scholarly journals Histone H3 K4 Demethylation during Activation and Attenuation of GAL1 Transcription in Saccharomyces cerevisiae

2007 ◽  
Vol 27 (22) ◽  
pp. 7856-7864 ◽  
Author(s):  
Kristin Ingvarsdottir ◽  
Chris Edwards ◽  
Min Gyu Lee ◽  
Jung Shin Lee ◽  
David C. Schultz ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mammalian Cells ◽  
Gene Activation ◽  
Histone H3 ◽  
Histone Lysine ◽  
Insight Into ◽  
Demethylation Activity

ABSTRACT In mammalian cells, histone lysine demethylation is carried out by two classes of enzymes, the LSD1/BHC110 class and the jumonji class. The enzymes of the jumonji class in the yeast Saccharomyces cerevisiae have recently also been shown to have lysine demethylation activity. Here we report that the protein encoded by YJR119c (termed KDM5), coding for one of five predicted jumonji domain proteins in yeast, specifically demethylates trimethylated histone H3 lysine 4 (H3K4me3), H3K4me2, and H3K4me1 in vitro. We found that loss of KDM5 increased mono-, di-, and trimethylation of lysine 4 during activation of the GAL1 gene. Interestingly, cells deleted of KDM5 also displayed a delayed reduction of K4me3 upon reestablishment of GAL1 repression. These results indicate that K4 demethylation has two roles at GAL1, first to establish appropriate levels of K4 methylation during gene activation and second to remove K4 trimethylation during the attenuation phase of transcription. Thus, analysis of lysine demethylation in yeast provides new insight into the physiological roles of jumonji demethylase enzymes.

scholarly journals Cells Expressing Murine RAD52 Splice Variants Favor Sister Chromatid Repair

2006 ◽  
Vol 26 (10) ◽  
pp. 3752-3763 ◽  
Author(s):  
Peter H. Thorpe ◽  
Vanessa A. Marrero ◽  
Margaret H. Savitzky ◽  
Ivana Sunjevaric ◽  
Tom C. Freeman ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Sister Chromatid ◽  
Mammalian Cells ◽  
Splice Variants ◽  
Protein A ◽  
Single Stranded Dna ◽  
Culture Cells ◽  
Dominant Phenotype ◽  
Mouse Tissues

ABSTRACT The RAD52 gene is essential for homologous recombination in the yeast Saccharomyces cerevisiae. RAD52 is the archetype in an epistasis group of genes essential for DNA damage repair. By catalyzing the replacement of replication protein A with Rad51 on single-stranded DNA, Rad52 likely promotes strand invasion of a double-stranded DNA molecule by single-stranded DNA. Although the sequence and in vitro functions of mammalian RAD52 are conserved with those of yeast, one difference is the presence of introns and consequent splicing of the mammalian RAD52 pre-mRNA. We identified two novel splice variants from the RAD52 gene that are expressed in adult mouse tissues. Expression of these splice variants in tissue culture cells elevates the frequency of recombination that uses a sister chromatid template. To characterize this dominant phenotype further, the RAD52 gene from the yeast Saccharomyces cerevisiae was truncated to model the mammalian splice variants. The same dominant sister chromatid recombination phenotype seen in mammalian cells was also observed in yeast. Furthermore, repair from a homologous chromatid is reduced in yeast, implying that the choice of alternative repair pathways may be controlled by these variants. In addition, a dominant DNA repair defect induced by one of the variants in yeast is suppressed by overexpression of RAD51, suggesting that the Rad51-Rad52 interaction is impaired.

Functional interaction between p21rap1A and components of the budding pathway in Saccharomyces cerevisiae

1992 ◽  
Vol 12 (9) ◽  
pp. 4084-4092
Author(s):  
P C McCabe ◽  
H Haubruck ◽  
P Polakis ◽  
F McCormick ◽  
M A Innis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mammalian Cells ◽  
Bound States ◽  
Yeast Cells ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Amino Terminal ◽  
Loop Region

The rap1A gene encodes a 21-kDa, ras-related GTP-binding protein (p21rap1A) of unknown function. A close structural homolog of p21rap1A (65% identity in the amino-terminal two-thirds) is the RSR1 gene product (Rsr1p) of Saccharomyces cerevisiae. Although Rsr1p is not essential for growth, its presence is required for nonrandom selection of bud sites. To assess the similarity of these proteins at the functional level, wild-type and mutant forms of p21rap1A were tested for complementation of activities known to be fulfilled by Rsr1p. Expression of p21rap1A, like multicopy expression of RSR1, suppressed the conditional lethality of a temperature-sensitive cdc24 mutation. Point mutations predicted to affect the localization of p21rap1A or its ability to cycle between GDP and GTP-bound states disrupted suppression of cdc24ts, while other mutations in the 61-65 loop region improved suppression. Expression of p21rap1A could not, however, suppress the random budding phenotype of rsr1 cells. p21rap1A also apparently interfered with the normal activity of Rsrlp, causing random budding in diploid wild-type cells, suggesting an inability of p21rap1A to interact appropriately with Rsr1p regulatory proteins. Consistent with this hypothesis, we found an Rsr1p-specific GTPase-activating protein (GAP) activity in yeast membranes which was not active toward p21rap1A, indicating that p21rap1A may be predominantly GTP bound in yeast cells. Coexpression of human Rap1-specific GAP suppressed the random budding due to expression of p21rap1A or its derivatives, including Rap1AVal-12. Although Rap1-specific GAP stimulated the GTPase of Rsr1p in vitro, it did not dominantly interfere with Rsr1p function in vivo. A chimera consisting of Rap1A1-165::Rsr1p166-272 did not exhibit normal Rsr1p function in the budding pathway. These results indicated that p21rap1A and Rsr1p share at least partial functional homology, which may have implications for p21rap1A function in mammalian cells.

