume, Peter J; Humphreys, Daniel

2011-01-01

The WAVE regulatory complex (WRC) is a critical element in the control of actin polymerization at the eukaryotic cell membrane, but how WRC is activated remains uncertain. While Rho GTPase Rac1 can bind and activate WRC in vitro, this interaction is of low affinity, suggesting other factors may...... be important. By reconstituting WAVE-dependent actin assembly on membrane-coated beads in mammalian cell extracts, we found that Rac1 was not sufficient to engender bead motility, and we uncovered a key requirement for Arf GTPases. In vitro, Rac1 and Arf1 were individually able to bind weakly to recombinant...... be central components in WAVE signalling, acting directly, alongside Rac1....

Structural Insights into Arl1-Mediated Targeting of the Arf-GEF BIG1 to the trans-Golgi

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Antonio Galindo

2016-07-01

Full Text Available The GTPase Arf1 is the major regulator of vesicle traffic at both the cis- and trans-Golgi. Arf1 is activated at the cis-Golgi by the guanine nucleotide exchange factor (GEF GBF1 and at the trans-Golgi by the related GEF BIG1 or its paralog, BIG2. The trans-Golgi-specific targeting of BIG1 and BIG2 depends on the Arf-like GTPase Arl1. We find that Arl1 binds to the dimerization and cyclophilin binding (DCB domain in BIG1 and report a crystal structure of human Arl1 bound to this domain. Residues in the DCB domain that bind Arl1 are required for BIG1 to locate to the Golgi in vivo. DCB domain-binding residues in Arl1 have a distinct conformation from those in known Arl1-effector complexes, and this plasticity allows Arl1 to interact with different effectors of unrelated structure. The findings provide structural insight into how Arf1 GEFs, and hence active Arf1, achieve their correct subcellular distribution.

Arf4 is required for Mammalian development but dispensable for ciliary assembly.

Directory of Open Access Journals (Sweden)

John A Follit

2014-02-01

Full Text Available The primary cilium is a sensory organelle, defects in which cause a wide range of human diseases including retinal degeneration, polycystic kidney disease and birth defects. The sensory functions of cilia require specific receptors to be targeted to the ciliary subdomain of the plasma membrane. Arf4 has been proposed to sort cargo destined for the cilium at the Golgi complex and deemed a key regulator of ciliary protein trafficking. In this work, we show that Arf4 binds to the ciliary targeting sequence (CTS of fibrocystin. Knockdown of Arf4 indicates that it is not absolutely required for trafficking of the fibrocystin CTS to cilia as steady-state CTS levels are unaffected. However, we did observe a delay in delivery of newly synthesized CTS from the Golgi complex to the cilium when Arf4 was reduced. Arf4 mutant mice are embryonic lethal and die at mid-gestation shortly after node formation. Nodal cilia appeared normal and functioned properly to break left-right symmetry in Arf4 mutant embryos. At this stage of development Arf4 expression is highest in the visceral endoderm but we did not detect cilia on these cells. In the visceral endoderm, the lack of Arf4 caused defects in cell structure and apical protein localization. This work suggests that while Arf4 is not required for ciliary assembly, it is important for the efficient transport of fibrocystin to cilia, and also plays critical roles in non-ciliary processes.

Eukaryotic transcription factors

DEFF Research Database (Denmark)

Staby, Lasse; O'Shea, Charlotte; Willemoës, Martin

2017-01-01

Gene-specific transcription factors (TFs) are key regulatory components of signaling pathways, controlling, for example, cell growth, development, and stress responses. Their biological functions are determined by their molecular structures, as exemplified by their structured DNA-binding domains...... regions with function-related, short sequence motifs and molecular recognition features with structural propensities. This review focuses on molecular aspects of TFs, which represent paradigms of ID-related features. Through specific examples, we review how the ID-associated flexibility of TFs enables....... It is furthermore emphasized how classic biochemical concepts like allostery, conformational selection, induced fit, and feedback regulation are undergoing a revival with the appreciation of ID. The review also describes the most recent advances based on computational simulations of ID-based interaction mechanisms...

DNA damage-inducible transcripts in mammalian cells

International Nuclear Information System (INIS)

Fornace, A.J. Jr.; Alamo, I. Jr.; Hollander, M.C.

1988-01-01

Hybridization subtraction at low ratios of RNA to cDNA was used to enrich for the cDNA of transcripts increased in Chinese hamster cells after UV irradiation. Forty-nine different cDNA clones were isolated. Most coded for nonabundant transcripts rapidly induced 2- to 10-fold after UV irradiation. Only 2 of the 20 cDNA clones sequenced matched known sequences (metallothionein I and II). The predicted amino acid sequence of one cDNA had two localized areas of homology with the rat helix-destabilizing protein. These areas of homology were at the two DNA-binding sites of this nucleic acid single-strand-binding protein. The induced transcripts were separated into two general classes. Class I transcripts were induced by UV radiation and not by the alkylating agent methyl methanesulfonate. Class II transcripts were induced by UV radiation and by methyl methanesulfonate. Many class II transcripts were induced also by H2O2 and various alkylating agents but not by heat shock, phorbol 12-tetradecanoate 13-acetate, or DNA-damaging agents which do not produce high levels of base damage. Since many of the cDNA clones coded for transcripts which were induced rapidly and only by certain types of DNA-damaging agents, their induction is likely a specific response to such damage rather than a general response to cell injury

Identification and Expression Profiling of the Auxin Response Factors in Capsicum annuum L. under Abiotic Stress and Hormone Treatments

Directory of Open Access Journals (Sweden)

Chenliang Yu

2017-12-01

Full Text Available Auxin response factors (ARFs play important roles in regulating plant growth and development and response to environmental stress. An exhaustive analysis of the CaARF family was performed using the latest publicly available genome for pepper (Capsicum annuum L.. In total, 22 non-redundant CaARF gene family members in six classes were analyzed, including chromosome locations, gene structures, conserved motifs of proteins, phylogenetic relationships and Subcellular localization. Phylogenetic analysis of the ARFs from pepper (Capsicum annuum L., tomato (Solanum lycopersicum L., Arabidopsis and rice (Oryza sativa L. revealed both similarity and divergence between the four ARF families, and aided in predicting biological functions of the CaARFs. Furthermore, expression profiling of CaARFs was obtained in various organs and tissues using quantitative real-time RT-PCR (qRT-PCR. Expression analysis of these genes was also conducted with various hormones and abiotic treatments using qRT-PCR. Most CaARF genes were regulated by exogenous hormone treatments at the transcriptional level, and many CaARF genes were altered by abiotic stress. Systematic analysis of CaARF genes is imperative to elucidate the roles of CaARF family members in mediating auxin signaling in the adaptation of pepper to a challenging environment.

Histone H1 and Chromosomal Protein HMGN2 Regulate Prolactin-induced STAT5 Transcription Factor Recruitment and Function in Breast Cancer Cells.

Science.gov (United States)

Schauwecker, Suzanne M; Kim, J Julie; Licht, Jonathan D; Clevenger, Charles V

2017-02-10

The hormone prolactin (PRL) contributes to breast cancer pathogenesis through various signaling pathways, one of the most notable being the JAK2/signal transducer and activator of transcription 5 (STAT5) pathway. PRL-induced activation of the transcription factor STAT5 results in the up-regulation of numerous genes implicated in breast cancer pathogenesis. However, the molecular mechanisms that enable STAT5 to access the promoters of these genes are not well understood. Here, we show that PRL signaling induces chromatin decompaction at promoter DNA, corresponding with STAT5 binding. The chromatin-modifying protein high mobility group nucleosomal binding domain 2 (HMGN2) specifically promotes STAT5 accessibility at promoter DNA by facilitating the dissociation of the linker histone H1 in response to PRL. Knockdown of H1 rescues the decrease in PRL-induced transcription following HMGN2 knockdown, and it does so by allowing increased STAT5 recruitment. Moreover, H1 and STAT5 are shown to function antagonistically in regulating PRL-induced transcription as well as breast cancer cell biology. While reduced STAT5 activation results in decreased PRL-induced transcription and cell proliferation, knockdown of H1 rescues both of these effects. Taken together, we elucidate a novel mechanism whereby the linker histone H1 prevents STAT5 binding at promoter DNA, and the PRL-induced dissociation of H1 mediated by HMGN2 is necessary to allow full STAT5 recruitment and promote the biological effects of PRL signaling. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Analysis of the stress-inducible transcription factor SsNAC23 in sugarcane plants

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Renata Fava Ditt

2011-08-01

Full Text Available Stresses such as cold and drought can impair plant yield and induce a highly complex array of responses. Sugarcane (Saccharum spp. is cultivated in tropical and subtropical areas and is considered a cold-sensitive plant. We previously showed that cold stress induces the expression of several genes in in vitro sugarcane plantlets. Here we characterize one of those genes, SsNAC23, a member of the NAC family of plant-specific transcription factors, which are induced by low temperature and other stresses in several plant species. The expression of SsNAC23 was induced in sugarcane plants exposed to low temperatures (4ºC. With the aim of further understanding the regulatory network in response to stress, we used the yeast two-hybrid system to identify sugarcane proteins that interact with SsNAC23. Using SsNAC23 as bait, we screened a cDNA expression library of sugarcane plants submitted to 4ºC for 48 h. Several interacting partners were identified, including stress-related proteins, increasing our knowledge on how sugarcane plants respond to cold stress. One of these interacting partners, a thioredoxin h1, offers insights into the regulation of SsNAC23 activity.

ARF6-dependent regulation of P2Y receptor traffic and function in human platelets.

Science.gov (United States)

Kanamarlapudi, Venkateswarlu; Owens, Sian E; Saha, Keya; Pope, Robert J; Mundell, Stuart J

2012-01-01

Adenosine diphosphate (ADP) is a critical regulator of platelet activation, mediating its actions through two G protein-coupled receptors, the P2Y(1) and P2Y(12) purinoceptors. Recently, we demonstrated that P2Y(1) and P2Y(12) purinoceptor activities are rapidly and reversibly modulated in human platelets, revealing that the underlying mechanism requires receptor internalization and subsequent trafficking as an essential part of this process. In this study we investigated the role of the small GTP-binding protein ADP ribosylation factor 6 (ARF6) in the internalization and function of P2Y(1) and P2Y(12) purinoceptors in human platelets. ARF6 has been implicated in the internalization of a number of GPCRs, although its precise molecular mechanism in this process remains unclear. In this study we show that activation of either P2Y(1) or P2Y(12) purinoceptors can stimulate ARF6 activity. Further blockade of ARF6 function either in cell lines or human platelets blocks P2Y purinoceptor internalization. This blockade of receptor internalization attenuates receptor resensitization. Furthermore, we demonstrate that Nm23-H1, a nucleoside diphosphate (NDP) kinase regulated by ARF6 which facilitates dynamin-dependent fission of coated vesicles during endocytosis, is also required for P2Y purinoceptor internalization. These data describe a novel function of ARF6 in the internalization of P2Y purinoceptors and demonstrate the integral importance of this small GTPase upon platelet ADP receptor function.

ARF6-dependent regulation of P2Y receptor traffic and function in human platelets.

Directory of Open Access Journals (Sweden)

Venkateswarlu Kanamarlapudi

Full Text Available Adenosine diphosphate (ADP is a critical regulator of platelet activation, mediating its actions through two G protein-coupled receptors, the P2Y(1 and P2Y(12 purinoceptors. Recently, we demonstrated that P2Y(1 and P2Y(12 purinoceptor activities are rapidly and reversibly modulated in human platelets, revealing that the underlying mechanism requires receptor internalization and subsequent trafficking as an essential part of this process. In this study we investigated the role of the small GTP-binding protein ADP ribosylation factor 6 (ARF6 in the internalization and function of P2Y(1 and P2Y(12 purinoceptors in human platelets. ARF6 has been implicated in the internalization of a number of GPCRs, although its precise molecular mechanism in this process remains unclear. In this study we show that activation of either P2Y(1 or P2Y(12 purinoceptors can stimulate ARF6 activity. Further blockade of ARF6 function either in cell lines or human platelets blocks P2Y purinoceptor internalization. This blockade of receptor internalization attenuates receptor resensitization. Furthermore, we demonstrate that Nm23-H1, a nucleoside diphosphate (NDP kinase regulated by ARF6 which facilitates dynamin-dependent fission of coated vesicles during endocytosis, is also required for P2Y purinoceptor internalization. These data describe a novel function of ARF6 in the internalization of P2Y purinoceptors and demonstrate the integral importance of this small GTPase upon platelet ADP receptor function.

Hypoxia-Inducible Factor 1 Is an Inductor of Transcription Factor Activating Protein 2 Epsilon Expression during Chondrogenic Differentiation

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Stephan Niebler

2015-01-01

Full Text Available The transcription factor AP-2ε (activating enhancer-binding protein epsilon is expressed in cartilage of humans and mice. However, knowledge about regulatory mechanisms influencing AP-2ε expression is limited. Using quantitative real time PCR, we detected a significant increase in AP-2ε mRNA expression comparing initial and late stages of chondrogenic differentiation processes in vitro and in vivo. Interestingly, in these samples the expression pattern of the prominent hypoxia marker gene angiopoietin-like 4 (Angptl4 strongly correlated with that of AP-2ε suggesting that hypoxia might represent an external regulator of AP-2ε expression in mammals. In order to show this, experiments directly targeting the activity of hypoxia-inducible factor-1 (HIF1, the complex mediating responses to oxygen deprivation, were performed. While the HIF1-activating compounds 2,2′-dipyridyl and desferrioxamine resulted in significantly enhanced mRNA concentration of AP-2ε, siRNA against HIF1α led to a significantly reduced expression rate of AP-2ε. Additionally, we detected a significant upregulation of the AP-2ε mRNA level after oxygen deprivation. In sum, these different experimental approaches revealed a novel role for the HIF1 complex in the regulation of the AP-2ε gene in cartilaginous cells and underlined the important role of hypoxia as an important external regulatory stimulus during chondrogenic differentiation modulating the expression of downstream transcription factors.

The Journey of a Transcription Factor

DEFF Research Database (Denmark)

Pireyre, Marie

Plants have developed astonishing networks regulating their metabolism to adapt to their environment. The complexity of these networks is illustrated by the expansion of families of regulators such as transcription factors in the plant kingdom. Transcription factors specifically impact...... transcriptional networks by integrating exogenous and endogenous stimuli and regulating gene expression accordingly. Regulation of transcription factors and their activation is thus highly important to modulate the transcriptional programs and increase fitness of the plant in a given environment. Plant metabolism....... The biosynthetic machinery of GLS is governed by interplay of six MYB and three bHLH transcription factors. MYB28, MYB29 and MYB76 regulate methionine-derived GLS, and MYB51, MYB34 and MYB122 regulate tryptophan-derived GLS. The three bHLH transcription factors MYC2, MYC3 and MYC4 physically interact with all six...

Role of ARF6 in internalization of metal-binding proteins, metallothionein and transferrin, and cadmium-metallothionein toxicity in kidney proximal tubule cells

International Nuclear Information System (INIS)

Wolff, Natascha A.; Lee, Wing-Kee; Abouhamed, Marouan; Thevenod, Frank

2008-01-01

Filtered metal-protein complexes, such as cadmium-metallothionein-1 (CdMT-1) or transferrin (Tf) are apically endocytosed partly via megalin/cubilin by kidney proximal tubule (PT) cells where CdMT-1 internalization causes apoptosis. Small GTPase ARF (ADP-ribosylation factor) proteins regulate endocytosis and vesicular trafficking. We investigated roles of ARF6, which has been shown to be involved in internalization of ligands and endocytic trafficking in PT cells, following MT-1/CdMT-1 and Tf uptake by PT cells. WKPT-0293 Cl.2 cells derived from rat PT S1 segment were transfected with hemagglutinin-tagged wild-type (ARF6-WT) or dominant negative (ARF6-T27N) forms of ARF6. Using immunofluorescence, endogenous ARF6 was associated with the plasma membrane (PM) as well as juxtanuclear and co-localized with Rab5a and Rab11 involved in early and recycling endosomal trafficking. Immunofluorescence staining of megalin showed reduced surface labelling in ARF6 dominant negative (ARF6-DN) cells. Intracellular Alexa Fluor 546-conjugated MT-1 uptake was reduced in ARF6-DN cells and CdMT-1 (14.8 μM for 24 h) toxicity was significantly attenuated from 27.3 ± 3.9% in ARF6-WT to 11.1 ± 4.0% in ARF6-DN cells (n = 6, P < 0.02). Moreover, reduced Alexa Fluor 546-conjugated Tf uptake was observed in ARF-DN cells (75.0 ± 4.6% versus 3.9 ± 3.9% of ARF6-WT cells, n = 3, P < 0.01) and/or remained near the PM (89.3 ± 5. 6% versus 45.2 ± 14.3% of ARF6-WT cells, n = 3, P < 0.05). In conclusion, the data support roles for ARF6 in receptor-mediated endocytosis and trafficking of MT-1/Tf to endosomes/lysosomes and CdMT-1 toxicity of PT cells

Genomewide identification and expression analysis of the ARF gene ...

Indian Academy of Sciences (India)

Figure 1. Phylogenetic relation of apple ARF genes. The phylogenetic tree was constructed based on a complete protein sequence align- ment of MdARFs by the neighbour-joining method with bootstrapping analysis (1000 replicates). The scale bar represents 0.05 amino acid substitutions per site. Paralogous gene pairs ...

Regulation of basophil and mast cell development by transcription factors

Directory of Open Access Journals (Sweden)

Haruka Sasaki

2016-04-01

Full Text Available Basophils and mast cells play important roles in host defense against parasitic infections and allergic responses. Several progenitor populations, either shared or specific, for basophils and/or mast cells have been identified, thus elucidating the developmental pathways of these cells. Multiple transcription factors essential for their development and the relationships between them have been also revealed. For example, IRF8 induces GATA2 expression to promote the generation of both basophils and mast cells. The STAT5-GATA2 axis induces C/EBPα and MITF expression, facilitating the differentiation into basophils and mast cells, respectively. In addition, C/EBPα and MITF mutually suppress each other's expression. This review provides an overview of recent advances in our understanding of how transcription factors regulate the development of basophils and mast cells.

Supra-optimal expression of the cold-regulated OsMyb4 transcription factor in transgenic rice changes the complexity of transcriptional network with major effects on stress tolerance and panicle development

KAUST Repository

Park, Myoungryoul

2010-09-28

The R2R3-type OsMyb4 transcription factor of rice has been shown to play a role in the regulation of osmotic adjustment in heterologous overexpression studies. However, the exact composition and organization of its underlying transcriptional network has not been established to be a robust tool for stress tolerance enhancement by regulon engineering. OsMyb4 network was dissected based on commonalities between the global chilling stress transcriptome and the transcriptome configured by OsMyb4 overexpression. OsMyb4 controls a hierarchical network comprised of several regulatory sub-clusters associated with cellular defense and rescue, metabolism and development. It regulates target genes either directly or indirectly through intermediary MYB, ERF, bZIP, NAC, ARF and CCAAT-HAP transcription factors. Regulatory sub-clusters have different combinations of MYB-like, GCC-box-like, ERD1-box-like, ABRE-like, G-box-like, as1/ocs/TGA-like, AuxRE-like, gibberellic acid response element (GARE)-like and JAre-like cis-elements. Cold-dependent network activity enhanced cellular antioxidant capacity through radical scavenging mechanisms and increased activities of phenylpropanoid and isoprenoid metabolic processes involving various abscisic acid (ABA), jasmonic acid (JA), salicylic acid (SA), ethylene and reactive oxygen species (ROS) responsive genes. OsMyb4 network is independent of drought response element binding protein/C-repeat binding factor (DREB/CBF) and its sub-regulons operate with possible co-regulators including nuclear factor-Y. Because of its upstream position in the network hierarchy, OsMyb4 functions quantitatively and pleiotrophically. Supra-optimal expression causes misexpression of alternative targets with costly trade-offs to panicle development. © 2010 Blackwell Publishing Ltd.

Supra-optimal expression of the cold-regulated OsMyb4 transcription factor in transgenic rice changes the complexity of transcriptional network with major effects on stress tolerance and panicle development.

Science.gov (United States)

Park, Myoung-Ryoul; Yun, Kil-Young; Mohanty, Bijayalaxmi; Herath, Venura; Xu, Fuyu; Wijaya, Edward; Bajic, Vladimir B; Yun, Song-Joong; De Los Reyes, Benildo G

2010-12-01

The R2R3-type OsMyb4 transcription factor of rice has been shown to play a role in the regulation of osmotic adjustment in heterologous overexpression studies. However, the exact composition and organization of its underlying transcriptional network has not been established to be a robust tool for stress tolerance enhancement by regulon engineering. OsMyb4 network was dissected based on commonalities between the global chilling stress transcriptome and the transcriptome configured by OsMyb4 overexpression. OsMyb4 controls a hierarchical network comprised of several regulatory sub-clusters associated with cellular defense and rescue, metabolism and development. It regulates target genes either directly or indirectly through intermediary MYB, ERF, bZIP, NAC, ARF and CCAAT-HAP transcription factors. Regulatory sub-clusters have different combinations of MYB-like, GCC-box-like, ERD1-box-like, ABRE-like, G-box-like, as1/ocs/TGA-like, AuxRE-like, gibberellic acid response element (GARE)-like and JAre-like cis-elements. Cold-dependent network activity enhanced cellular antioxidant capacity through radical scavenging mechanisms and increased activities of phenylpropanoid and isoprenoid metabolic processes involving various abscisic acid (ABA), jasmonic acid (JA), salicylic acid (SA), ethylene and reactive oxygen species (ROS) responsive genes. OsMyb4 network is independent of drought response element binding protein/C-repeat binding factor (DREB/CBF) and its sub-regulons operate with possible co-regulators including nuclear factor-Y. Because of its upstream position in the network hierarchy, OsMyb4 functions quantitatively and pleiotrophically. Supra-optimal expression causes misexpression of alternative targets with costly trade-offs to panicle development. © 2010 Blackwell Publishing Ltd.

EWS-FLI1 inhibits TNFα-induced NFκB-dependent transcription in Ewing sarcoma cells

International Nuclear Information System (INIS)

Lagirand-Cantaloube, Julie; Laud, Karine; Lilienbaum, Alain; Tirode, Franck; Delattre, Olivier; Auclair, Christian; Kryszke, Marie-Helene

2010-01-01

Research highlights: → EWS-FLI1 interferes with TNF-induced activation of NFκB in Ewing sarcoma cells. → EWS-FLI1 knockdown in Ewing sarcoma cells increases TNF-induced NFκB binding to DNA. → EWS-FLI1 reduces TNF-stimulated NFκB-dependent transcriptional activation. → Constitutive NFκB activity is not affected by EWS-FLI1. → EWS-FLI1 physically interacts with NFκB p65 in vivo. -- Abstract: Ewing sarcoma is primarily caused by a t(11;22) chromosomal translocation encoding the EWS-FLI1 fusion protein. To exert its oncogenic function, EWS-FLI1 acts as an aberrant transcription factor, broadly altering the gene expression profile of tumor cells. Nuclear factor-kappaB (NFκB) is a tightly regulated transcription factor controlling cell survival, proliferation and differentiation, as well as tumorigenesis. NFκB activity is very low in unstimulated Ewing sarcoma cells, but can be induced in response to tumor necrosis factor (TNF). We wondered whether NFκB activity could be modulated by EWS-FLI1 in Ewing sarcoma. Using a knockdown approach in Ewing sarcoma cells, we demonstrated that EWS-FLI1 has no influence on NFκB basal activity, but impairs TNF-induced NFκB-driven transcription, at least in part through inhibition of NFκB binding to DNA. We detected an in vivo physical interaction between the fusion protein and NFκB p65, which could mediate these effects. Our findings suggest that, besides directly controlling the activity of its primary target promoters, EWS-FLI1 can also indirectly influence gene expression in tumor cells by modulating the activity of key transcription factors such as NFκB.

Betulinic acid inhibits colon cancer cell and tumor growth and induces proteasome-dependent and -independent downregulation of specificity proteins (Sp transcription factors

Directory of Open Access Journals (Sweden)

Pathi Satya

2011-08-01

Full Text Available Abstract Background Betulinic acid (BA inhibits growth of several cancer cell lines and tumors and the effects of BA have been attributed to its mitochondriotoxicity and inhibition of multiple pro-oncogenic factors. Previous studies show that BA induces proteasome-dependent degradation of specificity protein (Sp transcription factors Sp1, Sp3 and Sp4 in prostate cancer cells and this study focused on the mechanism of action of BA in colon cancer cells. Methods The effects of BA on colon cancer cell proliferation and apoptosis and tumor growth in vivo were determined using standardized assays. The effects of BA on Sp proteins and Sp-regulated gene products were analyzed by western blots, and real time PCR was used to determine microRNA-27a (miR-27a and ZBTB10 mRNA expression. Results BA inhibited growth and induced apoptosis in RKO and SW480 colon cancer cells and inhibited tumor growth in athymic nude mice bearing RKO cells as xenograft. BA also decreased expression of Sp1, Sp3 and Sp4 transcription factors which are overexpressed in colon cancer cells and decreased levels of several Sp-regulated genes including survivin, vascular endothelial growth factor, p65 sub-unit of NFκB, epidermal growth factor receptor, cyclin D1, and pituitary tumor transforming gene-1. The mechanism of action of BA was dependent on cell context, since BA induced proteasome-dependent and proteasome-independent downregulation of Sp1, Sp3 and Sp4 in SW480 and RKO cells, respectively. In RKO cells, the mechanism of BA-induced repression of Sp1, Sp3 and Sp4 was due to induction of reactive oxygen species (ROS, ROS-mediated repression of microRNA-27a, and induction of the Sp repressor gene ZBTB10. Conclusions These results suggest that the anticancer activity of BA in colon cancer cells is due, in part, to downregulation of Sp1, Sp3 and Sp4 transcription factors; however, the mechanism of this response is cell context-dependent.

Betulinic acid inhibits colon cancer cell and tumor growth and induces proteasome-dependent and -independent downregulation of specificity proteins (Sp) transcription factors

International Nuclear Information System (INIS)

Chintharlapalli, Sudhakar; Papineni, Sabitha; Lei, Ping; Pathi, Satya; Safe, Stephen

2011-01-01

Betulinic acid (BA) inhibits growth of several cancer cell lines and tumors and the effects of BA have been attributed to its mitochondriotoxicity and inhibition of multiple pro-oncogenic factors. Previous studies show that BA induces proteasome-dependent degradation of specificity protein (Sp) transcription factors Sp1, Sp3 and Sp4 in prostate cancer cells and this study focused on the mechanism of action of BA in colon cancer cells. The effects of BA on colon cancer cell proliferation and apoptosis and tumor growth in vivo were determined using standardized assays. The effects of BA on Sp proteins and Sp-regulated gene products were analyzed by western blots, and real time PCR was used to determine microRNA-27a (miR-27a) and ZBTB10 mRNA expression. BA inhibited growth and induced apoptosis in RKO and SW480 colon cancer cells and inhibited tumor growth in athymic nude mice bearing RKO cells as xenograft. BA also decreased expression of Sp1, Sp3 and Sp4 transcription factors which are overexpressed in colon cancer cells and decreased levels of several Sp-regulated genes including survivin, vascular endothelial growth factor, p65 sub-unit of NFκB, epidermal growth factor receptor, cyclin D1, and pituitary tumor transforming gene-1. The mechanism of action of BA was dependent on cell context, since BA induced proteasome-dependent and proteasome-independent downregulation of Sp1, Sp3 and Sp4 in SW480 and RKO cells, respectively. In RKO cells, the mechanism of BA-induced repression of Sp1, Sp3 and Sp4 was due to induction of reactive oxygen species (ROS), ROS-mediated repression of microRNA-27a, and induction of the Sp repressor gene ZBTB10. These results suggest that the anticancer activity of BA in colon cancer cells is due, in part, to downregulation of Sp1, Sp3 and Sp4 transcription factors; however, the mechanism of this response is cell context-dependent

Sucrose-induced anthocyanin accumulation in vegetative tissue of Petunia plants requires anthocyanin regulatory transcription factors.

Science.gov (United States)

Ai, Trinh Ngoc; Naing, Aung Htay; Arun, Muthukrishnan; Lim, Sun-Hyung; Kim, Chang Kil

2016-11-01

The effects of three different sucrose concentrations on plant growth and anthocyanin accumulation were examined in non-transgenic (NT) and transgenic (T 2 ) specimens of the Petunia hybrida cultivar 'Mirage rose' that carried the anthocyanin regulatory transcription factors B-Peru+mPAP1 or RsMYB1. Anthocyanin accumulation was not observed in NT plants in any treatments, whereas a range of anthocyanin accumulation was observed in transgenic plants. The anthocyanin content detected in transgenic plants expressing the anthocyanin regulatory transcription factors (B-Peru+mPAP1 or RsMYB1) was higher than that in NT plants. In addition, increasing sucrose concentration strongly enhanced anthocyanin content as shown by quantitative real-time polymerase chain reaction (qRT-PCR) analysis, wherein increased concentrations of sucrose enhanced transcript levels of the transcription factors that are responsible for the induction of biosynthetic genes involved in anthocyanin synthesis; this pattern was not observed in NT plants. In addition, sucrose affected plant growth, although the effects were different between NT and transgenic plants. Taken together, the application of sucrose could enhance anthocyanin production in vegetative tissue of transgenic Petunia carrying anthocyanin regulatory transcription factors, and this study provides insights about interactive effects of sucrose and transcription factors in anthocyanin biosynthesis in the transgenic plant. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

An EAR-motif-containing ERF transcription factor affects herbivore-induced signaling, defense and resistance in rice.

Science.gov (United States)

Lu, Jing; Ju, Hongping; Zhou, Guoxin; Zhu, Chuanshu; Erb, Matthias; Wang, Xiaopeng; Wang, Peng; Lou, Yonggen

2011-11-01

Ethylene responsive factors (ERFs) are a large family of plant-specific transcription factors that are involved in the regulation of plant development and stress responses. However, little to nothing is known about their role in herbivore-induced defense. We discovered a nucleus-localized ERF gene in rice (Oryza sativa), OsERF3, that was rapidly up-regulated in response to feeding by the rice striped stem borer (SSB) Chilo suppressalis. Antisense and over-expression of OsERF3 revealed that it positively affects transcript levels of two mitogen-activated protein kinases (MAPKs) and two WRKY genes as well as concentrations of jasmonate (JA), salicylate (SA) and the activity of trypsin protease inhibitors (TrypPIs). OsERF3 was also found to mediate the resistance of rice to SSB. On the other hand, OsERF3 was slightly suppressed by the rice brown planthopper (BPH) Nilaparvata lugens (Stål) and increased susceptibility to this piercing sucking insect, possibly by suppressing H(2)O(2) biosynthesis. We propose that OsERF3 affects early components of herbivore-induced defense responses by suppressing MAPK repressors and modulating JA, SA, ethylene and H(2)O(2) pathways as well as plant resistance. Our results also illustrate that OsERF3 acts as a central switch that gears the plant's metabolism towards an appropriate response to chewing or piercing/sucking insects. © 2011 The Authors. The Plant Journal © 2011 Blackwell Publishing Ltd.

Activation of transcription factor AP-2 mediates UVA radiation- and singlet oxygen-induced expression of the human intercellular adhesion molecule 1 gene

International Nuclear Information System (INIS)

Grether-Beck, S.; Olaizola-Horn, S.; Schmitt, H.; Grewe, M.

1996-01-01

UVA radiation is the major component of the UV solar spectrum that reaches the earth, and the therapeutic application of UVA radiation is increasing in medicine. Analysis of the cellular effects of UVA radiation has revealed that exposure of human cells to UVA radiation at physiological doses leads to increased gene expression and that this UVA response is primarily mediated through the generation of singlet oxygen. In this study, the mechanisms by which UVA radiation induces transcriptional activation of the human intercellular adhesion molecule 1 (ICAM-1) were examined. UVA radiation was capable of inducing activation of the human ICAM-1 promoter and increasing OCAM-1 mRNA and protein expression. These UVA radiation effects were inhibited by singlet oxygen quenchers, augmented by enhancement of singlet oxygen life-time, and mimicked in unirradiated cells by a singlet oxygen-generating system. UVA radiation as well as singlet oxygen-induced ICAM-1 promoter activation required activation of the transcription factor AP-2. Accordingly, both stimuli activated AP-2, and deletion of the putative AP-2-binding site abrogated ICAM-1 promoter activation in this system. This study identified the AP-2 site as the UVA radiation- and singlet oxygen-responsive element of the human ICAM-1 gene. The capacity of UVA radiation and/or singlet oxygen to induce human gene expression through activation of AP-2 indicates a previously unrecognized role of this transcription factor in the mammalian stress response. 38 refs., 3 figs., 3 tabs

Endogenous target mimics down-regulate miR160 mediation of ARF10, -16 and -17 cleavage during somatic embryogenesis in Dimocarpus longan Lour.

Directory of Open Access Journals (Sweden)

yuling elin

2015-11-01

Full Text Available MicroRNA160 plays a critical role in plant development by negatively regulating the auxin response factors ARF10, -16 and -17. However, the ways in which miR160 expression is regulated at the transcriptional level, and how miR160 interacts with its targets during plant embryo development, remain unknown. Here, we studied the regulatory relationships among endogenous target mimics (eTMs, and miR160 and its targets, and their involvement in hormone signaling and somatic embryogenesis (SE in Dimocarpus longan. We identified miR160 family members and isolated the miR160 precursor, primary transcript, and promoter. The promoter contained cis-acting elements responsive to stimuli such as light, abscisic acid, salicylic acid and heat stress. The pri-miR160 was down-regulated in response to salicylic acid but up-regulated by gibberellic acid, ethylene, and methyl jasmonate treatment, suggesting that pri-miR160 was associated with hormone transduction. Dlo-miR160a, -a* and -d* reached expression peaks in torpedo-shaped embryos, globular embryos and cotyledonary embryos, respectively, but were barely detectable in embryogenic callus. This suggests that they have expression-related and functional diversity, especially during the middle and later developmental stages of SE. Four potential eTMs for miR160 were identified. Two of them, glucan endo-1,3-beta- glucosidase-like protein 2-like and calpain-type cysteine protease DEK1, were confirmed to control the corresponding dlo-miR160a* expression level. This suggests that they may function to abolish the binding between dlo-miR160a* and its targets. These two eTMs also participated in auxin and ABA signal transduction. DlARF10, -16, and -17 targeting by dlo-miR160a was confirmed; their expression levels were higher in friable-embryogenic callus and incomplete compact pro-embryogenic cultures and responded to 2,4-D, suggesting they may play a major role in the early stages of longan SE dependent on 2,4-D. The e

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

International Nuclear Information System (INIS)

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

2010-01-01

Research highlights: → FMDV L pro inhibits poly(I:C)-induced IFN-α1/β mRNA expression. → L pro inhibits MDA5-mediated activation of the IFN-α1/β promoter. → L pro significantly reduced the transcription of multiple IRF-responsive genes. → L pro inhibits IFN-α1/β promoter activation by decreasing IRF-3/7 in protein levels. → The ability to process eIF-4G of L pro is not necessary to inhibit IFN-α1/β activation. -- Abstract: The leader proteinase (L pro ) of foot-and-mouth disease virus (FMDV) has been identified as an interferon-β (IFN-β) antagonist that disrupts the integrity of transcription factor nuclear factor κB (NF-κB). In this study, we showed that the reduction of double stranded RNA (dsRNA)-induced IFN-α1/β expression caused by L pro was also associated with a decrease of interferon regulatory factor 3/7 (IRF-3/7) in protein levels, two critical transcription factors for activation of IFN-α/β. Furthermore, overexpression of L pro significantly reduced the transcription of multiple IRF-responsive genes including 2',5'-OAS, ISG54, IP-10, and RANTES. Screening L pro mutants indicated that the ability to process eIF-4G of L pro is not required for suppressing dsRNA-induced activation of the IFN-α1/β promoter and decreasing IRF-3/7 expression. Taken together, our results demonstrate that, in addition to disrupting NF-κB, L pro also decreases IRF-3/7 expression to suppress dsRNA-induced type I IFN production, suggesting multiple strategies used by FMDV to counteract the immune response to viral infection.

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

Energy Technology Data Exchange (ETDEWEB)

Wang, Dang; Fang, Liurong; Luo, Rui; Ye, Rui; Fang, Ying; Xie, Lilan; Chen, Huanchun [Division of Animal Infectious Diseases, State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); Xiao, Shaobo, E-mail: shaoboxiao@yahoo.com [Division of Animal Infectious Diseases, State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China)

2010-08-13

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

Mitochondrial Reactive Oxygen Species Trigger Hypoxia-Induced Transcription

Science.gov (United States)

Chandel, N. S.; Maltepe, E.; Goldwasser, E.; Mathieu, C. E.; Simon, M. C.; Schumacker, P. T.

1998-09-01

Transcriptional activation of erythropoietin, glycolytic enzymes, and vascular endothelial growth factor occurs during hypoxia or in response to cobalt chloride (CoCl2) in Hep3B cells. However, neither the mechanism of cellular O2 sensing nor that of cobalt is fully understood. We tested whether mitochondria act as O2 sensors during hypoxia and whether hypoxia and cobalt activate transcription by increasing generation of reactive oxygen species (ROS). Results show (i) wild-type Hep3B cells increase ROS generation during hypoxia (1.5% O2) or CoCl2 incubation, (ii) Hep3B cells depleted of mitochondrial DNA (ρ 0 cells) fail to respire, fail to activate mRNA for erythropoietin, glycolytic enzymes, or vascular endothelial growth factor during hypoxia, and fail to increase ROS generation during hypoxia; (iii) ρ 0 cells increase ROS generation in response to CoCl2 and retain the ability to induce expression of these genes; and (iv) the antioxidants pyrrolidine dithiocarbamate and ebselen abolish transcriptional activation of these genes during hypoxia or CoCl2 in wild-type cells, and abolish the response to CoCl2 in ρ 0 cells. Thus, hypoxia activates transcription via a mitochondria-dependent signaling process involving increased ROS, whereas CoCl2 activates transcription by stimulating ROS generation via a mitochondria-independent mechanism.

Involvement of WRKY Transcription Factors in Abscisic-Acid-Induced Cold Tolerance of Banana Fruit.

Science.gov (United States)

Luo, Dong-Lan; Ba, Liang-Jie; Shan, Wei; Kuang, Jian-Fei; Lu, Wang-Jin; Chen, Jian-Ye

2017-05-10

Phytohormone abscisic acid (ABA) and plant-specific WRKY transcription factors (TFs) have been implicated to play important roles in various stress responses. The involvement of WRKY TFs in ABA-mediated cold tolerance of economical fruits, such as banana fruit, however remains largely unknown. Here, we reported that ABA application could induce expressions of ABA biosynthesis-related genes MaNCED1 and MaNCED2, increase endogenous ABA contents, and thereby enhance cold tolerance in banana fruit. Four banana fruit WRKY TFs, designated as MaWRKY31, MaWRKY33, MaWRKY60, and MaWRKY71, were identified and characterized. All four of these MaWRKYs were nuclear-localized and displayed transactivation activities. Their expressions were induced by ABA treatment during cold storage. More importantly, the gel mobility shift assay and transient expression analysis revealed that MaWRKY31, MaWRKY33, MaWRKY60, and MaWRKY71 directly bound to the W-box elements in MaNCED1 and MaNCED2 promoters and activated their expressions. Taken together, our findings demonstrate that banana fruit WRKY TFs are involved in ABA-induced cold tolerance by, at least in part, increasing ABA levels via directly activating NECD expressions.

BDNF restores the expression of Jun and Fos inducible transcription factors in the rat brain following repetitive electroconvulsive seizures.

Science.gov (United States)

Hsieh, T F; Simler, S; Vergnes, M; Gass, P; Marescaux, C; Wiegand, S J; Zimmermann, M; Herdegen, T

1998-01-01

The expression of inducible transcription factors was studied following repetitive electroconvulsive seizures (ECS), c-Fos, c-Jun, JunB, and JunD immunoreactivities were investigated following a single (1 x ECS) or repetitive ECS evoked once per day for 4, 5, or 10 days (4 x ECS, 5 x ECS, or 10 x ECS). Animals were killed 3 or 12 h following the last ECS. Three hours after 1 x ECS, c-Fos was expressed throughout the cortex and hippocampus. After 5 x ECS and 10 x ECS, c-Fos was reexpressed in the CA4 area, but was completely absent in the other hippocampal areas and cortex. In these areas, c-Fos became only reinducible when the time lag between two ECS stimuli was 5 days. In contrast to c-Fos, intense JunB expression was inducible in the cortex and hippocampus, but not CA4 subfield, after 1 x ECS, 5 x ECS, and 10 x ECS. Repetitive ECS did not effect c-Jun and JunD expression. In a second model of systemic excitation of the brain, repetitive daily injection of kainic acid for 4 days completely failed to express c-Fos, c-Jun, and JunB after the last application whereas injection of kainic acid once per week did not alter the strong expressions compared to a single application of kainic acid. In order to study the maintenance of c-Fos expression during repetitive seizures, brain-derived neurotrophic factor (BDNF) was applied in parallel for 5 or 10 days via miniosmotic pumps and permanent cannula targeted at the hippocampus or the parietal cortex. Infusion of BDNF completely reinduced c-Fos expression during 5 x ECS or 10 x ECS in the cortex ipsilaterally to the cannula and, to a less extent, also increased the expression of c-Jun and JunB when compared to saline-treated controls. BDNF had no effect on the expression patterns in the hippocampus. ECS with or without BDNF infusion did not change the expression patterns of the constitutive transcription factors ATF-2, CREB, and SRF. These data demonstrate that various transcription factors substantially differ in their

An ABRE promoter sequence is involved in osmotic stress-responsive expression of the DREB2A gene, which encodes a transcription factor regulating drought-inducible genes in Arabidopsis.

Science.gov (United States)

Kim, June-Sik; Mizoi, Junya; Yoshida, Takuya; Fujita, Yasunari; Nakajima, Jun; Ohori, Teppei; Todaka, Daisuke; Nakashima, Kazuo; Hirayama, Takashi; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

2011-12-01

In plants, osmotic stress-responsive transcriptional regulation depends mainly on two major classes of cis-acting elements found in the promoter regions of stress-inducible genes: ABA-responsive elements (ABREs) and dehydration-responsive elements (DREs). ABRE has been shown to perceive ABA-mediated osmotic stress signals, whereas DRE is known to be involved in an ABA-independent pathway. Previously, we reported that the transcription factor DRE-BINDING PROTEIN 2A (DREB2A) regulates DRE-mediated transcription of target genes under osmotic stress conditions in Arabidopsis (Arabidopsis thaliana). However, the transcriptional regulation of DREB2A itself remains largely uncharacterized. To elucidate the transcriptional mechanism associated with the DREB2A gene under osmotic stress conditions, we generated a series of truncated and base-substituted variants of the DREB2A promoter and evaluated their transcriptional activities individually. We found that both ABRE and coupling element 3 (CE3)-like sequences located approximately -100 bp from the transcriptional initiation site are necessary for the dehydration-responsive expression of DREB2A. Coupling our transient expression analyses with yeast one-hybrid and chromatin immunoprecipitation (ChIP) assays indicated that the ABRE-BINDING PROTEIN 1 (AREB1), AREB2 and ABRE-BINDING FACTOR 3 (ABF3) bZIP transcription factors can bind to and activate the DREB2A promoter in an ABRE-dependent manner. Exogenous ABA application induced only a modest accumulation of the DREB2A transcript when compared with the osmotic stress treatment. However, the osmotic stress-induced DREB2A expression was found to be markedly impaired in several ABA-deficient and ABA-insensitive mutants. These results suggest that in addition to an ABA-independent pathway, the ABA-dependent pathway plays a positive role in the osmotic stress-responsive expression of DREB2A.

Transcript-specific effects of adrenalectomy on seizure-induced BDNF expression in rat hippocampus

DEFF Research Database (Denmark)

Lauterborn, J C; Poulsen, F R; Stinis, C T

1998-01-01

Activity-induced brain-derived neurotrophic factor (BDNF) expression is negatively modulated by circulating adrenal steroids. The rat BDNF gene gives rise to four major transcript forms that each contain a unique 5' exon (I-IV) and a common 3' exon (V) that codes for BDNF protein. Exon-specific i......Activity-induced brain-derived neurotrophic factor (BDNF) expression is negatively modulated by circulating adrenal steroids. The rat BDNF gene gives rise to four major transcript forms that each contain a unique 5' exon (I-IV) and a common 3' exon (V) that codes for BDNF protein. Exon...... and in exon II-containing mRNA with 30-days survival. In the dentate gyrus granule cells, adrenalectomy markedly potentiated increases in exon I and II cRNA labeling, but not increases in exon III and IV cRNA labeling, elicited by one hippocampal afterdischarge. Similarly, for the granule cells and CA1...... no effect on exon IV-containing mRNA content. These results demonstrate that the negative effects of adrenal hormones on activity-induced BDNF expression are by far the greatest for transcripts containing exons I and II. Together with evidence for region-specific transcript expression, these results suggest...

TG-interacting factor transcriptionally induced by AKT/FOXO3A is a negative regulator that antagonizes arsenic trioxide-induced cancer cell apoptosis

International Nuclear Information System (INIS)

Liu, Zi-Miao; Tseng, Hong-Yu; Cheng, Ya-Ling; Yeh, Bi-Wen; Wu, Wen-Jeng; Huang, Huei-Sheng

2015-01-01

Arsenic trioxide (ATO) is a multi-target drug approved by the Food and Drug Administration as the first-line chemotherapeutic agent for the treatment of acute promyelocytic leukemia. In addition, several clinical trials are being conducted with arsenic-based drugs for the treatment of other hematological malignancies and solid tumors. However, ATO's modest clinical efficacy on some cancers, and potential toxic effects on humans have been reported. Determining how best to reduce these adverse effects while increasing its therapeutic efficacy is obviously a critical issue. Previously, we demonstrated that the JNK-induced complex formation of phosphorylated c-Jun and TG-interacting factor (TGIF) antagonizes ERK-induced cyclin-dependent kinase inhibitor CDKN1A (p21 WAF1/CIP1 ) expression and resultant apoptosis in response to ATO in A431 cells. Surprisingly, at low-concentrations (0.1–0.2 μM), ATO increased cellular proliferation, migration and invasion, involving TGIF expression, however, at high-concentrations (5–20 μM), ATO induced cell apoptosis. Using a promoter analysis, TGIF was transcriptionally regulated by ATO at the FOXO3A binding site (− 1486 to − 1479 bp) via the c-Src/EGFR/AKT pathway. Stable overexpression of TGIF promoted advancing the cell cycle into the S phase, and attenuated 20 μM ATO-induced apoptosis. Furthermore, blockage of the AKT pathway enhanced ATO-induced CDKN1A expression and resultant apoptosis in cancer cells, but overexpression of AKT1 inhibited CDKN1A expression. Therefore, we suggest that TGIF is transcriptionally regulated by the c-Src/EGFR/AKT pathway, which plays a role as a negative regulator in antagonizing ATO-induced CDKN1A expression and resultant apoptosis. Suppression of these antagonistic effects might be a promising therapeutic strategy toward improving clinical efficacy of ATO. - Highlights: • ATO-induced biphasic survival responses of cancer cells depend on low- or high-concentrations. • TGIF mediates

TG-interacting factor transcriptionally induced by AKT/FOXO3A is a negative regulator that antagonizes arsenic trioxide-induced cancer cell apoptosis

Energy Technology Data Exchange (ETDEWEB)

Liu, Zi-Miao; Tseng, Hong-Yu; Cheng, Ya-Ling [Department of Medical Laboratory Science and Biotechnology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan (China); Yeh, Bi-Wen [Department of Medical Laboratory Science and Biotechnology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan (China); Department of Urology, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Department of Urology, School of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Wu, Wen-Jeng [Department of Urology, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Department of Urology, School of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Huang, Huei-Sheng, E-mail: huanghs@mail.ncku.edu.tw [Department of Medical Laboratory Science and Biotechnology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan (China)

2015-05-15

Arsenic trioxide (ATO) is a multi-target drug approved by the Food and Drug Administration as the first-line chemotherapeutic agent for the treatment of acute promyelocytic leukemia. In addition, several clinical trials are being conducted with arsenic-based drugs for the treatment of other hematological malignancies and solid tumors. However, ATO's modest clinical efficacy on some cancers, and potential toxic effects on humans have been reported. Determining how best to reduce these adverse effects while increasing its therapeutic efficacy is obviously a critical issue. Previously, we demonstrated that the JNK-induced complex formation of phosphorylated c-Jun and TG-interacting factor (TGIF) antagonizes ERK-induced cyclin-dependent kinase inhibitor CDKN1A (p21{sup WAF1/CIP1}) expression and resultant apoptosis in response to ATO in A431 cells. Surprisingly, at low-concentrations (0.1–0.2 μM), ATO increased cellular proliferation, migration and invasion, involving TGIF expression, however, at high-concentrations (5–20 μM), ATO induced cell apoptosis. Using a promoter analysis, TGIF was transcriptionally regulated by ATO at the FOXO3A binding site (− 1486 to − 1479 bp) via the c-Src/EGFR/AKT pathway. Stable overexpression of TGIF promoted advancing the cell cycle into the S phase, and attenuated 20 μM ATO-induced apoptosis. Furthermore, blockage of the AKT pathway enhanced ATO-induced CDKN1A expression and resultant apoptosis in cancer cells, but overexpression of AKT1 inhibited CDKN1A expression. Therefore, we suggest that TGIF is transcriptionally regulated by the c-Src/EGFR/AKT pathway, which plays a role as a negative regulator in antagonizing ATO-induced CDKN1A expression and resultant apoptosis. Suppression of these antagonistic effects might be a promising therapeutic strategy toward improving clinical efficacy of ATO. - Highlights: • ATO-induced biphasic survival responses of cancer cells depend on low- or high-concentrations. • TGIF

Transcriptional profiling of Medicago truncatula under salt stress identified a novel CBF transcription factor MtCBF4 that plays an important role in abiotic stress responses

Directory of Open Access Journals (Sweden)

Su Zhen

2011-07-01

Full Text Available Abstract Background Salt stress hinders the growth of plants and reduces crop production worldwide. However, different plant species might possess different adaptive mechanisms to mitigate salt stress. We conducted a detailed pathway analysis of transcriptional dynamics in the roots of Medicago truncatula seedlings under salt stress and selected a transcription factor gene, MtCBF4, for experimental validation. Results A microarray experiment was conducted using root samples collected 6, 24, and 48 h after application of 180 mM NaCl. Analysis of 11 statistically significant expression profiles revealed different behaviors between primary and secondary metabolism pathways in response to external stress. Secondary metabolism that helps to maintain osmotic balance was induced. One of the highly induced transcription factor genes was successfully cloned, and was named MtCBF4. Phylogenetic analysis revealed that MtCBF4, which belongs to the AP2-EREBP transcription factor family, is a novel member of the CBF transcription factor in M. truncatula. MtCBF4 is shown to be a nuclear-localized protein. Expression of MtCBF4 in M. truncatula was induced by most of the abiotic stresses, including salt, drought, cold, and abscisic acid, suggesting crosstalk between these abiotic stresses. Transgenic Arabidopsis over-expressing MtCBF4 enhanced tolerance to drought and salt stress, and activated expression of downstream genes that contain DRE elements. Over-expression of MtCBF4 in M. truncatula also enhanced salt tolerance and induced expression level of corresponding downstream genes. Conclusion Comprehensive transcriptomic analysis revealed complex mechanisms exist in plants in response to salt stress. The novel transcription factor gene MtCBF4 identified here played an important role in response to abiotic stresses, indicating that it might be a good candidate gene for genetic improvement to produce stress-tolerant plants.

Transcriptional profiling of Medicago truncatula under salt stress identified a novel CBF transcription factor MtCBF4 that plays an important role in abiotic stress responses

Science.gov (United States)

2011-01-01

Background Salt stress hinders the growth of plants and reduces crop production worldwide. However, different plant species might possess different adaptive mechanisms to mitigate salt stress. We conducted a detailed pathway analysis of transcriptional dynamics in the roots of Medicago truncatula seedlings under salt stress and selected a transcription factor gene, MtCBF4, for experimental validation. Results A microarray experiment was conducted using root samples collected 6, 24, and 48 h after application of 180 mM NaCl. Analysis of 11 statistically significant expression profiles revealed different behaviors between primary and secondary metabolism pathways in response to external stress. Secondary metabolism that helps to maintain osmotic balance was induced. One of the highly induced transcription factor genes was successfully cloned, and was named MtCBF4. Phylogenetic analysis revealed that MtCBF4, which belongs to the AP2-EREBP transcription factor family, is a novel member of the CBF transcription factor in M. truncatula. MtCBF4 is shown to be a nuclear-localized protein. Expression of MtCBF4 in M. truncatula was induced by most of the abiotic stresses, including salt, drought, cold, and abscisic acid, suggesting crosstalk between these abiotic stresses. Transgenic Arabidopsis over-expressing MtCBF4 enhanced tolerance to drought and salt stress, and activated expression of downstream genes that contain DRE elements. Over-expression of MtCBF4 in M. truncatula also enhanced salt tolerance and induced expression level of corresponding downstream genes. Conclusion Comprehensive transcriptomic analysis revealed complex mechanisms exist in plants in response to salt stress. The novel transcription factor gene MtCBF4 identified here played an important role in response to abiotic stresses, indicating that it might be a good candidate gene for genetic improvement to produce stress-tolerant plants. PMID:21718548

Resveratrol via sirtuin-1 downregulates RE1-silencing transcription factor (REST) expression preventing PCB-95-induced neuronal cell death.

Science.gov (United States)

Guida, Natascia; Laudati, Giusy; Anzilotti, Serenella; Secondo, Agnese; Montuori, Paolo; Di Renzo, Gianfranco; Canzoniero, Lorella M T; Formisano, Luigi

2015-11-01

Resveratrol (3,5,4'-trihydroxystilbene) (RSV), a polyphenol widely present in plants, exerts a neuroprotective function in several neurological conditions; it is an activator of class III histone deacetylase sirtuin1 (SIRT1), a crucial regulator in the pathophysiology of neurodegenerative diseases. By contrast, the RE1-silencing transcription factor (REST) is involved in the neurotoxic effects following exposure to polychlorinated biphenyl (PCB) mixture A1254. The present study investigated the effects of RSV-induced activation of SIRT1 on REST expression in SH-SY5Y cells. Further, we investigated the possible relationship between the non-dioxin-like (NDL) PCB-95 and REST through SIRT1 to regulate neuronal death in rat cortical neurons. Our results revealed that RSV significantly decreased REST gene and protein levels in a dose- and time-dependent manner. Interestingly, overexpression of SIRT1 reduced REST expression, whereas EX-527, an inhibitor of SIRT1, increased REST expression and blocked RSV-induced REST downregulation. These results suggest that RSV downregulates REST through SIRT1. In addition, RSV enhanced activator protein 1 (AP-1) transcription factor c-Jun expression and its binding to the REST promoter gene. Indeed, c-Jun knockdown reverted RSV-induced REST downregulation. Intriguingly, in SH-SY5Y cells and rat cortical neurons the NDL PCB-95 induced necrotic cell death in a concentration-dependent manner by increasing REST mRNA and protein expression. In addition, SIRT1 knockdown blocked RSV-induced neuroprotection in rat cortical neurons treated with PCB-95. Collectively, these results indicate that RSV via SIRT1 activates c-Jun, thereby reducing REST expression in SH-SY5Y cells under physiological conditions and blocks PCB-95-induced neuronal cell death by activating the same SIRT1/c-Jun/REST pathway. Copyright © 2015 Elsevier Inc. All rights reserved.

TcoF-DB: dragon database for human transcription co-factors and transcription factor interacting proteins

KAUST Repository

Schaefer, Ulf; Schmeier, Sebastian; Bajic, Vladimir B.

2010-01-01

The initiation and regulation of transcription in eukaryotes is complex and involves a large number of transcription factors (TFs), which are known to bind to the regulatory regions of eukaryotic DNA. Apart from TF-DNA binding, protein-protein interaction involving TFs is an essential component of the machinery facilitating transcriptional regulation. Proteins that interact with TFs in the context of transcription regulation but do not bind to the DNA themselves, we consider transcription co-factors (TcoFs). The influence of TcoFs on transcriptional regulation and initiation, although indirect, has been shown to be significant with the functionality of TFs strongly influenced by the presence of TcoFs. While the role of TFs and their interaction with regulatory DNA regions has been well-studied, the association between TFs and TcoFs has so far been given less attention. Here, we present a resource that is comprised of a collection of human TFs and the TcoFs with which they interact. Other proteins that have a proven interaction with a TF, but are not considered TcoFs are also included. Our database contains 157 high-confidence TcoFs and additionally 379 hypothetical TcoFs. These have been identified and classified according to the type of available evidence for their involvement in transcriptional regulation and their presence in the cell nucleus. We have divided TcoFs into four groups, one of which contains high-confidence TcoFs and three others contain TcoFs which are hypothetical to different extents. We have developed the Dragon Database for Human Transcription Co-Factors and Transcription Factor Interacting Proteins (TcoF-DB). A web-based interface for this resource can be freely accessed at http://cbrc.kaust.edu.sa/tcof/ and http://apps.sanbi.ac.za/tcof/. © The Author(s) 2010.

TcoF-DB: dragon database for human transcription co-factors and transcription factor interacting proteins

KAUST Repository

Schaefer, Ulf

2010-10-21

The initiation and regulation of transcription in eukaryotes is complex and involves a large number of transcription factors (TFs), which are known to bind to the regulatory regions of eukaryotic DNA. Apart from TF-DNA binding, protein-protein interaction involving TFs is an essential component of the machinery facilitating transcriptional regulation. Proteins that interact with TFs in the context of transcription regulation but do not bind to the DNA themselves, we consider transcription co-factors (TcoFs). The influence of TcoFs on transcriptional regulation and initiation, although indirect, has been shown to be significant with the functionality of TFs strongly influenced by the presence of TcoFs. While the role of TFs and their interaction with regulatory DNA regions has been well-studied, the association between TFs and TcoFs has so far been given less attention. Here, we present a resource that is comprised of a collection of human TFs and the TcoFs with which they interact. Other proteins that have a proven interaction with a TF, but are not considered TcoFs are also included. Our database contains 157 high-confidence TcoFs and additionally 379 hypothetical TcoFs. These have been identified and classified according to the type of available evidence for their involvement in transcriptional regulation and their presence in the cell nucleus. We have divided TcoFs into four groups, one of which contains high-confidence TcoFs and three others contain TcoFs which are hypothetical to different extents. We have developed the Dragon Database for Human Transcription Co-Factors and Transcription Factor Interacting Proteins (TcoF-DB). A web-based interface for this resource can be freely accessed at http://cbrc.kaust.edu.sa/tcof/ and http://apps.sanbi.ac.za/tcof/. © The Author(s) 2010.

Extracellular Matrix-Regulated Gene Expression RequiresCooperation of SWI/SNF and Transcription Factors

Energy Technology Data Exchange (ETDEWEB)

Xu, Ren; Spencer, Virginia A.; Bissell, Mina J.

2006-05-25

Extracellular cues play crucial roles in the transcriptional regulation of tissue-specific genes, but whether and how these signals lead to chromatin remodeling is not understood and subject to debate. Using chromatin immunoprecipitation (ChIP) assays and mammary-specific genes as models, we show here that extracellular matrix (ECM) molecules and prolactin cooperate to induce histone acetylation and binding of transcription factors and the SWI/SNF complex to the {beta}- and ?-casein promoters. Introduction of a dominant negative Brg1, an ATPase subunit of SWI/SNF complex, significantly reduced both {beta}- and ?-casein expression, suggesting that SWI/SNF-dependent chromatin remodeling is required for transcription of mammary-specific genes. ChIP analyses demonstrated that the ATPase activity of SWI/SNF is necessary for recruitment of RNA transcriptional machinery, but not for binding of transcription factors or for histone acetylation. Coimmunoprecipitation analyses showed that the SWI/SNF complex is associated with STAT5, C/EBP{beta}, and glucocorticoid receptor (GR). Thus, ECM- and prolactin-regulated transcription of the mammary-specific casein genes requires the concerted action of chromatin remodeling enzymes and transcription factors.

The WRKY transcription factor family in Brachypodium distachyon.

Science.gov (United States)

Tripathi, Prateek; Rabara, Roel C; Langum, Tanner J; Boken, Ashley K; Rushton, Deena L; Boomsma, Darius D; Rinerson, Charles I; Rabara, Jennifer; Reese, R Neil; Chen, Xianfeng; Rohila, Jai S; Rushton, Paul J

2012-06-22

A complete assembled genome sequence of wheat is not yet available. Therefore, model plant systems for wheat are very valuable. Brachypodium distachyon (Brachypodium) is such a system. The WRKY family of transcription factors is one of the most important families of plant transcriptional regulators with members regulating important agronomic traits. Studies of WRKY transcription factors in Brachypodium and wheat therefore promise to lead to new strategies for wheat improvement. We have identified and manually curated the WRKY transcription factor family from Brachypodium using a pipeline designed to identify all potential WRKY genes. 86 WRKY transcription factors were found, a total higher than all other current databases. We therefore propose that our numbering system (BdWRKY1-BdWRKY86) becomes the standard nomenclature. In the JGI v1.0 assembly of Brachypodium with the MIPS/JGI v1.0 annotation, nine of the transcription factors have no gene model and eleven gene models are probably incorrectly predicted. In total, twenty WRKY transcription factors (23.3%) do not appear to have accurate gene models. To facilitate use of our data, we have produced The Database of Brachypodium distachyon WRKY Transcription Factors. Each WRKY transcription factor has a gene page that includes predicted protein domains from MEME analyses. These conserved protein domains reflect possible input and output domains in signaling. The database also contains a BLAST search function where a large dataset of WRKY transcription factors, published genes, and an extensive set of wheat ESTs can be searched. We also produced a phylogram containing the WRKY transcription factor families from Brachypodium, rice, Arabidopsis, soybean, and Physcomitrella patens, together with published WRKY transcription factors from wheat. This phylogenetic tree provides evidence for orthologues, co-orthologues, and paralogues of Brachypodium WRKY transcription factors. The description of the WRKY transcription factor

Generating autologous hematopoietic cells from human-induced pluripotent stem cells through ectopic expression of transcription factors.

Science.gov (United States)

Hwang, Yongsung; Broxmeyer, Hal E; Lee, Man Ryul

2017-07-01

Hematopoietic cell transplantation (HCT) is a successful treatment modality for patients with malignant and nonmalignant disorders, usually when no other treatment option is available. The cells supporting long-term reconstitution after HCT are the hematopoietic stem cells (HSCs), which can be limited in numbers. Moreover, finding an appropriate human leukocyte antigen-matched donor can be problematic. If HSCs can be stably produced in large numbers from autologous or allogeneic cell sources, it would benefit HCT. Induced pluripotent stem cells (iPSCs) established from patients' own somatic cells can be differentiated into hematopoietic cells in vitro. This review will highlight recent methods for regulating human (h) iPSC production of HSCs and more mature blood cells. Advancements in transcription factor-mediated regulation of the developmental stages of in-vivo hematopoietic lineage commitment have begun to provide an understanding of the molecular mechanism of hematopoiesis. Such studies involve not only directed differentiation in which transcription factors, specifically expressed in hematopoietic lineage-specific cells, are overexpressed in iPSCs, but also direct conversion in which transcription factors are introduced into patient-derived somatic cells which are dedifferentiated to hematopoietic cells. As iPSCs derived from patients suffering from genetically mutated diseases would express the same mutated genetic information, CRISPR-Cas9 gene editing has been utilized to differentiate genetically corrected iPSCs into normal hematopoietic cells. IPSCs provide a model for molecular understanding of disease, and also may function as a cell population for therapy. Efficient differentiation of patient-specific iPSCs into HSCs and progenitor cells is a potential means to overcome limitations of such cells for HCT, as well as for providing in-vitro drug screening templates as tissue-on-a-chip models.

Journal of Genetics | Indian Academy of Sciences

Indian Academy of Sciences (India)

Auxin response factors (ARF) are transcription factors that regulate auxin responses in plants. Although the genomewide analysis of this family has been performed in some species, little is known regarding ARF genes in apple (Malus domestica). In this study, 31 putative apple ARF genes have been identified and located ...

Constitutive expression of a salinity-induced wheat WRKY transcription factor enhances salinity and ionic stress tolerance in transgenic Arabidopsis thaliana

Energy Technology Data Exchange (ETDEWEB)

Qin, Yuxiang, E-mail: yuxiangqin@126.com [Department of Biotechnology, University of Jinan, Jinan 250022 (China); Tian, Yanchen [The Key Laboratory of Plant Cell Engineering and Germplasm Innovation, Ministry of Education, School of Life Science, Shandong University, Jinan 250100 (China); Han, Lu; Yang, Xinchao [Department of Biotechnology, University of Jinan, Jinan 250022 (China)

2013-11-15

Highlights: •A class II WRKY transcription factor, TaWRKY79 was isolated and characterized. •TaWRKY79 was induced by NaCl or abscisic acid. •843 bp regulatory segment was sufficient to respond to ABA or NaCl treatment. •TaWRKY79 enhanced salinity and ionic tolerance while reduced sensitivity to ABA. •TaWRKY79 increased salinity and ionic tolerance in an ABA-dependent pathway. -- Abstract: The isolation and characterization of TaWRKY79, a wheat class II WRKY transcription factor, is described. Its 1297 bp coding region includes a 987 bp long open reading frame. TaWRKY79 was induced by stressing seedlings with either NaCl or abscisic acid (ABA). When a fusion between an 843 bp segment upstream of the TaWRKY79 coding sequence and GUS was introduced into Arabidopsis thaliana, GUS staining indicated that this upstream segment captured the sequence(s) required to respond to ABA or NaCl treatment. When TaWRKY79 was constitutively expressed as a transgene in A. thaliana, the transgenic plants showed an improved capacity to extend their primary root in the presence of either 100 mM NaCl, 10 mM LiCl or 2 μM ABA. The inference was that TaWRKY79 enhanced the level of tolerance to both salinity and ionic stress, while reducing the level of sensitivity to ABA. The ABA-related genes ABA1, ABA2 ABI1 and ABI5 were all up-regulated in the TaWRKY79 transgenic plants, suggesting that the transcription factor operates in an ABA-dependent pathway.

Constitutive expression of a salinity-induced wheat WRKY transcription factor enhances salinity and ionic stress tolerance in transgenic Arabidopsis thaliana

International Nuclear Information System (INIS)

Qin, Yuxiang; Tian, Yanchen; Han, Lu; Yang, Xinchao

2013-01-01

Highlights: •A class II WRKY transcription factor, TaWRKY79 was isolated and characterized. •TaWRKY79 was induced by NaCl or abscisic acid. •843 bp regulatory segment was sufficient to respond to ABA or NaCl treatment. •TaWRKY79 enhanced salinity and ionic tolerance while reduced sensitivity to ABA. •TaWRKY79 increased salinity and ionic tolerance in an ABA-dependent pathway. -- Abstract: The isolation and characterization of TaWRKY79, a wheat class II WRKY transcription factor, is described. Its 1297 bp coding region includes a 987 bp long open reading frame. TaWRKY79 was induced by stressing seedlings with either NaCl or abscisic acid (ABA). When a fusion between an 843 bp segment upstream of the TaWRKY79 coding sequence and GUS was introduced into Arabidopsis thaliana, GUS staining indicated that this upstream segment captured the sequence(s) required to respond to ABA or NaCl treatment. When TaWRKY79 was constitutively expressed as a transgene in A. thaliana, the transgenic plants showed an improved capacity to extend their primary root in the presence of either 100 mM NaCl, 10 mM LiCl or 2 μM ABA. The inference was that TaWRKY79 enhanced the level of tolerance to both salinity and ionic stress, while reducing the level of sensitivity to ABA. The ABA-related genes ABA1, ABA2 ABI1 and ABI5 were all up-regulated in the TaWRKY79 transgenic plants, suggesting that the transcription factor operates in an ABA-dependent pathway

Fatty Acid–Regulated Transcription Factors in the Liver

Science.gov (United States)

Jump, Donald B.; Tripathy, Sasmita; Depner, Christopher M.

2014-01-01

Fatty acid regulation of hepatic gene transcription was first reported in the early 1990s. Several transcription factors have been identified as targets of fatty acid regulation. This regulation is achieved by direct fatty acid binding to the transcription factor or by indirect mechanisms where fatty acids regulate signaling pathways controlling the expression of transcription factors or the phosphorylation, ubiquitination, or proteolytic cleavage of the transcription factor. Although dietary fatty acids are well-established regulators of hepatic transcription factors, emerging evidence indicates that endogenously generated fatty acids are equally important in controlling transcription factors in the context of glucose and lipid homeostasis. Our first goal in this review is to provide an up-to-date examination of the molecular and metabolic bases of fatty acid regulation of key transcription factors controlling hepatic metabolism. Our second goal is to link these mechanisms to nonalcoholic fatty liver disease (NAFLD), a growing health concern in the obese population. PMID:23528177

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

Science.gov (United States)

Artyomov, Maxim; Meissner, Alex; Chakraborty, Arup

2010-03-01

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

Wnt3a induces the expression of acetylcholinesterase during osteoblast differentiation via the Runx2 transcription factor.

Science.gov (United States)

Xu, Miranda L; Bi, Cathy W C; Liu, Etta Y L; Dong, Tina T X; Tsim, Karl W K

2017-07-28

Acetylcholinesterase (AChE) hydrolyzes acetylcholine to terminate cholinergic transmission in neurons. Apart from this AChE activity, emerging evidence suggests that AChE could also function in other, non-neuronal cells. For instance, in bone, AChE exists as a proline-rich membrane anchor (PRiMA)-linked globular form in osteoblasts, in which it is proposed to play a noncholinergic role in differentiation. However, this hypothesis is untested. Here, we found that in cultured rat osteoblasts, AChE expression was increased in parallel with osteoblastic differentiation. Because several lines of evidence indicate that AChE activity in osteoblast could be triggered by Wnt/β-catenin signaling, we added recombinant human Wnt3a to cultured osteoblasts and found that this addition induced expression of the ACHE gene and protein product. This Wnt3a-induced AChE expression was blocked by the Wnt-signaling inhibitor Dickkopf protein-1 (DKK-1). We hypothesized that the Runt-related transcription factor 2 (Runx2), a downstream transcription factor in Wnt/β-catenin signaling, is involved in AChE regulation in osteoblasts, confirmed by the identification of a Runx2-binding site in the ACHE gene promoter, further corroborated by ChIP. Of note, Runx2 overexpression in osteoblasts induced AChE expression and activity of the ACHE promoter tagged with the luciferase gene. Moreover, deletion of the Runx2-binding site in the ACHE promoter reduced its activity during osteoblastic differentiation, and addition of 5-azacytidine and trichostatin A to differentiating osteoblasts affected AChE expression, suggesting epigenetic regulation of the ACHE gene. We conclude that AChE plays a role in osteoblastic differentiation and is regulated by both Wnt3a and Runx2. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Rosetta comparative modeling for library design: Engineering alternative inducer specificity in a transcription factor.

Science.gov (United States)

Jha, Ramesh K; Chakraborti, Subhendu; Kern, Theresa L; Fox, David T; Strauss, Charlie E M

2015-07-01

Structure-based rational mutagenesis for engineering protein functionality has been limited by the scarcity and difficulty of obtaining crystal structures of desired proteins. On the other hand, when high-throughput selection is possible, directed evolution-based approaches for gaining protein functionalities have been random and fortuitous with limited rationalization. We combine comparative modeling of dimer structures, ab initio loop reconstruction, and ligand docking to select positions for mutagenesis to create a library focused on the ligand-contacting residues. The rationally reduced library requirement enabled conservative control of the substitutions by oligonucleotide synthesis and bounding its size within practical transformation efficiencies (∼ 10(7) variants). This rational approach was successfully applied on an inducer-binding domain of an Acinetobacter transcription factor (TF), pobR, which shows high specificity for natural effector molecule, 4-hydroxy benzoate (4HB), but no native response to 3,4-dihydroxy benzoate (34DHB). Selection for mutants with high transcriptional induction by 34DHB was carried out at the single-cell level under flow cytometry (via green fluorescent protein expression under the control of pobR promoter). Critically, this selection protocol allows both selection for induction and rejection of constitutively active mutants. In addition to gain-of-function for 34DHB induction, the selected mutants also showed enhanced sensitivity and response for 4HB (native inducer) while no sensitivity was observed for a non-targeted but chemically similar molecule, 2-hydroxy benzoate (2HB). This is unique application of the Rosetta modeling protocols for library design to engineer a TF. Our approach extends applicability of the Rosetta redesign protocol into regimes without a priori precision structural information. © 2015 Wiley Periodicals, Inc.

The Transcription Factor Encyclopedia

DEFF Research Database (Denmark)

Yusuf, Dimas; Butland, Stefanie L; Swanson, Magdalena I

2012-01-01

mini review articles on pertinent human, mouse and rat TFs. Notable features of the TFe website include a high-quality PDF generator and web API for programmatic data retrieval. TFe aims to rapidly educate scientists about the TFs they encounter through the delivery of succinct summaries written......ABSTRACT: Here we present the Transcription Factor Encyclopedia (TFe), a new web-based compendium of mini review articles on transcription factors (TFs) that is founded on the principles of open access and collaboration. Our consortium of over 100 researchers has collectively contributed over 130...

ArF photo resist pattern sample preparation method using FIB without protective coating

Science.gov (United States)

Okushima, Hirohisa; Onozuka, Toshihiko; Kuroda, Yasushi; Yaguchi, Toshie; Umemura, Kaoru; Tamochi, Ryuichiro; Watanabe, Kenji; Hasegawa, Norio; Kawata, Isao; Rijpers, Bart

2006-03-01

This paper presents a novel method of FIB (FIB: focused ion beam) sample preparation to accurately evaluate critical dimensions and profiles of ArF photo resist patterns without the use of a protective coating on the photo resist. In order to accomplish this, the FIB micro-sampling method that is one of effective FIB milling and fabrication method was employed. First a Si cap is picked up from a silicon wafer and fixed to ArF photo resist patterns to protect against ion beam irradiation. Then, a micro-sample, a piece of Si-capped ArF photo resist, was extracted from the bulk ArF photo resist. In this procedure, this silicon cap always protects ArF photo resist patterns against ion beam irradiation. For the next step, the micro-sample is fixed to a needle stub of the FIB-STEM (STEM: scanning transmission electron microscopy) compatible rotation holder. This sample on the needle stub was rotated 180 degrees and milled from the side of Si substrate. Lastly, the sample is milled to the thickness of 2μm. In this process, the ion beam is irradiating from the silicon substrate side to minimize the ion beam irradiation damages on the ArF photo resist patterns. EDX (EDX: Energy dispersive X-ray spectroscopy) analysis proved that no gallium ions were detected on the surface of the ArF photo resist patterns. The feasibility of high accelerating voltage observation of STEM to observe line edge roughness of a thick sample like 2μm without shrinkage has been demonstrated.

Transcription factor FoxO1 is essential for enamel biomineralization.

Directory of Open Access Journals (Sweden)

Ross A Poché

Full Text Available The Transforming growth factor β (Tgf-β pathway, by signaling via the activation of Smad transcription factors, induces the expression of many diverse downstream target genes thereby regulating a vast array of cellular events essential for proper development and homeostasis. In order for a specific cell type to properly interpret the Tgf-β signal and elicit a specific cellular response, cell-specific transcriptional co-factors often cooperate with the Smads to activate a discrete set of genes in the appropriate temporal and spatial manner. Here, via a conditional knockout approach, we show that mice mutant for Forkhead Box O transcription factor FoxO1 exhibit an enamel hypomaturation defect which phenocopies that of the Smad3 mutant mice. Furthermore, we determined that both the FoxO1 and Smad3 mutant teeth exhibit changes in the expression of similar cohort of genes encoding enamel matrix proteins required for proper enamel development. These data raise the possibility that FoxO1 and Smad3 act in concert to regulate a common repertoire of genes necessary for complete enamel maturation. This study is the first to define an essential role for the FoxO family of transcription factors in tooth development and provides a new molecular entry point which will allow researchers to delineate novel genetic pathways regulating the process of biomineralization which may also have significance for studies of human tooth diseases such as amelogenesis imperfecta.

Chemically Induced Degradation of the Oncogenic Transcription Factor BCL6

Directory of Open Access Journals (Sweden)

Nina Kerres

2017-09-01

Full Text Available The transcription factor BCL6 is a known driver of oncogenesis in lymphoid malignancies, including diffuse large B cell lymphoma (DLBCL. Disruption of its interaction with transcriptional repressors interferes with the oncogenic effects of BCL6. We used a structure-based drug design to develop highly potent compounds that block this interaction. A subset of these inhibitors also causes rapid ubiquitylation and degradation of BCL6 in cells. These compounds display significantly stronger induction of expression of BCL6-repressed genes and anti-proliferative effects than compounds that merely inhibit co-repressor interactions. This work establishes the BTB domain as a highly druggable structure, paving the way for the use of other members of this protein family as drug targets. The magnitude of effects elicited by this class of BCL6-degrading compounds exceeds that of our equipotent non-degrading inhibitors, suggesting opportunities for the development of BCL6-based lymphoma therapeutics.

Loss of Arf4 causes severe degeneration of the exocrine pancreas but not cystic kidney disease or retinal degeneration.

Directory of Open Access Journals (Sweden)

Jillian N Pearring

2017-04-01

Full Text Available Arf4 is proposed to be a critical regulator of membrane protein trafficking in early secretory pathway. More recently, Arf4 was also implicated in regulating ciliary trafficking, however, this has not been comprehensively tested in vivo. To directly address Arf4's role in ciliary transport, we deleted Arf4 specifically in either rod photoreceptor cells, kidney, or globally during the early postnatal period. Arf4 deletion in photoreceptors did not cause protein mislocalization or retinal degeneration, as expected if Arf4 played a role in protein transport to the ciliary outer segment. Likewise, Arf4 deletion in kidney did not cause cystic disease, as expected if Arf4 were involved in general ciliary trafficking. In contrast, global Arf4 deletion in the early postnatal period resulted in growth restriction, severe pancreatic degeneration and early death. These findings are consistent with Arf4 playing a critical role in endomembrane trafficking, particularly in the pancreas, but not in ciliary function.

Overexpression of a transcription factor LYL1 induces T- and B-cell lymphoma in mice.

Science.gov (United States)

Zhong, Y; Jiang, L; Hiai, H; Toyokuni, S; Yamada, Y

2007-10-18

LYL1, a member of the class II basic helix-loop-helix transcription factors, is aberrantly expressed in a fraction of human T-cell acute lymphoblastic leukemia. Here, we generated transgenic mice ubiquitously overexpressing LYL1 using a construct expressing full-length cDNA driven by a human elongation factor 1alpha promoter. Four independent lines exhibiting high LYL1 expression were established. Of these transgenic mice, 96% displayed loss of hair with a short kinked tail. Furthermore, 30% of them developed malignant lymphoma, with an average latent period of 352 days. In these mice, histological examination revealed tumor cell infiltration in multiple organs and immunohistochemical analysis showed that the infiltrated tumor cells were either CD3 or CD45R/B220-positive; fluorescence-activated cell sorter analysis indicated that each tumor consisted either of mainly CD4, CD8 double-positive T cells or mature B cells; the clonality of LYL1-induced lymphoma was confirmed by T-cell receptor rearrangement and immunoglobulin heavy-chain gene rearrangement analyses. Mammalian two-hybrid analysis and luciferase assay suggested that excess LYL1 blocked the dimerization of E2A and thus inhibited the regulatory activity of E2A on the CD4 promoter. Reverse transcription-polymerase chain reaction results showed that the expression of certain E2A/HEB target genes was downregulated. Taken together, our results provide direct evidence that aberrant expression of LYL1 plays a role in lymphomagenesis.

Intratumoral Immunization by p19Arf and Interferon-β Gene Transfer in a Heterotopic Mouse Model of Lung Carcinoma

Directory of Open Access Journals (Sweden)

João Paulo Portela Catani

2016-12-01

Full Text Available Therapeutic strategies that act by eliciting and enhancing antitumor immunity have been clinically validated as an effective treatment modality but may benefit from the induction of both cell death and immune activation as primary stimuli. Using our AdRGD-PG adenovector platform, we show here for the first time that in situ gene transfer of p19Arf and interferon-β (IFNβ in the LLC1 mouse model of lung carcinoma acts as an immunotherapy. Although p19Arf is sufficient to induce cell death, only its pairing with IFNβ significantly induced markers of immunogenic cell death. In situ gene therapy with IFNβ, either alone or in combination with p19Arf, could retard tumor progression, but only the combined treatment was associated with a protective immune response. Specifically in the case of combined intratumoral gene transfer, we identified 167 differentially expressed genes when using microarray to evaluate tumors that were treated in vivo and confirmed the activation of CCL3, CXCL3, IL1α, IL1β, CD274, and OSM, involved in immune response and chemotaxis. Histologic evaluation revealed significant tumor infiltration by neutrophils, whereas functional depletion of granulocytes ablated the antitumor effect of our approach. The association of in situ gene therapy with cisplatin resulted in synergistic elimination of tumor progression. In all, in situ gene transfer with p19Arf and IFNβ acts as an immunotherapy involving recruitment of neutrophils, a desirable but previously untested outcome, and this approach may be allied with chemotherapy, thus providing significant antitumor activity and warranting further development for the treatment of lung carcinoma.

A transcription factor for cold sensation!

Directory of Open Access Journals (Sweden)

Milbrandt Jeffrey

2005-03-01

Full Text Available Abstract The ability to feel hot and cold is critical for animals and human beings to survive in the natural environment. Unlike other sensations, the physiology of cold sensation is mostly unknown. In the present study, we use genetically modified mice that do not express nerve growth factor-inducible B (NGFIB to investigate the possible role of NGFIB in cold sensation. We found that genetic deletion of NGFIB selectively affected behavioral responses to cold stimuli while behavioral responses to noxious heat or mechanical stimuli were normal. Furthermore, behavioral responses remained reduced or blocked in NGFIB knockout mice even after repetitive application of cold stimuli. Our results provide strong evidence that the first transcription factor NGFIB determines the ability of animals to respond to cold stimulation.

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

Science.gov (United States)

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

2018-04-19

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

Loss of p19(Arf facilitates the angiogenic switch and tumor initiation in a multi-stage cancer model via p53-dependent and independent mechanisms.

Directory of Open Access Journals (Sweden)

Danielle B Ulanet

2010-08-01

Full Text Available The Arf tumor suppressor acts as a sensor of oncogenic signals, countering aberrant proliferation in large part via activation of the p53 transcriptional program, though a number of p53-independent functions have been described. Mounting evidence suggests that, in addition to promoting tumorigenesis via disruptions in the homeostatic balance between cell proliferation and apoptosis of overt cancer cells, genetic alterations leading to tumor suppressor loss of function or oncogene gain of function can also incite tumor development via effects on the tumor microenvironment. In a transgenic mouse model of multi-stage pancreatic neuroendocrine carcinogenesis (PNET driven by inhibition of the canonical p53 and Rb tumor suppressors with SV40 large T-antigen (Tag, stochastic progression to tumors is limited in part by a requirement for initiation of an angiogenic switch. Despite inhibition of p53 by Tag in this mouse PNET model, concomitant disruption of Arf via genetic knockout resulted in a significantly accelerated pathway to tumor formation that was surprisingly not driven by alterations in tumor cell proliferation or apoptosis, but rather via earlier activation of the angiogenic switch. In the setting of a constitutional p53 gene knockout, loss of Arf also accelerated tumor development, albeit to a lesser degree. These findings demonstrate that Arf loss of function can promote tumorigenesis via facilitating angiogenesis, at least in part, through p53-independent mechanisms.

Resveratrol via sirtuin-1 downregulates RE1-silencing transcription factor (REST) expression preventing PCB-95-induced neuronal cell death

International Nuclear Information System (INIS)

Guida, Natascia; Laudati, Giusy; Anzilotti, Serenella; Secondo, Agnese; Montuori, Paolo; Di Renzo, Gianfranco; Canzoniero, Lorella M.T.; Formisano, Luigi

2015-01-01

Resveratrol (3,5,4′-trihydroxystilbene) (RSV), a polyphenol widely present in plants, exerts a neuroprotective function in several neurological conditions; it is an activator of class III histone deacetylase sirtuin1 (SIRT1), a crucial regulator in the pathophysiology of neurodegenerative diseases. By contrast, the RE1-silencing transcription factor (REST) is involved in the neurotoxic effects following exposure to polychlorinated biphenyl (PCB) mixture A1254. The present study investigated the effects of RSV-induced activation of SIRT1 on REST expression in SH-SY5Y cells. Further, we investigated the possible relationship between the non-dioxin-like (NDL) PCB-95 and REST through SIRT1 to regulate neuronal death in rat cortical neurons. Our results revealed that RSV significantly decreased REST gene and protein levels in a dose- and time-dependent manner. Interestingly, overexpression of SIRT1 reduced REST expression, whereas EX-527, an inhibitor of SIRT1, increased REST expression and blocked RSV-induced REST downregulation. These results suggest that RSV downregulates REST through SIRT1. In addition, RSV enhanced activator protein 1 (AP-1) transcription factor c-Jun expression and its binding to the REST promoter gene. Indeed, c-Jun knockdown reverted RSV-induced REST downregulation. Intriguingly, in SH-SY5Y cells and rat cortical neurons the NDL PCB-95 induced necrotic cell death in a concentration-dependent manner by increasing REST mRNA and protein expression. In addition, SIRT1 knockdown blocked RSV-induced neuroprotection in rat cortical neurons treated with PCB-95. Collectively, these results indicate that RSV via SIRT1 activates c-Jun, thereby reducing REST expression in SH-SY5Y cells under physiological conditions and blocks PCB-95-induced neuronal cell death by activating the same SIRT1/c-Jun/REST pathway. - Highlights: • Resveratrol via SIRT1/c-Jun downregulates REST mRNA and protein in SH-SY5Y cells. • Non-dioxin-like (NDL) PCB-95 is cytotoxic to

Resveratrol via sirtuin-1 downregulates RE1-silencing transcription factor (REST) expression preventing PCB-95-induced neuronal cell death

Energy Technology Data Exchange (ETDEWEB)

Guida, Natascia [IRCSS SDN, Naples 80131 (Italy); Laudati, Giusy [Division of Pharmacology, Department of Neuroscience, Reproductive and Dentistry Sciences, School of Medicine, “Federico II” University of Naples, Via Pansini, 5, 80131 Naples (Italy); Anzilotti, Serenella [IRCSS SDN, Naples 80131 (Italy); Secondo, Agnese [Division of Pharmacology, Department of Neuroscience, Reproductive and Dentistry Sciences, School of Medicine, “Federico II” University of Naples, Via Pansini, 5, 80131 Naples (Italy); Montuori, Paolo [Department of Public Health, ‘Federico II’ University of Naples, Naples (Italy); Di Renzo, Gianfranco [Division of Pharmacology, Department of Neuroscience, Reproductive and Dentistry Sciences, School of Medicine, “Federico II” University of Naples, Via Pansini, 5, 80131 Naples (Italy); Canzoniero, Lorella M.T. [Division of Pharmacology, Department of Neuroscience, Reproductive and Dentistry Sciences, School of Medicine, “Federico II” University of Naples, Via Pansini, 5, 80131 Naples (Italy); Division of Pharmacology, Department of Science and Technology, University of Sannio, Via Port' Arsa 11, 82100 Benevento (Italy); Formisano, Luigi, E-mail: cformisa@unisannio.it [Division of Pharmacology, Department of Neuroscience, Reproductive and Dentistry Sciences, School of Medicine, “Federico II” University of Naples, Via Pansini, 5, 80131 Naples (Italy); Division of Pharmacology, Department of Science and Technology, University of Sannio, Via Port' Arsa 11, 82100 Benevento (Italy)

2015-11-01

Resveratrol (3,5,4′-trihydroxystilbene) (RSV), a polyphenol widely present in plants, exerts a neuroprotective function in several neurological conditions; it is an activator of class III histone deacetylase sirtuin1 (SIRT1), a crucial regulator in the pathophysiology of neurodegenerative diseases. By contrast, the RE1-silencing transcription factor (REST) is involved in the neurotoxic effects following exposure to polychlorinated biphenyl (PCB) mixture A1254. The present study investigated the effects of RSV-induced activation of SIRT1 on REST expression in SH-SY5Y cells. Further, we investigated the possible relationship between the non-dioxin-like (NDL) PCB-95 and REST through SIRT1 to regulate neuronal death in rat cortical neurons. Our results revealed that RSV significantly decreased REST gene and protein levels in a dose- and time-dependent manner. Interestingly, overexpression of SIRT1 reduced REST expression, whereas EX-527, an inhibitor of SIRT1, increased REST expression and blocked RSV-induced REST downregulation. These results suggest that RSV downregulates REST through SIRT1. In addition, RSV enhanced activator protein 1 (AP-1) transcription factor c-Jun expression and its binding to the REST promoter gene. Indeed, c-Jun knockdown reverted RSV-induced REST downregulation. Intriguingly, in SH-SY5Y cells and rat cortical neurons the NDL PCB-95 induced necrotic cell death in a concentration-dependent manner by increasing REST mRNA and protein expression. In addition, SIRT1 knockdown blocked RSV-induced neuroprotection in rat cortical neurons treated with PCB-95. Collectively, these results indicate that RSV via SIRT1 activates c-Jun, thereby reducing REST expression in SH-SY5Y cells under physiological conditions and blocks PCB-95-induced neuronal cell death by activating the same SIRT1/c-Jun/REST pathway. - Highlights: • Resveratrol via SIRT1/c-Jun downregulates REST mRNA and protein in SH-SY5Y cells. • Non-dioxin-like (NDL) PCB-95 is cytotoxic to

The Hv NAC6 transcription factor: a positive regulator of penetration resistance in barley and Arabidopsis

DEFF Research Database (Denmark)

Jensen, Michael Krogh; Rung, Jesper Henrik; Gregersen, Per Langkjaer

2007-01-01

Pathogens induce the expression of many genes encoding plant transcription factors, though specific knowledge of the biological function of individual transcription factors remains scarce. NAC transcription factors are encoded in plants by a gene family with proposed functions in both abiotic...... and biotic stress adaptation, as well as in developmental processes. In this paper, we provide convincing evidence that a barley NAC transcription factor has a direct role in regulating basal defence. The gene transcript was isolated by differential display from barley leaves infected with the biotrophic...... powdery mildew fungus, Blumeria graminis f.sp. hordei (Bgh). The full-length cDNA clone was obtained using 5'-RACE and termed HvNAC6, due to its high similarity to the rice homologue, OsNAC6. Gene silencing of HvNAC6 during Bgh inoculation compromises penetration resistance in barley epidermal cells...

Transcriptomic profiling-based mutant screen reveals three new transcription factors mediating menadione resistance in Neurospora crassa.

Science.gov (United States)

Zhu, Jufen; Yu, Xinxu; Xie, Baogui; Gu, Xiaokui; Zhang, Zhenying; Li, Shaojie

2013-06-01

To gain insight into the regulatory mechanisms of oxidative stress responses in filamentous fungi, the genome-wide transcriptional response of Neurospora crassa to menadione was analysed by digital gene expression (DGE) profiling, which identified 779 upregulated genes and 576 downregulated genes. Knockout mutants affecting 130 highly-upregulated genes were tested for menadione sensitivity, which revealed that loss of the transcription factor siderophore regulation (SRE) (a transcriptional repressor for siderophore biosynthesis), catatase-3, cytochrome c peroxidase or superoxide dismutase 1 copper chaperone causes hypersensitivity to menadione. Deletion of sre dramatically increased transcription of the siderophore biosynthesis gene ono and the siderophore iron transporter gene sit during menadione stress, suggesting that SRE is required for repression of iron uptake under oxidative stress conditions. Contrary to its phenotype, the sre deletion mutant showed higher transcriptional levels of genes encoding reactive oxygen species (ROS) scavengers than wild type during menadione stress, which implies that the mutant suffers a higher level of oxidative stress than wild type. Uncontrolled iron uptake in the sre mutant might exacerbate cellular oxidative stress. This is the first report of a negative regulator of iron assimilation participating in the fungal oxidative stress response. In addition to SRE, eight other transcription factor genes were also menadione-responsive but their single gene knockout mutants showed wild-type menadione sensitivity. Two of them, named as mit-2 (menadione induced transcription factor-2) and mit-4 (menadione induced transcription factor-4), were selected for double mutant analysis. The double mutant was hypersensitive to menadione. Similarly, the double mutation of mit-2 and sre also had additive effects on menadione sensitivity, suggesting multiple transcription factors mediate oxidative stress resistance in an additive manner

Arf6, Rab11 and transferrin receptor define distinct populations of recycling endosomes.

Science.gov (United States)

Kobayashi, Hotaka; Fukuda, Mitsunori

2013-09-01

Recycling endosomes are key platforms for endocytic recycling that return internalized molecules back to the plasma membrane. To determine how recycling endosomes perform their functions, searching for proteins and lipids that specifically localized at recycling endosomes has often been performed by colocalization analyses between candidate molecules and conventional recycling endosome markers. However, it remains unclear whether all the conventional markers have identical localizations. Here we report finding that three well-known recycling endosome markers, i.e., Arf6, Rab11 and transferrin receptor (TfR), have different intracellular localizations in PC12 cells. The results of immunofluorescence analyses showed that the signals of endogenous Arf6, Rab11 and TfR in nerve growth factor-stimulated PC12 cells generally differed, although there was some overlapping. Our findings provide new information about recycling endosome markers, and they highlight the heterogeneity of recycling endosomes.

Acute interstitial nephritis induced by intermittent use of Rifampicin in patient with Brucellosis

International Nuclear Information System (INIS)

Salih, S. Bin; Kharal, M.; Qahtani, M.; Dahneem, L.; Nohair, S.

2008-01-01

Acute oliguric renal failure (ARF) developed in a patient 2 days after she was started on intermittent anti-Brucella therapy including rifampicin. The clinical picture was compatible with acute allergic interstitial nephritis. Renal histology revealed mainly acute tubular necrosis with mild tubulo-intertitial mononuclear cellular infiltrate. Intermittent therapy, as in our patient, has been the major factor in the development of rifampicin induced ARF in cases reviewed in literature. (author)

Inducible, tunable and multiplex human gene regulation using CRISPR-Cpf1-based transcription factors | Office of Cancer Genomics

Science.gov (United States)

Targeted and inducible regulation of mammalian gene expression is a broadly important research capability that may also enable development of novel therapeutics for treating human diseases. Here we demonstrate that a catalytically inactive RNA-guided CRISPR-Cpf1 nuclease fused to transcriptional activation domains can up-regulate endogenous human gene expression. We engineered drug-inducible Cpf1-based activators and show how this system can be used to tune the regulation of endogenous gene transcription in human cells.

The dynamic changes of DNA methylation and histone modifications of salt responsive transcription factor genes in soybean.

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Yuguang Song

Full Text Available Epigenetic modification contributes to the regulation of gene expression and plant development under salinity stress. Here we describe the identification of 49 soybean transcription factors by microarray analysis as being inducible by salinity stress. A semi-quantitative RT-PCR-based expression assay confirmed the salinity stress inducibility of 45 of these 49 transcription factors, and showed that ten of them were up-regulated when seedlings were exposed to the demethylation agent 5-aza-2-deoxycytidine. Salinity stress was shown to affect the methylation status of four of these ten transcription factors (one MYB, one b-ZIP and two AP2/DREB family members using a combination of bisulfite sequencing and DNA methylation-sensitive DNA gel blot analysis. ChIP analysis indicated that the activation of three of the four DNA methylated transcription factors was correlated with an increased level of histone H3K4 trimethylation and H3K9 acetylation, and/or a reduced level of H3K9 demethylation in various parts of the promoter or coding regions. Our results suggest a critical role for some transcription factors' activation/repression by DNA methylation and/or histone modifications in soybean tolerance to salinity stress.

A population-based study on the association between acute renal failure (ARF and the duration of polypharmacy

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Chang Yi-Ping

2012-08-01

Full Text Available Abstract Background Because of the rapid growth in elderly population, polypharmacy has become a serious public health issue worldwide. Although acute renal failure (ARF is one negative consequence of polypharmacy, the association between the duration of polypharmacy and ARF remains unclear. We therefore assessed this association using a population-based database. Methods Data were collected from the Taiwan National Health Insurance Research Database (NHIRD from 2003 through 2006. The case group included patients hospitalized for ARF during 2006, but not admitted due to trauma, surgery, burn trauma, car accident, transplantation, or infectious diseases; the control group included patients hospitalized without ARF. The cumulative number of days of polypharmacy (defined as more than 5 prescriptions per day for 1 year prior to admission was determined, with patients further subdivided into 4 categories: less than 30 days, 31–90 days, 91–180 days, and over 181 days. The dependent variable was ARF, and the control variables were age, gender, comorbidities in patients hospitalized for ARF, stay in ICUs during ARF hospitalization and site of operation for prior admissions within one month of ARF hospitalization. Results Of 20,790 patients who were admitted to hospitals for ARF in 2006, 12,314 (59.23 % were male and more than 60 % were older than 65 years. Of patients with and without ARF, 16.14 % and 10.61 %, respectively, received polypharmacy for 91–180 days and 50.22 % and 24.12 %, respectively, for over 181 days. A statistical model indicated that, relative to patients who received polypharmacy for less than 30 days, those who received polypharmacy for 31–90, 91–180 and over 181 days had odds ratios of developing ARF of 1.33 (p Conclusions We observed statistically significant associations between the duration of polypharmacy and the occurrence of ARF.

Nucleophosmin is required for DNA integrity and p19Arf protein stability

DEFF Research Database (Denmark)

Colombo, Emanuela; Bonetti, Paola; Lazzerini Denchi, Eros

2005-01-01

, such as mutated Ras or overexpressed Myc. In the absence of NPM, Arf protein is excluded from nucleoli and is markedly less stable. Our data demonstrate that NPM regulates DNA integrity and, through Arf, inhibits cell proliferation and are consistent with a putative tumor-suppressive function of NPM....

Members of miR-169 family are induced by high salinity and transiently inhibit the NF-YA transcription factor

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Lin Hongxuan

2009-04-01

Full Text Available Abstract Background MicroRNAs (miRNAs are endogenously expressed small RNAs with a length of about 21 nt. MiRNAs silence their target genes at the post-transcriptional level. In plants, miRNAs play various developmental and physiological roles by cleavaging mRNAs predominantly. Drought and high salinity are the most severe environmental abiotic stresses and cause crop losses all over the world. Results In this study, we identified miR-169g and miR-169n (o as high salinity-responsive miRNAs in rice. MiR-169n and miR169o were in a miRNA cluster with a distance of 3707 base pairs (bp. The high degree of conservation and close phylogenic distance of pre-miR-169n and pre-miR-169o indicated that they were derived from a very recent tandem duplication evolutionary event. The existence of a cis-acting abscisic acid responsive element (ABRE in the upstream region of miR-169n (o suggested that miR-169n (o may be regulated by ABA. In our previous study, we found that miR-169g was induced by the osmotic stress caused by drought via a dehydration-responsive element (DRE. Thus, our data showed that there were both overlapping and distinct responses of the miR-169 family to drought and salt stresses. We also showed that these miR-169 members selectively cleaved one of the NF-YA genes, Os03g29760, which is a CCAAT-box binding transcription factor and participates in transcriptional regulation of large number genes. Finally, we found one or more ath-miR-169 member that was also induced by high salinity. Conclusion We identified members of the miR-169 family as salt-induced miRNAs and analyzed their evolution, gene organization, expression, transcriptional regulation motif and target gene. Our data also indicated that the salt-induction of some miR-169 members was a general property in plants.

Members of miR-169 family are induced by high salinity and transiently inhibit the NF-YA transcription factor.

Science.gov (United States)

Zhao, Botao; Ge, Liangfa; Liang, Ruqiang; Li, Wei; Ruan, Kangcheng; Lin, Hongxuan; Jin, Youxin

2009-04-08

MicroRNAs (miRNAs) are endogenously expressed small RNAs with a length of about 21 nt. MiRNAs silence their target genes at the post-transcriptional level. In plants, miRNAs play various developmental and physiological roles by cleavaging mRNAs predominantly. Drought and high salinity are the most severe environmental abiotic stresses and cause crop losses all over the world. In this study, we identified miR-169g and miR-169n (o) as high salinity-responsive miRNAs in rice. MiR-169n and miR169o were in a miRNA cluster with a distance of 3707 base pairs (bp). The high degree of conservation and close phylogenic distance of pre-miR-169n and pre-miR-169o indicated that they were derived from a very recent tandem duplication evolutionary event. The existence of a cis-acting abscisic acid responsive element (ABRE) in the upstream region of miR-169n (o) suggested that miR-169n (o) may be regulated by ABA. In our previous study, we found that miR-169g was induced by the osmotic stress caused by drought via a dehydration-responsive element (DRE). Thus, our data showed that there were both overlapping and distinct responses of the miR-169 family to drought and salt stresses. We also showed that these miR-169 members selectively cleaved one of the NF-YA genes, Os03g29760, which is a CCAAT-box binding transcription factor and participates in transcriptional regulation of large number genes. Finally, we found one or more ath-miR-169 member that was also induced by high salinity. We identified members of the miR-169 family as salt-induced miRNAs and analyzed their evolution, gene organization, expression, transcriptional regulation motif and target gene. Our data also indicated that the salt-induction of some miR-169 members was a general property in plants.

The Transcription Factor OsWRKY45 Negatively Modulates the Resistance of Rice to the Brown Planthopper Nilaparvata lugens

OpenAIRE

Huangfu, Jiayi; Li, Jiancai; Li, Ran; Ye, Meng; Kuai, Peng; Zhang, Tongfang; Lou, Yonggen

2016-01-01

WRKY transcription factors play a central role not only in plant growth and development but also in plant stress responses. However, the role of WRKY transcription factors in herbivore-induced plant defenses and their underlying mechanisms, especially in rice, remains largely unclear. Here, we cloned a rice WRKY gene OsWRKY45, whose expression was induced by mechanical wounding, by infestation of the brown planthopper (BPH, Nilaparvata lugens) and by treatment with jasmonic acid (JA) or salic...

Jasmonate-responsive transcription factors regulating plant secondary metabolism.

Science.gov (United States)

Zhou, Meiliang; Memelink, Johan

2016-01-01

Plants produce a large variety of secondary metabolites including alkaloids, glucosinolates, terpenoids and phenylpropanoids. These compounds play key roles in plant-environment interactions and many of them have pharmacological activity in humans. Jasmonates (JAs) are plant hormones which induce biosynthesis of many secondary metabolites. JAs-responsive transcription factors (TFs) that regulate the JAs-induced accumulation of secondary metabolites belong to different families including AP2/ERF, bHLH, MYB and WRKY. Here, we give an overview of the types and functions of TFs that have been identified in JAs-induced secondary metabolite biosynthesis, and highlight their similarities and differences in regulating various biosynthetic pathways. We review major recent developments regarding JAs-responsive TFs mediating secondary metabolite biosynthesis, and provide suggestions for further studies. Copyright © 2016 Elsevier Inc. All rights reserved.

Comparison of KrF and ArF excimer laser treatment of biopolymer surface

Energy Technology Data Exchange (ETDEWEB)

Michaljaničová, I. [Department of Solid State Engineering, University of Chemistry and Technology, 166 28 Prague (Czech Republic); Slepička, P., E-mail: petr.slepicka@vscht.cz [Department of Solid State Engineering, University of Chemistry and Technology, 166 28 Prague (Czech Republic); Heitz, J.; Barb, R.A. [Institute of Applied Physics, Johannes Kepler University Linz, A-4040 Linz (Austria); Sajdl, P. [Department of Power Engineering, University of Chemistry and Technology, 166 28 Prague (Czech Republic); Švorčík, V. [Department of Solid State Engineering, University of Chemistry and Technology, 166 28 Prague (Czech Republic)

2015-06-01

Highlights: • The influence of ArF and KrF laser on biopolymer surface was determined. • ArF laser acts predominantly on biopolymer surface. • PHB roughness is increased similarly for both applied wavelengths. • Roughness of nanostructures can be precisely controlled. • ArF laser introduces nitrogen on PHB surface. - Abstract: The goal of this work was the investigation of the impact of two different excimer lasers on two biocompatible and biodegradable polymers (poly-L-lactide and poly hydroxybutyrate). Both polymers find usage in medical and pharmaceutical fields. The polymers were modified by KrF and ArF excimer lasers. Subsequently the impact on surface morphology, surface chemistry changes, and thermal properties was studied by means of confocal and AFM microscopy, FTIR and XPS spectroscopy and DSC calorimetry. Under the same conditions of laser treatment it was observed that ArF laser causes more significant changes on surface chemistry, surface morphology and pattern formation on the polymers under investigation. The data obtained in this work can be used for a wide range of possible applications, in tissue engineering or in combination with metallization in electronics, e.g. for biosensors.

MiT/TFE transcription factors are activated during mitophagy downstream of Parkin and Atg5.

Science.gov (United States)

Nezich, Catherine L; Wang, Chunxin; Fogel, Adam I; Youle, Richard J

2015-08-03

The kinase PINK1 and ubiquitin ligase Parkin can regulate the selective elimination of damaged mitochondria through autophagy (mitophagy). Because of the demand on lysosomal function by mitophagy, we investigated a role for the transcription factor EB (TFEB), a master regulator of lysosomal biogenesis, in this process. We show that during mitophagy TFEB translocates to the nucleus and displays transcriptional activity in a PINK1- and Parkin-dependent manner. MITF and TFE3, homologues of TFEB belonging to the same microphthalmia/transcription factor E (MiT/TFE) family, are similarly regulated during mitophagy. Unlike TFEB translocation after starvation-induced mammalian target of rapamycin complex 1 inhibition, Parkin-mediated TFEB relocalization required Atg9A and Atg5 activity. However, constitutively active Rag guanosine triphosphatases prevented TFEB translocation during mitophagy, suggesting cross talk between these two MiT/TFE activation pathways. Analysis of clustered regularly interspaced short palindromic repeats-generated TFEB/MITF/TFE3/TFEC single, double, and triple knockout cell lines revealed that these proteins partly facilitate Parkin-mediated mitochondrial clearance. These results illuminate a pathway leading to MiT/TFE transcription factor activation, distinct from starvation-induced autophagy, which occurs during mitophagy.

Cloning of an ADP-ribosylation factor gene from banana (Musa acuminata) and its expression patterns in postharvest ripening fruit.

Science.gov (United States)

Wang, Yuan; Wu, Jing; Xu, Bi-Yu; Liu, Ju-Hua; Zhang, Jian-Bin; Jia, Cai-Hong; Jin, Zhi-Qiang

2010-08-15

A full-length cDNA encoding an ADP-ribosylation factor (ARF) from banana (Musa acuminata) fruit was cloned and named MaArf. It contains an open reading frame encoding a 181-amino-acid polypeptide. Sequence analysis showed that MaArf shared high similarity with ARF of other plant species. The genomic sequence of MaArf was also obtained using polymerase chain reaction (PCR). Sequence analysis showed that MaArf was a split gene containing five exons and four introns in genomic DNA. Reverse-transcriptase PCR was used to analyze the spatial expression of MaArf. The results showed that MaArf was expressed in all the organs examined: root, rhizome, leaf, flower and fruit. Real-time quantitative PCR was used to explore expression patterns of MaArf in postharvest banana. There was differential expression of MaArf associated with ethylene biosynthesis. In naturally ripened banana, expression of MaArf was in accordance with ethylene biosynthesis. However, in 1-methylcyclopropene-treated banana, the expression of MaArf was inhibited and changed little. When treated with ethylene, MaArf expression in banana fruit significantly increased in accordance with ethylene biosynthesis; the peak of MaArf was 3 d after harvest, 11 d earlier than for naturally ripened banana fruits. These results suggest that MaArf is induced by ethylene in regulating postharvest banana ripening. Finally, subcellular localization assays showed the MaArf protein in the cytoplasm. Copyright 2010 Elsevier GmbH. All rights reserved.

The DNA damage checkpoint precedes activation of ARF in response to escalating oncogenic stress during tumorigenesis

Czech Academy of Sciences Publication Activity Database

Evangelou, K.; Bartkova, J.; Kotsinas, A.; Pateras, I.S.; Liontos, M.; Velimezi, G.; Košař, Martin; Liloglou, T.; Trougakos, I.P.; Dyrskjot, L.; Andersen, C.J.; Papaioannou, G.; Drosos, A.; Papafotiou, M.; Hodný, Zdeněk; Sosa-Pineda, B.; Wu, X.R.; Klinakis, A.; Orntoft, T.; Lukas, J.; Bartek, Jiří; Gorgoulis, V.G.

2013-01-01

Roč. 20, č. 11 (2013), s. 1485-1497 ISSN 1350-9047 Institutional support: RVO:68378050 Keywords : ARF * carcinogenesis * DDR * E2F1 * p16(INK4A) Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 8.385, year: 2013

Arabidopsis R2R3-MYB transcription factor AtMYB60 functions as a transcriptional repressor of anthocyanin biosynthesis in lettuce (Lactuca sativa).

Science.gov (United States)

Park, Jong-Sug; Kim, Jung-Bong; Cho, Kang-Jin; Cheon, Choong-Ill; Sung, Mi-Kyung; Choung, Myoung-Gun; Roh, Kyung-Hee

2008-06-01

The MYB transcription factors play important roles in the regulation of many secondary metabolites at the transcriptional level. We evaluated the possible roles of the Arabidopsis R2R3-MYB transcription factors in flavonoid biosynthesis because they are induced by UV-B irradiation but their associated phenotypes are largely unexplored. We isolated their genes by RACE-PCR, and performed transgenic approach and metabolite analyses in lettuce (Lactuca sativa). We found that one member of this protein family, AtMYB60, inhibits anthocyanin biosynthesis in the lettuce plant. Wild-type lettuce normally accumulates anthocyanin, predominantly cyanidin and traces of delphinidin, and develops a red pigmentation. However, the production and accumulation of anthocyanin pigments in AtMYB60-overexpressing lettuce was inhibited. Using RT-PCR analysis, we also identified the complete absence or reduction of dihydroflavonol 4-reductase (DFR) transcripts in AtMYB60- overexpressing lettuce (AtMYB60-117 and AtMYB60-112 lines). The correlation between the overexpression of AtMYB60 and the inhibition of anthocyanin accumulation suggests that the transcription factorAtMYB60 controls anthocyanin biosynthesis in the lettuce leaf. Clarification of the roles of the AtMYB60 transcription factor will facilitate further studies and provide genetic tools to better understand the regulation in plants of the genes controlled by the MYB-type transcription factors. Furthermore, the characterization of AtMYB60 has implications for the development of new varieties of lettuce and other commercially important plants with metabolic engineering approaches.

Increased accumulation of hypoxia-inducible factor-1α with reduced transcriptional activity mediates the antitumor effect of triptolide

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Li Zheng

2010-10-01

Full Text Available Abstract Background Hypoxia-inducible factor-1α (HIF-1α, a critical transcription factor to reduced O2 availability, has been demonstrated to be extensively involved in tumor survival, aggressive progression, drug resistance and angiogenesis. Thus it has been considered as a potential anticancer target. Triptolide is the main principle responsible for the biological activities of the Traditional Chinese Medicine tripterygium wilfordii Hook F. Triptolide possesses great chemotherapy potential for cancer with its broad-spectrum anticancer, antiangiogenesis, and drug-resistance circumvention activities. Numerous biological molecules inhibited by triptolide have been viewed as its possible targets. However, the anticancer action mechanisms of triptolide remains to be further investigated. Here we used human ovarian SKOV-3 cancer cells as a model to probe the effect of triptolide on HIF-1α. Results Triptolide was observed to inhibit the proliferation of SKOV-3 cells, and meanwhile, to enhance the accumulation of HIF-1α protein in SKOV-3, A549 and DU145 cells under different conditions. Triptolide did not change the kinetics or nuclear localization of HIF-1α protein or the 26 S proteasome activity in SKOV-3 cells. However, triptolide was found to increase the levels of HIF-1α mRNA. Unexpectedly, the HIF-1α protein induced by triptolide appeared to lose its transcriptional activity, as evidenced by the decreased mRNA levels of its target genes including VEGF, BNIP3 and CAIX. The results were further strengthened by the lowered secretion of VEGF protein, the reduced sprout outgrowth from the rat aorta rings and the inhibitory expression of the hypoxia responsive element-driven luciferase reporter gene. Moreover, the silencing of HIF-1α partially prevented the cytotoxicity and apoptosis triggered by triptolide. Conclusions The potent induction of HIF-1α protein involved in its cytotoxicity, together with the suppression of HIF-1 transcriptional

Specificity versus redundancy in the RAP2.4 transcription factor family of Arabidopsis thaliana: transcriptional regulation of genes for chloroplast peroxidases.

Science.gov (United States)

Rudnik, Radoslaw; Bulcha, Jote Tafese; Reifschneider, Elena; Ellersiek, Ulrike; Baier, Margarete

2017-08-23

The Arabidopsis ERFIb / RAP2.4 transcription factor family consists of eight members with highly conserved DNA binding domains. Selected members have been characterized individually, but a systematic comparison is pending. The redox-sensitive transcription factor RAP2.4a mediates chloroplast-to-nucleus redox signaling and controls induction of the three most prominent chloroplast peroxidases, namely 2-Cys peroxiredoxin A (2CPA) and thylakoid- and stromal ascorbate peroxidase (tAPx and sAPx). To test the specificity and redundancy of RAP2.4 transcription factors in the regulation of genes for chloroplast peroxidases, we compared the DNA-binding sites of the transcription factors in tertiary structure models, analyzed transcription factor and target gene regulation by qRT-PCR in RAP2.4, 2-Cys peroxiredoxin and ascorbate peroxidase T-DNA insertion lines and RAP2.4 overexpressing lines of Arabidopsis thaliana and performed promoter binding studies. All RAP2.4 proteins bound the tAPx promoter, but only the four RAP2.4 proteins with identical DNA contact sites, namely RAP2.4a, RAP2.4b, RAP2.4d and RAP2.4h, interacted stably with the redox-sensitive part of the 2CPA promoter. Gene expression analysis in RAP2.4 knockout lines revealed that RAP2.4a is the only one supporting 2CPA and chloroplast APx expression. Rap2.4h binds to the same promoter region as Rap2.4a and antagonizes 2CPA expression. Like the other six RAP2.4 proteins, Rap2.4 h promotes APx mRNA accumulation. Chloroplast ROS signals induced RAP2.4b and RAP2.4d expression, but these two transcription factor genes are (in contrast to RAP2.4a) insensitive to low 2CP availability, and their expression decreased in APx knockout lines. RAP2.4e and RAP2.4f gradually responded to chloroplast APx availability and activated specifically APx expression. These transcription factors bound, like RAP2.4c and RAP2.4g, the tAPx promoter, but hardly the 2CPA promoter. The RAP2.4 transcription factors form an environmentally and

Roles of Arabidopsis WRKY3 and WRKY4 Transcription Factors in Plant Responses to Pathogens

Directory of Open Access Journals (Sweden)

Fan Baofang

2008-06-01

Full Text Available Abstract Background Plant WRKY DNA-binding transcription factors are involved in plant responses to biotic and abiotic responses. It has been previously shown that Arabidopsis WRKY3 and WRKY4, which encode two structurally similar WRKY transcription factors, are induced by pathogen infection and salicylic acid (SA. However, the role of the two WRKY transcription factors in plant disease resistance has not been directly analyzed. Results Both WRKY3 and WRKY4 are nuclear-localized and specifically recognize the TTGACC W-box sequences in vitro. Expression of WRKY3 and WRKY4 was induced rapidly by stress conditions generated by liquid infiltration or spraying. Stress-induced expression of WRKY4 was further elevated by pathogen infection and SA treatment. To determine directly their role in plant disease resistance, we have isolated T-DNA insertion mutants and generated transgenic overexpression lines for WRKY3 and WRKY4. Both the loss-of-function mutants and transgenic overexpression lines were examined for responses to the biotrophic bacterial pathogen Pseudomonas syringae and the necrotrophic fungal pathogen Botrytis cinerea. The wrky3 and wrky4 single and double mutants exhibited more severe disease symptoms and support higher fungal growth than wild-type plants after Botrytis infection. Although disruption of WRKY3 and WRKY4 did not have a major effect on plant response to P. syringae, overexpression of WRKY4 greatly enhanced plant susceptibility to the bacterial pathogen and suppressed pathogen-induced PR1 gene expression. Conclusion The nuclear localization and sequence-specific DNA-binding activity support that WRKY3 and WRKY4 function as transcription factors. Functional analysis based on T-DNA insertion mutants and transgenic overexpression lines indicates that WRKY3 and WRKY4 have a positive role in plant resistance to necrotrophic pathogens and WRKY4 has a negative effect on plant resistance to biotrophic pathogens.

Cisplatin Induces Cytotoxicity through the Mitogen-Activated Protein Kinase Pathways ana Activating Transcription Factor 3

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Carly St. Germain

2010-07-01

Full Text Available The mechanisms underlying the proapoptotic effect of the chemotherapeutic agent, cisplatin, are largely undefined. Understanding the mechanisms regulating cisplatin cytotoxicity may uncover strategies to enhance the efficacy of this important therapeutic agent. This study evaluates the role of activating transcription factor 3 (ATF3 as a mediator of cisplatin-induced cytotoxicity. Cytotoxic doses of cisplatin and carboplatin treatments consistently induced ATF3 expression in five tumor-derived cell lines. Characterization of this induction revealed a p53, BRCA1, and integrated stress response-independent mechanism, all previously implicated in stress-mediated ATF3 induction. Analysis of mitogenactivated protein kinase (MAPK pathway involvement in ATF3 induction by cisplatin revealed a MAPK-dependent mechanism. Cisplatin treatment combined with specific inhibitors to each MAPK pathway (c-Jun N-terminal kinase, extracellularsignal-regulated kinase, and p38 resulted in decreasedATF3 induction at the protein level. MAPK pathway inhibition led to decreased ATF3 messenger RNA expression and reduced cytotoxic effects of cisplatin as measured by the 3-(4,5-dimethylthiazol-2-ylF2,5-diphenyltetrazolium bromide cell viability assay. In A549 lung carcinoma cells, targeting ATF3 with specific small hairpin RNA also attenuated the cytotoxic effects of cisplatin. Similarly, ATF3-/murine embryonic fibroblasts (MEFs were shown to be less sensitive to cisplatin-induced cytotoxicity compared with ATF3+/+ MEFs. This study identifies cisplatin as a MAPK pathway-dependent inducer of ATF3, whose expression influences cisplatin’s cytotoxic effects.

Triptolide inhibits transcription of hTERT through down-regulation of transcription factor specificity protein 1 in primary effusion lymphoma cells

Energy Technology Data Exchange (ETDEWEB)

Long, Cong; Wang, Jingchao [Department of Pathogen Biology, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071 (China); Guo, Wei [Department of Pathology and Physiology, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071 (China); Wang, Huan; Wang, Chao; Liu, Yu [Department of Pathogen Biology, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071 (China); Sun, Xiaoping, E-mail: xsun6@whu.edu.cn [Department of Pathogen Biology, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071 (China); State Key Laboratory of Virology, Wuhan University, Wuhan, 430072 (China)

2016-01-01

Primary effusion lymphoma (PEL) is a rare and aggressive non-Hodgkin's lymphoma. Human telomerase reverse transcriptase (hTERT), a key component responsible for the regulation of telomerase activity, plays important roles in cellular immortalization and cancer development. Triptolide purified from Tripterygium extracts displays a broad-spectrum bioactivity profile, including immunosuppressive, anti-inflammatory, and anti-tumor. In this study, it is investigated whether triptolide reduces hTERT expression and suppresses its activity in PEL cells. The mRNA and protein levels of hTERT were examined by real time-PCR and Western blotting, respectively. The activity of hTERT promoter was determined by Dual luciferase reporter assay. Our results demonstrated that triptolide decreased expression of hTERT at both mRNA and protein levels. Further gene sequence analysis indicated that the activity of hTERT promoter was suppressed by triptolide. Triptolide also reduced the half-time of hTERT. Additionally, triptolide inhibited the expression of transcription factor specificity protein 1(Sp1) in PEL cells. Furthermore, knock-down of Sp1 by using specific shRNAs resulted in down-regulation of hTERT transcription and protein expression levels. Inhibition of Sp1 by specific shRNAs enhanced triptolide-induced cell growth inhibition and apoptosis. Collectively, our results demonstrate that the inhibitory effect of triptolide on hTERT transcription is possibly mediated by inhibition of transcription factor Sp1 in PEL cells. - Highlights: • Triptolide reduces expression of hTERT by decreasing its transcription level. • Triptolide reduces promoter activity and stability of hTERT. • Triptolide down-regulates expression of Sp1. • Special Sp1 shRNAs inhibit transcription and protein expression of hTERT. • Triptolide and Sp1 shRNA2 induce cell proliferation inhibition and apoptosis.

NAC transcription factors: structurally distinct, functionally diverse

DEFF Research Database (Denmark)

Olsen, Addie Nina; Ernst, Heidi A; Leggio, Leila Lo

2005-01-01

level and localization, and to the first indications of NAC participation in transcription factor networks. The recent determination of the DNA and protein binding NAC domain structure offers insight into the molecular functions of the protein family. Research into NAC transcription factors has......NAC proteins constitute one of the largest families of plant-specific transcription factors, and the family is present in a wide range of land plants. Here, we summarize the biological and molecular functions of the NAC family, paying particular attention to the intricate regulation of NAC protein...

Assessing the Role of ETHYLENE RESPONSE FACTOR Transcriptional Repressors in Salicylic Acid-Mediated Suppression of Jasmonic Acid-Responsive Genes.

Science.gov (United States)

Caarls, Lotte; Van der Does, Dieuwertje; Hickman, Richard; Jansen, Wouter; Verk, Marcel C Van; Proietti, Silvia; Lorenzo, Oscar; Solano, Roberto; Pieterse, Corné M J; Van Wees, Saskia C M

2017-02-01

Salicylic acid (SA) and jasmonic acid (JA) cross-communicate in the plant immune signaling network to finely regulate induced defenses. In Arabidopsis, SA antagonizes many JA-responsive genes, partly by targeting the ETHYLENE RESPONSE FACTOR (ERF)-type transcriptional activator ORA59. Members of the ERF transcription factor family typically bind to GCC-box motifs in the promoters of JA- and ethylene-responsive genes, thereby positively or negatively regulating their expression. The GCC-box motif is sufficient for SA-mediated suppression of JA-responsive gene expression. Here, we investigated whether SA-induced ERF-type transcriptional repressors, which may compete with JA-induced ERF-type activators for binding at the GCC-box, play a role in SA/JA antagonism. We selected ERFs that are transcriptionally induced by SA and/or possess an EAR transcriptional repressor motif. Several of the 16 ERFs tested suppressed JA-dependent gene expression, as revealed by enhanced JA-induced PDF1.2 or VSP2 expression levels in the corresponding erf mutants, while others were involved in activation of these genes. However, SA could antagonize JA-induced PDF1.2 or VSP2 in all erf mutants, suggesting that the tested ERF transcriptional repressors are not required for SA/JA cross-talk. Moreover, a mutant in the co-repressor TOPLESS, that showed reduction in repression of JA signaling, still displayed SA-mediated antagonism of PDF1.2 and VSP2. Collectively, these results suggest that SA-regulated ERF transcriptional repressors are not essential for antagonism of JA-responsive gene expression by SA. We further show that de novo SA-induced protein synthesis is required for suppression of JA-induced PDF1.2, pointing to SA-stimulated production of an as yet unknown protein that suppresses JA-induced transcription. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

A transcription factor active on the epidermal growth factor receptor gene

International Nuclear Information System (INIS)

Kageyama, R.; Merlino, G.T.; Pastan, I.

1988-01-01

The authors have developed an in vitro transcription system for the epidermal growth factor receptor (EGFR) oncogene by using nuclear extracts of A431 human epidermoid carcinoma cells, which overproduce EGFR. They found that a nuclear factor, termed EGFR-specific transcription factor (ETF), specifically stimulated EGFR transcription by 5- to 10-fold. In this report, ETF, purified by using sequence-specific oligonucleotide affinity chromatography, is shown by renaturing material eluted from a NaDodSO 4 /polyacrylamide gel to be a protein with a molecular mass of 120 kDa. ETF binds to the promoter region, as measured by DNase I footprinting and gel-mobility-shift assays, and specifically stimulates the transcription of the EGFR gene in a reconstituted in vitro transcription system. These results suggest that ETF could play a role in the overexpression of the cellular oncogene EGFR

Interaction between the triglyceride lipase ATGL and the arf1 activator GBF1

KAUST Repository

Ellong, Emy Njoh

2011-07-18

The Arf1 exchange factor GBF1 (Golgi Brefeldin A resistance factor 1) and its effector COPI are required for delivery of ATGL (adipose triglyceride lipase) to lipid droplets (LDs). Using yeast two hybrid, co-immunoprecipitation in mammalian cells and direct protein binding approaches, we report here that GBF1 and ATGL interact directly and in cells, through multiple contact sites on each protein. The C-terminal region of ATGL interacts with N-terminal domains of GBF1, including the catalytic Sec7 domain, but not with full-length GBF1 or its entire N-terminus. The N-terminal lipase domain of ATGL (called the patatin domain) interacts with two C-terminal domains of GBF1, HDS (Homology downstream of Sec7) 1 and HDS2. These two domains of GBF1 localize to lipid droplets when expressed alone in cells, but not to the Golgi, unlike the full-length GBF1 protein, which localizes to both. We suggest that interaction of GBF1 with ATGL may be involved in the membrane trafficking pathway mediated by GBF1, Arf1 and COPI that contributes to the localization of ATGL to lipid droplets.

Interaction between the triglyceride lipase ATGL and the arf1 activator GBF1

KAUST Repository

Ellong, Emy Njoh; Soni, Krishnakant G.; Bui, Quynh-Trang; Sougrat, Rachid; Golinelli-Cohen, Marie-Pierre; Jackson, Catherine L.

2011-01-01

The Arf1 exchange factor GBF1 (Golgi Brefeldin A resistance factor 1) and its effector COPI are required for delivery of ATGL (adipose triglyceride lipase) to lipid droplets (LDs). Using yeast two hybrid, co-immunoprecipitation in mammalian cells and direct protein binding approaches, we report here that GBF1 and ATGL interact directly and in cells, through multiple contact sites on each protein. The C-terminal region of ATGL interacts with N-terminal domains of GBF1, including the catalytic Sec7 domain, but not with full-length GBF1 or its entire N-terminus. The N-terminal lipase domain of ATGL (called the patatin domain) interacts with two C-terminal domains of GBF1, HDS (Homology downstream of Sec7) 1 and HDS2. These two domains of GBF1 localize to lipid droplets when expressed alone in cells, but not to the Golgi, unlike the full-length GBF1 protein, which localizes to both. We suggest that interaction of GBF1 with ATGL may be involved in the membrane trafficking pathway mediated by GBF1, Arf1 and COPI that contributes to the localization of ATGL to lipid droplets.

The H3K27me3 demethylase JMJD3 contributes to the activation of the INK4A-ARF locus in response to oncogene- and stress-induced senescence

DEFF Research Database (Denmark)

Agger, Karl; Cloos, Paul A C; Rudkjaer, Lise

2009-01-01

The tumor suppressor proteins p16INK4A and p14ARF, encoded by the INK4A-ARF locus, are key regulators of cellular senescence. The locus is epigenetically silenced by the repressive H3K27me3 mark in normally growing cells, but becomes activated in response to oncogenic stress. Here, we show that e...... in mouse embryonic fibroblasts results in suppression of p16Ink4a and p19Arf expression and in their immortalization....

ATF1 Modulates the Heat Shock Response by Regulating the Stress-Inducible Heat Shock Factor 1 Transcription Complex

Science.gov (United States)

Takii, Ryosuke; Fujimoto, Mitsuaki; Tan, Ke; Takaki, Eiichi; Hayashida, Naoki; Nakato, Ryuichiro; Shirahige, Katsuhiko

2014-01-01

The heat shock response is an evolutionally conserved adaptive response to high temperatures that controls proteostasis capacity and is regulated mainly by an ancient heat shock factor (HSF). However, the regulation of target genes by the stress-inducible HSF1 transcription complex has not yet been examined in detail in mammalian cells. In the present study, we demonstrated that HSF1 interacted with members of the ATF1/CREB family involved in metabolic homeostasis and recruited them on the HSP70 promoter in response to heat shock. The HSF1 transcription complex, including the chromatin-remodeling factor BRG1 and lysine acetyltransferases p300 and CREB-binding protein (CBP), was formed in a manner that was dependent on the phosphorylation of ATF1. ATF1-BRG1 promoted the establishment of an active chromatin state and HSP70 expression during heat shock, whereas ATF1-p300/CBP accelerated the shutdown of HSF1 DNA-binding activity during recovery from acute stress, possibly through the acetylation of HSF1. Furthermore, ATF1 markedly affected the resistance to heat shock. These results revealed the unanticipated complexity of the primitive heat shock response mechanism, which is connected to metabolic adaptation. PMID:25312646

Mechanisms of inhibition of zinc-finger transcription factors by selenium compounds ebselen and selenite.

Science.gov (United States)

Larabee, Jason L; Hocker, James R; Hanas, Jay S

2009-03-01

The anti-inflammatory selenium compounds, ebselen (2-phenyl-1,2-benzisoselenazol-3[2H]-one) and selenite, were found to alter the DNA binding mechanisms and structures of cysteine-rich zinc-finger transcription factors. As assayed by DNase I protection, DNA binding by TFIIIA (transcription factor IIIA, prototypical Cys(2)His(2) zinc finger protein), was inhibited by micromolar amounts of ebselen. In a gel shift assay, ebselen inhibited the Cys(2)His(2) zinc finger-containing DNA binding domain (DBD) of the NF-kappaB mediated transcription factor Sp1. Ebselen also inhibited DNA binding by the p50 subunit of the pro-inflammatory Cys-containing NF-kappaB transcription factor. Electrospray ionization mass spectrometry (ESI-MS) was utilized to elucidate mechanisms of chemical interaction between ebselen and a zinc-bound Cys(2)His(2) zinc finger polypeptide modeled after the third finger of Sp1 (Sp1-3). Exposing Sp1-3 to micromolar amounts of ebselen resulted in Zn(2+) release from this peptide and the formation of a disulfide bond by oxidation of zinc finger SH groups, the likely mechanism for DNA binding inhibition. Selenite was shown by ESI-MS to also eject zinc from Sp1-3 as well as induce disulfide bond formation through SH oxidation. The selenite-dependent inhibition/oxidation mechanism differed from that of ebselen by inducing the formation of a stable selenotrisulfide bond. Selenite-induced selenotrisulfide formation was dependent upon the structure of the Cys(2)His(2) zinc finger as alteration in the finger structure enhanced this reaction as well as selenite-dependent zinc release. Ebselen and selenite-dependent inhibition/oxidation of Cys-rich zinc finger proteins, with concomitant release of zinc and finger structural changes, points to mechanisms at the atomic and protein level for selenium-induced alterations in Cys-rich proteins, and possible amelioration of certain inflammatory, neurodegenerative, and oncogenic responses.

Expression of transcription factor Pokemon in non-small cell lung cancer and its clinical significance.

Science.gov (United States)

Zhao, Zhi-hong; Wang, Sheng-fa; Yu, Liang; Wang, Ju; Chang, Hao; Yan, Wei-li; Fu, Kai; Zhang, Jian

2008-03-05

Transcription factor Pokemon, a central regulation gene of the important tumor suppressor ARF gene, exerted its activity by acting upstream of many tumor-suppressing genes and proto-oncogenes. Its expression in non-small cell lung cancer (NSCLC) and its clinical significance remains unclear. The aim of this study was to investigate the expression of Pokemon in NSCLC and to explore its correlation with the clinical pathological characteristics and its influence on patients' prognosis. Fifty-five cases of NSCLC were involved in this study. The expression of Pokemon in the tumor tissue, the corresponding tumor adjacent tissue and the surrounding tissue was detected via reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, with the aim of investigating the correlation between the expression of Pokemon in tumor tissue of NSCLC and its clinical pathological characteristics. Moreover, a prognostic analysis was carried out based upon the immunohistochemical (IHC) detection of the expression of Pokemon gene in archival tumor specimens (5 years ago) of 62 cases of NSCLC. Statistical significance of the expression of Pokemon mRNA and protein was determined in the tumor tissue, the tumor adjacent tissue and the surrounding tissue (PPokemon was determined not to be associated with the patients' sex, age, smoking condition, tumor differentiation degree, histology and lymph node metastasis condition. However, its relationship with TNM staging was established (PPokemon expression was significantly higher than that of those with positive Pokemon expression (P=0.004), therefore, the expression of Pokemon is believed to be an independent factor affecting prognosis (P=0.034). Pokemon was over-expressed in NSCLC tissue and the expression of Pokemon might be of clinical significance in non-small cell lung cancer prognostic evaluation.

Acute Alcohol Intoxication Exacerbates Rhabdomyolysis-Induced Acute Renal Failure in Rats.

Science.gov (United States)

Tsai, Jen-Pi; Lee, Chung-Jen; Subeq, Yi-Maun; Lee, Ru-Ping; Hsu, Bang-Gee

2017-01-01

Traumatic and nontraumatic rhabdomyolysis can lead to acute renal failure (ARF), and acute alcohol intoxication can lead to multiple abnormalities of the renal tubules. We examined the effect of acute alcohol intoxication in a rat model of rhabdomyolysis and ARF. Intravenous injections of 5 g/kg ethanol were given to rats over 3 h, followed by glycerol-induced rhabdomyolysis. Biochemical parameters, including blood urea nitrogen (BUN), creatinine (Cre), glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), and creatine phosphokinase (CPK), were measured before and after induction of rhabdomyolysis. Renal tissue injury score, renal tubular cell expression of E-cadherin, nuclear factor-κB (NF-κB), and inducible nitric oxide synthase (iNOS) were determined. Relative to rats in the vehicle group, rats in the glycerol-induced rhabdomyolysis group had significantly increased serum levels of BUN, Cre, GOT, GPT, and CPK, elevated renal tissue injury scores, increased expression of NF-κB and iNOS, and decreased expression of E-cadherin. Ethanol exacerbated all of these pathological responses. Our results suggest that acute alcohol intoxication exacerbates rhabdomyolysis-induced ARF through its pro-oxidant and inflammatory effects.

Activating transcription factor 6 mediates oxidized LDL-induced cholesterol accumulation and apoptosis in macrophages by up-regulating CHOP expression.

Science.gov (United States)

Yao, Shutong; Zong, Chuanlong; Zhang, Ying; Sang, Hui; Yang, Mingfeng; Jiao, Peng; Fang, Yongqi; Yang, Nana; Song, Guohua; Qin, Shucun

2013-01-01

This study was to explore whether activating transcription factor 6 (ATF6), an important sensor to endoplasmic reticulum (ER) stress, would mediate oxidized low-density lipoprotein (ox-LDL)- induced cholesterol accumulation and apoptosis in cultured macrophages and the underlying molecular mechanisms. Intracellular lipid droplets and total cholesterol levels were assayed by oil red O staining and enzymatic colorimetry, respectively. Cell viability and apoptosis were determined using MTT assay and AnnexinV-FITC apoptosis detection kit, respectively. The nuclear translocation of ATF6 in cells was detected by immunofluorescence analysis. Protein and mRNA levels were examined by Western blot analysis and real time-PCR, respectively. ATF6 siRNA was transfected to RAW264.7 cells by lipofectamin. Exposure of cells to ox-LDL induced glucose-regulated protein 78 (GRP78). C/EBP homologous protein (CHOP), a key-signaling component of ER stress-induced apoptosis, was up-regulated in ox-LDL-treated cells. ATF6, a factor that positively regulates CHOP expression, was activated by ox-LDL in a concentration- and time- dependent manner. The role of the ATF6-mediated ER stress pathway was further confirmed through the siRNA-mediated knockdown of ATF6, which attenuated ox-LDL-induced upregulation of CHOP, cholesterol accumulation and apoptosis in macrophages. In addition, the phosphorylation of double-stranded RNA-activated protein kinase-like endoplasmic reticulum kinase (PERK), another factor that positively regulates CHOP expression, was induced in the presence of ox-LDL, and PERK-specific siRNA also inhibited the ox-LDL-induced upregulation of CHOP and apoptosis in RAW264.7 cells. These results demonstrate that ER stress-related proteins, particularly ATF6 and its downstream molecule CHOP, are involved in ox-LDL-induced cholesterol accumulation and apoptosis in macrophages.

Transcription Factors Expressed in Lateral Organ Boundaries: Identification of Downstream Targets

Energy Technology Data Exchange (ETDEWEB)

Springer, Patricia S

2010-07-12

The processes of lateral organ initiation and patterning are central to the generation of mature plant form. Characterization of the molecular mechanisms underlying these processes is essential to our understanding of plant development. Communication between the shoot apical meristem and initiating organ primordia is important both for functioning of the meristem and for proper organ patterning, and very little is known about this process. In particular, the boundary between meristem and leaf is emerging as a critical region that is important for SAM maintenance and regulation of organogenesis. The goal of this project was to characterize three boundary-expressed genes that encode predicted transcription factors. Specifically, we have studied LATERAL ORGAN BOUNDARIES (LOB), LATERAL ORGAN FUSION1 (LOF1), and LATERAL ORGAN FUSION2 (LOF2). LOB encodes the founding member of the LOB-DOMAIN (LBD) plant-specific DNA binding transcription factor family and LOF1 and LOF2 encode paralogous MYB-domain transcription factors. We characterized the genetic relationship between these three genes and other boundary and meristem genes. We also used an ectopic inducible expression system to identify direct targets of LOB.

Inducer-independent production of pectinases in Aspergillus niger by overexpression of the D-galacturonic acid-responsive transcription factor gaaR.

Science.gov (United States)

Alazi, Ebru; Knetsch, Tim; Di Falco, Marcos; Reid, Ian D; Arentshorst, Mark; Visser, Jaap; Tsang, Adrian; Ram, Arthur F J

2018-03-01

The transcription factor GaaR is needed for the expression of genes required for pectin degradation and transport and catabolism of the main degradation product, D-galacturonic acid (GA) in Aspergillus niger. In this study, we used the strong constitutive gpdA promoter of Aspergillus nidulans to overexpress gaaR in A. niger. Overexpression of gaaR resulted in an increased transcription of the genes encoding pectinases, (putative) GA transporters, and catabolic pathway enzymes even under non-inducing conditions, i.e., in the absence of GA. Exoproteome analysis of a strain overexpressing gaaR showed that this strain secretes highly elevated levels of pectinases when grown in fructose. The genes encoding exo-polygalacturonases were found to be subjected to CreA-mediated carbon catabolite repression, even in the presence of fructose. Deletion of creA in the strain overexpressing gaaR resulted in a further increase in pectinase production in fructose. We showed that GaaR localizes mainly in the nucleus regardless of the presence of an inducer, and that overexpression of gaaR leads to an increased concentration of GaaR in the nucleus.

The transcription factor ATF3 is upregulated during chondrocyte differentiation and represses cyclin D1 and A gene transcription

Directory of Open Access Journals (Sweden)

James Claudine G

2006-09-01

Full Text Available Abstract Background Coordinated chondrocyte proliferation and differentiation are required for normal endochondral bone growth. Transcription factors binding to the cyclicAMP response element (CRE are known to regulate these processes. One member of this family, Activating Tanscription Factor 3 (ATF3, is expressed during skeletogenesis and acts as a transcriptional repressor, but the function of this protein in chondrogenesis is unknown. Results Here we demonstrate that Atf3 mRNA levels increase during mouse chondrocyte differentiation in vitro and in vivo. In addition, Atf3 mRNA levels are increased in response to cytochalasin D treatment, an inducer of chondrocyte maturation. This is accompanied by increased Atf3 promoter activity in cytochalasin D-treated chondrocytes. We had shown earlier that transcription of the cell cycle genes cyclin D1 and cyclin A in chondrocytes is dependent on CREs. Here we demonstrate that overexpression of ATF3 in primary mouse chondrocytes results in reduced transcription of both genes, as well as decreased activity of a CRE reporter plasmid. Repression of cyclin A transcription by ATF3 required the CRE in the cyclin A promoter. In parallel, ATF3 overexpression reduces the activity of a SOX9-dependent promoter and increases the activity of a RUNX2-dependent promoter. Conclusion Our data suggest that transcriptional induction of the Atf3 gene in maturing chondrocytes results in down-regulation of cyclin D1 and cyclin A expression as well as activation of RUNX2-dependent transcription. Therefore, ATF3 induction appears to facilitate cell cycle exit and terminal differentiation of chondrocytes.

Aberrant DNA methylation of ESR1 and p14ARF genes could be useful as prognostic indicators in osteosarcoma

Directory of Open Access Journals (Sweden)

Sonaglio V

2013-06-01

Full Text Available Viviane Sonaglio,1 Ana C de Carvalho,2 Silvia R C Toledo,3,4 Carolina Salinas-Souza,3,4 André L Carvalho,5 Antonio S Petrilli,3 Beatriz de Camargo,6 André L Vettore21Pediatrics Department, A C Camargo Hospital, São Paulo, Brazil; 2Biological Science Department, Federal University of São Paulo, Diadema, Brazil; 3Department of Pediatrics, Pediatric Oncology Institute, GRAACC/Federal University of São Paulo, São Paulo, Brazil; 4Department of Morphology and Genetics, Federal University of São Paulo, São Paulo, Brazil; 5Department of Head and Neck Surgery, PIO XII Foundation, Barretos Cancer Hospital, Barretos, São Paulo, Brazil; 6Research Program Pediatric Oncology Program, CPNq, Instituto Nacional do Cancer, Rio de Janeiro, BrazilAbstract: Osteosarcoma (OS is the eighth most common form of childhood and adolescence cancer. Approximately 10%–20% of patients present metastatic disease at diagnosis and the 5-year overall survival remains around 70% for nonmetastatic patients and around 30% for metastatic patients. Metastatic disease at diagnosis and the necrosis grade induced by preoperative treatment are the only well-established prognostic factors for osteosarcoma. The DNA aberrant methylation is a frequent epigenetic alteration in humans and has been described as a molecular marker in different tumor types. This study evaluated the DNA aberrant methylation status of 18 genes in 34 OS samples without previous chemotherapy treatment and in four normal bone specimens and compared the methylation profile with clinicopathological characteristics of the patients. We were able to define a three-gene panel (AIM1, p14ARF, and ESR1 in which methylation was correlated with OS cases. The hypermethylation of p14ARF showed a significant association with the absence of metastases at diagnoses, while ESR1 hypermethylation was marginally associated with worse overall survival. This study demonstrated that aberrant promoter methylation is a common event

DNA residence time is a regulatory factor of transcription repression

Science.gov (United States)

Clauß, Karen; Popp, Achim P.; Schulze, Lena; Hettich, Johannes; Reisser, Matthias; Escoter Torres, Laura; Uhlenhaut, N. Henriette

2017-01-01

Abstract Transcription comprises a highly regulated sequence of intrinsically stochastic processes, resulting in bursts of transcription intermitted by quiescence. In transcription activation or repression, a transcription factor binds dynamically to DNA, with a residence time unique to each factor. Whether the DNA residence time is important in the transcription process is unclear. Here, we designed a series of transcription repressors differing in their DNA residence time by utilizing the modular DNA binding domain of transcription activator-like effectors (TALEs) and varying the number of nucleotide-recognizing repeat domains. We characterized the DNA residence times of our repressors in living cells using single molecule tracking. The residence times depended non-linearly on the number of repeat domains and differed by more than a factor of six. The factors provoked a residence time-dependent decrease in transcript level of the glucocorticoid receptor-activated gene SGK1. Down regulation of transcription was due to a lower burst frequency in the presence of long binding repressors and is in accordance with a model of competitive inhibition of endogenous activator binding. Our single molecule experiments reveal transcription factor DNA residence time as a regulatory factor controlling transcription repression and establish TALE-DNA binding domains as tools for the temporal dissection of transcription regulation. PMID:28977492

Runx transcription factors in neuronal development

Directory of Open Access Journals (Sweden)

Shiga Takashi

2008-08-01

Full Text Available Abstract Runt-related (Runx transcription factors control diverse aspects of embryonic development and are responsible for the pathogenesis of many human diseases. In recent years, the functions of this transcription factor family in the nervous system have just begun to be understood. In dorsal root ganglion neurons, Runx1 and Runx3 play pivotal roles in the development of nociceptive and proprioceptive sensory neurons, respectively. Runx appears to control the transcriptional regulation of neurotrophin receptors, numerous ion channels and neuropeptides. As a consequence, Runx contributes to diverse aspects of the sensory system in higher vertebrates. In this review, we summarize recent progress in determining the role of Runx in neuronal development.

Repression of Meiotic Genes by Antisense Transcription and by Fkh2 Transcription Factor in Schizosaccharomyces pombe

OpenAIRE

Chen, Huei-Mei; Rosebrock, Adam P.; Khan, Sohail R.; Futcher, Bruce; Leatherwood, Janet K.

2012-01-01

In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription ...

Precise integration of inducible transcriptional elements (PrIITE) enables absolute control of gene expression

DEFF Research Database (Denmark)

Pinto, Rita; Hansen, Lars; Hintze, John

2017-01-01

to be a limitation. Here, we report that the combined use of genome editing tools and last generation Tet-On systems can resolve these issues. Our principle is based on precise integration of inducible transcriptional elements (coined PrIITE) targeted to: (i) exons of an endogenous gene of interest (GOI) and (ii......Tetracycline-based inducible systems provide powerful methods for functional studies where gene expression can be controlled. However, the lack of tight control of the inducible system, leading to leakiness and adverse effects caused by undesirable tetracycline dosage requirements, has proven......) a safe harbor locus. Using PrIITE cells harboring a GFP reporter or CDX2 transcription factor, we demonstrate discrete inducibility of gene expression with complete abrogation of leakiness. CDX2 PrIITE cells generated by this approach uncovered novel CDX2 downstream effector genes. Our results provide...

The microRNA390/TRANS ACTING SHORT INTERFERING RNA3 module mediates lateral root growth under salt stress via the auxin pathway.

Science.gov (United States)

He, Fu; Xu, Changzheng; Fu, Xiaokang; Shen, Yun; Guo, Li; Leng, Mi; Luo, Keming

2018-05-01

Salt-induced developmental plasticity in a plant root system strongly depends on auxin signaling. However, the molecular events underlying this process are poorly understood. MicroRNA390 (miR390), trans-acting small interference RNAs (tasiRNAs) and AUXIN RESPONSE FACTORs (ARFs) form a regulatory module involved in controlling lateral root (LR) growth. Here, we found that miR390 expression was strongly induced by exposure to salt during LR formation in poplar (Populus spp.) plants. miR390 overexpression stimulated LR development and increased salt tolerance, whereas miR390 knockdown caused by a short tandem target mimic repressed LR growth and compromised salt resistance. ARF3.1, ARF3.2, and ARF4 expression was significantly inhibited by the presence of salt, and transcript abundance was dramatically decreased in the miR390-overexpressing line but increased in the miR390-knockdown line. Constitutive expression of ARF4m harboring mutated trans-acting small interference ARF-binding sites removed the salt resistance of the miR390 overexpressors. miR390 positively regulated auxin signaling in LRs subjected to salt but ARF4 inhibited auxin signaling. Salinity stabilized the poplar Aux/IAA repressor INDOLE-3-ACETIC ACID17.1, and overexpression of an auxin/salt resistant form of this repressor suppressed LR growth in miR390-overexpressing and ARF4-RNAi lines in the presence of salt. Thus, the miR390/TAS3/ARFs module is a key regulator, via modulating the auxin pathway, of LR growth in poplar subjected to salt stress. {copyright, serif} 2018 American Society of Plant Biologists. All rights reserved.

Potential Role of Activating Transcription Factor 5 during Osteogenesis

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Luisa Vicari

2016-01-01

Full Text Available Human adipose-derived stem cells are an abundant population of stem cells readily isolated from human adipose tissue that can differentiate into connective tissue lineages including bone, cartilage, fat, and muscle. Activating transcription factor 5 is a transcription factor of the ATF/cAMP response element-binding protein (CREB family. It is transcribed in two types of mRNAs (activating transcription factor 5 isoform 1 and activating transcription factor 5 isoform 2, encoding the same single 30-kDa protein. Although it is well demonstrated that it regulates the proliferation, differentiation, and apoptosis, little is known about its potential role in osteogenic differentiation. The aim of this study was to evaluate the expression levels of the two isoforms and protein during osteogenic differentiation of human adipose-derived stem cells. Our data indicate that activating transcription factor 5 is differentially expressed reaching a peak of expression at the stage of bone mineralization. These findings suggest that activating transcription factor 5 could play an interesting regulatory role during osteogenesis, which would provide a powerful tool to study bone physiology.

Potential Role of Activating Transcription Factor 5 during Osteogenesis.

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Vicari, Luisa; Calabrese, Giovanna; Forte, Stefano; Giuffrida, Raffaella; Colarossi, Cristina; Parrinello, Nunziatina Laura; Memeo, Lorenzo

2016-01-01

Human adipose-derived stem cells are an abundant population of stem cells readily isolated from human adipose tissue that can differentiate into connective tissue lineages including bone, cartilage, fat, and muscle. Activating transcription factor 5 is a transcription factor of the ATF/cAMP response element-binding protein (CREB) family. It is transcribed in two types of mRNAs (activating transcription factor 5 isoform 1 and activating transcription factor 5 isoform 2), encoding the same single 30-kDa protein. Although it is well demonstrated that it regulates the proliferation, differentiation, and apoptosis, little is known about its potential role in osteogenic differentiation. The aim of this study was to evaluate the expression levels of the two isoforms and protein during osteogenic differentiation of human adipose-derived stem cells. Our data indicate that activating transcription factor 5 is differentially expressed reaching a peak of expression at the stage of bone mineralization. These findings suggest that activating transcription factor 5 could play an interesting regulatory role during osteogenesis, which would provide a powerful tool to study bone physiology.

Analysis of miRNAs targeting transcription factors in Persicaria minor induced by Fusarium oxysporum

Science.gov (United States)

Samad, Abdul Fatah A.; Ali, Nazaruddin Muhammad; Ismail, Ismanizan; Murad, Abdul Munir Abdul

2016-11-01

A recent discovery showed small non-coding RNA known as microRNA has a crucial role in plant development and plant survival in extreme condition. In the past few years, researchers have managed to identify the various families of transcription factors that play a crucial role in regulating plant development and plant responses to stresses. This study focuses on the expression pattern of miRNA targeted transcription factor under biotic stress in a plant rich with secondary metabolite, Persicaria minor. A pathogenic fungus, Fusarium oxysporum was used in the biotic stress treatment since the previous study revealed this fungus could trigger plant defense system. Two small RNA libraries were constructed which consist of control and treated samples. In order to identify the potential target, psRobot target prediction software was used for each miRNA that shows significant change due to the infection. The result showed miR156b/c, miR172a, miR319, miR858, and miR894 were found to be targeting a wide range of transcription factors that involve in plant development and plant response towards stresses. The expression of miR156b/c and miR172 were up-regulated while the expression of miR319, miR858, and miR894 was found to be down-regulated. These results may provide a certain level of networking between those two regulatory molecules in plant genetic system under biotic stress.

The obesity-associated transcription factor ETV5 modulates circulating glucocorticoids

Science.gov (United States)

Gutierrez-Aguilar, Ruth; Thompson, Abigail; Marchand, Nathalie; Dumont, Patrick; Woods, Stephen C.; de Launoit, Yvan; Seeley, Randy J.; Ulrich-Lai, Yvonne M.

2015-01-01

The transcription factor E-twenty-six version 5 (ETV5) has been linked with obesity in genome-wide association studies. Moreover, ETV5-deficient mice (knockout; KO) have reduced body weight, lower fat mass, and are resistant to diet-induced obesity, directly linking ETV5 to the regulation of energy balance and metabolism. ETV5 is expressed in hypothalamic brain regions that regulate both metabolism and HPA axis activity, suggesting that ETV5 may also modulate HPA axis function. In order to test this possibility, plasma corticosterone levels were measured in ETV5 KO and wildtype (WT) mice before (pre-stress) and after (post-stress) a mild stressor (intraperitoneal injection). ETV5 deficiency increased both pre- and post-stress plasma corticosterone, suggesting that loss of ETV5 elevated glucocorticoid tone. Consistent with this idea, ETV5 KO mice have reduced thymus weight, suggestive of increased glucocorticoid-induced thymic involution. ETV5 deficiency also decreased the mRNA expression of glucocorticoid receptor (GR), mineralocorticoid receptor (MR), and vasopressin receptor 1A in the hypothalamus, without altering vasopressin, corticotropin-releasing hormone, or oxytocin mRNA expression. In order to test whether reduced MR and GR expression affected glucocorticoid negative feedback, a dexamethasone suppression test was performed. Dexamethasone reduced plasma corticosterone in both ETV5 KO and WT mice, suggesting that glucocorticoid negative feedback was unaltered by ETV5 deficiency. In summary, these data suggest that the obesity-associated transcription factor ETV5 normally acts to diminish circulating glucocorticoids. This might occur directly via ETV5 actions on HPA-regulatory brain circuitry, and/or indirectly via ETV5-induced alterations in metabolic factors that then influence the HPA axis. PMID:25813907

Polyphenol Compound as a Transcription Factor Inhibitor.

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Park, Seyeon

2015-10-30

A target-based approach has been used to develop novel drugs in many therapeutic fields. In the final stage of intracellular signaling, transcription factor-DNA interactions are central to most biological processes and therefore represent a large and important class of targets for human therapeutics. Thus, we focused on the idea that the disruption of protein dimers and cognate DNA complexes could impair the transcriptional activation and cell transformation regulated by these proteins. Historically, natural products have been regarded as providing the primary leading compounds capable of modulating protein-protein or protein-DNA interactions. Although their mechanism of action is not fully defined, polyphenols including flavonoids were found to act mostly as site-directed small molecule inhibitors on signaling. There are many reports in the literature of screening initiatives suggesting improved drugs that can modulate the transcription factor interactions responsible for disease. In this review, we focus on polyphenol compound inhibitors against dimeric forms of transcription factor components of intracellular signaling pathways (for instance, c-jun/c-fos (Activator Protein-1; AP-1), c-myc/max, Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and β-catenin/T cell factor (Tcf)).

NF-kappaB and p53 are the dominant apoptosis-inducing transcription factors elicited by the HIV-1 envelope.

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Perfettini, Jean-Luc; Roumier, Thomas; Castedo, Maria; Larochette, Nathanael; Boya, Patricia; Raynal, Brigitte; Lazar, Vladimir; Ciccosanti, Fabiola; Nardacci, Roberta; Penninger, Josef; Piacentini, Mauro; Kroemer, Guido

2004-03-01

The coculture of cells expressing the HIV-1 envelope glycoprotein complex (Env) with cells expressing CD4 results into cell fusion, deregulated mitosis, and subsequent cell death. Here, we show that NF-kappaB, p53, and AP1 are activated in Env-elicited apoptosis. The nuclear factor kappaB (NF-kappaB) super repressor had an antimitotic and antiapoptotic effect and prevented the Env-elicited phosphorylation of p53 on serine 15 and 46, as well as the activation of AP1. Transfection with dominant-negative p53 abolished apoptosis and AP1 activation. Signs of NF-kappaB and p53 activation were also detected in lymph node biopsies from HIV-1-infected individuals. Microarrays revealed that most (85%) of the transcriptional effects of HIV-1 Env were blocked by the p53 inhibitor pifithrin-alpha. Macroarrays led to the identification of several Env-elicited, p53-dependent proapoptotic transcripts, in particular Puma, a proapoptotic "BH3-only" protein from the Bcl-2 family known to activate Bax/Bak. Down modulation of Puma by antisense oligonucleotides, as well as RNA interference of Bax and Bak, prevented Env-induced apoptosis. HIV-1-infected primary lymphoblasts up-regulated Puma in vitro. Moreover, circulating CD4+ lymphocytes from untreated, HIV-1-infected donors contained enhanced amounts of Puma protein, and these elevated Puma levels dropped upon antiretroviral therapy. Altogether, these data indicate that NF-kappaB and p53 cooperate as the dominant proapoptotic transcription factors participating in HIV-1 infection.

SoyDB: a knowledge database of soybean transcription factors

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Valliyodan Babu

2010-01-01

Full Text Available Abstract Background Transcription factors play the crucial rule of regulating gene expression and influence almost all biological processes. Systematically identifying and annotating transcription factors can greatly aid further understanding their functions and mechanisms. In this article, we present SoyDB, a user friendly database containing comprehensive knowledge of soybean transcription factors. Description The soybean genome was recently sequenced by the Department of Energy-Joint Genome Institute (DOE-JGI and is publicly available. Mining of this sequence identified 5,671 soybean genes as putative transcription factors. These genes were comprehensively annotated as an aid to the soybean research community. We developed SoyDB - a knowledge database for all the transcription factors in the soybean genome. The database contains protein sequences, predicted tertiary structures, putative DNA binding sites, domains, homologous templates in the Protein Data Bank (PDB, protein family classifications, multiple sequence alignments, consensus protein sequence motifs, web logo of each family, and web links to the soybean transcription factor database PlantTFDB, known EST sequences, and other general protein databases including Swiss-Prot, Gene Ontology, KEGG, EMBL, TAIR, InterPro, SMART, PROSITE, NCBI, and Pfam. The database can be accessed via an interactive and convenient web server, which supports full-text search, PSI-BLAST sequence search, database browsing by protein family, and automatic classification of a new protein sequence into one of 64 annotated transcription factor families by hidden Markov models. Conclusions A comprehensive soybean transcription factor database was constructed and made publicly accessible at http://casp.rnet.missouri.edu/soydb/.

Melanogenesis-Inducing Effect of Cirsimaritin through Increases in Microphthalmia-Associated Transcription Factor and Tyrosinase Expression

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Hyo Jung Kim

2015-04-01

Full Text Available The melanin-inducing properties of cirsimaritin were investigated in murine B16F10 cells. Cirsimaritin is an active flavone with methoxy groups, which is isolated from the branches of Lithocarpus dealbatus. Tyrosinase activity and melanin content in murine B16F10 melanoma cells were increased by cirsimaritin in a dose-dependent manner. Western blot analysis revealed that tyrosinase, tyrosinase-related protein (TRP 1, TRP2 protein levels were enhanced after treatment with cirsimaritin for 48 h. Cirsimaritin also upregulated the expression of microphthalmia-associated transcription factor (MITF after 24 h of treatment. Furthermore, cirsimaritin induced phosphorylation of cyclic adenosine monophosphate (cAMP response element-binding protein (CREB in a dose-dependent manner after treatment for 15 min. The cirsimaritin-mediated increase of tyrosinase activity was significantly attenuated by H89, a cAMP-dependent protein kinase A inhibitor. These findings indicate that cirsimaritin stimulates melanogenesis in B16F10 cells by activation of CREB as well as upregulation of MITF and tyrosinase expression, which was activated by cAMP signaling. Finally, the melanogenic effect of cirsimaritin was confirmed in human epidermal melanocytes. These results support the putative application of cirsimaritin in ultraviolet photoprotection and hair coloration treatments.

Hepcidin mediates transcriptional changes that modulate acute cytokine-induced inflammatory responses in mice.

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De Domenico, Ivana; Zhang, Tian Y; Koening, Curry L; Branch, Ryan W; London, Nyall; Lo, Eric; Daynes, Raymond A; Kushner, James P; Li, Dean; Ward, Diane M; Kaplan, Jerry

2010-07-01

Hepcidin is a peptide hormone that regulates iron homeostasis and acts as an antimicrobial peptide. It is expressed and secreted by a variety of cell types in response to iron loading and inflammation. Hepcidin mediates iron homeostasis by binding to the iron exporter ferroportin, inducing its internalization and degradation via activation of the protein kinase Jak2 and the subsequent phosphorylation of ferroportin. Here we have shown that hepcidin-activated Jak2 also phosphorylates the transcription factor Stat3, resulting in a transcriptional response. Hepcidin treatment of ferroportin-expressing mouse macrophages showed changes in mRNA expression levels of a wide variety of genes. The changes in transcript levels for half of these genes were a direct effect of hepcidin, as shown by cycloheximide insensitivity, and dependent on the presence of Stat3. Hepcidin-mediated transcriptional changes modulated LPS-induced transcription in both cultured macrophages and in vivo mouse models, as demonstrated by suppression of IL-6 and TNF-alpha transcript and secreted protein. Hepcidin-mediated transcription in mice also suppressed toxicity and morbidity due to single doses of LPS, poly(I:C), and turpentine, which is used to model chronic inflammatory disease. Most notably, we demonstrated that hepcidin pretreatment protected mice from a lethal dose of LPS and that hepcidin-knockout mice could be rescued from LPS toxicity by injection of hepcidin. The results of our study suggest a new function for hepcidin in modulating acute inflammatory responses.

Transcription Factor Amr1 Induces Melanin Biosynthesis and Suppresses Virulence in Alternaria brassicicola

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Cho, Yangrae; Srivastava, Akhil; Ohm, Robin A.; Lawrence, Christopher B.; Wang, Koon-Hui; Grigoriev, Igor V.; Marahatta, Sharadchandra P.

2012-05-01

Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. Several A. brassicicola genes have been characterized as affecting pathogenesis of Brassica species. To study regulatory mechanisms of pathogenesis, we mined 421 genes in silico encoding putative transcription factors in a machine-annotated, draft genome sequence of A. brassicicola. In this study, targeted gene disruption mutants for 117 of the transcription factor genes were produced and screened. Three of these genes were associated with pathogenesis. Disruption mutants of one gene (AbPacC) were nonpathogenic and another gene (AbVf8) caused lesions less than half the diameter of wild-type lesions. Unexpectedly, mutants of the third gene, Amr1, caused lesions with a two-fold larger diameter than the wild type and complementation mutants. Amr1 is a homolog of Cmr1, a transcription factor that regulates melanin biosynthesis in several fungi. We created gene deletion mutants of ?amr1 and characterized their phenotypes. The ?amr1 mutants used pectin as a carbon source more efficiently than the wild type, were melanin-deficient, and more sensitive to UV light and glucanase digestion. The AMR1 protein was localized in the nuclei of hyphae and in highly melanized conidia during the late stage of plant pathogenesis. RNA-seq analysis revealed that three genes in the melanin biosynthesis pathway, along with the deleted Amr1 gene, were expressed at low levels in the mutants. In contrast, many hydrolytic enzyme-coding genes were expressed at higher levels in the mutants than in the wild type during pathogenesis. The results of this study suggested that a gene important for survival in nature negatively affected virulence, probably by a less efficient use of plant cell-wall materials. We speculate that the functions of the Amr1 gene are important to the success of A. brassicicola as a competitive saprophyte and plant parasite.

Tobacco TTG2 and ARF8 function concomitantly to control flower colouring by regulating anthocyanin synthesis genes.

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Li, P; Chen, X; Sun, F; Dong, H

2017-07-01

Recently we elucidated that tobacco TTG2 cooperates with ARF8 to regulate the vegetative growth and seed production. Here we show that TTG2 and ARF8 control flower colouring by regulating expression of ANS and DFR genes, which function in anthocyanin biosynthesis. Genetic modifications that substantially altered expression levels of the TTG2 gene and production quantities of TTG2 protein were correlated with flower development and colouring. Degrees of flower colour were increased by TTG2 overexpression but decreased through TTG2 silencing, in coincidence with high and low concentrations of anthocyanins in flowers. Of five genes involved in the anthocyanin biosynthesis pathway, only ANS and DFR were TTG2-regulated and displayed enhancement and diminution of expression with TTG2 overexpression and silencing, respectively. The floral expression of ANS and DFR also needed a functional ARF8 gene, as ANS and DFR expression were attenuated by ARF8 silencing, which concomitantly diminished the role of TTG2 in anthocyanin production. While ARF8 required TTG2 to be expressed by itself and to regulate ANS and DFR expression, the concurrent presence of normally functional TTG2 and ARF8 was critical for floral production of anthocyanins and also for flower colouration. Our data suggest that TTG2 functions concomitantly with ARF8 to control degrees of flower colour by regulating expression of ANS and DFR, which are involved in the anthocyanin biosynthesis pathway. ARF8 depends on TTG2 to regulate floral expression of ANS and DFR with positive effects on anthocyanin production and flower colour. © 2017 German Botanical Society and The Royal Botanical Society of the Netherlands.

PtrWRKY73, a salicylic acid-inducible poplar WRKY transcription factor, is involved in disease resistance in Arabidopsis thaliana.

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Duan, Yanjiao; Jiang, Yuanzhong; Ye, Shenglong; Karim, Abdul; Ling, Zhengyi; He, Yunqiu; Yang, Siqi; Luo, Keming

2015-05-01

A salicylic acid-inducible WRKY gene, PtrWRKY73, from Populus trichocarpa , was isolated and characterized. Overexpression of PtrWRKY73 in Arabidopsis thaliana increased resistance to biotrophic pathogens but reduced resistance against necrotrophic pathogens. WRKY transcription factors are commonly involved in plant defense responses. However, limited information is available about the roles of the WRKY genes in poplar defense. In this study, we isolated a salicylic acid (SA)-inducible WRKY gene, PtrWRKY73, from Populus trichocarpa, belonging to group I family and containing two WRKY domains, a D domain and an SP cluster. PtrWRKY73 was expressed predominantly in roots, old leaves, sprouts and stems, especially in phloem and its expression was induced in response to treatment with exogenous SA. PtrWRKY73 was localized to the nucleus of plant cells and exhibited transcriptional activation. Overexpression of PtrWRKY73 in Arabidopsis thaliana resulted in increased resistance to a virulent strain of the bacterial pathogen Pseudomonas syringae (PstDC3000), but more sensitivity to the necrotrophic fungal pathogen Botrytis cinerea. The SA-mediated defense-associated genes, such as PR1, PR2 and PAD4, were markedly up-regulated in transgenic plants overexpressing PtrWRKY73. Arabidopsis non-expressor of PR1 (NPR1) was not affected, whereas a defense-related gene PAL4 had reduced in PtrWRKY73 overexpressor plants. Together, these results indicated that PtrWRKY73 plays a positive role in plant resistance to biotrophic pathogens but a negative effect on resistance against necrotrophic pathogens.

Comparison of Transcription Factor Binding Site Models

KAUST Repository

Bhuyan, Sharifulislam

2012-05-01

Modeling of transcription factor binding sites (TFBSs) and TFBS prediction on genomic sequences are important steps to elucidate transcription regulatory mechanism. Dependency of transcription regulation on a great number of factors such as chemical specificity, molecular structure, genomic and epigenetic characteristics, long distance interaction, makes this a challenging problem. Different experimental procedures generate evidence that DNA-binding domains of transcription factors show considerable DNA sequence specificity. Probabilistic modeling of TFBSs has been moderately successful in identifying patterns from a family of sequences. In this study, we compare performances of different probabilistic models and try to estimate their efficacy over experimental TFBSs data. We build a pipeline to calculate sensitivity and specificity from aligned TFBS sequences for several probabilistic models, such as Markov chains, hidden Markov models, Bayesian networks. Our work, containing relevant statistics and evaluation for the models, can help researchers to choose the most appropriate model for the problem at hand.

Structures and short linear motif of disordered transcription factor regions provide clues to the interactome of the cellular hub radical-induced cell death1

DEFF Research Database (Denmark)

O'Shea, Charlotte; Staby, Lasse; Bendsen, Sidsel Krogh

2017-01-01

Intrinsically disordered protein regions (IDRs) lack a well-defined three-dimensional structure, but often facilitate key protein functions. Some interactions between IDRs and folded protein domains rely on short linear motifs (SLiMs). These motifs are challenging to identify, but once found can...... point to larger networks of interactions, such as with proteins that serve as hubs for essential cellular functions. The stress-associated plant protein Radical-Induced Cell Death1 (RCD1) is one such hub, interacting with many transcription factors via their flexible IDRs. To identify the SLiM bound......046 formed different structures or were fuzzy in the complexes. These findings allow us to present a model of the stress-associated RCD1-transcription factor interactome and to contribute to the emerging understanding of the interactions between folded hubs and their intrinsically disordered partners....

Age-dependent regulation of ERF-VII transcription factor activity in Arabidopsis thaliana.

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Giuntoli, Beatrice; Shukla, Vinay; Maggiorelli, Federica; Giorgi, Federico M; Lombardi, Lara; Perata, Pierdomenico; Licausi, Francesco

2017-10-01

The Group VII Ethylene Responsive Factors (ERFs-VII) RAP2.2 and RAP2.12 have been mainly characterized with regard to their contribution as activators of fermentation in plants. However, transcriptional changes measured in conditions that stabilize these transcription factors exceed the mere activation of this biochemical pathway, implying additional roles performed by the ERF-VIIs in other processes. We evaluated gene expression in transgenic Arabidopsis lines expressing a stabilized form of RAP2.12, or hampered in ERF-VII activity, and identified genes affected by this transcriptional regulator and its homologs, including some involved in oxidative stress response, which are not universally induced under anaerobic conditions. The contribution of the ERF-VIIs in regulating this set of genes in response to chemically induced or submergence-stimulated mitochondria malfunctioning was found to depend on the plant developmental stage. A similar age-dependent mechanism also restrained ERF-VII activity upon the core-hypoxic genes, independently of the N-end rule pathway, which is accounted for the control of the anaerobic response. To conclude, this study shed new light on a dual role of ERF-VII proteins under submergence: as positive regulators of the hypoxic response and as repressors of oxidative-stress related genes, depending on the developmental stage at which plants are challenged by stress conditions. © 2017 John Wiley & Sons Ltd.

Mycobacterium leprae induces NF-κB-dependent transcription repression in human Schwann cells

International Nuclear Information System (INIS)

Pereira, Renata M.S.; Calegari-Silva, Teresa Cristina; Hernandez, Maristela O.; Saliba, Alessandra M.; Redner, Paulo; Pessolani, Maria Cristina V.; Sarno, Euzenir N.; Sampaio, Elizabeth P.; Lopes, Ulisses G.

2005-01-01

Mycobacterium leprae, the causative agent of leprosy, invades peripheral nerve Schwann cells, resulting in deformities associated with this disease. NF-κB is an important transcription factor involved in the regulation of host immune antimicrobial responses. We aimed in this work to investigate NF-κB signaling pathways in the human ST88-14 Schwannoma cell line infected with M. leprae. Gel shift and supershift assays indicate that two NF-κB dimers, p65/p50 and p50/p50, translocate to the nucleus in Schwann cells treated with lethally irradiated M. leprae. Consistent with p65/p50 and p50/p50 activation, we observed IκB-α degradation and reduction of p105 levels. The nuclear translocation of p50/p50 complex due to M. leprae treatment correlated with repression of NF-κB-driven transcription induced by TNF-α. Moreover, thalidomide inhibited p50 homodimer nuclear translocation induced by M. leprae and consequently rescues Schwann cells from NF-κB-dependent transcriptional repression. Here, we report for the first time that M. leprae induces NF-κB activation in Schwann cells and thalidomide is able to modulate this activation

Induction of specific neuron types by overexpression of single transcription factors.

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Teratani-Ota, Yusuke; Yamamizu, Kohei; Piao, Yulan; Sharova, Lioudmila; Amano, Misa; Yu, Hong; Schlessinger, David; Ko, Minoru S H; Sharov, Alexei A

2016-10-01

Specific neuronal types derived from embryonic stem cells (ESCs) can facilitate mechanistic studies and potentially aid in regenerative medicine. Existing induction methods, however, mostly rely on the effects of the combined action of multiple added growth factors, which generally tend to result in mixed populations of neurons. Here, we report that overexpression of specific transcription factors (TFs) in ESCs can rather guide the differentiation of ESCs towards specific neuron lineages. Analysis of data on gene expression changes 2 d after induction of each of 185 TFs implicated candidate TFs for further ESC differentiation studies. Induction of 23 TFs (out of 49 TFs tested) for 6 d facilitated neural differentiation of ESCs as inferred from increased proportion of cells with neural progenitor marker PSA-NCAM. We identified early activation of the Notch signaling pathway as a common feature of most potent inducers of neural differentiation. The majority of neuron-like cells generated by induction of Ascl1, Smad7, Nr2f1, Dlx2, Dlx4, Nr2f2, Barhl2, and Lhx1 were GABA-positive and expressed other markers of GABAergic neurons. In the same way, we identified Lmx1a and Nr4a2 as inducers for neurons bearing dopaminergic markers and Isl1, Fezf2, and St18 for cholinergic motor neurons. A time-course experiment with induction of Ascl1 showed early upregulation of most neural-specific messenger RNA (mRNA) and microRNAs (miRNAs). Sets of Ascl1-induced mRNAs and miRNAs were enriched in Ascl1 targets. In further studies, enrichment of cells obtained with the induction of Ascl1, Smad7, and Nr2f1 using microbeads resulted in essentially pure population of neuron-like cells with expression profiles similar to neural tissues and expressed markers of GABAergic neurons. In summary, this study indicates that induction of transcription factors is a promising approach to generate cultures that show the transcription profiles characteristic of specific neural cell types.

Keap1 silencing boosts lipopolysaccharide-induced transcription of interleukin 6 via activation of nuclear factor κB in macrophages

International Nuclear Information System (INIS)

Lv, Peng; Xue, Peng; Dong, Jian; Peng, Hui; Clewell, Rebecca; Wang, Aiping; Wang, Yue; Peng, Shuangqing; Qu, Weidong; Zhang, Qiang; Andersen, Melvin E.; Pi, Jingbo

2013-01-01

Interleukin-6 (IL6) is a multifunctional cytokine that regulates immune and inflammatory responses. Multiple transcription factors, including nuclear factor κB (NF-κB) and nuclear factor E2-related factor 2 (Nrf2), regulate IL6 transcription. Kelch-like ECH-associated protein 1 (Keap1) is a substrate adaptor protein for the Cullin 3-dependent E3 ubiquitin ligase complex, which regulates the degradation of many proteins, including Nrf2 and IκB kinase β (IKKβ). Here, we found that stable knockdown of Keap1 (Keap1-KD) in RAW 264.7 (RAW) mouse macrophages and human monocyte THP-1 cells significantly increased expression of Il6, and Nrf2-target genes, under basal and lipopolysaccharide (LPS, 0.001–0.1 μg/ml)-challenged conditions. However, Nrf2 activation alone, by tert-butylhydroquinone treatment of RAW cells, did not increase expression of Il6. Compared to cells transduced with scrambled non-target negative control shRNA, Keap1-KD RAW cells showed enhanced protein levels of IKKβ and increased expression and phosphorylation of NF-κB p65 under non-stressed and LPS-treated conditions. Because the expression of Il6 in Keap1-KD RAW cells was significantly attenuated by silencing of Ikkβ, but not Nrf2, it appears that stabilized IKKβ is responsible for the enhanced transactivation of Il6 in Keap1-KD cells. This study demonstrated that silencing of Keap1 in macrophages boosts LPS-induced transcription of Il6 via NF-κB activation. Given the importance of IL6 in the inflammatory response, the Keap1–IKKβ–NF-κB pathway may be a novel target for treatment and prevention of inflammation and associated disorders. - Highlights: • Knockdown of Keap1 increases expression of Il6 in macrophages. • Silencing of Keap1 results in protein accumulation of IKKβ and NF-κB p65. • Induction of Il6 resulting from Keap1 silencing is attributed to NF-κB activation

Keap1 silencing boosts lipopolysaccharide-induced transcription of interleukin 6 via activation of nuclear factor κB in macrophages

Energy Technology Data Exchange (ETDEWEB)

Lv, Peng [Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Xue, Peng; Dong, Jian [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Peng, Hui [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Evaluation and Research Center for Toxicology, Institute of Disease Control and Prevention, Academy of Military Medical Sciences (China); Clewell, Rebecca [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Wang, Aiping [Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Wang, Yue [Institute for Medical Device Standardization Administration, National Institutes for Food and Drug Control, Beijing (China); Peng, Shuangqing [Evaluation and Research Center for Toxicology, Institute of Disease Control and Prevention, Academy of Military Medical Sciences (China); Qu, Weidong [Key Laboratory of the Public Health Safety, Ministry of Education, School of Public Health, Fudan University, Shanghai (China); Zhang, Qiang; Andersen, Melvin E. [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); Pi, Jingbo, E-mail: jpi@thehamner.org [Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States)

2013-11-01

Interleukin-6 (IL6) is a multifunctional cytokine that regulates immune and inflammatory responses. Multiple transcription factors, including nuclear factor κB (NF-κB) and nuclear factor E2-related factor 2 (Nrf2), regulate IL6 transcription. Kelch-like ECH-associated protein 1 (Keap1) is a substrate adaptor protein for the Cullin 3-dependent E3 ubiquitin ligase complex, which regulates the degradation of many proteins, including Nrf2 and IκB kinase β (IKKβ). Here, we found that stable knockdown of Keap1 (Keap1-KD) in RAW 264.7 (RAW) mouse macrophages and human monocyte THP-1 cells significantly increased expression of Il6, and Nrf2-target genes, under basal and lipopolysaccharide (LPS, 0.001–0.1 μg/ml)-challenged conditions. However, Nrf2 activation alone, by tert-butylhydroquinone treatment of RAW cells, did not increase expression of Il6. Compared to cells transduced with scrambled non-target negative control shRNA, Keap1-KD RAW cells showed enhanced protein levels of IKKβ and increased expression and phosphorylation of NF-κB p65 under non-stressed and LPS-treated conditions. Because the expression of Il6 in Keap1-KD RAW cells was significantly attenuated by silencing of Ikkβ, but not Nrf2, it appears that stabilized IKKβ is responsible for the enhanced transactivation of Il6 in Keap1-KD cells. This study demonstrated that silencing of Keap1 in macrophages boosts LPS-induced transcription of Il6 via NF-κB activation. Given the importance of IL6 in the inflammatory response, the Keap1–IKKβ–NF-κB pathway may be a novel target for treatment and prevention of inflammation and associated disorders. - Highlights: • Knockdown of Keap1 increases expression of Il6 in macrophages. • Silencing of Keap1 results in protein accumulation of IKKβ and NF-κB p65. • Induction of Il6 resulting from Keap1 silencing is attributed to NF-κB activation.

B-GATA transcription factors - insights into their structure, regulation and role in plant development

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Claus eSchwechheimer

2015-02-01

Full Text Available GATA transcription factors are evolutionarily conserved transcriptional regulators that recognize promoter elements with a G-A-T-A core sequence. In comparison to animal genomes, the GATA transcription factor family in plants is comparatively large with approximately 30 members. In spite of a long-standing interest of plant molecular biologists in GATA factors, only research conducted in the last years has led to reliable insights into their functions during plant development. Here, we review the current knowledge on B-GATAs, one of four GATA factor subfamilies from Arabidopsis thaliana. We show that B-GATAs can be subdivided based on structural features and their biological function into family members with a C-terminal LLM- (leucine-leucine-methionine domain or an N-terminal HAN- (HANABA TARANU domain. The paralogous GNC (GATA, NITRATE-INDUCIBLE, CARBON-METABOLISM INVOLVED and CGA1/GNL (CYTOKININ-INDUCED GATA1/GNC-LIKE are introduced as LLM-domain containing B-GATAs from Arabidopsis that control germination, greening, senescence and flowering time downstream from several growth regulatory signals including light and the hormones gibberellin, auxin, and cytokinin. Arabidopsis HAN and its monocot-specific paralogs from rice (NECK LEAF1, maize (TASSEL SHEATH1, and barley (THIRD OUTER GLUME are HAN-domain-containing B-GATAs with a predominant role in embryo development and floral development. We also review GATA23, a regulator of lateral root initiation from Arabidopsis, that is closely related to GNC and GNL but has a degenerate LLM-domain that is seemingly specific for the Brassicaceae family. The Brassicaceae-specific GATA23 together with the above-mentioned monocot-specific HAN-domain GATAs provide evidence that neofunctionalization of the B-GATAs was used during plant evolution to expand the functional repertoire of these transcription factors.

Role of the GRAS transcription factor ATA/RAM1 in the transcriptional reprogramming of arbuscular mycorrhiza in Petunia hybrida.

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Rich, Mélanie K; Courty, Pierre-Emmanuel; Roux, Christophe; Reinhardt, Didier

2017-08-08

Development of arbuscular mycorrhiza (AM) requires a fundamental reprogramming of root cells for symbiosis. This involves the induction of hundreds of genes in the host. A recently identified GRAS-type transcription factor in Petunia hybrida, ATA/RAM1, is required for the induction of host genes during AM, and for morphogenesis of the fungal endosymbiont. To better understand the role of RAM1 in symbiosis, we set out to identify all genes that depend on activation by RAM1 in mycorrhizal roots. We have carried out a transcript profiling experiment by RNAseq of mycorrhizal plants vs. non-mycorrhizal controls in wild type and ram1 mutants. The results show that the expression of early genes required for AM, such as the strigolactone biosynthetic genes and the common symbiosis signalling genes, is independent of RAM1. In contrast, genes that are involved at later stages of symbiosis, for example for nutrient exchange in cortex cells, require RAM1 for induction. RAM1 itself is highly induced in mycorrhizal roots together with many other transcription factors, in particular GRAS proteins. Since RAM1 has previously been shown to be directly activated by the common symbiosis signalling pathway through CYCLOPS, we conclude that it acts as an early transcriptional switch that induces many AM-related genes, among them genes that are essential for the development of arbuscules, such as STR, STR2, RAM2, and PT4, besides hundreds of additional RAM1-dependent genes the role of which in symbiosis remains to be explored. Taken together, these results indicate that the defect in the morphogenesis of the fungal arbuscules in ram1 mutants may be an indirect consequence of functional defects in the host, which interfere with nutrient exchange and possibly other functions on which the fungus depends.

The influence of partial renal function impairment on thyroid hormones level in acute renal failure-ARF patients

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Ilic, S.; Vlajkovic, M.; Rajic, M.; Bogicevic, M.

2002-01-01

The aim of this study was to assess the relationship of thyroid hormone levels with glomerular and tubular renal function in ARF. In thirty one patients with different underlying diseases, glomerular filtration (GFR) was estimated by measuring 99mTc-DTPA clearance and tubular function by 131I-OIH clearance. The thyroid status was evaluated by radioimmunoassays of T4, FT4, T3, FT3, rT3 and TSH, while the cortisol level was used to indicate the stress role. All the measurements were performed within 7 days as well after 1,3,6 and 12 months from the ARF onset. The results obtained were compared with those of 30 healthy subjects. At the ARF beginning, the impairment of renal function was manifested by the fall of 99mTc-DTPA clearance to 21∫7ml/min and 131I-OIH clearance to 172±34ml/min, while end values were 77±19ml/min and 395±70ml/min, respectively. The hormone analysis within 7 days, showed T3 decrease to 1.21±0.16nmol/l and the increase rT3 to 38±7ng/1ooml, as well as cortisol to 350±71ng/ml, but at the end of the study these hormone levels were normalized. Other hormone levels didn't differ significantly from control values. Regression analysis established significant correlation of 99mTc-DTPA clearance with T3 (r=0.693, p 100, OIH>500), partial recovery (DTPA 51-100, OIH 250-500) and without any recovery (DTPA<50,OIH<250), showed hormone disturbances proportional to the degree of renal function damage. The most severe disorders were found in the third group, where both total and free T4 and T3 were lowered, while rT3 and cortisol elevated at 7th day from ARF onset. After 12 months all these values associated with that of TSH were observed as declined.This study suggests thyroid hormone levels to be dependent on the renal function in ARF. Greater impairment of glomerular related to tubular function appears to be more responsible for thyroid dysfunction, due to uremic toxins retention as well as stress induced by the extreme function failure

Hyperosmotic stress regulates the distribution and stability of myocardin-related transcription factor, a key modulator of the cytoskeleton

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Ly, Donald L.; Waheed, Faiza; Lodyga, Monika

2013-01-01

Hyperosmotic stress initiates several adaptive responses, including the remodeling of the cytoskeleton. Besides maintaining structural integrity, the cytoskeleton has emerged as an important regulator of gene transcription. Myocardin-related transcription factor (MRTF), an actin-regulated coactiv......Hyperosmotic stress initiates several adaptive responses, including the remodeling of the cytoskeleton. Besides maintaining structural integrity, the cytoskeleton has emerged as an important regulator of gene transcription. Myocardin-related transcription factor (MRTF), an actin......-regulated coactivator of serum response factor, is a major link between the actin skeleton and transcriptional control. We therefore investigated whether MRTF is regulated by hyperosmotic stress. Here we show that hypertonicity induces robust, rapid, and transient translocation of MRTF from the cytosol to the nucleus...... in kidney tubular cells. We found that the hyperosmolarity-triggered MRTF translocation is mediated by the RhoA/Rho kinase (ROK) pathway. Moreover, the Rho guanine nucleotide exchange factor GEF-H1 is activated by hyperosmotic stress, and it is a key contributor to the ensuing RhoA activation and MRTF...

Expression of the Transcription Factor E4BP4 in Human Basophils

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Jensen, Bettina Margrethe; Gohr, Maria; Poulsen, Lars Kærgaard

2014-01-01

Rationale The cytokine IL-3 plays an important role for human basophil development, function and survival. IL-3 is also reported to induce the expression of the transcription factor E4BP4, but it is not known whether E4BP4 is expressed in basophils and influences basophil responsiveness. The aim...... by Alcian blue. RNA was extracted (0.005-0.02 µg RNA from 0.5 - 1 x 106 cells), and the corresponding cDNA analyzed by real-time PCR where E4BP4 expression was calculated as 2-(CT(E4BP4) - CT(β-actin)). E4BP4 protein expression was visualized in basophil lysates (107 cells/ml) by Western blot followed...... the transcription factor E4BP4 which might have an impact on basophil histamine release....

Dysregulated ΔNp63α inhibits expression of Ink4a/arf, blocks senescence, and promotes malignant conversion of keratinocytes.

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Linan Ha

Full Text Available p63 is critical for squamous epithelial development, and elevated levels of the ΔNp63α isoform are seen in squamous cell cancers of various organ sites. However, significant controversy exists regarding the role of p63 isoforms as oncoproteins or tumor suppressors. Here, lentiviruses were developed to drive long-term overexpression of ΔNp63α in primary keratinocytes. Elevated levels of ΔNp63α in vitro promote long-term survival and block both replicative and oncogene-induced senescence in primary keratinocytes, as evidenced by the expression of SA-β-gal and the presence of nuclear foci of heterochromatin protein 1γ. The contribution of ΔNp63α to cancer development was assessed using an in vivo grafting model of experimental skin tumorigenesis that allows distinction between benign and malignant tumors. Grafted lenti-ΔNp63α keratinocytes do not form tumors, whereas lenti-GFP/v-ras(Ha keratinocytes develop well-differentiated papillomas. Lenti-ΔNp63α/v-ras(Ha keratinocytes form undifferentiated carcinomas. The average volume of lenti-ΔNp63α/v-ras(Ha tumors was significantly higher than those in the lenti-GFP/v-ras(Ha group, consistent with increased BrdU incorporation detected by immunohistochemistry. The block in oncogene-induced senescence corresponds to sustained levels of E2F1 and phosphorylated AKT, and is associated with loss of induction of p16(ink4a/p19(arf. The relevance of p16(ink4a/p19(arf loss was demonstrated in grafting studies of p19(arf-null keratinocytes, which develop malignant carcinomas in the presence of v-ras(Ha similar to those arising in wildtype keratinocytes that express lenti-ΔNp63α and v-ras(Ha. Our findings establish that ΔNp63α has oncogenic activity and its overexpression in human squamous cell carcinomas contributes to the malignant phenotype, and implicate its ability to regulate p16(ink4a/p19(arf in the process.

Polyphenol Compound as a Transcription Factor Inhibitor

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Seyeon Park

2015-10-01

Full Text Available A target-based approach has been used to develop novel drugs in many therapeutic fields. In the final stage of intracellular signaling, transcription factor–DNA interactions are central to most biological processes and therefore represent a large and important class of targets for human therapeutics. Thus, we focused on the idea that the disruption of protein dimers and cognate DNA complexes could impair the transcriptional activation and cell transformation regulated by these proteins. Historically, natural products have been regarded as providing the primary leading compounds capable of modulating protein–protein or protein-DNA interactions. Although their mechanism of action is not fully defined, polyphenols including flavonoids were found to act mostly as site-directed small molecule inhibitors on signaling. There are many reports in the literature of screening initiatives suggesting improved drugs that can modulate the transcription factor interactions responsible for disease. In this review, we focus on polyphenol compound inhibitors against dimeric forms of transcription factor components of intracellular signaling pathways (for instance, c-jun/c-fos (Activator Protein-1; AP-1, c-myc/max, Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB and β-catenin/T cell factor (Tcf.

High promoter hypermethylation frequency of p14/ARF in supratentorial PNET but not in medulloblastoma.

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Inda, M M; Muñoz, J; Coullin, P; Fauvet, D; Danglot, G; Tuñón, T; Bernheim, A; Castresana, J S

2006-04-01

Medulloblastoma (MB) is the most common primitive neuroectodermal tumour (PNET) of the central nervous system. Although supratentorial PNET (sPNET) and MB are histologically similar, their clinical behaviour differs, sPNET being more aggressive than MB. The aim of this study was to determine whether sPNET and MB are genetically different entities. We investigated 32 PNET primary tumour samples (23 MB and nine sPNET) and four PNET cell lines, for the presence of CDKN2A homozygous deletions at exon 1-alpha of p16/INK4 and exon 1-beta of p14/ARF, and promoter hypermethylation of both genes. No homozygous deletion of either p16/INK4 or p14/ARF was demonstrated in any of the PNET primary tumour samples. Methylation of p16/INK4 was found in one of six sPNET and in one of 23 MB, while p14/ARF methylation was observed in three of six sPNET and in three of 21 MB. No methylation of p16/INK4 or p14/ARF was found in any of the PNET cell lines analysed. The three MB cell lines did not show p16/INK4 expression, and only the MB Daoy cell line (homozygously deleted at CDKN2A) presented loss of p14/ARF expression. Our results in this limited series of central PNET show that p14/ARF is frequently involved in PNET carcinogenesis, with a higher frequency, but not statistically significant, for sPNET than for MB.

Role of arachidonic acid metabolism in transcriptional induction of tumor necrosis factor gene expression by phorbol ester

Energy Technology Data Exchange (ETDEWEB)

Horiguchi, J.; Spriggs, D.; Imamura, K.; Stone, R.; Luebbers, R.; Kufe, D.

1989-01-01

The treatment of human HL-60 promyelocytic leukemia cells with 12-0 tetradecanoylphorbol-13-acetate (TPA) is associated with induction of tumor necrosis factor (TNF) transcripts. The study reported here has examined TPA-induced signaling mechanisms responsible for the regulation of TNF gene expression in these cells. Run-on assays demonstrated that TPA increases TNS mRNA levels by transcriptional activation of this gene. The induction of TNF transcripts by TPA was inhibited by the isoquinolinesulfonamide derivative H7 but not by HA1004, suggesting that this effect of TPA is mediated by activation of protein kinase C. TPA treatment also resulted in increased arachidonic acid release. Moreover, inhibitors of phospholipase, A/sub 2/ blocked both the increase in arachidonic acid release and the induction of TNF transcripts. These findings suggest that TPA induces TNF gene expression through the formation of arachidonic acid metabolites. Although indomethacin had no detectable effect on this induction of TNF transcripts, ketoconazole, an inhibitor of 5-lipoxygenase, blocked TPA-induced increases in TNF mRNA levels. Moreover, TNF mRNA levels were increased by the 5-lipoxygenase metabolite leukotriene B/sub 4/. In contrast, the cyclooxygenase metabolite prostaglandin E/sub 2/ inhibited the induction of TNF transcripts by TPA. Taken together, these results suggest that TPA induces TNF gene expression through the arachidonic acid cascade and that the level of TNF transcripts is regulated by metabolites of the pathway, leukotriene B/sub 4/ and prostaglandin E/sub 2/.

How salicylic acid takes transcriptional control over jasmonic acid signaling

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Lotte eCaarls

2015-03-01

Full Text Available Transcriptional regulation is a central process in plant immunity. The induction or repression of defense genes is orchestrated by signaling networks that are directed by plant hormones of which salicylic acid (SA and jasmonic acid (JA are the major players. Extensive cross-communication between the hormone signaling pathways allows for fine tuning of transcriptional programs, determining resistance to invaders and trade-offs with plant development. Here, we give an overview of how SA can control transcriptional reprogramming of JA-induced genes in Arabidopsis thaliana. SA can influence activity and/or localization of transcriptional regulators by post-translational modifications of transcription factors and co-regulators. SA-induced redox changes, mediated by thioredoxins and glutaredoxins, modify transcriptional regulators that are involved in suppression of JA-dependent genes, such as NPR1 and TGA transcription factors, which affects their localization or DNA binding activity. Furthermore, SA can mediate sequestering of JA-responsive transcription factors away from their target genes by stalling them in the cytosol or in complexes with repressor proteins in the nucleus. SA also affects JA-induced transcription by inducing degradation of transcription factors with an activating role in JA signaling, as was shown for the ERF transcription factor ORA59. Additionally, SA can induce negative regulators, among which WRKY transcription factors, that can directly or indirectly inhibit JA-responsive gene expression. Finally, at the DNA level, modification of histones by SA-dependent factors can result in repression of JA-responsive genes. These diverse and complex regulatory mechanisms affect important signaling hubs in the integration of hormone signaling networks. Some pathogens have evolved effectors that highjack hormone crosstalk mechanisms for their own good, which are described in this review as well.

Hypoxia-Inducible Factor-1α in carcinogenesis and progression of breast cancer

NARCIS (Netherlands)

Bos, R.

2004-01-01

This thesis is primarily focused on the previously hardly explored role of HIF-1 in breast cancer. HIF-1 is a transcription factor induced by hypoxia, but also by some oncogenes, tumor suppressor genes and growth factors. Activated HIF-1 can induce angiogenesis, glycolysis, erythropoiesis, and other

Using network component analysis to dissect regulatory networks mediated by transcription factors in yeast.

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Chun Ye

2009-03-01

Full Text Available Understanding the relationship between genetic variation and gene expression is a central question in genetics. With the availability of data from high-throughput technologies such as ChIP-Chip, expression, and genotyping arrays, we can begin to not only identify associations but to understand how genetic variations perturb the underlying transcription regulatory networks to induce differential gene expression. In this study, we describe a simple model of transcription regulation where the expression of a gene is completely characterized by two properties: the concentrations and promoter affinities of active transcription factors. We devise a method that extends Network Component Analysis (NCA to determine how genetic variations in the form of single nucleotide polymorphisms (SNPs perturb these two properties. Applying our method to a segregating population of Saccharomyces cerevisiae, we found statistically significant examples of trans-acting SNPs located in regulatory hotspots that perturb transcription factor concentrations and affinities for target promoters to cause global differential expression and cis-acting genetic variations that perturb the promoter affinities of transcription factors on a single gene to cause local differential expression. Although many genetic variations linked to gene expressions have been identified, it is not clear how they perturb the underlying regulatory networks that govern gene expression. Our work begins to fill this void by showing that many genetic variations affect the concentrations of active transcription factors in a cell and their affinities for target promoters. Understanding the effects of these perturbations can help us to paint a more complete picture of the complex landscape of transcription regulation. The software package implementing the algorithms discussed in this work is available as a MATLAB package upon request.

Embryonic maturation of epidermal Merkel cells is controlled by a redundant transcription factor network.

Science.gov (United States)

Perdigoto, Carolina N; Bardot, Evan S; Valdes, Victor J; Santoriello, Francis J; Ezhkova, Elena

2014-12-01

Merkel cell-neurite complexes are located in touch-sensitive areas of the mammalian skin and are involved in recognition of the texture and shape of objects. Merkel cells are essential for these tactile discriminations, as they generate action potentials in response to touch stimuli and induce the firing of innervating afferent nerves. It has been shown that Merkel cells originate from epidermal stem cells, but the cellular and molecular mechanisms of their development are largely unknown. In this study, we analyzed Merkel cell differentiation during development and found that it is a temporally regulated maturation process characterized by a sequential activation of Merkel cell-specific genes. We uncovered key transcription factors controlling this process and showed that the transcription factor Atoh1 is required for initial Merkel cell specification. The subsequent maturation steps of Merkel cell differentiation are controlled by cooperative function of the transcription factors Sox2 and Isl1, which physically interact and work to sustain Atoh1 expression. These findings reveal the presence of a robust transcriptional network required to produce functional Merkel cells that are required for tactile discrimination. © 2014. Published by The Company of Biologists Ltd.

The transcription factor ETS-1 regulates angiotensin II-stimulated fibronectin production in mesangial cells.

Science.gov (United States)

Hua, Ping; Feng, Wenguang; Rezonzew, Gabriel; Chumley, Phillip; Jaimes, Edgar A

2012-06-01

Angiotensin II (ANG II) produced as result of activation of the renin-angiotensin system (RAS) plays a critical role in the pathogenesis of chronic kidney disease via its hemodynamic effects on the renal microcirculation as well as by its nonhemodynamic actions including the production of extracellular matrix proteins such as fibronectin, a multifunctional extracellular matrix protein that plays a major role in cell adhesion and migration as well as in the development of glomerulosclerosis. ETS-1 is an important transcription factor essential for normal kidney development and glomerular integrity. We previously showed that ANG II increases ETS-1 expression and is required for fibronectin production in mesangial cells. In these studies, we determined that ANG II induces phosphorylation of ETS-1 via activation of the type 1 ANG II receptor and that Erk1/2 and Akt/PKB phosphorylation are required for these effects. In addition, we characterized the role of ETS-1 on the transcriptional activation of fibronectin production in mesangial cells. We determined that ETS-1 directly activates the fibronectin promoter and by utilizing gel shift assays and chromatin immunoprecipitation assays identified two different ETS-1 binding sites that promote the transcriptional activation of fibronectin in response to ANG II. In addition, we identified the essential role of CREB and its coactivator p300 on the transcriptional activation of fibronectin by ETS-1. These studies unveil novel mechanisms involved in RAS-induced production of the extracellular matrix protein fibronectin in mesangial cells and establish the role of the transcription factor ETS-1 as a direct mediator of these effects.

Protein-protein interactions in the regulation of WRKY transcription factors.

Science.gov (United States)

Chi, Yingjun; Yang, Yan; Zhou, Yuan; Zhou, Jie; Fan, Baofang; Yu, Jing-Quan; Chen, Zhixiang

2013-03-01

It has been almost 20 years since the first report of a WRKY transcription factor, SPF1, from sweet potato. Great progress has been made since then in establishing the diverse biological roles of WRKY transcription factors in plant growth, development, and responses to biotic and abiotic stress. Despite the functional diversity, almost all analyzed WRKY proteins recognize the TTGACC/T W-box sequences and, therefore, mechanisms other than mere recognition of the core W-box promoter elements are necessary to achieve the regulatory specificity of WRKY transcription factors. Research over the past several years has revealed that WRKY transcription factors physically interact with a wide range of proteins with roles in signaling, transcription, and chromatin remodeling. Studies of WRKY-interacting proteins have provided important insights into the regulation and mode of action of members of the important family of transcription factors. It has also emerged that the slightly varied WRKY domains and other protein motifs conserved within each of the seven WRKY subfamilies participate in protein-protein interactions and mediate complex functional interactions between WRKY proteins and between WRKY and other regulatory proteins in the modulation of important biological processes. In this review, we summarize studies of protein-protein interactions for WRKY transcription factors and discuss how the interacting partners contribute, at different levels, to the establishment of the complex regulatory and functional network of WRKY transcription factors.

Emerging Functions of Transcription Factors in Malaria Parasite

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Renu Tuteja

2011-01-01

Full Text Available Transcription is a process by which the genetic information stored in DNA is converted into mRNA by enzymes known as RNA polymerase. Bacteria use only one RNA polymerase to transcribe all of its genes while eukaryotes contain three RNA polymerases to transcribe the variety of eukaryotic genes. RNA polymerase also requires other factors/proteins to produce the transcript. These factors generally termed as transcription factors (TFs are either associated directly with RNA polymerase or add in building the actual transcription apparatus. TFs are the most common tools that our cells use to control gene expression. Plasmodium falciparum is responsible for causing the most lethal form of malaria in humans. It shows most of its characteristics common to eukaryotic transcription but it is assumed that mechanisms of transcriptional control in P. falciparum somehow differ from those of other eukaryotes. In this article we describe the studies on the main TFs such as myb protein, high mobility group protein and ApiA2 family proteins from malaria parasite. These studies show that these TFs are slowly emerging to have defined roles in the regulation of gene expression in the parasite.

JUNGBRUNNEN1, a Reactive Oxygen Species–Responsive NAC Transcription Factor, Regulates Longevity in Arabidopsis

NARCIS (Netherlands)

Wu, A.; Devi Allu, A.; Garapati, P.; Siddiqui, H.; Dortay, H.; Zanor, M.I.; Amparo Asensi-Fabado, M.; Munne´ -Bosch, S.; Antonio, C.; Tohge, T.; Fernie, A.R.; Kaufmann, K.; Xue, G.P.; Mueller-Roeber, B.; Balazadeh, S.

2012-01-01

The transition from juvenility through maturation to senescence is a complex process that involves the regulation of longevity. Here, we identify JUNGBRUNNEN1 (JUB1), a hydrogen peroxide (H2O2)-induced NAC transcription factor, as a central longevity regulator in Arabidopsis thaliana. JUB1

Transcription-induced DNA supercoiling: New roles of intranucleosomal DNA loops in DNA repair and transcription.

Science.gov (United States)

Gerasimova, N S; Pestov, N A; Kulaeva, O I; Clark, D J; Studitsky, V M

2016-05-26

RNA polymerase II (Pol II) transcription through chromatin is accompanied by formation of small intranucleosomal DNA loops. Pol II captured within a small loop drives accumulation of DNA supercoiling, facilitating further transcription. DNA breaks relieve supercoiling and induce Pol II arrest, allowing detection of DNA damage hidden in chromatin structure.

Capsella rubella TGA4, a bZIP transcription factor, causes delayed flowering in Arabidopsis thaliana

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Li Maofu

2016-01-01

Full Text Available Flowering time is usually regulated by many environmental factors and endogenous signals. TGA family members are bZIP transcription factors that bind to the octopine synthase element, which has been closely linked to defense/stress responses. Most TGA factors interact with non-expressor of PR1 (NPR1 and plant defense responses are strengthened by this interaction. TGA1and TGA4factors bind to NPR1 only in salicylic acid (SA-induced leaves, suggesting that TGA4 has another function during plant development. Here, we isolated a bZIP transcription factor gene, TGA4, from Capsella rubella. TGA4transcripts were detected in most tissues, with high expression in leaves, low expression in stems and flowering buds, and undetectable in siliques. CruTGA4was over expressed in Arabidopsis thaliana wild typeCol-0 plants. Flowering time and total leaf number in the transgenic plants showed that overexpression of CruTGA4could delay flowering in A. thaliana. Our findings suggest that TGA4 may act as flowering regulator that controls plant flowering.

Targeting HOX and PBX transcription factors in ovarian cancer

International Nuclear Information System (INIS)

Morgan, Richard; Plowright, Lynn; Harrington, Kevin J; Michael, Agnieszka; Pandha, Hardev S

2010-01-01

Ovarian cancer still has a relatively poor prognosis due to the frequent occurrence of drug resistance, making the identification of new therapeutic targets an important goal. We have studied the role of HOX genes in the survival and proliferation of ovarian cancer cells. These are a family of homeodomain-containing transcription factors that determine cell and tissue identity in the early embryo, and have an anti-apoptotic role in a number of malignancies including lung and renal cancer. We used QPCR to determine HOX gene expression in normal ovary and in the ovarian cancer cell lines SK-OV3 and OV-90. We used a short peptide, HXR9, to disrupt the formation of HOX/PBX dimers and alter transcriptional regulation by HOX proteins. In this study we show that the ovarian cancer derived line SK-OV3, but not OV-90, exhibits highly dysregulated expression of members of the HOX gene family. Disrupting the interaction between HOX proteins and their co-factor PBX induces apoptosis in SK-OV3 cells and retards tumour growth in vivo. HOX/PBX binding is a potential target in ovarian cancer

Transcription Factor Functional Protein-Protein Interactions in Plant Defense Responses

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Murilo S. Alves

2014-03-01

Full Text Available Responses to biotic stress in plants lead to dramatic reprogramming of gene expression, favoring stress responses at the expense of normal cellular functions. Transcription factors are master regulators of gene expression at the transcriptional level, and controlling the activity of these factors alters the transcriptome of the plant, leading to metabolic and phenotypic changes in response to stress. The functional analysis of interactions between transcription factors and other proteins is very important for elucidating the role of these transcriptional regulators in different signaling cascades. In this review, we present an overview of protein-protein interactions for the six major families of transcription factors involved in plant defense: basic leucine zipper containing domain proteins (bZIP, amino-acid sequence WRKYGQK (WRKY, myelocytomatosis related proteins (MYC, myeloblastosis related proteins (MYB, APETALA2/ ETHYLENE-RESPONSIVE ELEMENT BINDING FACTORS (AP2/EREBP and no apical meristem (NAM, Arabidopsis transcription activation factor (ATAF, and cup-shaped cotyledon (CUC (NAC. We describe the interaction partners of these transcription factors as molecular responses during pathogen attack and the key components of signal transduction pathways that take place during plant defense responses. These interactions determine the activation or repression of response pathways and are crucial to understanding the regulatory networks that modulate plant defense responses.

Pdlim7 Regulates Arf6-Dependent Actin Dynamics and Is Required for Platelet-Mediated Thrombosis in Mice.

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Alexander E Urban

Full Text Available Upon vessel injury, platelets become activated and rapidly reorganize their actin cytoskeleton to adhere to the site of endothelial damage, triggering the formation of a fibrin-rich plug to prevent further blood loss. Inactivation of Pdlim7 provides the new perspective that regulation of actin cytoskeletal changes in platelets is dependent on the encoded PDZ-LIM protein. Loss-of-function of Pdlim7 triggers hypercoagulopathy and causes significant perinatal lethality in mice. Our in vivo and in vitro studies reveal that Pdlim7 is dynamically distributed along actin fibers, and lack of Pdlim7 leads to a marked inability to rearrange the actin cytoskeleton. Specifically, the absence of Pdlim7 prevents platelets from bundling actin fibers into a concentric ring that defines the round spread shape of activated platelets. Similarly, in mouse embryonic fibroblasts, loss of Pdlim7 abolishes the formation of stress fibers needed to adopt the typical elongated fibroblast shape. In addition to revealing a fundamental cell biological role in actin cytoskeletal organization, we also demonstrate a function of Pdlim7 in regulating the cycling between the GTP/GDP-bound states of Arf6. The small GTPase Arf6 is an essential factor required for actin dynamics, cytoskeletal rearrangements, and platelet activation. Consistent with our findings of significantly elevated initial F-actin ratios and subsequent morphological aberrations, loss of Pdlim7 causes a shift in balance towards an increased Arf6-GTP level in resting platelets. These findings identify a new Pdlim7-Arf6 axis controlling actin dynamics and implicate Pdlim7 as a primary endogenous regulator of platelet-dependent hemostasis.

Transcription factor Amr1 induces melanin biosynthesis and suppresses virulence in Alternaria brassicicola.

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Yangrae Cho

Full Text Available Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. Several A. brassicicola genes have been characterized as affecting pathogenesis of Brassica species. To study regulatory mechanisms of pathogenesis, we mined 421 genes in silico encoding putative transcription factors in a machine-annotated, draft genome sequence of A. brassicicola. In this study, targeted gene disruption mutants for 117 of the transcription factor genes were produced and screened. Three of these genes were associated with pathogenesis. Disruption mutants of one gene (AbPacC were nonpathogenic and another gene (AbVf8 caused lesions less than half the diameter of wild-type lesions. Unexpectedly, mutants of the third gene, Amr1, caused lesions with a two-fold larger diameter than the wild type and complementation mutants. Amr1 is a homolog of Cmr1, a transcription factor that regulates melanin biosynthesis in several fungi. We created gene deletion mutants of Δamr1 and characterized their phenotypes. The Δamr1 mutants used pectin as a carbon source more efficiently than the wild type, were melanin-deficient, and more sensitive to UV light and glucanase digestion. The AMR1 protein was localized in the nuclei of hyphae and in highly melanized conidia during the late stage of plant pathogenesis. RNA-seq analysis revealed that three genes in the melanin biosynthesis pathway, along with the deleted Amr1 gene, were expressed at low levels in the mutants. In contrast, many hydrolytic enzyme-coding genes were expressed at higher levels in the mutants than in the wild type during pathogenesis. The results of this study suggested that a gene important for survival in nature negatively affected virulence, probably by a less efficient use of plant cell-wall materials. We speculate that the functions of the Amr1 gene are important to the success of A. brassicicola as a competitive saprophyte and plant parasite.

Nfatc1 Is a Functional Transcriptional Factor Mediating Nell-1-Induced Runx3 Upregulation in Chondrocytes

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Chenshuang Li

2018-01-01

Full Text Available Neural EGFL like 1 (Nell-1 is essential for chondrogenic differentiation, maturation, and regeneration. Our previous studies have demonstrated that Nell-1’s pro-chondrogenic activities are predominantly reliant upon runt-related transcription factor 3 (Runx3-mediated Indian hedgehog (Ihh signaling. Here, we identify the nuclear factor of activated T-cells 1 (Nfatc1 as the key transcriptional factor mediating the Nell-1 → Runx3 signal transduction in chondrocytes. Using chromatin immunoprecipitation assay, we were able to determine that Nfatc1 binds to the −833–−810 region of the Runx3-promoter in response to Nell-1 treatment. By revealing the Nell-1 → Nfatc1 → Runx3 → Ihh cascade, we demonstrate the involvement of Nfatc1, a nuclear factor of activated T-cells, in chondrogenesis, while providing innovative insights into developing a novel therapeutic strategy for cartilage regeneration and other chondrogenesis-related conditions.

The obesity-induced transcriptional regulator TRIP-Br2 mediates visceral fat endoplasmic reticulum stress-induced inflammation.

Science.gov (United States)

Qiang, Guifen; Kong, Hyerim Whang; Fang, Difeng; McCann, Maximilian; Yang, Xiuying; Du, Guanhua; Blüher, Matthias; Zhu, Jinfang; Liew, Chong Wee

2016-04-25

The intimate link between location of fat accumulation and metabolic disease risk and depot-specific differences is well established, but how these differences between depots are regulated at the molecular level remains largely unclear. Here we show that TRIP-Br2 mediates endoplasmic reticulum (ER) stress-induced inflammatory responses in visceral fat. Using in vitro, ex vivo and in vivo approaches, we demonstrate that obesity-induced circulating factors upregulate TRIP-Br2 specifically in visceral fat via the ER stress pathway. We find that ablation of TRIP-Br2 ameliorates both chemical and physiological ER stress-induced inflammatory and acute phase response in adipocytes, leading to lower circulating levels of inflammatory cytokines. Using promoter assays, as well as molecular and pharmacological experiments, we show that the transcription factor GATA3 is responsible for the ER stress-induced TRIP-Br2 expression in visceral fat. Taken together, our study identifies molecular regulators of inflammatory response in visceral fat that-given that these pathways are conserved in humans-might serve as potential therapeutic targets in obesity.

Nerve growth factor enhances the CRE-dependent transcriptional activity activated by nobiletin in PC12 cells.

Science.gov (United States)

Takito, Jiro; Kimura, Junko; Kajima, Koji; Uozumi, Nobuyuki; Watanabe, Makoto; Yokosuka, Akihito; Mimaki, Yoshihiro; Nakamura, Masanori; Ohizumi, Yasushi

2016-07-01

Prevention and treatment of Alzheimer disease are urgent problems for elderly people in developed countries. We previously reported that nobiletin, a poly-methoxylated flavone from the citrus peel, improved the symptoms in various types of animal models of memory loss and activated the cAMP responsive element (CRE)-dependent transcription in PC12 cells. Nobiletin activated the cAMP/PKA/MEK/Erk/MAPK signaling pathway without using the TrkA signaling activated by nerve growth factor (NGF). Here, we examined the effect of combination of nobiletin and NGF on the CRE-dependent transcription in PC12 cells. Although NGF alone had little effect on the CRE-dependent transcription, NGF markedly enhanced the CRE-dependent transcription induced by nobiletin. The NGF-induced enhancement was neutralized by a TrkA antagonist, K252a. This effect of NGF was effective on the early signaling event elicited by nobiletin. These results suggested that there was crosstalk between NGF and nobiletin signaling in activating the CRE-dependent transcription in PC12 cells.

Role of WRKY Transcription Factors in Arabidopsis Development and Stress Responses

OpenAIRE

Li, Jing

2014-01-01

It has been well established that environmentally induced alterations in gene expression are mediated by transcription factors (TFs). One of the important plant-specific TF groups is the WRKY (TFs containing a highly conserved WRKY domain) family, which is involved in regulation of various physiological programs including biotic and abiotic defenses, senescence and trichome development. Two members of WRKY group III in Arabidopsis thaliana, WRKY54 and WRKY70, are demonstrated in this study to...

Structurally distinct polycyclic aromatic hydrocarbons induce differential transcriptional responses in developing zebrafish

International Nuclear Information System (INIS)

Goodale, Britton C.; Tilton, Susan C.; Corvi, Margaret M.; Wilson, Glenn R.; Janszen, Derek B.; Anderson, Kim A.; Waters, Katrina M.; Tanguay, Robert L.

2013-01-01

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous in the environment as components of fossil fuels and by-products of combustion. These multi-ring chemicals differentially activate the aryl hydrocarbon receptor (AHR) in a structurally dependent manner, and induce toxicity via both AHR-dependent and -independent mechanisms. PAH exposure is known to induce developmental malformations in zebrafish embryos, and recent studies have shown cardiac toxicity induced by compounds with low AHR affinity. Unraveling the potentially diverse molecular mechanisms of PAH toxicity is essential for understanding the hazard posed by complex PAH mixtures present in the environment. We analyzed transcriptional responses to PAH exposure in zebrafish embryos exposed to benz(a)anthracene (BAA), dibenzothiophene (DBT) and pyrene (PYR) at concentrations that induced developmental malformations by 120 h post-fertilization (hpf). Whole genome microarray analysis of mRNA expression at 24 and 48 hpf identified genes that were differentially regulated over time and in response to the three PAH structures. PAH body burdens were analyzed at both time points using GC–MS, and demonstrated differences in PAH uptake into the embryos. This was important for discerning dose-related differences from those that represented unique molecular mechanisms. While BAA misregulated the least number of transcripts, it caused strong induction of cyp1a and other genes known to be downstream of the AHR, which were not induced by the other two PAHs. Analysis of functional roles of misregulated genes and their predicted regulatory transcription factors also distinguished the BAA response from regulatory networks disrupted by DBT and PYR exposure. These results indicate that systems approaches can be used to classify the toxicity of PAHs based on the networks perturbed following exposure, and may provide a path for unraveling the toxicity of complex PAH mixtures. - Highlights: • Defined global mRNA expression

Fusarium oxysporum Triggers Tissue-Specific Transcriptional Reprogramming in Arabidopsis thaliana

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Lyons, Rebecca; Stiller, Jiri; Powell, Jonathan; Rusu, Anca; Manners, John M.; Kazan, Kemal

2015-01-01

Some of the most devastating agricultural diseases are caused by root-infecting pathogens, yet the majority of studies on these interactions to date have focused on the host responses of aerial tissues rather than those belowground. Fusarium oxysporum is a root-infecting pathogen that causes wilt disease on several plant species including Arabidopsis thaliana. To investigate and compare transcriptional changes triggered by F. oxysporum in different Arabidopsis tissues, we infected soil-grown plants with F. oxysporum and subjected root and leaf tissue harvested at early and late timepoints to RNA-seq analyses. At least half of the genes induced or repressed by F. oxysporum showed tissue-specific regulation. Regulators of auxin and ABA signalling, mannose binding lectins and peroxidases showed strong differential expression in root tissue. We demonstrate that ARF2 and PRX33, two genes regulated in the roots, promote susceptibility to F. oxysporum. In the leaves, defensins and genes associated with the response to auxin, cold and senescence were strongly regulated while jasmonate biosynthesis and signalling genes were induced throughout the plant. PMID:25849296

Fusarium oxysporum triggers tissue-specific transcriptional reprogramming in Arabidopsis thaliana.

Directory of Open Access Journals (Sweden)

Rebecca Lyons

Full Text Available Some of the most devastating agricultural diseases are caused by root-infecting pathogens, yet the majority of studies on these interactions to date have focused on the host responses of aerial tissues rather than those belowground. Fusarium oxysporum is a root-infecting pathogen that causes wilt disease on several plant species including Arabidopsis thaliana. To investigate and compare transcriptional changes triggered by F. oxysporum in different Arabidopsis tissues, we infected soil-grown plants with F. oxysporum and subjected root and leaf tissue harvested at early and late timepoints to RNA-seq analyses. At least half of the genes induced or repressed by F. oxysporum showed tissue-specific regulation. Regulators of auxin and ABA signalling, mannose binding lectins and peroxidases showed strong differential expression in root tissue. We demonstrate that ARF2 and PRX33, two genes regulated in the roots, promote susceptibility to F. oxysporum. In the leaves, defensins and genes associated with the response to auxin, cold and senescence were strongly regulated while jasmonate biosynthesis and signalling genes were induced throughout the plant.

In Vitro Culture Conditions and OeARF and OeH3 Expressions Modulate Adventitious Root Formation from Oleaster (Olea europaea L. subsp. europaea var. sylvestris) Cuttings

Science.gov (United States)

Gagliardi, Cinzia; Bruno, Leonardo; Bitonti, Maria Beatrice

2014-01-01

Olea europaea L. subsp. europaea var. sylvestris, also named oleaster, is the wild form of olive and it is used as rootstock and pollen donor for many cultivated varieties. An efficient procedure for in vitro propagation of oleaster was established in this study. A zeatin concentration of 2.5 mg/L was effective to induce an appreciable vegetative growth. Also high rooting efficiency was obtained by using a short IBA pulse, followed by two different IBA concentrations in the culture medium. With the aim to enlarge knowledge on the molecular aspects of adventitious rooting, we also evaluated the transcriptional modulation of an ARFs member and HISTONE H3 genes, involved in auxin signaling and cell replication, respectively, during the root induction phase of cuttings. The obtained results suggest that the selected genes, as markers of the induction phase, could be very useful for setting up efficient culture conditions along the rooting process, thus increasing micropropagation efficiency. PMID:24587768

In vitro culture conditions and OeARF and OeH3 expressions modulate adventitious root formation from oleaster (Olea europaea L. subsp. europaea var. sylvestris) cuttings.

Science.gov (United States)

Chiappetta, Adriana; Gagliardi, Cinzia; Bruno, Leonardo; Bitonti, Maria Beatrice

2014-01-01

Olea europaea L. subsp. europaea var. sylvestris, also named oleaster, is the wild form of olive and it is used as rootstock and pollen donor for many cultivated varieties. An efficient procedure for in vitro propagation of oleaster was established in this study. A zeatin concentration of 2.5 mg/L was effective to induce an appreciable vegetative growth. Also high rooting efficiency was obtained by using a short IBA pulse, followed by two different IBA concentrations in the culture medium. With the aim to enlarge knowledge on the molecular aspects of adventitious rooting, we also evaluated the transcriptional modulation of an ARFs member and HISTONE H3 genes, involved in auxin signaling and cell replication, respectively, during the root induction phase of cuttings. The obtained results suggest that the selected genes, as markers of the induction phase, could be very useful for setting up efficient culture conditions along the rooting process, thus increasing micropropagation efficiency.

Molecular cloning and characterization of a novel freezing-inducible DREB1/CBF transcription factor gene in boreal plant Iceland poppy (Papaver nudicaule

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Zhuo Huang

Full Text Available Abstract DREB1 of the AP2/ERF superfamily plays a key role in the regulation of plant response to low temperatures. In this study, a novel DREB1/CBF transcription factor, PnDREB1, was isolated from Iceland poppy (Papaver nudicaule, a plant adaptive to low temperature environments. It is homologous to the known DREB1s of Arabidopsis and other plant species. It also shares similar 3D structure, and conserved and functionally important motifs with DREB1s of Arabidopsis. The phylogenetic analysis indicated that the AP2 domain of PnDREB1 is similar to those of Glycine max, Medicago truncatula, and M. sativa. PnDREB1 is constitutively expressed in diverse tissues and is increased in roots. qPCR analyses indicated that PnDREB1 is significantly induced by freezing treatment as well as by abscissic acid. The expression levels induced by freezing treatment were higher in the variety with higher degree of freezing tolerance. These results suggested that PnDREB1 is a novel and functional DREB1 transcription factor involved in freezing response and possibly in other abiotic stresses. Furthermore, the freezing-induction could be suppressed by exogenous gibberellins acid, indicating that PnDREB1 might play some role in the GA signaling transduction pathway. This study provides a basis for better understanding the roles of DREB1 in adaption of Iceland poppy to low temperatures.

Renin-angiotensin system inhibition ameliorates CCl4-induced liver fibrosis in mice through the inactivation of nuclear transcription factor kappa B.

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Saber, Sameh; Mahmoud, Amr A A; Helal, Noha S; El-Ahwany, Eman; Abdelghany, Rasha H

2018-06-01

Therapeutic interventions for liver fibrosis are still limited due to the complicated molecular pathogenesis. Renin-angiotensin system (RAS) seems to contribute to the development of hepatic fibrosis. Therefore, we aimed to examine the effect of RAS inhibition on CCl 4 -induced liver fibrosis. Mice were treated with silymarin (30 mg·kg -1 ), perindopril (1 mg·kg -1 ), fosinopril (2 mg·kg -1 ), or losartan (10 mg·kg -1 ). The administration of RAS inhibitors improved liver histology and decreased protein expression of alpha smooth muscle actin (α-SMA) and hepatic content of hydroxyproline. These effects found to be mediated via inactivation of nuclear transcription factor kappa B (NFκB) pathway by the inhibition of NFκB p65 phosphorylation at the Ser536 residue and phosphorylation-induced degradation of nuclear factor kappa-B inhibitor alpha (NFκBia) subsequently inhibited NFκB-induced TNF-α and TGF-β1, leading to lower levels of tissue inhibitor of metalloproteinase-1 (TIMP-1) and vascular endothelial growth factor (VEGF). We concluded that the tissue affinity of the angiotensin converting enzyme inhibitors (ACEIs) has no impact on its antifibrotic activity and that interfering the RAS either through the inhibition of ACE or the blockade of AT1R has the same therapeutic benefit. These results suggest RAS inhibitors as promising candidates for further clinical trials in the management of hepatic fibrosis.

A deeper look into transcription regulatory code by preferred pair distance templates for transcription factor binding sites

KAUST Repository

Kulakovskiy, Ivan V.; Belostotsky, A. A.; Kasianov, Artem S.; Esipova, Natalia G.; Medvedeva, Yulia; Eliseeva, Irina A.; Makeev, Vsevolod J.

2011-01-01

Motivation: Modern experimental methods provide substantial information on protein-DNA recognition. Studying arrangements of transcription factor binding sites (TFBSs) of interacting transcription factors (TFs) advances understanding

Palmitoylation regulates 17β-estradiol-induced estrogen receptor-α degradation and transcriptional activity.

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La Rosa, Piergiorgio; Pesiri, Valeria; Leclercq, Guy; Marino, Maria; Acconcia, Filippo

2012-05-01

The estrogen receptor-α (ERα) is a transcription factor that regulates gene expression through the binding to its cognate hormone 17β-estradiol (E2). ERα transcriptional activity is regulated by E2-evoked 26S proteasome-mediated ERα degradation and ERα serine (S) residue 118 phosphorylation. Furthermore, ERα mediates fast cell responses to E2 through the activation of signaling cascades such as the MAPK/ERK and phosphoinositide-3-kinase/v-akt murine thymoma viral oncogene homolog 1 pathways. These E2 rapid effects require a population of the ERα located at the cell plasma membrane through palmitoylation, a dynamic enzymatic modification mediated by palmitoyl-acyl-transferases. However, whether membrane-initiated and transcriptional ERα activities integrate in a unique picture or represent parallel pathways still remains to be firmly clarified. Hence, we evaluated here the impact of ERα palmitoylation on E2-induced ERα degradation and S118 phosphorylation. The lack of palmitoylation renders ERα more susceptible to E2-dependent degradation, blocks ERα S118 phosphorylation and prevents E2-induced ERα estrogen-responsive element-containing promoter occupancy. Consequently, ERα transcriptional activity is prevented and the receptor addressed to the nuclear matrix subnuclear compartment. These data uncover a circuitry in which receptor palmitoylation links E2-dependent ERα degradation, S118 phosphorylation, and transcriptional activity in a unique molecular mechanism. We propose that rapid E2-dependent signaling could be considered as a prerequisite for ERα transcriptional activity and suggest an integrated model of ERα intracellular signaling where E2-dependent early extranuclear effects control late receptor-dependent nuclear actions.

Factor requirements for transcription in the Archaeon Sulfolobus shibatae.

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Qureshi, S A; Bell, S D; Jackson, S P

1997-05-15

Archaea (archaebacteria) constitute a domain of life that is distinct from Bacteria (eubacteria) and Eucarya (eukaryotes). Although archaeal cells share many morphological features with eubacteria, their transcriptional apparatus is more akin to eukaryotic RNA polymerases I, II and III than it is to eubacterial transcription systems. Thus, in addition to possessing a 10 subunit RNA polymerase and a homologue of the TATA-binding protein (TBP), Archaea possess a polypeptide termed TFB that is homologous to eukaryotic TFIIB. Here, we investigate the factor requirements for transcription of several promoters of the archaeon Sulfolobus shibatae and its associated virus SSV. Through in vitro transcription and immunodepletion, we demonstrate that S. shibatae TBP, TFB and RNA polymerase are not complexed tightly with one another and that each is required for efficient transcription of all promoters tested. Furthermore, full transcription is restored by supplementing respective depleted extracts with recombinant TBP or TFB, indicating that TBP-associated factors or TFB-associated factors are not required. Indeed, gel-filtration suggests that Sulfolobus TBP and TFB are not associated stably with other proteins. Finally, all promoters analysed are transcribed accurately and efficiently in an in vitro system comprising recombinant TBP and TFB, together with essentially homogeneous preparation of RNA polymerase. Transcription in Archaea is therefore fundamentally homologous to that in eukaryotes, although factor requirements appear to be much less complex.

Mechanisms of bee venom-induced acute renal failure.

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Grisotto, Luciana S D; Mendes, Glória E; Castro, Isac; Baptista, Maria A S F; Alves, Venancio A; Yu, Luis; Burdmann, Emmanuel A

2006-07-01

The spread of Africanized bees in the American continent has increased the number of severe envenomation after swarm attacks. Acute renal failure (ARF) is one of the major hazards in surviving patients. To assess the mechanisms of bee venom-induced ARF, rats were evaluated before, up to 70 min and 24h after 0.5mg/kg of venom injection. Control rats received saline. Bee venom caused an early and significant reduction in glomerular filtration rate (GFR, inulin clearance, 0.84+/-0.05 to 0.40+/-0.08 ml/min/100g, pbee venom-induced ARF that may occur even without hemolysis or hypotension.

Transcription factor-based biosensor

Science.gov (United States)

Dietrich, Jeffrey A; Keasling, Jay D

2013-10-08

The present invention provides for a system comprising a BmoR transcription factor, a .sigma..sup.54-RNA polymerase, and a pBMO promoter operatively linked to a reporter gene, wherein the pBMO promoter is capable of expression of the reporter gene with an activated form of the BmoR and the .sigma..sup.54-RNA polymerase.

Detecting Differential Transcription Factor Activity from ATAC-Seq Data

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Ignacio J. Tripodi

2018-05-01

Full Text Available Transcription factors are managers of the cellular factory, and key components to many diseases. Many non-coding single nucleotide polymorphisms affect transcription factors, either by directly altering the protein or its functional activity at individual binding sites. Here we first briefly summarize high-throughput approaches to studying transcription factor activity. We then demonstrate, using published chromatin accessibility data (specifically ATAC-seq, that the genome-wide profile of TF recognition motifs relative to regions of open chromatin can determine the key transcription factor altered by a perturbation. Our method of determining which TFs are altered by a perturbation is simple, is quick to implement, and can be used when biological samples are limited. In the future, we envision that this method could be applied to determine which TFs show altered activity in response to a wide variety of drugs and diseases.

Functional Profiling of Transcription Factor Genes in Neurospora crassa

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Alexander J. Carrillo

2017-09-01

Full Text Available Regulation of gene expression by DNA-binding transcription factors is essential for proper control of growth and development in all organisms. In this study, we annotate and characterize growth and developmental phenotypes for transcription factor genes in the model filamentous fungus Neurospora crassa. We identified 312 transcription factor genes, corresponding to 3.2% of the protein coding genes in the genome. The largest class was the fungal-specific Zn2Cys6 (C6 binuclear cluster, with 135 members, followed by the highly conserved C2H2 zinc finger group, with 61 genes. Viable knockout mutants were produced for 273 genes, and complete growth and developmental phenotypic data are available for 242 strains, with 64% possessing at least one defect. The most prominent defect observed was in growth of basal hyphae (43% of mutants analyzed, followed by asexual sporulation (38%, and the various stages of sexual development (19%. Two growth or developmental defects were observed for 21% of the mutants, while 8% were defective in all three major phenotypes tested. Analysis of available mRNA expression data for a time course of sexual development revealed mutants with sexual phenotypes that correlate with transcription factor transcript abundance in wild type. Inspection of this data also implicated cryptic roles in sexual development for several cotranscribed transcription factor genes that do not produce a phenotype when mutated.

Id-1 is induced in MDCK epithelial cells by activated Erk/MAPK pathway in response to expression of the Snail and E47 transcription factors

International Nuclear Information System (INIS)

Jorda, Mireia; Vinyals, Antonia; Marazuela, Anna; Cubillo, Eva; Olmeda, David; Valero, Eva; Cano, Amparo; Fabra, Angels

2007-01-01

Id-1, a member of the helix-loop-helix transcription factor family has been shown to be involved in cell proliferation, angiogenesis and invasion of many types of human cancers. We have previously shown that stable expression of E47 and Snail repressors of the E-cadherin promoter in MDCK epithelial cell line triggers epithelial mesenchymal transition (EMT) concomitantly with changes in gene expression. We show here that both factors activate the Id-1 gene promoter and induce Id-1 mRNA and protein. The upregulation of the Id-1 gene occurs through the transactivation of the promoter by the Erk/MAPK signaling pathway. Moreover, oncogenic Ras is also able to activate Id-1 promoter in MDCK cells in the absence of both E47 and Snail transcription factors. Several transcriptionally active regulatory elements have been identified in the proximal promoter, including AP-1, Sp1 and four putative E-boxes. By EMSA, we only detected an increased binding to Sp1 and AP-1 elements in E47- and Snail-expressing cells. Binding is affected by the treatment of cells with PD 98059 MEK inhibitor, suggesting that MAPK/Erk contributes to the recruitment or assembly of proteins to Id-1 promoter. Small interfering RNA directed against Sp1 reduced Id-1 expression and the upregulation of the promoter, indicating that Sp1 is required for Id-1 induction in E47- and Snail-expressing cells. Our results provide new insights into how some target genes are activated during and/or as a consequence of the EMT triggered by both E47 and Snail transcription factors

Genomewide identification and expression analysis of the ARF gene ...

Indian Academy of Sciences (India)

2014-12-26

Dec 26, 2014 ... In this study, 31 putative apple ARF genes have been identified and located within the apple genome. ... including growth and development of the root and stem, for- ..... Script 1st Strand cDNA Synthesis Kit (Takara, Dalian,.

Cyclin D3 interacts with human activating transcription factor 5 and potentiates its transcription activity

International Nuclear Information System (INIS)

Liu Wenjin; Sun Maoyun; Jiang Jianhai; Shen Xiaoyun; Sun Qing; Liu Weicheng; Shen Hailian; Gu Jianxin

2004-01-01

The Cyclin D3 protein is a member of the D-type cyclins. Besides serving as cell cycle regulators, D-type cyclins have been reported to be able to interact with several transcription factors and modulate their transcriptional activations. Here we report that human activating transcription factor 5 (hATF5) is a new interacting partner of Cyclin D3. The interaction was confirmed by in vivo coimmunoprecipitation and in vitro binding analysis. Neither interaction between Cyclin D1 and hATF5 nor interaction between Cyclin D2 and hATF5 was observed. Confocal microscopy analysis showed that Cyclin D3 could colocalize with hATF5 in the nuclear region. Cyclin D3 could potentiate hATF5 transcriptional activity independently of its Cdk4 partner. But Cyclin D1 and Cyclin D2 had no effect on hATF5 transcriptional activity. These data provide a new clue to understand the new role of Cyclin D3 as a transcriptional regulator

Reconsolidation or extinction: transcription factor switch in the determination of memory course after retrieval.

Science.gov (United States)

de la Fuente, Verónica; Freudenthal, Ramiro; Romano, Arturo

2011-04-13

In fear conditioning, aversive stimuli are readily associated with contextual features. A brief reexposure to the training context causes fear memory reconsolidation, whereas a prolonged reexposure induces memory extinction. The regulation of hippocampal gene expression plays a key role in contextual memory consolidation and reconsolidation. However, the mechanisms that determine whether memory will reconsolidate or extinguish are not known. Here, we demonstrate opposing roles for two evolutionarily related transcription factors in the mouse hippocampus. We found that nuclear factor-κB (NF-κB) is required for fear memory reconsolidation. Conversely, calcineurin phosphatase inhibited NF-κB and induced nuclear factor of activated T-cells (NFAT) nuclear translocation in the transition between reconsolidation and extinction. Accordingly, the hippocampal inhibition of both calcineurin and NFAT independently impaired memory extinction, whereas inhibition of NF-κB enhanced memory extinction. These findings represent the first insight into the molecular mechanisms that determine memory reprocessing after retrieval, supporting a transcriptional switch that directs memory toward reconsolidation or extinction. The precise molecular characterization of postretrieval processes has potential importance to the development of therapeutic strategies for fear memory disorders.

Characterization of the transcriptional profile in primary astrocytes after oxidative stress induced by Paraquat

DEFF Research Database (Denmark)

Olesen, Birgitte S. M. Thuesen; Clausen, Jørgen; Vang, Ole

2008-01-01

the antioxidative enzymes Mn- and CuZn superoxide dismutase (SOD) and catalase as well as the transcription factor component AP-1. Paraquat induced the expression of Mn- and CuZn SOD, catalase and decreases the expression of c-jun (a part of AP-1). Furthermore, the gene expression profiles were investigated after...

Bypass of senescence by the polycomb group protein CBX8 through direct binding to the INK4A-ARF locus

DEFF Research Database (Denmark)

Dietrich, Nikolaj; Bracken, Adrian P; Trinh, Emmanuelle

2007-01-01

-ARF, and that ectopic expression of CBX8 leads to repression of the Ink4a-Arf locus and bypass of senescence, leading to cellular immortalization. Gene expression and location analysis demonstrate that besides the INK4A-ARF locus, CBX8 also regulates a number of other genes important for cell growth and survival...

Modulation of DNA binding by gene-specific transcription factors.

Science.gov (United States)

Schleif, Robert F

2013-10-01

The transcription of many genes, particularly in prokaryotes, is controlled by transcription factors whose activity can be modulated by controlling their DNA binding affinity. Understanding the molecular mechanisms by which DNA binding affinity is regulated is important, but because forming definitive conclusions usually requires detailed structural information in combination with data from extensive biophysical, biochemical, and sometimes genetic experiments, little is truly understood about this topic. This review describes the biological requirements placed upon DNA binding transcription factors and their consequent properties, particularly the ways that DNA binding affinity can be modulated and methods for its study. What is known and not known about the mechanisms modulating the DNA binding affinity of a number of prokaryotic transcription factors, including CAP and lac repressor, is provided.

Effect of Cuscuta chinensis on renal function in ischemia/reperfusion-induced acute renal failure rats.

Science.gov (United States)

Shin, Sun; Lee, Yun Jung; Kim, Eun Ju; Lee, An Sook; Kang, Dae Gill; Lee, Ho Sub

2011-01-01

The kidneys play a central role in regulating water, ion composition and excretion of metabolic waste products in the urine. Cuscuta chinensis has been known as an important traditional Oriental medicine for the treatment of liver and kidney disorders. Thus, we studied whether an aqueous extract of Cuscuta chinensis (ACC) seeds has an effect on renal function parameters in ischemia/reperfusion-induced acute renal failure (ARF) rats. Administration of 250 mg/kg/day ACC showed that renal functional parameters including urinary excretion rate, osmolality, Na(+), K(+), Cl(-), creatinine clearance, solute-free water reabsorption were significantly recovered in ischemia/reperfusion-induced ARF. Periodic acid Schiff staining showed that administration of ACC improved tubular damage in ischemia/reperfusion-induced ARF. In immunoblot and immunohistological examinations, ischemia/reperfusion-induced ARF decreased the expressions of water channel AQP 2, 3 and sodium potassium pump Na,K-ATPase in the renal medulla. However, administration of ACC markedly incremented AQP 2, 3 and Na,K-ATPase expressions. Therefore, these data indicate that administration of ACC ameliorates regulation of the urine concentration and renal functions in rats with ischemia/reperfusion-induced ARF.

Hypoxia-inducible factor-1α induces multidrug resistance protein in colon cancer

Directory of Open Access Journals (Sweden)

Lv Y

2015-07-01

Full Text Available Yingqian Lv, Shan Zhao, Jinzhu Han, Likang Zheng, Zixin Yang, Li Zhao Department of Oncology, The Second Hospital, Hebei Medical University, Shijiazhuang, Hebei Province, People’s Republic of China Abstract: Multidrug resistance is the major cause of chemotherapy failure in many solid tumors, including colon cancer. Hypoxic environment is a feature for all solid tumors and is important for the development of tumor resistance to chemotherapy. Hypoxia-inducible factor (HIF-1α is the key transcription factor that mediates cellular response to hypoxia. HIF-1α has been shown to play an important role in tumor resistance; however, the mechanism is still not fully understood. Here, we found that HIF-1α and the drug resistance-associated gene multidrug resistance associated protein 1 (MRP1 were induced by treatment of colon cancer cells with the hypoxia-mimetic agent cobalt chloride. Inhibition of HIF-1α by RNA interference and dominant-negative protein can significantly reduce the induction of MRP1 by hypoxia. Bioinformatics analysis showed that a hypoxia response element is located at -378 to -373 bp upstream of the transcription start site of MRP1 gene. Luciferase reporter assay combined with mutation analysis confirmed that this element is essential for hypoxia-mediated activation of MRP gene. Furthermore, RNA interference revealed that HIF-1α is necessary for this hypoxia-driven activation of MRP1 promoter. Importantly, chromatin immunoprecipitation analysis demonstrated that HIF-1α could directly bind to this HRE site in vivo. Together, these data suggest that MRP1 is a downstream target gene of HIF-1α, which provides a potential novel mechanism for HIF-1α-mediated drug resistance in colon cancer and maybe other solid tumors as well. Keywords: hypoxia, hypoxia-inducible factor-1α, multidrug resistance associated protein, transcriptional regulation, chemotherapy tolerance

Activation-Inactivation Cycling of Rab35 and ARF6 Is Required for Phagocytosis of Zymosan in RAW264 Macrophages

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Youhei Egami

2015-01-01

Full Text Available Phagocytosis of zymosan by phagocytes is a widely used model of microbial recognition by the innate immune system. Live-cell imaging showed that fluorescent protein-fused Rab35 accumulated in the membranes of phagocytic cups and then dissociated from the membranes of newly formed phagosomes. By our novel pull-down assay for Rab35 activity, we found that Rab35 is deactivated immediately after zymosan internalization into the cells. Phagosome formation was inhibited in cells expressing the GDP- or GTP-locked Rab35 mutant. Moreover, the simultaneous expression of ACAP2—a Rab35 effector protein—with GTP-locked Rab35 or the expression of plasma membrane-targeted ACAP2 showed a marked inhibitory effect on phagocytosis through ARF6 inactivation by the GAP activity of ACAP2. ARF6, a substrate for ACAP2, was also localized on the phagocytic cups and dissociated from the membranes of internalized phagosomes. In support of the microscopic observations, ARF6-GTP pull-down experiments showed that ARF6 is transiently activated during phagosome formation. Furthermore, the expression of GDP- or GTP-locked ARF6 mutants also suppresses the uptake of zymosan. These data suggest that the activation-inactivation cycles of Rab35 and ARF6 are required for the uptake of zymosan and that ACAP2 is an important component that links Rab35/ARF6 signaling during phagocytosis of zymosan.

A Systematic Approach to Identify Candidate Transcription Factors that Control Cell Identity

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Ana C. D’Alessio

2015-11-01

Full Text Available Hundreds of transcription factors (TFs are expressed in each cell type, but cell identity can be induced through the activity of just a small number of core TFs. Systematic identification of these core TFs for a wide variety of cell types is currently lacking and would establish a foundation for understanding the transcriptional control of cell identity in development, disease, and cell-based therapy. Here, we describe a computational approach that generates an atlas of candidate core TFs for a broad spectrum of human cells. The potential impact of the atlas was demonstrated via cellular reprogramming efforts where candidate core TFs proved capable of converting human fibroblasts to retinal pigment epithelial-like cells. These results suggest that candidate core TFs from the atlas will prove a useful starting point for studying transcriptional control of cell identity and reprogramming in many human cell types.

A critique on nuclear factor-kappa B and signal transducer and activator of transcription 3: The key transcription factors in periodontal pathogenesis

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Ranjith Ambili

2017-01-01

Full Text Available Periodontal disease is initiated by microorganisms in dental plaque, and host immunoinflammatory response to the microbial challenge helps in disease progression. Conventional periodontal therapy was mainly targeted on the elimination of microbial component. However, a better understanding of molecular aspects in host response will enable the clinicians to formulate effective host modulation therapy (HMT for the periodontal management. Inflammatory mediators were the main targets for HMT in the past. Transcription factors can regulate the production of multiple mediators simultaneously, and inhibition of these factors will be more beneficial than blocking individual molecule. Two important transcription factors implicated in chronic inflammatory diseases are nuclear factor kappa B (NF-κB and signal transducers and activators of transcription 3. The role of these factors in periodontal disease is a less explored area. This comprehensive review is aimed at unveiling the critical role of NF-κB and signal transducers and activators of transcription 3 in periodontal pathogenesis. An online search was performed using MEDLINE/PubMed database. All publications till 2016 related to NF-κB, signal transducer and activator of transcription 3 (STAT3, and inflammation were included in writing this review. A total of 27,390 references were published based on the search terms used. Out of these, 507 were related to the periodontal research published in English till 2016. Relevant papers were chosen after carefully reading the abstract. This review has attempted to comprehend the existing knowledge regarding the role of transcription factors NF-κB and STAT3 in periodontal disease. Moreover, it also provides a connecting molecular link for the periodontal medicine concept.

Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes.

Science.gov (United States)

Riechmann, J L; Heard, J; Martin, G; Reuber, L; Jiang, C; Keddie, J; Adam, L; Pineda, O; Ratcliffe, O J; Samaha, R R; Creelman, R; Pilgrim, M; Broun, P; Zhang, J Z; Ghandehari, D; Sherman, B K; Yu, G

2000-12-15

The completion of the Arabidopsis thaliana genome sequence allows a comparative analysis of transcriptional regulators across the three eukaryotic kingdoms. Arabidopsis dedicates over 5% of its genome to code for more than 1500 transcription factors, about 45% of which are from families specific to plants. Arabidopsis transcription factors that belong to families common to all eukaryotes do not share significant similarity with those of the other kingdoms beyond the conserved DNA binding domains, many of which have been arranged in combinations specific to each lineage. The genome-wide comparison reveals the evolutionary generation of diversity in the regulation of transcription.

Endocytosis of HERG is clathrin-independent and involves arf6.

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Rucha Karnik

Full Text Available The hERG potassium channel is critical for repolarisation of the cardiac action potential. Reduced expression of hERG at the plasma membrane, whether caused by hereditary mutations or drugs, results in long QT syndrome and increases the risk of ventricular arrhythmias. Thus, it is of fundamental importance to understand how the density of this channel at the plasma membrane is regulated. We used antibodies to an extracellular native or engineered epitope, in conjunction with immunofluorescence and ELISA, to investigate the mechanism of hERG endocytosis in recombinant cells and validated the findings in rat neonatal cardiac myocytes. The data reveal that this channel undergoes rapid internalisation, which is inhibited by neither dynasore, an inhibitor of dynamin, nor a dominant negative construct of Rab5a, into endosomes that are largely devoid of the transferrin receptor. These results support a clathrin-independent mechanism of endocytosis and exclude involvement of dynamin-dependent caveolin and RhoA mechanisms. In agreement, internalised hERG displayed marked overlap with glycosylphosphatidylinositol-anchored GFP, a clathrin-independent cargo. Endocytosis was significantly affected by cholesterol extraction with methyl-β-cyclodextrin and inhibition of Arf6 function with dominant negative Arf6-T27N-eGFP. Taken together, we conclude that hERG undergoes clathrin-independent endocytosis via a mechanism involving Arf6.

E2F1 transcription is induced by genotoxic stress through ATM/ATR activation.

Science.gov (United States)

Carcagno, Abel L; Ogara, María F; Sonzogni, Silvina V; Marazita, Mariela C; Sirkin, Pablo F; Ceruti, Julieta M; Cánepa, Eduardo T

2009-05-01

E2F1, a member of the E2F family of transcription factors, plays a critical role in controlling both cell cycle progression and apoptotic cell death in response to DNA damage and oncogene activation. Following genotoxic stresses, E2F1 protein is stabilized by phosphorylation and acetylation driven to its accumulation. The aim of the present work was to examine whether the increase in E2F1 protein levels observed after DNA damage is only a reflection of an increase in E2F1 protein stability or is also the consequence of enhanced transcription of the E2F1 gene. The data presented here demonstrates that UV light and other genotoxics induce the transcription of E2F1 gene in an ATM/ATR dependent manner, which results in increasing E2F1 mRNA and protein levels. After genotoxic stress, transcription of cyclin E, an E2F1 target gene, was significantly induced. This induction was the result of two well-differentiated effects, one of them dependent on de novo protein synthesis and the other on the protein stabilization. Our results strongly support a transcriptional effect of DNA damaging agents on E2F1 expression. The results presented herein uncover a new mechanism involving E2F1 in response to genotoxic stress.

Investigation on gas medium parameters for an ArF excimer laser through orthogonal experimental design

Science.gov (United States)

Song, Xingliang; Sha, Pengfei; Fan, Yuanyuan; Jiang, R.; Zhao, Jiangshan; Zhou, Yi; Yang, Junhong; Xiong, Guangliang; Wang, Yu

2018-02-01

Due to complex kinetics of formation and loss mechanisms, such as ion-ion recombination reaction, neutral species harpoon reaction, excited state quenching and photon absorption, as well as their interactions, the performance behavior of different laser gas medium parameters for excimer laser varies greatly. Therefore, the effects of gas composition and total gas pressure on excimer laser performance attract continual research studies. In this work, orthogonal experimental design (OED) is used to investigate quantitative and qualitative correlations between output laser energy characteristics and gas medium parameters for an ArF excimer laser with plano-plano optical resonator operation. Optimized output laser energy with good pulse to pulse stability can be obtained effectively by proper selection of the gas medium parameters, which makes the most of the ArF excimer laser device. Simple and efficient method for gas medium optimization is proposed and demonstrated experimentally, which provides a global and systematic solution. By detailed statistical analysis, the significance sequence of relevant parameter factors and the optimized composition for gas medium parameters are obtained. Compared with conventional route of varying single gas parameter factor sequentially, this paper presents a more comprehensive way of considering multivariables simultaneously, which seems promising in striking an appropriate balance among various complicated parameters for power scaling study of an excimer laser.

Inhibition of FoxO1 acetylation by INHAT subunit SET/TAF-Iβ induces p21 transcription.

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Chae, Yun-Cheol; Kim, Kee-Beom; Kang, Joo-Young; Kim, Se-Ryeon; Jung, Hyeon-Soo; Seo, Sang-Beom

2014-08-25

Post-translational modification of forkhead family transcription factor, FoxO1, is an important regulatory mode for its diverse activities. FoxO1 is acetylated by HAT coactivators and its transcriptional activity is decreased via reduced DNA binding affinity. Here, we report that SET/TAF-Iβ inhibited p300-mediated FoxO1 acetylation in an INHAT domain-dependent manner. SET/TAF-Iβ interacted with FoxO1 and activated transcription of FoxO1 target gene, p21. Moreover, SET/TAF-Iβ inhibited acetylation of FoxO1 and increased p21 transcription induced by oxidative stress. Our results suggest that SET/TAF-Iβ inhibits FoxO1 acetylation and activates its transcriptional activity toward p21. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

BACH transcription factors in innate and adaptive immunity.

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Igarashi, Kazuhiko; Kurosaki, Tomohiro; Roychoudhuri, Rahul

2017-07-01

BTB and CNC homology (BACH) proteins are transcriptional repressors of the basic region leucine zipper (bZIP) transcription factor family. Recent studies indicate widespread roles of BACH proteins in controlling the development and function of the innate and adaptive immune systems, including the differentiation of effector and memory cells of the B and T cell lineages, CD4 + regulatory T cells and macrophages. Here, we emphasize similarities at a molecular level in the cell-type-specific activities of BACH factors, proposing that competitive interactions of BACH proteins with transcriptional activators of the bZIP family form a common mechanistic theme underlying their diverse actions. The findings contribute to a general understanding of how transcriptional repressors shape lineage commitment and cell-type-specific functions through repression of alternative lineage programmes.

Hypoxia-inducible transcription factor-1α promotes hypoxia-induced A549 apoptosis via a mechanism that involves the glycolysis pathway

International Nuclear Information System (INIS)

Luo, FengMing; Liu, XiaoJing; Yan, NaiHong; Li, ShuangQing; Cao, GuiQun; Cheng, QingYing; Xia, QingJie; Wang, HongJing

2006-01-01

Hypoxia-inducible transcription factor-1α (HIF-1α), which plays an important role in controlling the hypoxia-induced glycolysis pathway, is a 'master' gene in the tissue hypoxia response during tumor development. However, its role in the apoptosis of non-small cell lung cancer remains unknown. Here, we have studied the effects of HIF-1α on apoptosis by modulating HIF-1α gene expression in A549 cells through both siRNA knock-down and over-expression. A549 cells were transfected with a HIF-1α siRNA plasmid or a HIF-1α expression vector. Transfected cells were exposed to a normoxic or hypoxic environment in the presence or absence of 25 mM HEPES and 2-deoxyglucose (2-DG) (5 mM). The expression of three key genes of the glycolysis pathway, glucose transporter type 1(GLUT1), phosphoglycerate kinase 1(PGK1), and hexokinase 1(HK1), were measured using real-time RT-PCR. Glycolysis was monitored by measuring changes of pH and lactate concentration in the culture medium. Apoptosis was detected by TUNEL assay and flow cytometry. Knocking down expression of HIF-1α inhibited the glycolysis pathway, increased the pH of the culture medium, and protected the cells from hypoxia-induced apoptosis. In contrast, over-expression of HIF-1α accelerated glycolysis in A549 cells, decreased the pH of the culture medium, and enhanced hypoxia-induced apoptosis. These effects of HIF-1α on glycolysis, pH of the medium, and apoptosis were reversed by treatment with the glycolytic inhibitor, 2-DG. Apoptosis induced by HIF-1α over-expression was partially inhibited by increasing the buffering capacity of the culture medium by adding HEPES. During hypoxia in A549 cells, HIF-1α promotes activity of the glycolysis pathway and decreases the pH of the culture medium, resulting in increased cellular apoptosis

Rice homeobox transcription factor HOX1a positively regulates gibberellin responses by directly suppressing EL1.

Science.gov (United States)

Wen, Bi-Qing; Xing, Mei-Qing; Zhang, Hua; Dai, Cheng; Xue, Hong-Wei

2011-11-01

Homeobox transcription factors are involved in various aspects of plant development, including maintenance of the biosynthesis and signaling pathways of different hormones. However, few direct targets of homeobox proteins have been identified. We here show that overexpression of rice homeobox gene HOX1a resulted in enhanced gibberellin (GA) response, indicating a positive effect of HOX1a in GA signaling. HOX1a is induced by GA and encodes a homeobox transcription factor with transcription repression activity. In addition, HOX1a suppresses the transcription of early flowering1 (EL1), a negative regulator of GA signaling, and further electrophoretic mobility shift assay and chromatin immunoprecipitation analysis revealed that HOX1a directly bound to the promoter region of EL1 to suppress its expression and stimulate GA signaling. These results demonstrate that HOX1a functions as a positive regulator of GA signaling by suppressing EL1, providing informative hints on the study of GA signaling. © 2011 Institute of Botany, Chinese Academy of Sciences.

Thirty-seven transcription factor genes differentially respond to a ...

Indian Academy of Sciences (India)

Plant transcription factors and insect defence si. Thirty-seven transcription factor genes differentially respond to a harpin protein and affect resistance to the green peach aphid in Arabidopsis. HUNLIN. PIN. RUOXUE LIŲ, BEIBEI LÜ, XIAOMENG WANG, CHUNLING ZHANG, SHUPING ZHANG, JUN QIAN, LEI CHEN,.

Structural Fingerprints of Transcription Factor Binding Site Regions

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Peter Willett

2009-03-01

Full Text Available Fourier transforms are a powerful tool in the prediction of DNA sequence properties, such as the presence/absence of codons. We have previously compiled a database of the structural properties of all 32,896 unique DNA octamers. In this work we apply Fourier techniques to the analysis of the structural properties of human chromosomes 21 and 22 and also to three sets of transcription factor binding sites within these chromosomes. We find that, for a given structural property, the structural property power spectra of chromosomes 21 and 22 are strikingly similar. We find common peaks in their power spectra for both Sp1 and p53 transcription factor binding sites. We use the power spectra as a structural fingerprint and perform similarity searching in order to find transcription factor binding site regions. This approach provides a new strategy for searching the genome data for information. Although it is difficult to understand the relationship between specific functional properties and the set of structural parameters in our database, our structural fingerprints nevertheless provide a useful tool for searching for function information in sequence data. The power spectrum fingerprints provide a simple, fast method for comparing a set of functional sequences, in this case transcription factor binding site regions, with the sequences of whole chromosomes. On its own, the power spectrum fingerprint does not find all transcription factor binding sites in a chromosome, but the results presented here show that in combination with other approaches, this technique will improve the chances of identifying functional sequences hidden in genomic data.

Alteration of BRCA1 expression affects alcohol-induced transcription of RNA Pol III-dependent genes.

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Zhong, Qian; Shi, Ganggang; Zhang, Yanmei; Lu, Lei; Levy, Daniel; Zhong, Shuping

2015-02-01

Emerging evidence has indicated that alcohol consumption is an established risk factor for breast cancer. Deregulation of RNA polymerase III (Pol III) transcription enhances cellular Pol III gene production, leading to an increase in translational capacity to promote cell transformation and tumor formation. We have reported that alcohol intake increases Pol III gene transcription to promote cell transformation and tumor formation in vitro and in vivo. Studies revealed that tumor suppressors, pRb, p53, PTEN and Maf1 repress the transcription of Pol III genes. BRCA1 is a tumor suppressor and its mutation is tightly related to breast cancer development. However, it is not clear whether BRCA1 expression affects alcohol-induced transcription of Pol III genes. At the present studies, we report that restoring BRCA1 in HCC 1937 cells, which is a BRCA1 deficient cell line, represses Pol III gene transcription. Expressing mutant or truncated BRCA1 in these cells does not affect the ability of repression on Pol III genes. Our analysis has demonstrated that alcohol induces Pol III gene transcription. More importantly, overexpression of BRCA1 in estrogen receptor positive (ER+) breast cancer cells (MCF-7) decreases the induction of tRNA(Leu) and 5S rRNA genes by alcohol, whereas reduction of BRCA1 by its siRNA slightly increases the transcription of the class of genes. This suggests that BRCA1 is associated with alcohol-induced deregulation of Pol III genes. These studies for the first time demonstrate the role of BRCA1 in induction of Pol III genes by alcohol and uncover a novel mechanism of alcohol-associated breast cancer. Copyright © 2014 Elsevier B.V. All rights reserved.

Hypoxia Inducible Factors and Hypertension: Lessons from Sleep Apnea Syndrome

Science.gov (United States)

Nanduri, Jayasri; Peng, Ying-Jie; Yuan, Guoxiang; Kumar, Ganesh K.; Prabhakar, Nanduri R.

2015-01-01

Systemic hypertension is one of the most prevalent cardiovascular diseases. Sleep disordered breathing (SDB) with recurrent apnea is a major risk factor for developing essential hypertension. Chronic intermittent hypoxia (CIH) is a hallmark manifestation of recurrent apnea. Rodent models patterned after the O2 profiles seen with SDB patients showed that CIH is the major stimulus for causing systemic hypertension. This article reviews the physiological and molecular basis of CIH-induced hypertension. Physiological studies have identified that augmented carotid body chemosensory reflex and the resulting increase in sympathetic nerve activity is a major contributor to CIH-induced hypertension. Analysis of molecular mechanisms revealed that CIH activates hypoxia-inducible factor (HIF)-1 and suppresses HIF-2- mediated transcription. Dysregulation of HIF-1- and HIF-2- mediated transcription leads to imbalance of pro-oxidant and anti-oxidant enzyme gene expression resulting in increased reactive species (ROS) generation in the chemosensory reflex which is central for developing hypertension. PMID:25772710

Integration and diversity of the regulatory network composed of Maf and CNC families of transcription factors.

Science.gov (United States)

Motohashi, Hozumi; O'Connor, Tania; Katsuoka, Fumiki; Engel, James Douglas; Yamamoto, Masayuki

2002-07-10

Recent progress in the analysis of transcriptional regulation has revealed the presence of an exquisite functional network comprising the Maf and Cap 'n' collar (CNC) families of regulatory proteins, many of which have been isolated. Among Maf factors, large Maf proteins are important in the regulation of embryonic development and cell differentiation, whereas small Maf proteins serve as obligatory heterodimeric partner molecules for members of the CNC family. Both Maf homodimers and CNC-small Maf heterodimers bind to the Maf recognition element (MARE). Since the MARE contains a consensus TRE sequence recognized by AP-1, Jun and Fos family members may act to compete or interfere with the function of CNC-small Maf heterodimers. Overall then, the quantitative balance of transcription factors interacting with the MARE determines its transcriptional activity. Many putative MARE-dependent target genes such as those induced by antioxidants and oxidative stress are under concerted regulation by the CNC family member Nrf2, as clearly proven by mouse germline mutagenesis. Since these genes represent a vital aspect of the cellular defense mechanism against oxidative stress, Nrf2-null mutant mice are highly sensitive to xenobiotic and oxidative insults. Deciphering the molecular basis of the regulatory network composed of Maf and CNC families of transcription factors will undoubtedly lead to a new paradigm for the cooperative function of transcription factors.

Radiation-induced alternative transcripts as detected in total and polysome-bound mRNA.

Science.gov (United States)

Wahba, Amy; Ryan, Michael C; Shankavaram, Uma T; Camphausen, Kevin; Tofilon, Philip J

2018-01-02

Alternative splicing is a critical event in the posttranscriptional regulation of gene expression. To investigate whether this process influences radiation-induced gene expression we defined the effects of ionizing radiation on the generation of alternative transcripts in total cellular mRNA (the transcriptome) and polysome-bound mRNA (the translatome) of the human glioblastoma stem-like cell line NSC11. For these studies, RNA-Seq profiles from control and irradiated cells were compared using the program SpliceSeq to identify transcripts and splice variations induced by radiation. As compared to the transcriptome (total RNA) of untreated cells, the radiation-induced transcriptome contained 92 splice events suggesting that radiation induced alternative splicing. As compared to the translatome (polysome-bound RNA) of untreated cells, the radiation-induced translatome contained 280 splice events of which only 24 were overlapping with the radiation-induced transcriptome. These results suggest that radiation not only modifies alternative splicing of precursor mRNA, but also results in the selective association of existing mRNA isoforms with polysomes. Comparison of radiation-induced alternative transcripts to radiation-induced gene expression in total RNA revealed little overlap (about 3%). In contrast, in the radiation-induced translatome, about 38% of the induced alternative transcripts corresponded to genes whose expression level was affected in the translatome. This study suggests that whereas radiation induces alternate splicing, the alternative transcripts present at the time of irradiation may play a role in the radiation-induced translational control of gene expression and thus cellular radioresponse.

NAC Transcription Factors in Stress Responses and Senescence

DEFF Research Database (Denmark)

O'Shea, Charlotte

Plant-specific NAM/ATAF/CUC (NAC) transcription factors have recently received considerable attention due to their significant roles in plant development and stress signalling. This interest has resulted in a number of physiological, genetic and cell biological studies of their functions. Some...... of these studies have also revealed emerging gene regulatory networks and protein-protein interaction networks. However, structural studies relating structure to function are lagging behind. Structure-function analysis of the NAC transcription factors has therefore been the main focus of this PhD thesis...... not involve significant folding-upon-binding but fuzziness or an extended ANAC046 region. The ANAC046 regulatory domain functions as an entropic chain with a bait for interactions with for example RCD1. RCD1 interacts with transcription factors from several different families, and the large stress...

Furan- and Thiophene-2-Carbonyl Amino Acid Derivatives Activate Hypoxia-Inducible Factor via Inhibition of Factor Inhibiting Hypoxia-Inducible Factor-1

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Shin-ichi Kawaguchi

2018-04-01

Full Text Available Induction of a series of anti-hypoxic proteins protects cells during exposure to hypoxic conditions. Hypoxia-inducible factor-α (HIF-α is a major transcription factor that orchestrates this protective effect. To activate HIF exogenously, without exposing cells to hypoxic conditions, many small-molecule inhibitors targeting prolyl hydroxylase domain-containing protein have been developed. In addition, suppression of factor inhibiting HIF-1 (FIH-1 has also been shown to have the potential to activate HIF-α. However, few small-molecule inhibitors of FIH-1 have been developed. In this study, we synthesized a series of furan- and thiophene-2-carbonyl amino acid derivatives having the potential to inhibit FIH-1. The inhibitory activities of these compounds were evaluated in SK-N-BE(2c cells by measuring HIF response element (HRE promoter activity. Several furan- and thiophene-2-carbonyl amino acid derivatives inhibited FIH-1 based on correlations among the docking score of the FIH-1 active site, the chemical structure of the compounds, and biological HIF-α/HRE transcriptional activity.

Transcription factor levels enable metabolic diversification of single cells of environmental bacteria.

Science.gov (United States)

Guantes, Raúl; Benedetti, Ilaria; Silva-Rocha, Rafael; de Lorenzo, Víctor

2016-05-01

Transcriptional noise is a necessary consequence of the molecular events that drive gene expression in prokaryotes. However, some environmental microorganisms that inhabit polluted sites, for example, the m-xylene degrading soil bacterium Pseudomonas putida mt-2 seem to have co-opted evolutionarily such a noise for deploying a metabolic diversification strategy that allows a cautious exploration of new chemical landscapes. We have examined this phenomenon under the light of deterministic and stochastic models for activation of the main promoter of the master m-xylene responsive promoter of the system (Pu) by its cognate transcriptional factor (XylR). These analyses consider the role of co-factors for Pu activation and determinants of xylR mRNA translation. The model traces the onset and eventual disappearance of the bimodal distribution of Pu activity along time to the growth-phase dependent abundance of XylR itself, that is, very low in exponentially growing cells and high in stationary. This tenet was validated by examining the behaviour of a Pu-GFP fusion in a P. putida strain in which xylR expression was engineered under the control of an IPTG-inducible system. This work shows how a relatively simple regulatory scenario (for example, growth-phase dependent expression of a limiting transcription factor) originates a regime of phenotypic diversity likely to be advantageous in competitive environmental settings.

TDP2 suppresses chromosomal translocations induced by DNA topoisomerase II during gene transcription.

Science.gov (United States)

Gómez-Herreros, Fernando; Zagnoli-Vieira, Guido; Ntai, Ioanna; Martínez-Macías, María Isabel; Anderson, Rhona M; Herrero-Ruíz, Andrés; Caldecott, Keith W

2017-08-10

DNA double-strand breaks (DSBs) induced by abortive topoisomerase II (TOP2) activity are a potential source of genome instability and chromosome translocation. TOP2-induced DNA double-strand breaks are rejoined in part by tyrosyl-DNA phosphodiesterase 2 (TDP2)-dependent non-homologous end-joining (NHEJ), but whether this process suppresses or promotes TOP2-induced translocations is unclear. Here, we show that TDP2 rejoins DSBs induced during transcription-dependent TOP2 activity in breast cancer cells and at the translocation 'hotspot', MLL. Moreover, we find that TDP2 suppresses chromosome rearrangements induced by TOP2 and reduces TOP2-induced chromosome translocations that arise during gene transcription. Interestingly, however, we implicate TDP2-dependent NHEJ in the formation of a rare subclass of translocations associated previously with therapy-related leukemia and characterized by junction sequences with 4-bp of perfect homology. Collectively, these data highlight the threat posed by TOP2-induced DSBs during transcription and demonstrate the importance of TDP2-dependent non-homologous end-joining in protecting both gene transcription and genome stability.DNA double-strand breaks (DSBs) induced by topoisomerase II (TOP2) are rejoined by TDP2-dependent non-homologous end-joining (NHEJ) but whether this promotes or suppresses translocations is not clear. Here the authors show that TDP2 suppresses chromosome translocations from DSBs introduced during gene transcription.

Cross-Family Transcription Factor Interactions

NARCIS (Netherlands)

Bemer, Marian; Dijk, van Aalt-Jan; Immink, Richard G.H.; Angenent, Gerco C.

2017-01-01

Specific and dynamic gene expression strongly depends on transcription factor (TF) activity and most plant TFs function in a combinatorial fashion. They can bind to DNA and control the expression of the corresponding gene in an additive fashion or cooperate by physical interactions, forming larger

Transcription factor interplay in T helper cell differentiation

Science.gov (United States)

Evans, Catherine M.

2013-01-01

The differentiation of CD4 helper T cells into specialized effector lineages has provided a powerful model for understanding immune cell differentiation. Distinct lineages have been defined by differential expression of signature cytokines and the lineage-specifying transcription factors necessary and sufficient for their production. The traditional paradigm of differentiation towards Th1 and Th2 subtypes driven by T-bet and GATA3, respectively, has been extended to incorporate additional T cell lineages and transcriptional regulators. Technological advances have expanded our view of these lineage-specifying transcription factors to the whole genome and revealed unexpected interplay between them. From these data, it is becoming clear that lineage specification is more complex and plastic than previous models might have suggested. Here, we present an overview of the different forms of transcription factor interplay that have been identified and how T cell phenotypes arise as a product of this interplay within complex regulatory networks. We also suggest experimental strategies that will provide further insight into the mechanisms that underlie T cell lineage specification and plasticity. PMID:23878131

Transcription factor interplay in T helper cell differentiation.

Science.gov (United States)

Evans, Catherine M; Jenner, Richard G

2013-11-01

The differentiation of CD4 helper T cells into specialized effector lineages has provided a powerful model for understanding immune cell differentiation. Distinct lineages have been defined by differential expression of signature cytokines and the lineage-specifying transcription factors necessary and sufficient for their production. The traditional paradigm of differentiation towards Th1 and Th2 subtypes driven by T-bet and GATA3, respectively, has been extended to incorporate additional T cell lineages and transcriptional regulators. Technological advances have expanded our view of these lineage-specifying transcription factors to the whole genome and revealed unexpected interplay between them. From these data, it is becoming clear that lineage specification is more complex and plastic than previous models might have suggested. Here, we present an overview of the different forms of transcription factor interplay that have been identified and how T cell phenotypes arise as a product of this interplay within complex regulatory networks. We also suggest experimental strategies that will provide further insight into the mechanisms that underlie T cell lineage specification and plasticity.

In vivo evidence suggesting reciprocal renal hypoxia-inducible factor-1 upregulation and signal transducer and activator of transcription 3 activation in response to hypoxic and non-hypoxic stimuli.

Science.gov (United States)

Nechemia-Arbely, Yael; Khamaisi, Mogher; Rosenberger, Christian; Koesters, Robert; Shina, Ahuva; Geva, Carmit; Shriki, Anat; Klaus, Stephen; Rosen, Seymour; Rose-John, Stefan; Galun, Eithan; Axelrod, Jonathan H; Heyman, Samuel N

2013-04-01

In vitro studies suggest that combined activation of hypoxia-inducible factor (HIF) and signal transducer and activator of transcription 3 (STAT3) promotes the hypoxia response. However, their interrelationship in vivo remains poorly defined. The present study investigated the possible relationship between HIF-1 upregulation and STAT3 activation in the rodent kidney in vivo. Activation of HIF-1 and STAT3 was analysed by immunohistochemical staining and western blot analysis in: (i) models of hypoxia-associated kidney injury induced by radiocontrast media or rhabdomyolysis; (ii) following activation of STAT3 by the interleukin (IL)-6-soluble IL-6 receptor complex; or (iii) following HIF-1α stabilization using hypoxic and non-hypoxic stimuli (mimosine, FG-4497, CO, CoCl(2)) and in targeted von Hippel-Lindau-knockout mice. Western blot analysis and immunostaining revealed marked induction of both transcription factors under all conditions tested, suggesting that in vivo STAT3 can trigger HIF and vice versa. Colocalization of HIF-1α and phosphorylated STAT3 was detected in some, but not all, renal cell types, suggesting that in some cells a paracrine mechanism may be responsible for the reciprocal activation of the two transcription factors. Nevertheless, in several cell types spatial concordance was observed under the majority of conditions tested, suggesting that HIF-1 and STAT3 may act as cotranscription factors. These in vivo studies suggest that, in response to renal hypoxic-stress, upregulation of HIF-1 and activation of STAT3 may be both reciprocal and cell type dependent. © 2013 The Authors Clinical and Experimental Pharmacology and Physiology © 2013 Wiley Publishing Asia Pty Ltd.

Molecular cloning of transcripts induced by UV-radiation in rodent cells

International Nuclear Information System (INIS)

Fornace, A.J. Jr.; Mitchell, J.B.

1987-01-01

Several inducible DNA repair genes have been well characterized in bacteria. In eukaryotes including mammalian cells, there is increasing evidence that similar events may occur. Recently, the authors have shown that hybridization subtraction can be used to enrich for sequences induced only several fold by a particular cell treatment such as heat shock. Chinese hamster V79 cells were UV-irradiated with 17 Jm/sup -2/ and cDNA was synthesized from the polyadenylated (poly A) RNA. This ''UV'' cDNA was hybridized with a 3 fold excess of polyA RNA from unirradiated cells and the nonhybridizing cDNA was isolated. With this approach, UV-induced sequences were enriched over 20 fold. This enriched cDNA was cloned into a high copy number plasmid and a cDNA library was constructed. By RNA dot blot and northern analysis, 42 clones from this library were found to represent transcripts induced 3 to 25 fold by UV. The most common isolates were found to be metallothionein transcripts by DNA sequencing. The metallothionein transcripts were found to be induced 10 to 25 fold by UV with maximum induction at 4-8 h after 10 Jm/sup -2/. A similar approach was also used with a Chinese hamster ovary line which does not express metallothionein and multiple clones were isolated which represented transcripts induced 3-15 fold by UV. Except for the metallothionein clones, the other Chinese hamster cDNA clones have not been identified, but it is probable that the protein products of at least some of these transcripts play a role in the cellular response to UV damage

Evolutionarily conserved transcription factor Apontic controls the G1/S progression by inducing cyclin e during eye development

KAUST Repository

Liu, Qingxin

2014-06-16

During Drosophila eye development, differentiation initiates in the posterior region of the eye disk and progresses anteriorly as a wave marked by the morphogenetic furrow (MF), which demarcates the boundary between anterior undifferentiated cells and posterior differentiated photoreceptors. However, the mechanism underlying the regulation of gene expression immediately before the onset of differentiation remains unclear. Here, we show that Apontic (Apt), which is an evolutionarily conserved transcription factor, is expressed in the differentiating cells posterior to the MF. Moreover, it directly induces the expression of cyclin E and is also required for the G1-to-S phase transition, which is known to be essential for the initiation of cell differentiation at the MF. These observations identify a pathway crucial for eye development, governed by a mechanism in which Cyclin E promotes the G1-to-S phase transition when regulated by Apt.

Evolutionarily conserved transcription factor Apontic controls the G1/S progression by inducing cyclin e during eye development

KAUST Repository

Liu, Qingxin; Wang, Xianfeng; Ikeo, Kazuho; Hirose, Susumu; Gehring, Walter Jakob; Gojobori, Takashi

2014-01-01

During Drosophila eye development, differentiation initiates in the posterior region of the eye disk and progresses anteriorly as a wave marked by the morphogenetic furrow (MF), which demarcates the boundary between anterior undifferentiated cells and posterior differentiated photoreceptors. However, the mechanism underlying the regulation of gene expression immediately before the onset of differentiation remains unclear. Here, we show that Apontic (Apt), which is an evolutionarily conserved transcription factor, is expressed in the differentiating cells posterior to the MF. Moreover, it directly induces the expression of cyclin E and is also required for the G1-to-S phase transition, which is known to be essential for the initiation of cell differentiation at the MF. These observations identify a pathway crucial for eye development, governed by a mechanism in which Cyclin E promotes the G1-to-S phase transition when regulated by Apt.

Heat shock factor 1 upregulates transcription of Epstein–Barr Virus nuclear antigen 1 by binding to a heat shock element within the BamHI-Q promoter

International Nuclear Information System (INIS)

Wang, Feng-Wei; Wu, Xian-Rui; Liu, Wen-Ju; Liao, Yi-Ji; Lin, Sheng; Zong, Yong-Sheng; Zeng, Mu-Sheng; Zeng, Yi-Xin; Mai, Shi-Juan; Xie, Dan

2011-01-01

Epstein–Barr virus (EBV) nuclear antigen 1 (EBNA1) is essential for maintenance of the episome and establishment of latency. In this study, we observed that heat treatment effectively induced EBNA1 transcription in EBV-transformed B95-8 and human LCL cell lines. Although Cp is considered as the sole promoter used for the expression of EBNA1 transcripts in the lymphoblastoid cell lines, the RT-PCR results showed that the EBNA1 transcripts induced by heat treatment arise from Qp-initiated transcripts. Using bioinformatics, a high affinity and functional heat shock factor 1 (HSF1)-binding element within the − 17/+4 oligonucleotide of the Qp was found, and was determined by electrophoretic mobility shift assay and chromatin immunoprecipitation assay. Moreover, heat shock and exogenous HSF1 expression induced Qp activity in reporter assays. Further, RNA interference-mediated HSF1 gene silencing attenuated heat-induced EBNA1 expression in B95-8 cells. These results provide evidence that EBNA1 is a new target for the transcription factor HSF1.

Heat shock factor 1 upregulates transcription of Epstein-Barr Virus nuclear antigen 1 by binding to a heat shock element within the BamHI-Q promoter

Energy Technology Data Exchange (ETDEWEB)

Wang, Feng-Wei [The State Key Laboratory of Oncology in South China, Cancer Center, Sun Yat-Sen University, Guangzhou (China); Wu, Xian-Rui [Department of Surgery, Sixth Affiliated Hospital, Sun Yat-sen University, Guangzhou (China); Liu, Wen-Ju; Liao, Yi-Ji [The State Key Laboratory of Oncology in South China, Cancer Center, Sun Yat-Sen University, Guangzhou (China); Lin, Sheng [Laboratory of Integrated Biosciences, School of Life Science, Sun Yat-sen University, Guangzhou (China); Zong, Yong-Sheng; Zeng, Mu-Sheng; Zeng, Yi-Xin [The State Key Laboratory of Oncology in South China, Cancer Center, Sun Yat-Sen University, Guangzhou (China); Mai, Shi-Juan, E-mail: maishj@sysucc.org.cn [The State Key Laboratory of Oncology in South China, Cancer Center, Sun Yat-Sen University, Guangzhou (China); Xie, Dan, E-mail: xied@mail.sysu.edu.cn [The State Key Laboratory of Oncology in South China, Cancer Center, Sun Yat-Sen University, Guangzhou (China)

2011-12-20

Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) is essential for maintenance of the episome and establishment of latency. In this study, we observed that heat treatment effectively induced EBNA1 transcription in EBV-transformed B95-8 and human LCL cell lines. Although Cp is considered as the sole promoter used for the expression of EBNA1 transcripts in the lymphoblastoid cell lines, the RT-PCR results showed that the EBNA1 transcripts induced by heat treatment arise from Qp-initiated transcripts. Using bioinformatics, a high affinity and functional heat shock factor 1 (HSF1)-binding element within the - 17/+4 oligonucleotide of the Qp was found, and was determined by electrophoretic mobility shift assay and chromatin immunoprecipitation assay. Moreover, heat shock and exogenous HSF1 expression induced Qp activity in reporter assays. Further, RNA interference-mediated HSF1 gene silencing attenuated heat-induced EBNA1 expression in B95-8 cells. These results provide evidence that EBNA1 is a new target for the transcription factor HSF1.

DNA Binding by the Ribosomal DNA Transcription Factor Rrn3 Is Essential for Ribosomal DNA Transcription*

Science.gov (United States)

Stepanchick, Ann; Zhi, Huijun; Cavanaugh, Alice H.; Rothblum, Katrina; Schneider, David A.; Rothblum, Lawrence I.

2013-01-01

The human homologue of yeast Rrn3 is an RNA polymerase I-associated transcription factor that is essential for ribosomal DNA (rDNA) transcription. The generally accepted model is that Rrn3 functions as a bridge between RNA polymerase I and the transcription factors bound to the committed template. In this model Rrn3 would mediate an interaction between the mammalian Rrn3-polymerase I complex and SL1, the rDNA transcription factor that binds to the core promoter element of the rDNA. In the course of studying the role of Rrn3 in recruitment, we found that Rrn3 was in fact a DNA-binding protein. Analysis of the sequence of Rrn3 identified a domain with sequence similarity to the DNA binding domain of heat shock transcription factor 2. Randomization, or deletion, of the amino acids in this region in Rrn3, amino acids 382–400, abrogated its ability to bind DNA, indicating that this domain was an important contributor to DNA binding by Rrn3. Control experiments demonstrated that these mutant Rrn3 constructs were capable of interacting with both rpa43 and SL1, two other activities demonstrated to be essential for Rrn3 function. However, neither of these Rrn3 mutants was capable of functioning in transcription in vitro. Moreover, although wild-type human Rrn3 complemented a yeast rrn3-ts mutant, the DNA-binding site mutant did not. These results demonstrate that DNA binding by Rrn3 is essential for transcription by RNA polymerase I. PMID:23393135

DNA binding by the ribosomal DNA transcription factor rrn3 is essential for ribosomal DNA transcription.

Science.gov (United States)

Stepanchick, Ann; Zhi, Huijun; Cavanaugh, Alice H; Rothblum, Katrina; Schneider, David A; Rothblum, Lawrence I

2013-03-29

The human homologue of yeast Rrn3 is an RNA polymerase I-associated transcription factor that is essential for ribosomal DNA (rDNA) transcription. The generally accepted model is that Rrn3 functions as a bridge between RNA polymerase I and the transcription factors bound to the committed template. In this model Rrn3 would mediate an interaction between the mammalian Rrn3-polymerase I complex and SL1, the rDNA transcription factor that binds to the core promoter element of the rDNA. In the course of studying the role of Rrn3 in recruitment, we found that Rrn3 was in fact a DNA-binding protein. Analysis of the sequence of Rrn3 identified a domain with sequence similarity to the DNA binding domain of heat shock transcription factor 2. Randomization, or deletion, of the amino acids in this region in Rrn3, amino acids 382-400, abrogated its ability to bind DNA, indicating that this domain was an important contributor to DNA binding by Rrn3. Control experiments demonstrated that these mutant Rrn3 constructs were capable of interacting with both rpa43 and SL1, two other activities demonstrated to be essential for Rrn3 function. However, neither of these Rrn3 mutants was capable of functioning in transcription in vitro. Moreover, although wild-type human Rrn3 complemented a yeast rrn3-ts mutant, the DNA-binding site mutant did not. These results demonstrate that DNA binding by Rrn3 is essential for transcription by RNA polymerase I.

Transcription factor fos-related antigen-2 induces progressive peripheral vasculopathy in mice closely resembling human systemic sclerosis.

Science.gov (United States)

Maurer, Britta; Busch, Nicole; Jüngel, Astrid; Pileckyte, Margarita; Gay, Renate E; Michel, Beat A; Schett, Georg; Gay, Steffen; Distler, Jörg; Distler, Oliver

2009-12-08

Microvascular damage is one of the first pathological changes in systemic sclerosis. In this study, we investigated the role of Fos-related antigen-2 (Fra-2), a transcription factor of the activator protein-1 family, in the peripheral vasculopathy of systemic sclerosis and examined the underlying mechanisms. Expression of Fra-2 protein was significantly increased in skin biopsies of systemic sclerosis patients compared with healthy controls, especially in endothelial and vascular smooth muscle cells. Fra-2 transgenic mice developed a severe loss of small blood vessels in the skin that was paralleled by progressive skin fibrosis at 12 weeks of age. The reduction in capillary density was preceded by a significant increase in apoptosis in endothelial cells at week 9 as detected by immunohistochemistry. Similarly, suppression of Fra-2 by small interfering RNA prevented human microvascular endothelial cells from staurosporine-induced apoptosis and improved both the number of tubes and the cumulative tube lengths in the tube formation assay. In addition, cell migration in the scratch assay and vascular endothelial growth factor-dependent chemotaxis in a modified Boyden chamber assay were increased after transfection of human microvascular endothelial cells with Fra-2 small interfering RNA, whereas proliferation was not affected. Fra-2 is present in human systemic sclerosis and may contribute to the development of microvasculopathy by inducing endothelial cell apoptosis and by reducing endothelial cell migration and chemotaxis. Fra-2 transgenic mice are a promising preclinical model to study the mechanisms and therapeutic approaches of the peripheral vasculopathy in systemic sclerosis.

Characteristics and Efficacy of a Sterile Hyphomycete (ARF18), a New Biocontrol Agent for Heterodera glycines and Other Nematodes

Science.gov (United States)

Kim, D. G.; Riggs, R. D.

1991-01-01

A filamentous, nonsporulating fungus, designated Arkansas Fungus 18 (ARF18), was isolated from 9 of 95 populations of Heterodera glycines, the soybean cyst nematode, in Arkansas. In petri dishes, ARF18 parasitized 89% of H. glycines eggs in cysts. The fungus also infected eggs of Meloidogyne incognita and eggs in cysts of Cactodera betulae, H. graminophila, H. lespedezae, H. leuceilyma, H. schachtii, and H. trifolii. In pot tests, reproduction of SCN was 70% less in untreated field soil that was naturally infested by ARF18 than in autoclaved field soil. Although ARF18 grew well at 25 C on cornmeal agar over a wide pH range, it did not sporulate on 28 media and thus could not be identified to genus or species. PMID:19283127

Characteristics and Efficacy of a Sterile Hyphomycete (ARF18), a New Biocontrol Agent for Heterodera glycines and Other Nematodes.

Science.gov (United States)

Kim, D G; Riggs, R D

1991-07-01

A filamentous, nonsporulating fungus, designated Arkansas Fungus 18 (ARF18), was isolated from 9 of 95 populations of Heterodera glycines, the soybean cyst nematode, in Arkansas. In petri dishes, ARF18 parasitized 89% of H. glycines eggs in cysts. The fungus also infected eggs of Meloidogyne incognita and eggs in cysts of Cactodera betulae, H. graminophila, H. lespedezae, H. leuceilyma, H. schachtii, and H. trifolii. In pot tests, reproduction of SCN was 70% less in untreated field soil that was naturally infested by ARF18 than in autoclaved field soil. Although ARF18 grew well at 25 C on cornmeal agar over a wide pH range, it did not sporulate on 28 media and thus could not be identified to genus or species.

Transcription elongation factors are involved in programming hormone production in pituitary neuroendocrine GH4C1 cells.

Science.gov (United States)

Fujita, Toshitsugu; Piuz, Isabelle; Schlegel, Werner

2010-05-05

Transcription elongation of many eukaryotic genes is regulated. Two negative transcription elongation factors, 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) sensitivity-inducing factor (DSIF) and negative elongation factor (NELF) are known to stall collaboratively RNA polymerase II promoter proximally. We discovered that DSIF and NELF are linked to hormone expression in rat pituitary GH4C1 cells. When NELF-E, a subunit of NELF or Spt5, a subunit of DSIF was stably knocked-down, prolactin (PRL) expression was increased both at the mRNA and protein levels. In contrast, stable knock-down of only Spt5 abolished growth hormone (GH) expression. Transient NELF-E knock-down increased coincidentally PRL expression and enhanced transcription of a PRL-promoter reporter gene. However, no direct interaction of NELF with the PRL gene could be demonstrated by chromatin immuno-precipitation. Thus, NELF suppressed PRL promoter activity indirectly. In conclusion, transcription regulation by NELF and DSIF is continuously involved in the control of hormone production and may contribute to neuroendocrine cell differentiation. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

Melanoma cells revive an embryonic transcriptional network to dictate phenotypic heterogeneity.

Science.gov (United States)

Vandamme, Niels; Berx, Geert

2014-01-01

Compared to the overwhelming amount of literature describing how epithelial-to-mesenchymal transition (EMT)-inducing transcription factors orchestrate cellular plasticity in embryogenesis and epithelial cells, the functions of these factors in non-epithelial contexts, such as melanoma, are less clear. Melanoma is an aggressive tumor arising from melanocytes, endowed with unique features of cellular plasticity. The reversible phenotype-switching between differentiated and invasive phenotypes is increasingly appreciated as a mechanism accounting for heterogeneity in melanoma and is driven by oncogenic signaling and environmental cues. This phenotypic switch is coupled with an intriguing and somewhat counterintuitive signaling switch of EMT-inducing transcription factors. In contrast to carcinomas, different EMT-inducing transcription factors have antagonizing effects in melanoma. Balancing between these different EMT transcription factors is likely the key to successful metastatic spread of melanoma.

Ionizing radiation induced transcriptional changes in the developing mouse brain. Doctoral Thesis Prepared at SCK-CEN and Defended in 2006

International Nuclear Information System (INIS)

Verheyde, J.

2007-01-01

radiation exposure. The Trp53 null mutants at this stage exhibited a decreased expression profile for various Cyclins and Cyclin-dependent kinases, suggesting the induction of a Trp53 independent cell cycle arrest. The observed absence of high levels of apoptosis is in concordance with the described phenotype of Trp53 null mutant mice that show an increased risk for tumour formation. Next, we questioned if the developing brain is characterized by a regional dependent differences on radiation sensitivity. Therefore, additional transcriptional analyses of the different regions (hippocampus, pallidal neuroepithelium and cortex of control and irradiated brain embryos) of the ventral wild type brain at E15 were performed. In these three regions of the developing brain, the differential expression of Trp53inp1 and Ccng1 by in situ hybridization suggested that the ventral brain shows a specific regional dependent expression pattern. This was further evidenced by real time qPCR, by which it also became clear that the strongest radiation induced effect can be observed in the cerebral cortex. However, this experiment didn't allow making a distinction between neural cells in this experiment, and different cell types contribute to correct neural network formation. Detailed analysis of cultured and irradiated neural cell types indicated that the expression of the Trp53inp1/Hipk2 and Ccng1/Mdm2/P19Arf signalling pathways are activated in a cell-type dependent matter. From this data it appears that, depending on the cell type and the stage of differentiation, two possible mechanisms for the induction of apoptosis are activated. In one hand cell cycle arrest occurs via P21 (Cdkn1a) induction and to certain extends via P19ARF in the mitotic astrocytes, while in short-term neurons this mechanism seems to be preferentially induced by Cyclin G1. This arrest is then followed by the induction of the mitochondrial proapoptotic pathway. Interestingly, long term cultured neurons show also an

miRNA Enriched in Human Neuroblast Nuclei Bind the MAZ Transcription Factor and Their Precursors Contain the MAZ Consensus Motif.

Science.gov (United States)

Goldie, Belinda J; Fitzsimmons, Chantel; Weidenhofer, Judith; Atkins, Joshua R; Wang, Dan O; Cairns, Murray J

2017-01-01

While the cytoplasmic function of microRNA (miRNA) as post-transcriptional regulators of mRNA has been the subject of significant research effort, their activity in the nucleus is less well characterized. Here we use a human neuronal cell model to show that some mature miRNA are preferentially enriched in the nucleus. These molecules were predominantly primate-specific and contained a sequence motif with homology to the consensus MAZ transcription factor binding element. Precursor miRNA containing this motif were shown to have affinity for MAZ protein in nuclear extract. We then used Ago1/2 RIP-Seq to explore nuclear miRNA-associated mRNA targets. Interestingly, the genes for Ago2-associated transcripts were also significantly enriched with MAZ binding sites and neural function, whereas Ago1-transcripts were associated with general metabolic processes and localized with SC35 spliceosomes. These findings suggest the MAZ transcription factor is associated with miRNA in the nucleus and may influence the regulation of neuronal development through Ago2-associated miRNA induced silencing complexes. The MAZ transcription factor may therefore be important for organizing higher order integration of transcriptional and post-transcriptional processes in primate neurons.

Environmental contaminants and microRNA regulation: Transcription factors as regulators of toxicant-altered microRNA expression

Energy Technology Data Exchange (ETDEWEB)

Sollome, James; Martin, Elizabeth [Department of Environmental Science & Engineering, Gillings School of Global Public Health, University of North Carolina, Chapel Hill (United States); Sethupathy, Praveen [Department of Genetics, School of Medicine, University of North Carolina, Chapel Hill, NC (United States); Fry, Rebecca C., E-mail: rfry@unc.edu [Department of Environmental Science & Engineering, Gillings School of Global Public Health, University of North Carolina, Chapel Hill (United States); Curriculum in Toxicology, School of Medicine, University of North Carolina, Chapel Hill, NC (United States)

2016-12-01

MicroRNAs (miRNAs) regulate gene expression by binding mRNA and inhibiting translation and/or inducing degradation of the associated transcripts. Expression levels of miRNAs have been shown to be altered in response to environmental toxicants, thus impacting cellular function and influencing disease risk. Transcription factors (TFs) are known to be altered in response to environmental toxicants and play a critical role in the regulation of miRNA expression. To date, environmentally-responsive TFs that are important for regulating miRNAs remain understudied. In a state-of-the-art analysis, we utilized an in silico bioinformatic approach to characterize potential transcriptional regulators of environmentally-responsive miRNAs. Using the miRStart database, genomic sequences of promoter regions for all available human miRNAs (n = 847) were identified and promoter regions were defined as − 1000/+500 base pairs from the transcription start site. Subsequently, the promoter region sequences of environmentally-responsive miRNAs (n = 128) were analyzed using enrichment analysis to determine overrepresented TF binding sites (TFBS). While most (56/73) TFs differed across environmental contaminants, a set of 17 TFs was enriched for promoter binding among miRNAs responsive to numerous environmental contaminants. Of these, one TF was common to miRNAs altered by the majority of environmental contaminants, namely SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily A, member 3 (SMARCA3). These identified TFs represent candidate common transcriptional regulators of miRNAs perturbed by environmental toxicants. - Highlights: • Transcription factors that regulate environmentally-modulated miRNA expression are understudied • Transcription factor binding sites (TFBS) located within DNA promoter regions of miRNAs were identified. • Specific transcription factors may serve as master regulators of environmentally-mediated microRNA expression.

Environmental contaminants and microRNA regulation: Transcription factors as regulators of toxicant-altered microRNA expression

International Nuclear Information System (INIS)

Sollome, James; Martin, Elizabeth; Sethupathy, Praveen; Fry, Rebecca C.

2016-01-01

MicroRNAs (miRNAs) regulate gene expression by binding mRNA and inhibiting translation and/or inducing degradation of the associated transcripts. Expression levels of miRNAs have been shown to be altered in response to environmental toxicants, thus impacting cellular function and influencing disease risk. Transcription factors (TFs) are known to be altered in response to environmental toxicants and play a critical role in the regulation of miRNA expression. To date, environmentally-responsive TFs that are important for regulating miRNAs remain understudied. In a state-of-the-art analysis, we utilized an in silico bioinformatic approach to characterize potential transcriptional regulators of environmentally-responsive miRNAs. Using the miRStart database, genomic sequences of promoter regions for all available human miRNAs (n = 847) were identified and promoter regions were defined as − 1000/+500 base pairs from the transcription start site. Subsequently, the promoter region sequences of environmentally-responsive miRNAs (n = 128) were analyzed using enrichment analysis to determine overrepresented TF binding sites (TFBS). While most (56/73) TFs differed across environmental contaminants, a set of 17 TFs was enriched for promoter binding among miRNAs responsive to numerous environmental contaminants. Of these, one TF was common to miRNAs altered by the majority of environmental contaminants, namely SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily A, member 3 (SMARCA3). These identified TFs represent candidate common transcriptional regulators of miRNAs perturbed by environmental toxicants. - Highlights: • Transcription factors that regulate environmentally-modulated miRNA expression are understudied • Transcription factor binding sites (TFBS) located within DNA promoter regions of miRNAs were identified. • Specific transcription factors may serve as master regulators of environmentally-mediated microRNA expression

A Transcriptional Regulatory Network Containing Nuclear Receptors and Long Noncoding RNAs Controls Basal and Drug-Induced Expression of Cytochrome P450s in HepaRG Cells.

Science.gov (United States)

Chen, Liming; Bao, Yifan; Piekos, Stephanie C; Zhu, Kexin; Zhang, Lirong; Zhong, Xiao-Bo

2018-07-01

Cytochrome P450 (P450) enzymes are responsible for metabolizing drugs. Expression of P450s can directly affect drug metabolism, resulting in various outcomes in therapeutic efficacy and adverse effects. Several nuclear receptors are transcription factors that can regulate expression of P450s at both basal and drug-induced levels. Some long noncoding RNAs (lncRNAs) near a transcription factor are found to participate in the regulatory functions of the transcription factors. The aim of this study is to determine whether there is a transcriptional regulatory network containing nuclear receptors and lncRNAs controlling both basal and drug-induced expression of P450s in HepaRG cells. Small interfering RNAs or small hairpin RNAs were applied to knock down four nuclear receptors [hepatocyte nuclear factor 1 α (HNF1 α ), hepatocyte nuclear factor 4 α (HNF4 α ), pregnane X receptor (PXR), and constitutive androstane receptor (CAR)] as well as two lncRNAs [HNF1 α antisense RNA 1 (HNF1 α -AS1) and HNF4 α antisense RNA 1 (HNF4 α -AS1)] in HepaRG cells with or without treatment of phenobarbital or rifampicin. Expression of eight P450 enzymes was examined in both basal and drug-induced levels. CAR and PXR mainly regulated expression of specific P450s. HNF1 α and HNF4 α affected expression of a wide range of P450s as well as other transcription factors. HNF1 α and HNF4 α controlled the expression of their neighborhood lncRNAs, HNF1 α -AS1 and HNF4 α -AS1, respectively. HNF1 α -AS1 and HNF4 α -AS1 was also involved in the regulation of P450s and transcription factors in diverse manners. Altogether, our study concludes that a transcription regulatory network containing the nuclear receptors and lncRNAs controls both basal and drug-induced expression of P450s in HepaRG cells. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

TrSDB: a proteome database of transcription factors

Science.gov (United States)

Hermoso, Antoni; Aguilar, Daniel; Aviles, Francesc X.; Querol, Enrique

2004-01-01

TrSDB—TranScout Database—(http://ibb.uab.es/trsdb) is a proteome database of eukaryotic transcription factors based upon predicted motifs by TranScout and data sources such as InterPro and Gene Ontology Annotation. Nine eukaryotic proteomes are included in the current version. Extensive and diverse information for each database entry, different analyses considering TranScout classification and similarity relationships are offered for research on transcription factors or gene expression. PMID:14681387

Using TESS to predict transcription factor binding sites in DNA sequence.

Science.gov (United States)

Schug, Jonathan

2008-03-01

This unit describes how to use the Transcription Element Search System (TESS). This Web site predicts transcription factor binding sites (TFBS) in DNA sequence using two different kinds of models of sites, strings and positional weight matrices. The binding of transcription factors to DNA is a major part of the control of gene expression. Transcription factors exhibit sequence-specific binding; they form stronger bonds to some DNA sequences than to others. Identification of a good binding site in the promoter for a gene suggests the possibility that the corresponding factor may play a role in the regulation of that gene. However, the sequences transcription factors recognize are typically short and allow for some amount of mismatch. Because of this, binding sites for a factor can typically be found at random every few hundred to a thousand base pairs. TESS has features to help sort through and evaluate the significance of predicted sites.

Microarray-Based Identification of Transcription Factor Target Genes

NARCIS (Netherlands)

Gorte, M.; Horstman, A.; Page, R.B.; Heidstra, R.; Stromberg, A.; Boutilier, K.A.

2011-01-01

Microarray analysis is widely used to identify transcriptional changes associated with genetic perturbation or signaling events. Here we describe its application in the identification of plant transcription factor target genes with emphasis on the design of suitable DNA constructs for controlling TF

STAT3 induces transcription of the DNA methyltransferase 1 gene (DNMT1) in malignant T lymphocytes

DEFF Research Database (Denmark)

Zhang, Qian; Wang, Hong Y; Woetmann, Anders

2006-01-01

In this study, we demonstrated that STAT3, a well-characterized transcription factor expressed in continuously activated oncogenic form in the large spectrum of cancer types, induces in malignant T lymphocytes the expression of DNMT1, the key effector of epigenetic gene silencing. STAT3 binds in ...

Anthocyanin biosynthesis in pears is regulated by a R2R3-MYB transcription factor PyMYB10.

Science.gov (United States)

Feng, Shouqian; Wang, Yanling; Yang, Song; Xu, Yuting; Chen, Xuesen

2010-06-01

Skin color is an important factor in pear breeding programs. The degree of red coloration is determined by the content and composition of anthocyanins. In plants, many MYB transcriptional factors are involved in regulating anthocyanin biosynthesis. In this study, a R2R3-MYB transcription factor gene, PyMYB10, was isolated from Asian pear (Pyrus pyrifolia) cv. 'Aoguan'. Sequence analysis suggested that the PyMYB10 gene was an ortholog of MdMYB10 gene, which regulates anthocyanin biosynthesis in red fleshed apple (Malus x domestica) cv. 'Red Field'. PyMYB10 was identified at the genomic level and had three exons, with its upstream sequence containing core sequences of cis-acting regulatory elements involved in light responsiveness. Fruit bagging showed that light could induce expression of PyMYB10 and anthocyanin biosynthesis. Quantitative real-time PCR revealed that PyMYB10 was predominantly expressed in pear skins, buds, and young leaves, and the level of transcription in buds was higher than in skin and young leaves. In ripening fruits, the transcription of PyMYB10 in the skin was positively correlated with genes in the anthocyanin pathway and with anthocyanin biosynthesis. In addition, the transcription of PyMYB10 and genes of anthocyanin biosynthesis were more abundant in red-skinned pear cultivars compared to blushed cultivars. Transgenic Arabidopsis plants overexpressing PyMYB10 exhibited ectopic pigmentation in immature seeds. The study suggested that PyMYB10 plays a role in regulating anthocyanin biosynthesis and the overexpression of PyMYB10 was sufficient to induce anthocyanin accumulation.

Exploring the utility of organo-polyoxometalate hybrids to inhibit SOX transcription factors.

Science.gov (United States)

Narasimhan, Kamesh; Micoine, Kevin; Lacôte, Emmanuel; Thorimbert, Serge; Cheung, Edwin; Hasenknopf, Bernold; Jauch, Ralf

2014-01-01

SOX transcription factors constitute an attractive target class for intervention with small molecules as they play a prominent role in the field of regenerative biomedicine and cancer biology. However, rationally engineering specific inhibitors that interfere with transcription factor DNA interfaces continues to be a monumental challenge in the field of transcription factor chemical biology. Polyoxometalates (POMs) are inorganic compounds that were previously shown to target the high-mobility group (HMG) of SOX proteins at nanomolar concentrations. In continuation of this work, we carried out an assessment of the selectivity of a panel of newly synthesized organo-polyoxometalate hybrids in targeting different transcription factor families to enable the usage of polyoxometalates as specific SOX transcription factor drugs. The residual DNA-binding activities of 15 different transcription factors were measured after treatment with a panel of diverse polyoxometalates. Polyoxometalates belonging to the Dawson structural class were found to be more potent inhibitors than the Keggin class. Further, organically modified Dawson polyoxometalates were found to be the most potent in inhibiting transcription factor DNA binding activity. The size of the polyoxometalates and its derivitization were found to be the key determinants of their potency. Polyoxometalates are highly potent, nanomolar range inhibitors of the DNA binding activity of the Sox-HMG family. However, binding assays involving a limited subset of structurally diverse polyoxometalates revealed a low selectivity profile against different transcription factor families. Further progress in achieving selectivity and deciphering structure-activity relationship of POMs require the identification of POM binding sites on transcription factors using elaborate approaches like X-ray crystallography and multidimensional NMR. In summary, our report reaffirms that transcription factors are challenging molecular architectures

TcoF-DB v2: update of the database of human and mouse transcription co-factors and transcription factor interactions

KAUST Repository

Schmeier, Sebastian; Alam, Tanvir; Essack, Magbubah; Bajic, Vladimir B.

2016-01-01

Transcription factors (TFs) play a pivotal role in transcriptional regulation, making them crucial for cell survival and important biological functions. For the regulation of transcription, interactions of different regulatory proteins known as transcription co-factors (TcoFs) and TFs are essential in forming necessary protein complexes. Although TcoFs themselves do not bind DNA directly, their influence on transcriptional regulation and initiation, although indirect, has been shown to be significant, with the functionality of TFs strongly influenced by the presence of TcoFs. In the TcoF-DB v2 database, we collect information on TcoFs. In this article, we describe updates and improvements implemented in TcoF-DB v2. TcoF-DB v2 provides several new features that enables exploration of the roles of TcoFs. The content of the database has significantly expanded, and is enriched with information from Gene Ontology, biological pathways, diseases and molecular signatures. TcoF-DB v2 now includes many more TFs; has substantially increased the number of human TcoFs to 958, and now includes information on mouse (418 new TcoFs). TcoF-DB v2 enables the exploration of information on TcoFs and allows investigations into their influence on transcriptional regulation in humans and mice. TcoF-DB v2 can be accessed at http://tcofdb.org/.

TcoF-DB v2: update of the database of human and mouse transcription co-factors and transcription factor interactions

KAUST Repository

Schmeier, Sebastian

2016-10-17

Transcription factors (TFs) play a pivotal role in transcriptional regulation, making them crucial for cell survival and important biological functions. For the regulation of transcription, interactions of different regulatory proteins known as transcription co-factors (TcoFs) and TFs are essential in forming necessary protein complexes. Although TcoFs themselves do not bind DNA directly, their influence on transcriptional regulation and initiation, although indirect, has been shown to be significant, with the functionality of TFs strongly influenced by the presence of TcoFs. In the TcoF-DB v2 database, we collect information on TcoFs. In this article, we describe updates and improvements implemented in TcoF-DB v2. TcoF-DB v2 provides several new features that enables exploration of the roles of TcoFs. The content of the database has significantly expanded, and is enriched with information from Gene Ontology, biological pathways, diseases and molecular signatures. TcoF-DB v2 now includes many more TFs; has substantially increased the number of human TcoFs to 958, and now includes information on mouse (418 new TcoFs). TcoF-DB v2 enables the exploration of information on TcoFs and allows investigations into their influence on transcriptional regulation in humans and mice. TcoF-DB v2 can be accessed at http://tcofdb.org/.

The cellular transcription factor CREB corresponds to activating transcription factor 47 (ATF-47) and forms complexes with a group of polypeptides related to ATF-43.

Science.gov (United States)

Hurst, H C; Masson, N; Jones, N C; Lee, K A

1990-12-01

Promoter elements containing the sequence motif CGTCA are important for a variety of inducible responses at the transcriptional level. Multiple cellular factors specifically bind to these elements and are encoded by a multigene family. Among these factors, polypeptides termed activating transcription factor 43 (ATF-43) and ATF-47 have been purified from HeLa cells and a factor referred to as cyclic AMP response element-binding protein (CREB) has been isolated from PC12 cells and rat brain. We demonstrated that CREB and ATF-47 are identical and that CREB and ATF-43 form protein-protein complexes. We also found that the cis requirements for stable DNA binding by ATF-43 and CREB are different. Using antibodies to ATF-43 we have identified a group of polypeptides (ATF-43) in the size range from 40 to 43 kDa. ATF-43 polypeptides are related by their reactivity with anti-ATF-43, DNA-binding specificity, complex formation with CREB, heat stability, and phosphorylation by protein kinase A. Certain cell types vary in their ATF-43 complement, suggesting that CREB activity is modulated in a cell-type-specific manner through interaction with ATF-43. ATF-43 polypeptides do not appear simply to correspond to the gene products of the ATF multigene family, suggesting that the size of the ATF family at the protein level is even larger than predicted from cDNA-cloning studies.

The hematopoietic transcription factor PU.1 regulates RANK gene expression in myeloid progenitors

International Nuclear Information System (INIS)

Kwon, Oh Hyung; Lee, Chong-Kil; Lee, Young Ik; Paik, Sang-Gi; Lee, Hyun-Jun

2005-01-01

Osteoclasts are bone resorbing cells of hematopoietic origin. The hematopoietic transcription factor PU.1 is critical for osteoclastogenesis; however, the molecular mechanisms of PU.1-regulated osteoclastogenesis have not been explored. Here, we present evidence that the receptor activator of nuclear factor κB (RANK) gene that has been shown to be crucial for osteoclastogenesis is a transcriptional target of PU.1. The PU.1 -/- progenitor cells failed to express the RANK gene and reconstitution of PU.1 in these cells induced RANK expression. Treatment of the PU.1 reconstituted cells with M-CSF and RANKL further augmented the RANK gene expression. To explore the regulatory mechanism of the RANK gene expression by PU.1, we have cloned the human RANK promoter. Transient transfection assays have revealed that the 2.2-kb RANK promoter was functional in a monocyte line RAW264.7, whereas co-transfection of PU.1 transactivated the RANK promoter in HeLa cells. Taken together, these results suggest that PU.1 regulates the RANK gene transcription and this may represent one of the key roles of PU.1 in osteoclast differentiation

Analysis of the ARF/p53 Pathway During Oncogenic Stimulation

National Research Council Canada - National Science Library

Nahle, Zaher

2003-01-01

... or deficient for the ARF and/or p53 genes. We found that the ElA oncoprotein regulates the expression of a myriad of targets involved in a diversity of functions such as apoptosis, cell cycle progression, checkpoint control, DNA replication...

Stress-inducible expression of AtDREB1A transcription factor greatly improves drought stress tolerance in transgenic indica rice.

Science.gov (United States)

Ravikumar, G; Manimaran, P; Voleti, S R; Subrahmanyam, D; Sundaram, R M; Bansal, K C; Viraktamath, B C; Balachandran, S M

2014-06-01

The cultivation of rice (Oryza sativa L.), a major food crop, requires ample water (30 % of the fresh water available worldwide), and its productivity is greatly affected by drought, the most significant environmental factor. Much research has focussed on identifying quantitative trait loci, stress-regulated genes and transcription factors that will contribute towards the development of climate-resilient/tolerant crop plants in general and rice in particular. The transcription factor DREB1A, identified from the model plant Arabidopsis thaliana, has been reported to enhance stress tolerance against drought stress. We developed transgenic rice plants with AtDREB1A in the background of indica rice cultivar Samba Mahsuri through Agrobacterium-mediated transformation. The AtDREB1A gene was stably inherited and expressed in T1 and T2 plants and in subsequent generations, as indicated by the results of PCR, Southern blot and RT-PCR analyses. Expression of AtDREB1A was induced by drought stress in transgenic rice lines, which were highly tolerant to severe water deficit stress in both the vegetative and reproductive stages without affecting their morphological or agronomic traits. The physiological studies revealed that the expression of AtDREB1A was associated with an increased accumulation of the osmotic substance proline, maintenance of chlorophyll, increased relative water content and decreased ion leakage under drought stress. Most of the homozygous lines were highly tolerant to drought stress and showed significantly a higher grain yield and spikelet fertility relative to the nontransgenic control plants under both stressed and unstressed conditions. The improvement in drought stress tolerance in combination with agronomic traits is very essential in high premium indica rice cultivars, such as Samba Mahsuri, so that farmers can benefit in times of seasonal droughts and water scarcity.

Demonstrating Interactions of Transcription Factors with DNA by Electrophoretic Mobility Shift Assay.

Science.gov (United States)

Yousaf, Nasim; Gould, David

2017-01-01

Confirming the binding of a transcription factor with a particular DNA sequence may be important in characterizing interactions with a synthetic promoter. Electrophoretic mobility shift assay is a powerful approach to demonstrate the specific DNA sequence that is bound by a transcription factor and also to confirm the specific transcription factor involved in the interaction. In this chapter we describe a method we have successfully used to demonstrate interactions of endogenous transcription factors with sequences derived from endogenous and synthetic promoters.

Characterization of a novel radiation-inducible transcript, uscA, and analysis of its transcriptional regulation

Energy Technology Data Exchange (ETDEWEB)

Lim, Sang Yong; Kim, Dong Ho; Joe, Min Ho

2010-03-15

The transcriptional expression of the uscA promote (P{sub uscA}) only occurred under aerobic conditions and a dose of 2Gy maximally activated transcription of P{sub uscA}. However, various environmental stress including physical shocks (pH, temperature, osmotic shock), DNA damaging agents (UV and MMC) or oxidative stressagents (paraquat, menadione, and H{sub 2}O{sub 2}) didn't cause the transcriptional activationof P{sub uscA}. The transcription of uscA was initiated at 170 bp upstream of the cyoA start codon, and ended around the ampG stop codon. The size of uscA was determined through reverse transcription assay, approximately 250 bp. The deletion analysis of uscA promoter demonstrates that radiation inducibility of P{sub uscA} is mediated by sequences present between -20 and +111 relativeto +1 of P{sub uscA} and radiation causes P{sub uscA} activation thorough permitting the expression that is repressed under non-irradiated conditions

Characterization of a novel radiation-inducible transcript, uscA, and analysis of its transcriptional regulation

International Nuclear Information System (INIS)

Lim, Sang Yong; Kim, Dong Ho; Joe, Min Ho

2010-03-01

The transcriptional expression of the uscA promote (P uscA ) only occurred under aerobic conditions and a dose of 2Gy maximally activated transcription of P uscA . However, various environmental stress including physical shocks (pH, temperature, osmotic shock), DNA damaging agents (UV and MMC) or oxidative stressagents (paraquat, menadione, and H 2 O 2 ) didn't cause the transcriptional activationof P uscA . The transcription of uscA was initiated at 170 bp upstream of the cyoA start codon, and ended around the ampG stop codon. The size of uscA was determined through reverse transcription assay, approximately 250 bp. The deletion analysis of uscA promoter demonstrates that radiation inducibility of P uscA is mediated by sequences present between -20 and +111 relativeto +1 of P uscA and radiation causes P uscA activation thorough permitting the expression that is repressed under non-irradiated conditions

Expression of HSF2 decreases in mitosis to enable stress-inducible transcription and cell survival

Science.gov (United States)

Elsing, Alexandra N.; Aspelin, Camilla; Björk, Johanna K.; Bergman, Heidi A.; Himanen, Samu V.; Kallio, Marko J.; Roos-Mattjus, Pia

2014-01-01

Unless mitigated, external and physiological stresses are detrimental for cells, especially in mitosis, resulting in chromosomal missegregation, aneuploidy, or apoptosis. Heat shock proteins (Hsps) maintain protein homeostasis and promote cell survival. Hsps are transcriptionally regulated by heat shock factors (HSFs). Of these, HSF1 is the master regulator and HSF2 modulates Hsp expression by interacting with HSF1. Due to global inhibition of transcription in mitosis, including HSF1-mediated expression of Hsps, mitotic cells are highly vulnerable to stress. Here, we show that cells can counteract transcriptional silencing and protect themselves against proteotoxicity in mitosis. We found that the condensed chromatin of HSF2-deficient cells is accessible for HSF1 and RNA polymerase II, allowing stress-inducible Hsp expression. Consequently, HSF2-deficient cells exposed to acute stress display diminished mitotic errors and have a survival advantage. We also show that HSF2 expression declines during mitosis in several but not all human cell lines, which corresponds to the Hsp70 induction and protection against stress-induced mitotic abnormalities and apoptosis. PMID:25202032

Endocytosis of hERG Is Clathrin-Independent and Involves Arf6

Science.gov (United States)

Abuarab, Nada; Smith, Andrew J.; Hardy, Matthew E. L.; Elliott, David J. S.; Sivaprasadarao, Asipu

2013-01-01

The hERG potassium channel is critical for repolarisation of the cardiac action potential. Reduced expression of hERG at the plasma membrane, whether caused by hereditary mutations or drugs, results in long QT syndrome and increases the risk of ventricular arrhythmias. Thus, it is of fundamental importance to understand how the density of this channel at the plasma membrane is regulated. We used antibodies to an extracellular native or engineered epitope, in conjunction with immunofluorescence and ELISA, to investigate the mechanism of hERG endocytosis in recombinant cells and validated the findings in rat neonatal cardiac myocytes. The data reveal that this channel undergoes rapid internalisation, which is inhibited by neither dynasore, an inhibitor of dynamin, nor a dominant negative construct of Rab5a, into endosomes that are largely devoid of the transferrin receptor. These results support a clathrin-independent mechanism of endocytosis and exclude involvement of dynamin-dependent caveolin and RhoA mechanisms. In agreement, internalised hERG displayed marked overlap with glycosylphosphatidylinositol-anchored GFP, a clathrin-independent cargo. Endocytosis was significantly affected by cholesterol extraction with methyl-β-cyclodextrin and inhibition of Arf6 function with dominant negative Arf6-T27N-eGFP. Taken together, we conclude that hERG undergoes clathrin-independent endocytosis via a mechanism involving Arf6. PMID:24392021

Intragenic suppressor of Osiaa23 revealed a conserved tryptophan residue crucial for protein-protein interactions.

Directory of Open Access Journals (Sweden)

Jun Ni

Full Text Available The Auxin/Indole-3-Acetic Acid (Aux/IAA and Auxin Response Factor (ARF are two important families that play key roles in auxin signal transduction. Both of the families contain a similar carboxyl-terminal domain (Domain III/IV that facilitates interactions between these two families. In spite of the importance of protein-protein interactions among these transcription factors, the mechanisms involved in these interactions are largely unknown. In this study, we isolated six intragenic suppressors of an auxin insensitive mutant, Osiaa23. Among these suppressors, Osiaa23-R5 successfully rescued all the defects of the mutant. Sequence analysis revealed that an amino acid substitution occurred in the Tryptophan (W residue in Domain IV of Osiaa23. Yeast two-hybrid experiments showed that the mutation in Domain IV prevents the protein-protein interactions between Osiaa23 and OsARFs. Phylogenetic analysis revealed that the W residue is conserved in both OsIAAs and OsARFs. Next, we performed site-specific amino acid substitutions within Domain IV of OsARFs, and the conserved W in Domain IV was exchanged by Serine (S. The mutated OsARF(WSs can be released from the inhibition of Osiaa23 and maintain the transcriptional activities. Expression of OsARF(WSs in Osiaa23 mutant rescued different defects of the mutant. Our results suggest a previously unknown importance of Domain IV in both families and provide an indirect way to investigate functions of OsARFs.

A putative functional MYB transcription factor induced by low temperature regulates anthocyanin biosynthesis in purple kale (Brassica Oleracea var. acephala f. tricolor).