The Yeast Protein Complex Containing Cdc68 and Pob3 Mediates Core-Promoter Repression Through the Cdc68 N-Terminal Domain

Genetics ◽  
1998 ◽  
Vol 150 (4) ◽  
pp. 1393-1405 ◽  
Author(s):  
David R H Evans ◽  
Neil K Brewster ◽  
Qunli Xu ◽  
Adele Rowley ◽  
Brent A Altheim ◽  
...  
Keyword(s):  
Core Promoter ◽  
Kluyveromyces Lactis ◽  
Gene Activation ◽  
Mutant Form ◽  
Nuclear Complex ◽  
Terminal Domain ◽  
Chromatin Remodeling Complex ◽  
Functional Homolog ◽  
Repression Domain ◽  
Independent Mutant

Abstract Transcription of nuclear genes usually involves trans-activators, whereas repression is exerted by chromatin. For several genes the transcription mediated by trans-activators and the repression mediated by chromatin depend on the CP complex, a recently described abundant yeast nuclear complex of the Pob3 and Cdc68/Spt16 proteins. We report that the N-terminal third of the Saccharomyces cerevisiae Cdc68 protein is dispensable for gene activation but necessary for the maintenance of chromatin repression. The absence of this 300-residue N-terminal domain also decreases the need for the Swi/Snf chromatin-remodeling complex in transcription and confers an Spt- effect characteristic of chromatin alterations. The repression domain, and indeed the entire Cdc68 protein, is highly conserved, as shown by the sequence of the Cdc68 functional homolog from the yeast Kluyveromyces lactis and by database searches. The repression-defective (truncated) form of Cdc68 is stable but less active at high temperatures, whereas the known point-mutant form of Cdc68, encoded by three independent mutant alleles, alters the N-terminal repression domain and destabilizes the mutant protein.

scholarly journals FunctionalDomains of the RhlR Transcriptional Regulator ofPseudomonasaeruginosa

2003 ◽  
Vol 185 (24) ◽  
pp. 7129-7139 ◽  
Author(s):  
Janet R. Lamb ◽  
Hetal Patel ◽  
Timothy Montminy ◽  
Victoria E. Wagner ◽  
Barbara H. Iglewski
Keyword(s):  
Amino Acid ◽  
Transcriptional Activation ◽  
Gene Activation ◽  
Transcriptional Regulator ◽  
Homoserine Lactone ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Mutant Form ◽  
E Coli ◽  
Terminal Domain

ABSTRACT The RhlR transcriptional regulator of Pseudomonas aeruginosa, along with its cognate autoinducer, N-butyryl homoserine lactone (C4-HSL), regulates gene expression in response to cell density. With an Escherichia coli LexA-based protein interaction system, we demonstrated that RhlR multimerized and that the degree of multimerization was dependent on the C4-HSL concentration. Studies with an E. coli lasB::lacZ lysogen demonstrated that RhlR multimerization was necessary for it to function as a transcriptional activator. Deletion analysis of RhlR indicated that the N-terminal domain of the protein is necessary for C4-HSL binding. Single amino acid substitutions in the C-terminal domain of RhlR generated mutant RhlR proteins that had the ability to bind C4-HSL and multimerize but were unable to activate lasB expression, demonstrating that the C-terminal domain is important for target gene activation. Single amino acid substitutions in both the N-terminal and C-terminal domains of RhlR demonstrated that both domains possess residues involved in multimerization. RhlR with a C-terminal deletion and an RhlR site-specific mutant form that possessed multimerization but not transcriptional activation capabilities were able to inhibit the ability of wild-type RhlR to activate rhlA expression in P. aeruginosa. We conclude that C4-HSL binding is necessary for RhlR multimerization and that RhlR functions as a multimer in P. aeruginosa.

scholarly journals A Novel Transcriptional Repression Domain Mediates p21WAF1/CIP1 Induction of p300 Transactivation

2000 ◽  
Vol 20 (8) ◽  
pp. 2676-2686 ◽  
Author(s):  
Andrew W. Snowden ◽  
Lisa A. Anderson ◽  
Gill A. Webster ◽  
Neil D. Perkins
Keyword(s):  
Cell Cycle ◽  
Transcriptional Activation ◽  
Transcriptional Repression ◽  
Kinase Inhibitor ◽  
Core Promoter ◽  
Dependent Manner ◽  
Creb Binding Protein ◽  
Cycle Dependent ◽  
Repression Domain

ABSTRACT The transcriptional coactivators p300 and CREB binding protein (CBP) are important regulators of the cell cycle, differentiation, and tumorigenesis. Both p300 and CBP are targeted by viral oncoproteins, are mutated in certain forms of cancer, are phosphorylated in a cell cycle-dependent manner, interact with transcription factors such as p53 and E2F, and can be found complexed with cyclinE-Cdk2 in vivo. Moreover, p300-deficient cells show defects in proliferation. Here we demonstrate that transcriptional activation by both p300 and CBP is stimulated by coexpression of the cyclin-dependent kinase inhibitor p21WAF/CIP1. Significantly this stimulation is independent of both the inherent histone acetyltransferase (HAT) activity of p300 and CBP and of the previously reported carboxyl-terminal binding site for cyclinE-Cdk2. Rather, we describe a previously uncharacterized transcriptional repression domain (CRD1) within p300. p300 transactivation is stimulated through derepression of CRD1 by p21. Significantly p21 regulation of CRD1 is dependent on the nature of the core promoter. We suggest that CRD1 provides a novel mechanism through which p300 and CBP can switch activities between the promoters of genes that stimulate growth and those that enhance cell cycle arrest.

scholarly journals AP-2/Eps15 Interaction Is Required for Receptor-mediated Endocytosis

1998 ◽  
Vol 140 (5) ◽  
pp. 1055-1062 ◽  
Author(s):  
Alexandre Benmerah ◽  
Christophe Lamaze ◽  
Bernadette Bègue ◽  
Sandra L. Schmid ◽  
Alice Dautry-Varsat ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Fusion Protein ◽  
Binding Sites ◽  
Fluorescent Protein ◽  
Coated Vesicle ◽  
Mutant Form ◽  
A431 Cells ◽  
Coated Pits ◽  
Receptor Mediated Endocytosis ◽  
Terminal Domain

We have previously shown that the protein Eps15 is constitutively associated with the plasma membrane adaptor complex, AP-2, suggesting its possible role in endocytosis. To explore the role of Eps15 and the function of AP-2/Eps15 association in endocytosis, the Eps15 binding domain for AP-2 was precisely delineated. The entire COOH-terminal domain of Eps15 or a mutant form lacking all the AP-2–binding sites was fused to the green fluorescent protein (GFP), and these constructs were transiently transfected in HeLa cells. Overexpression of the fusion protein containing the entire COOH-terminal domain of Eps15 strongly inhibited endocytosis of transferrin, whereas the fusion protein in which the AP-2–binding sites had been deleted had no effect. These results were confirmed in a cell-free assay that uses perforated A431 cells to follow the first steps of coated vesicle formation at the plasma membrane. Addition of Eps15-derived glutathione-S-transferase fusion proteins containing the AP-2–binding site in this assay inhibited not only constitutive endocytosis of transferrin but also ligand-induced endocytosis of epidermal growth factor. This inhibition could be ascribed to a competition between the fusion protein and endogenous Eps15 for AP-2 binding. Altogether, these results show that interaction of Eps15 with AP-2 is required for efficient receptor-mediated endocytosis and thus provide the first evidence that Eps15 is involved in the function of plasma membrane–coated pits.

scholarly journals A Virus-Virus Interaction Circumvents the Virus Receptor Requirement for Infection by Pathogenic Retroviruses

2003 ◽  
Vol 77 (6) ◽  
pp. 3460-3469 ◽  
Author(s):  
David L. Wensel ◽  
Weihua Li ◽  
James M. Cunningham
Keyword(s):  
Cell Surface ◽  
Envelope Glycoprotein ◽  
Gene Activation ◽  
Host Cells ◽  
Virus Binding ◽  
Virus Envelope ◽  
Host Interactions ◽  
Terminal Domain ◽  
Retrovirus Infection ◽  
Ecotropic Virus

ABSTRACT During ongoing C-type retrovirus infection, the probability of leukemia caused by insertional gene activation is markedly increased by the emergence of recombinant retroviruses that repeatedly infect host cells. The murine mink cell focus-inducing (MCF) viruses with this property have acquired characteristic changes in the N-terminal domain of their envelope glycoprotein that specify binding to a different receptor than the parental ecotropic virus. In this report, we show that MCF virus infection occurs through binding to this receptor (termed Syg1) and, remarkably, by a second mechanism that does not utilize the Syg1 receptor. By the latter route, the N-terminal domain of the ecotropic virus glycoprotein expressed on the cell surface in a complex with its receptor activates the fusion mechanism of the MCF virus in trans. The rate of MCF virus spread through a population of permissive human cells was increased by establishment of trans activation, indicating that Syg1 receptor-dependent and -independent pathways function in parallel. Also, trans activation shortened the interval between initial infection and onset of cell-cell fusion associated with repeated infection of the same cell. Our findings indicate that pathogenic retrovirus infection may be initiated by virus binding to cell receptors or to the virus envelope glycoprotein of other viruses expressed on the cell surface. Also, they support a broader principle: that cooperative virus-virus interactions, as well as virus-host interactions, shape the composition and properties of the retrovirus quasispecies.

Nuclear receptor coactivators: the key to unlock chromatin

2005 ◽  
Vol 83 (4) ◽  
pp. 418-428 ◽  
Author(s):  
Wei Xu
Keyword(s):  
Nuclear Receptors ◽  
Nuclear Receptor ◽  
Chromatin Remodeling ◽  
Transcriptional Activation ◽  
Transcriptional Control ◽  
Biological Effects ◽  
Gene Activation ◽  
Nuclear Hormone Receptor ◽  
Nuclear Receptor Coactivators ◽  
Chromatin Remodeling Complex

The biological effects of hormones, ranging from organogenesis, metabolism, and proliferation, are transduced through nuclear receptors (NRs). Over the last decade, NRs have been used as a model to study transcriptional control. The conformation of activated NRs is favorable for the recruitment of coactivators, which promote transcriptional activation by directly communicating with chromatin. This review will focus on the function of different classes of coactivators and associated complexes, and on progress in our understanding of gene activation by NRs through chromatin remodeling.Key words: nuclear hormone receptor, p160 family of coactivators, histone modification, chromatin remodeling complex.

scholarly journals Promyelocytic Leukemia Protein Isoform II Promotes Transcription Factor Recruitment To Activate Interferon Beta and Interferon-Responsive Gene Expression

2015 ◽  
Vol 35 (10) ◽  
pp. 1660-1672 ◽  
Author(s):  
Yixiang Chen ◽  
Jordan Wright ◽  
Xueqiong Meng ◽  
Keith N. Leppard
Keyword(s):  
Target Genes ◽  
Gene Activation ◽  
Promyelocytic Leukemia ◽  
Type I ◽  
Gene Promoters ◽  
Creb Binding Protein ◽  
Terminal Domain ◽  
Efficient Induction ◽  
Transcription Complexes ◽  
Transcriptional Complex

To trigger type I interferon (IFN) responses, pattern recognition receptors activate signaling cascades that lead to transcription of IFN and IFN-stimulated genes (ISGs). The promyelocytic leukemia (PML) protein has been implicated in these responses, although its role has not been defined. Here, we show that PML isoform II (PML-II) is specifically required for efficient induction of IFN-β transcription and of numerous ISGs, acting at the point of transcriptional complex assembly on target gene promoters. PML-II associated with specific transcription factors NF-κB and STAT1, as well as the coactivator CREB-binding protein (CBP), to facilitate transcriptional complex formation. The absence of PML-II substantially reduced binding of these factors and IFN regulatory factor 3 (IRF3) to IFN-β or ISGs promoters and sharply reduced gene activation. The unique C-terminal domain of PML-II was essential for its activity, while the N-terminal RBCC motif common to all PML isoforms was dispensable. We propose a model in which PML-II contributes to the transcription of multiple genes via the association of its C-terminal domain with relevant transcription complexes, which promotes the stable assembly of these complexes at promoters/enhancers of target genes, and that in this way PML-II plays a significant role in the development of type I IFN responses.

scholarly journals Cryo-electron microscopy structure of a nucleosome-bound SWI/SNF chromatin remodeling complex

2019 ◽  
Author(s):  
Yan Han ◽  
Alexis A Reyes ◽  
Sara Malik ◽  
Yuan He
Keyword(s):  
Electron Microscopy ◽  
Chromatin Remodeling ◽  
Atp Hydrolysis ◽  
Protein Complexes ◽  
Gene Activation ◽  
The Body ◽  
Cellular Processes ◽  
Cryo Electron Microscopy ◽  
Chromatin Remodeling Complex ◽  
High Resolution Structure

AbstractThe multi-subunit chromatin remodeling complex SWI/SNF1–3 is highly conserved from yeast to humans and plays critical roles in various cellular processes including transcription and DNA damage repair4, 5. It uses the energy from ATP hydrolysis to remodel chromatin structure by sliding and evicting the histone octamer6–10, creating DNA regions that become accessible to other essential protein complexes. However, our mechanistic understanding of the chromatin remodeling activity is largely hindered by the lack of a high-resolution structure of any complex from this family. Here we report the first structure of SWI/SNF from the yeast S. cerevisiae bound to a nucleosome at near atomic resolution determined by cryo-electron microscopy (cryo-EM). In the structure, the Arp module is sandwiched between the ATPase and the Body module of the complex, with the Snf2 HSA domain connecting all modules. The HSA domain also extends into the Body and anchors at the opposite side of the complex. The Body contains an assembly scaffold composed of conserved subunits Snf12 (SMARCD/BAF60), Snf5 (SMARCB1/BAF47/ INI1) and an asymmetric dimer of Swi3 (SMARCC/BAF155/170). Another conserved subunit Swi1 (ARID1/BAF250) folds into an Armadillo (ARM) repeat domain that resides in the core of the SWI/SNF Body, acting as a molecular hub. In addition to the interaction between Snf2 and the nucleosome, we also observed interactions between the conserved Snf5 subunit and the histones at the acidic patch, which could serve as an anchor point during active DNA translocation. Our structure allows us to map and rationalize a subset of cancer-related mutations in the human SWI/SNF complex and propose a model of how SWI/SNF recognizes and remodels the +1 nucleosome to generate nucleosome-depleted regions during gene activation11–13.

scholarly journals Recruitment of the SWI-SNF Chromatin Remodeling Complex as a Mechanism of Gene Activation by the Glucocorticoid Receptor τ1 Activation Domain

2000 ◽  
Vol 20 (6) ◽  
pp. 2004-2013 ◽  
Author(s):  
Annika E. Wallberg ◽  
Kristen E. Neely ◽  
Ahmed H. Hassan ◽  
Jan-Åke Gustafsson ◽  
Jerry L. Workman ◽  
...  
Keyword(s):  
Glucocorticoid Receptor ◽  
Mammalian Cells ◽  
Gene Activation ◽  
Dependent Manner ◽  
Transactivation Domain ◽  
Chromatin Remodeling Complex ◽  
Nucleosomal Dna ◽  
Target Promoters

ABSTRACT The SWI-SNF complex has been shown to alter nucleosome conformation in an ATP-dependent manner, leading to increased accessibility of nucleosomal DNA to transcription factors. In this study, we show that the SWI-SNF complex can potentiate the activity of the glucocorticoid receptor (GR) through the N-terminal transactivation domain, τ1, in both yeast and mammalian cells. GR-τ1 can directly interact with purified SWI-SNF complex, and mutations in τ1 that affect the transactivation activity in vivo also directly affect τ1 interaction with SWI-SNF. Furthermore, the SWI-SNF complex can stimulate τ1-driven transcription from chromatin templates in vitro. Taken together, these results support a model in which the GR can directly recruit the SWI-SNF complex to target promoters during glucocorticoid-dependent gene activation. We also provide evidence that the SWI-SNF and SAGA complexes represent independent pathways of τ1-mediated activation but play overlapping roles that are able to compensate for one another under some conditions.

scholarly journals Molecular Genetic Analysis of the Yeast Repressor Rfx1/Crt1 Reveals a Novel Two-Step Regulatory Mechanism

2005 ◽  
Vol 25 (17) ◽  
pp. 7399-7411 ◽  
Author(s):  
Zhengjian Zhang ◽  
Joseph C. Reese
Keyword(s):  
Dna Damage ◽  
Histone Deacetylases ◽  
Molecular Genetic ◽  
Gene Activation ◽  
Molecular Genetic Analysis ◽  
Transcription Activator ◽  
N Terminus ◽  
Inducible Genes ◽  
Repression Domain

ABSTRACT In Saccharomyces cerevisiae, the repressor Crt1 and the global corepressor Ssn6-Tup1 repress the DNA damage-inducible ribonucleotide reductase (RNR) genes. Initiation of DNA damage signals causes the release of Crt1 and Ssn6-Tup1 from the promoter, coactivator recruitment, and derepression of transcription, indicating that Crt1 plays a crucial role in the switch between gene repression and activation. Here we have mapped the functional domains of Crt1 and identified two independent repression domains and a region required for gene activation. The N terminus of Crt1 is the major repression domain, it directly binds to the Ssn6-Tup1 complex, and its repression activities are dependent upon Ssn6-Tup1 and histone deacetylases (HDACs). In addition, we identified a C-terminal repression domain, which is independent of Ssn6-Tup1 and HDACs and functions at native genes in vivo. Furthermore, we show that TFIID and SWI/SNF bind to a region within the N terminus of Crt1, overlapping with but distinct from the Ssn6-Tup1 binding and repression domain, suggesting that Crt1 may have activator functions. Crt1 mutants were constructed to dissect its activator and repressor functions. All of the mutants were competent for repression of the DNA damage-inducible genes, but a majority were “derepression-defective” mutants. Further characterization of these mutants indicated that they are capable of receiving DNA damage signals and releasing the Ssn6-Tup1 complex from the promoter but are selectively impaired for TFIID and SWI/SNF recruitment. These results imply a two-step activation model of the DNA damage-inducible genes and that Crt1 functions as a signal-dependent dual-transcription activator and repressor that acts in a transient manner.

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