scholarly journals Mutational Analysis of the Sir3 BAH Domain Reveals Multiple Points of Interaction with Nucleosomes

2009 ◽  
Vol 29 (10) ◽  
pp. 2532-2545 ◽  
Author(s):  
Vinaya Sampath ◽  
Peihua Yuan ◽  
Isabel X. Wang ◽  
Evelyn Prugar ◽  
Fred van Leeuwen ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mutational Analysis ◽  
Histone H3 ◽  
Transcriptional Silencing ◽  
In Vitro Experiments ◽  
Multiple Points ◽  
Alanine Residue ◽  
Bah Domain ◽  
Multiple Domains

ABSTRACT Sir3, a component of the transcriptional silencing complex in the yeast Saccharomyces cerevisiae, has an N-terminal BAH domain that is crucial for the protein's silencing function. Previous work has shown that the N-terminal alanine residue of Sir3 (Ala2) and its acetylation play an important role in silencing. Here we show that the silencing defects of Sir3 Ala2 mutants can be suppressed by mutations in histones H3 and H4, specifically, by H3 D77N and H4 H75Y mutations. Additionally, a mutational analysis demonstrates that three separate regions of the Sir3 BAH domain are important for its role in silencing. Many of these BAH mutations also can be suppressed by the H3 D77N and H4 H75Y mutations. In agreement with the results of others, in vitro experiments show that the Sir3 BAH domain can interact with partially purified nucleosomes. The silencing-defective BAH mutants are defective for this interaction. These results, together with the previously characterized interaction between the C-terminal region of Sir3 and the histone H3/H4 tails, suggest that Sir3 utilizes multiple domains to interact with nucleosomes.

scholarly journals Probe the function of histone lysine 36 methylation using histone H3 lysine 36 to methionine mutant transgene in mammalian cells

Cell Cycle ◽  
2017 ◽  
Vol 16 (19) ◽  
pp. 1781-1789 ◽  
Author(s):  
Dong Fang ◽  
Haiyun Gan ◽  
Heping Wang ◽  
Hui Zhou ◽  
Zhiguo Zhang
Keyword(s):  
Mammalian Cells ◽  
Histone H3 ◽  
Histone Lysine

scholarly journals Molecular basis for phosphospecific recognition of histone H3 tails by Survivin paralogues at inner centromeres

2012 ◽  
Vol 23 (8) ◽  
pp. 1457-1466 ◽  
Author(s):  
Ewa Niedzialkowska ◽  
Fangwei Wang ◽  
Przemyslaw J. Porebski ◽  
Wladek Minor ◽  
Jonathan M. G. Higgins ◽  
...  
Keyword(s):  
Histone H3 ◽  
Histone Tails ◽  
X Ray Crystallography ◽  
Class A ◽  
Class B ◽  
A Subunit ◽  
Chromosome Passenger Complex ◽  
Different Characteristics ◽  
Insight Into

Survivin, a subunit of the chromosome passenger complex (CPC), binds the N-terminal tail of histone H3, which is phosphorylated on T3 by Haspin kinase, and localizes the complex to the inner centromeres. We used x-ray crystallography to determine the residues of Survivin that are important in binding phosphomodified histone H3. Mutation of amino acids that interact with the histone N-terminus lowered in vitro tail binding affinity and reduced CPC recruitment to the inner centromere in cells, validating our solved structures. Phylogenetic analysis shows that nonmammalian vertebrates have two Survivin paralogues, which we name class A and B. A distinguishing feature of these paralogues is an H-to-R change in an amino acid that interacts with the histone T3 phosphate. The binding to histone tails of the human class A paralogue, which has a histidine at this position, is sensitive to changes around physiological pH, whereas Xenopus Survivin class B is less so. Our data demonstrate that Survivin paralogues have different characteristics of phosphospecific binding to threonine-3 of histone H3, providing new insight into the biology of the inner centromere.

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 The histone H3-H4 tetramer is a copper reductase enzyme

Science ◽  
2020 ◽  
Vol 369 (6499) ◽  
pp. 59-64 ◽  
Author(s):  
Narsis Attar ◽  
Oscar A. Campos ◽  
Maria Vogelauer ◽  
Chen Cheng ◽  
Yong Xue ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Active Site ◽  
Histone H3 ◽  
Copper Binding ◽  
Active Site Residues ◽  
Chromatin Compaction ◽  
Putative Active Site ◽  
Dimerization Interface ◽  
Cellular Copper

Eukaryotic histone H3-H4 tetramers contain a putative copper (Cu2+) binding site at the H3-H3′ dimerization interface with unknown function. The coincident emergence of eukaryotes with global oxygenation, which challenged cellular copper utilization, raised the possibility that histones may function in cellular copper homeostasis. We report that the recombinant Xenopus laevis H3-H4 tetramer is an oxidoreductase enzyme that binds Cu2+ and catalyzes its reduction to Cu1+ in vitro. Loss- and gain-of-function mutations of the putative active site residues correspondingly altered copper binding and the enzymatic activity, as well as intracellular Cu1+ abundance and copper-dependent mitochondrial respiration and Sod1 function in the yeast Saccharomyces cerevisiae. The histone H3-H4 tetramer, therefore, has a role other than chromatin compaction or epigenetic regulation and generates biousable Cu1+ ions in eukaryotes.

scholarly journals Apoptotic induction induces Leishmania aethiopica and L. mexicana spreading in terminally differentiated THP-1 cells

Parasitology ◽  
2017 ◽  
Vol 144 (14) ◽  
pp. 1912-1921 ◽  
Author(s):  
RAJEEV RAI ◽  
PAUL DYER ