Interaction of Yna1 and Yna2 Is Required for Nuclear Accumulation and Transcriptional Activation of the Nitrate Assimilation Pathway in the Yeast Hansenula polymorpha

The genes encoding the nitrate transporter (YNT1), nitrite reductase (YNI1) and nitrate reductase (YNR1) are clustered in the yeast Hansenula polymorpha. In addition, DNA sequencing of the region containing these genes demonstrated that a new open reading frame called YNA1 (yeast nitrate assimilation) was located between YNR1 and YNI1. The YNA1 gene encodes a protein of 529 residues belonging to the family of Zn(II)2Cys6 fungal transcriptional factors, and has the highest similarity to the transcriptional factors encoded by nirA, and to a smaller extent to nit-4, involved in the nitrate induction of the gene involved in the assimilation of this compound in filamentous fungi. Northern blot analysis showed the presence of the YNA1 transcript in cells incubated in nitrate, nitrate plus ammonium, ammonium, and nitrogen-free media, with a decrease in its levels in those cells incubated in ammonium. In nitrate the strain Δyna1::URA3, with a disrupted YNA1 gene, neither grew nor expressed the genes YNT1, YNI1 and YNR1. In the gene cluster YNT1-YNI1-YNA1-YNR1, the four genes were transcribed independently in the YNT1 → YNR1 direction and the transcription start sites were determined by primer extension.

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Chromosome Y pericentric heterochromatin is a primary target of HSF1 in male cells

Chromosoma ◽

10.1007/s00412-021-00751-2 ◽

2021 ◽

Vol 130 (1) ◽

pp. 53-60

Author(s):

Jessica Penin ◽

Solenne Dufour ◽

Virginie Faure ◽

Sabrina Fritah ◽

Daphné Seigneurin-Berny ◽

...

Keyword(s):

Heat Shock ◽

Transcriptional Activation ◽

Human Fibroblasts ◽

Pericentric Heterochromatin ◽

Chromosome 9 ◽

Nuclear Accumulation ◽

Critical Element ◽

Heat Shock Factor 1 ◽

Primary Target ◽

Chromosome Y

AbstractThe heat shock factor 1 (HSF1)-dependent transcriptional activation of human pericentric heterochromatin in heat-shocked cells is the most striking example of transcriptional activation of heterochromatin. Until now, pericentric heterochromatin of chromosome 9 has been identified as the primary target of HSF1, in both normal and tumor heat-shocked cells. Transcriptional awakening of this large genomic region results in the nuclear accumulation of satellite III (SATIII) noncoding RNAs (ncRNAs) and the formation in cis of specific structures known as nuclear stress bodies (nSBs). Here, we show that, in four different male cell lines, including primary human fibroblasts and amniocytes, pericentric heterochromatin of chromosome Y can also serve as a unique primary site of HSF1-dependent heterochromatin transcriptional activation, production of SATIII ncRNA, and nucleation of nuclear stress bodies (nSBs) upon heat shock. Our observation suggests that the chromosomal origin of SATIII transcripts in cells submitted to heat shock is not a determinant factor as such, but that transcription of SATIII repetitive units or the SATIII ncRNA molecules is the critical element of HSF1-dependent transcription activation of constitutive heterochromatin.

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The Ebola Virus VP35 Protein Inhibits Activation of Interferon Regulatory Factor 3

Journal of Virology ◽

10.1128/jvi.77.14.7945-7956.2003 ◽

2003 ◽

Vol 77 (14) ◽

pp. 7945-7956 ◽

Cited By ~ 332

Author(s):

Christopher F. Basler ◽

Andrea Mikulasova ◽

Luis Martinez-Sobrido ◽

Jason Paragas ◽

Elke Mühlberger ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Transcriptional Activation ◽

Ebola Virus ◽

Nuclear Translocation ◽

Sendai Virus ◽

Vero Cells ◽

Nuclear Accumulation ◽

Regulatory Factor ◽

Cellular Transcription Factor

ABSTRACT The Ebola virus VP35 protein was previously found to act as an interferon (IFN) antagonist which could complement growth of influenza delNS1 virus, a mutant influenza virus lacking the influenza virus IFN antagonist protein, NS1. The Ebola virus VP35 could also prevent the virus- or double-stranded RNA-mediated transcriptional activation of both the beta IFN (IFN-β) promoter and the IFN-stimulated ISG54 promoter (C. Basler et al., Proc. Natl. Acad. Sci. USA 97:12289-12294, 2000). We now show that VP35 inhibits virus infection-induced transcriptional activation of IFN regulatory factor 3 (IRF-3)-responsive mammalian promoters and that VP35 does not block signaling from the IFN-α/β receptor. The ability of VP35 to inhibit this virus-induced transcription correlates with its ability to block activation of IRF-3, a cellular transcription factor of central importance in initiating the host cell IFN response. We demonstrate that VP35 blocks the Sendai virus-induced activation of two promoters which can be directly activated by IRF-3, namely, the ISG54 promoter and the ISG56 promoter. Further, expression of VP35 prevents the IRF-3-dependent activation of the IFN-α4 promoter in response to viral infection. The inhibition of IRF-3 appears to occur through an inhibition of IRF-3 phosphorylation. VP35 blocks virus-induced IRF-3 phosphorylation and subsequent IRF-3 dimerization and nuclear translocation. Consistent with these observations, Ebola virus infection of Vero cells activated neither transcription from the ISG54 promoter nor nuclear accumulation of IRF-3. These data suggest that in Ebola virus-infected cells, VP35 inhibits the induction of antiviral genes, including the IFN-β gene, by blocking IRF-3 activation.

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RSPO2 gene rearrangement: a powerful driver of β-catenin activation in liver tumours

Gut ◽

10.1136/gutjnl-2018-317632 ◽

2019 ◽

Vol 68 (7) ◽

pp. 1287-1296 ◽

Cited By ~ 9

Author(s):

Thomas Longerich ◽

Volker Endris ◽

Olaf Neumann ◽

Eugen Rempel ◽

Martina Kirchner ◽

...

Keyword(s):

Malignant Transformation ◽

Transcriptional Activation ◽

Gene Rearrangement ◽

Target Genes ◽

Integration Site ◽

Genetic Alterations ◽

Wnt Signalling ◽

Nuclear Accumulation ◽

Alternative Mechanism ◽

Mutational Status

ObjectiveWe aimed at the identification of genetic alterations that may functionally substitute for CTNNB1 mutation in ß-catenin-activated hepatocellular adenomas (HCAs) and hepatocellular carcinoma (HCC).DesignLarge cohorts of HCA (n=185) and HCC (n=468) were classified using immunohistochemistry. The mutational status of the CTNNB1 gene was determined in ß-catenin-activated HCA (b-HCA) and HCC with at least moderate nuclear CTNNB1 accumulation. Ultra-deep sequencing was used to characterise CTNNB1wild-type and ß-catenin-activated HCA and HCC. Expression profiling of HCA subtypes was performed.ResultsA roof plate-specific spondin 2 (RSPO2) gene rearrangement resulting from a 46.4 kb microdeletion on chromosome 8q23.1 was detected as a new morphomolecular driver of β-catenin-activated HCA. RSPO2 fusion positive HCA displayed upregulation of RSPO2 protein, nuclear accumulation of β-catenin and transcriptional activation of β-catenin-target genes indicating activation of Wingless-Type MMTV Integration Site Family (WNT) signalling. Architectural and cytological atypia as well as interstitial invasion indicated malignant transformation in one of the RSPO2 rearranged b-HCAs. The RSPO2 gene rearrangement was also observed in three β-catenin-activated HCCs developing in context of chronic liver disease. Mutations of the human telomerase reverse transcriptase promoter—known to drive malignant transformation of CTNNB1-mutated HCA—seem to be dispensable for RSPO2 rearranged HCA and HCC.ConclusionThe RSPO2 gene rearrangement leads to oncogenic activation of the WNT signalling pathway in HCA and HCC, represents an alternative mechanism for the development of b-HCA and may drive malignant transformation without additional TERT promoter mutation.

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Modulating PKCα Activity to Target Wnt/β-Catenin Signaling in Colon Cancer

Cancers ◽

10.3390/cancers11050693 ◽

2019 ◽

Vol 11 (5) ◽

pp. 693 ◽

Cited By ~ 5

Author(s):

Sébastien Dupasquier ◽

Philippe Blache ◽

Laurence Picque Lasorsa ◽

Han Zhao ◽

Jean-Daniel Abraham ◽

...

Keyword(s):

Transcriptional Activation ◽

Transcriptional Activity ◽

Target Genes ◽

Tumor Development ◽

Orphan Receptor ◽

Nuclear Accumulation ◽

Kinase C ◽

Extracellular Signal

Inactivating mutations of the tumor suppressor Adenomatosis Polyposis Coli (APC), which are found in familial adenomatosis polyposis and in 80% of sporadic colorectal cancers (CRC), result in constitutive activation of the Wnt/β-catenin pathway and tumor development in the intestine. These mutations disconnect the Wnt/β-catenin pathway from its Wnt extracellular signal by inactivating the APC/GSK3-β/axin destruction complex of β-catenin. This results in sustained nuclear accumulation of β-catenin, followed by β-catenin-dependent co-transcriptional activation of Wnt/β-catenin target genes. Thus, mechanisms acting downstream of APC, such as those controlling β-catenin stability and/or co-transcriptional activity, are attractive targets for CRC treatment. Protein Kinase C-α (PKCα) phosphorylates the orphan receptor RORα that then inhibits β-catenin co-transcriptional activity. PKCα also phosphorylates β-catenin, leading to its degradation by the proteasome. Here, using both in vitro (DLD-1 cells) and in vivo (C57BL/6J mice) PKCα knock-in models, we investigated whether enhancing PKCα function could be beneficial in CRC treatment. We found that PKCα is infrequently mutated in CRC samples, and that inducing PKCα function is not deleterious for the normal intestinal epithelium. Conversely, di-terpene ester-induced PKCα activity triggers CRC cell death. Together, these data indicate that PKCα is a relevant drug target for CRC treatment.

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Acetylation by PCAF Enhances CIITA Nuclear Accumulation and Transactivation of Major Histocompatibility Complex Class II Genes

Molecular and Cellular Biology ◽

10.1128/mcb.20.22.8489-8498.2000 ◽

2000 ◽

Vol 20 (22) ◽

pp. 8489-8498 ◽

Cited By ~ 103

Author(s):

Charalambos Spilianakis ◽

Joseph Papamatheakis ◽

Androniki Kretsovali

Keyword(s):

Major Histocompatibility Complex ◽

Major Histocompatibility Complex Class ◽

Transcriptional Activation ◽

Trichostatin A ◽

Class Ii ◽

Nuclear Accumulation ◽

Complex Class ◽

Major Histocompatibility ◽

Histocompatibility Complex

ABSTRACT The class II transactivator (CIITA), the master regulator of the tissue-specific and interferon gamma-inducible expression of major histocompatibility complex class II genes, synergizes with the histone acetylase coactivator CBP to activate gene transcription. Here we demonstrate that in addition to CBP, PCAF binds to CIITA both in vivo and in vitro and enhances CIITA-dependent transcriptional activation of class II promoters. Accordingly, E1A mutants defective for PCAF or CBP interaction show reduced ability in suppressing CIITA activity. Interestingly, CBP and PCAF acetylate CIITA at lysine residues within a nuclear localization signal. We show that CIITA is shuttling between the nucleus and cytoplasm. The shuttling behavior and activity of the protein are regulated by acetylation: overexpression of PCAF or inhibition of cellular deacetylases by trichostatin A increases the nuclear accumulation of CIITA in a manner determined by the presence of the acetylation target lysines. Furthermore, mutagenesis of the acetylated residues reduces the transactivation ability of CIITA. These results support a novel function for acetylation, i.e., to regulate gene expression by stimulating the nuclear accumulation of an activator.

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MondoA, a Novel Basic Helix-Loop-Helix–Leucine Zipper Transcriptional Activator That Constitutes a Positive Branch of a Max-Like Network

Molecular and Cellular Biology ◽

10.1128/mcb.20.23.8845-8854.2000 ◽

2000 ◽

Vol 20 (23) ◽

pp. 8845-8854 ◽

Cited By ~ 77

Author(s):

Andrew N. Billin ◽

Alanna L. Eilers ◽

Kathryn L. Coulter ◽

Jennifer S. Logan ◽

Donald E. Ayer

Keyword(s):

Transcriptional Activation ◽

Nuclear Export ◽

Mammalian Cells ◽

Leucine Zipper ◽

Functional Characterization ◽

Nuclear Accumulation ◽

Cytoplasmic Localization ◽

Basic Helix Loop Helix ◽

Helix Loop Helix

ABSTRACT Max is a common dimerization partner for a family of transcription factors (Myc, Mad [or Mxi]), and Mnt [or Rox] proteins) that regulate cell growth, proliferation, and apoptosis. We recently characterized a novel Max-like protein, Mlx, which interacts with Mad1 and Mad4. Here we describe the cloning and functional characterization of a new family of basic helix-loop-helix–leucine zipper heterodimeric partners for Mlx termed the Mondo family. MondoA forms homodimers weakly and does not interact with Max or members of the Myc or Mad families. MondoA and Mlx associate in vivo, and surprisingly, they are localized primarily to the cytoplasm of cultured mammalian cells. Treatment of cells with the nuclear export inhibitor leptomycin B results in the nuclear accumulation of MondoA and Mlx, demonstrating that they shuttle between the cytoplasmic and nuclear compartments rather than having exclusively cytoplasmic localization. MondoA preferentially forms heterodimers with Mlx, and this heterocomplex can bind to, and activate transcription from, CACGTG E-boxes when targeted to the nucleus via a heterologous nuclear localization signal. The amino termini of the Mondo proteins are highly conserved among family members and contain separable and autonomous cytoplasmic localization and transcription activation domains. Therefore, Mlx can mediate transcriptional repression in conjunction with the Mad family and can mediate transcriptional activation via the Mondo family. We propose that Mlx, like Max, functions as the center of a transcription factor network.

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Molecular Components of Nitrate and Nitrite Efflux in Yeast

Eukaryotic Cell ◽

10.1128/ec.00268-13 ◽

2013 ◽

Vol 13 (2) ◽

pp. 267-278 ◽

Cited By ~ 16

Author(s):

Elisa Cabrera ◽

Rafaela González-Montelongo ◽

Teresa Giraldez ◽

Diego Alvarez de la Rosa ◽

José M. Siverio

Keyword(s):

Nitrate Reductase ◽

Nitrate Uptake ◽

Reductase Activity ◽

Hansenula Polymorpha ◽

Nitrate Assimilation ◽

Nitrate Reductase Gene ◽

Content Type ◽

Nitrite Toxicity ◽

Essential Components ◽

Nitrate And Nitrite

ABSTRACTSome eukaryotes, such as plant and fungi, are capable of utilizing nitrate as the sole nitrogen source. Once transported into the cell, nitrate is reduced to ammonium by the consecutive action of nitrate and nitrite reductase. How nitrate assimilation is balanced with nitrate and nitrite efflux is unknown, as are the proteins involved. The nitrate assimilatory yeastHansenula polymorphawas used as a model to dissect these efflux systems. We identified the sulfite transporters Ssu1 and Ssu2 as effective nitrate exporters, Ssu2 being quantitatively more important, and we characterize the Nar1 protein as a nitrate/nitrite exporter. The use of strains lacking eitherSSU2orNAR1along with the nitrate reductase geneYNR1showed that nitrate reductase activity is not required for net nitrate uptake. Growth test experiments indicated that Ssu2 and Nar1 exporters allow yeast to cope with nitrite toxicity. We also have shown that the well-knownSaccharomyces cerevisiaesulfite efflux permease Ssu1 is also able to excrete nitrite and nitrate. These results characterize for the first time essential components of the nitrate/nitrite efflux system and their impact on net nitrate uptake and its regulation.

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Multi-parameter analysis of the kinetics of NF-κB signalling and transcription in single living cells

Journal of Cell Science ◽

10.1242/jcs.115.6.1137 ◽

2002 ◽

Vol 115 (6) ◽

pp. 1137-1148 ◽

Cited By ~ 1

Author(s):

Glyn Nelson ◽

Luminita Paraoan ◽

David G. Spiller ◽

Geraint J. C. Wilde ◽

Mark A. Browne ◽

...

Keyword(s):

Transcription Factor ◽

Transcriptional Activation ◽

Nuclear Export ◽

Mammalian Cells ◽

Nuclear Translocation ◽

Fluorescent Proteins ◽

Parameter Analysis ◽

Nuclear Accumulation ◽

Fusion Protein Expression ◽

Kinetics Of

Proteins of the NF-κB transcription factor family normally reside in the cytoplasm of cells in a complex with IκB inhibitor proteins. Stimulation with TNFα leads to proteosomal degradation of the IκB proteins and nuclear translocation of the NF-κB proteins. Expression of p65 and IκBα fused to fluorescent proteins was used to measure the dynamics of these processes in transfected HeLa cells. Simultaneous visualisation of p65-dsRed translocation and IκBα-EGFP degradation indicated that in the presence of dual fluorescent fusion protein expression,the half-time of IκBα-EGFP degradation was reduced and that of p65 translocation was significantly increased when compared with cells expressing the single fluorescent fusion proteins. These results suggest that the ratio of IκBα and p65 determine the kinetics of transcription factor translocation into the nucleus and indicate that the complex of p65 and IκBα is the true substrate for TNFα stimulation in mammalian cells. When cells were treated with the CRM-1-dependent nuclear export inhibitor,leptomycin B (LMB), there was nuclear accumulation of IκBα-EGFP and p65-dsRed, with IκBα-EGFP accumulating more rapidly. No NF-κB-dependent transcriptional activation was seen in response to LMB treatment. Following 1 hour treatment with LMB, significant IκBα-EGFP nuclear accumulation, but low levels of p65-dsRed nuclear accumulation, was observed. When these cells were stimulated with TNFα, degradation of IκBα-EGFP was observed in both the cytoplasm and nucleus. A normal transient transcription response was observed in the same cells using luminescence imaging of NF-κB-dependent transcription. These observations suggest that both normal activation and post-induction repression of NF-κB-dependent transcription occur even when nuclear export of NF-κB is inhibited. The results provide functional evidence that other factors, such as modification of p65 by phosphorylation, or interaction with other proteins such as transcriptional co-activators/co-repressors, may critically modulate the kinetics of transcription through this signalling pathway.

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Cyto-nuclear shuttling of afadin is required for rapid estradiol-mediated modifications of histone H3

10.1101/284422 ◽

2018 ◽

Author(s):

Katherine J. Sellers ◽

Iain A. Watson ◽

Deepak P. Srivastava

Keyword(s):

Learning And Memory ◽

Transcriptional Activation ◽

Histone H3 ◽

Neuronal Population ◽

Nuclear Accumulation ◽

Estradiol Treatment ◽

Nuclear Trafficking ◽

17Β Estradiol ◽

Nuclear Shuttling ◽

And Function

AbstractEstrogens have been shown to rapidly regulate local signalling events at both synapses and within the nucleus. The result of these signalling events is to rapidly modulate synapse structure and function, as well as epigenetic mechanisms including histone modifications. Ultimately these mechanisms are thought to contribute to long-lasting changes in neural circuitry, and thus influences cognitive functions such as learning and memory. However, the mechanisms by which estrogen-mediated local synaptic and nuclear signalling events are coordinated are not well understood. In this study we have found that the scaffold protein afadin, (also known as AF-6), undergoes a bi-directional trafficking to both synaptic and nuclear compartment in response to acute 17β-estradiol (estradiol) treatment. Interestingly, nuclear accumulation of afadin was coincidental with an increase in the phosphorylation of histone H3 at serine 10 (H3S10p). This histone modification is associated with the remodelling of chromatin into an open euchromatin state, allowing for transcriptional activation and related learning and memory processes. Critically, the cyto-nuclear trafficking of afadin was required for estradiol-dependent H3S10p. We further determined that nuclear accumulation of afadin is sufficient to induce phosphorylation of the mitogentic kinases ERK1/2 (pERK1/2) within the nucleus. Moreover, nuclear pERK1/2 was required for estradiol-dependent H3S10p. Taken together, we propose a model whereby estradiol incudes the bi-directional trafficking of afadin to synaptic and nuclear sub-compartments. Within the nucleus, afadin is required for increased pERK1/2 which in turn is required for H3S10p. Therefore this represents a mechanism through which estrogens may be able to coordinate both synaptic and nucleosomal events within the same neuronal population.

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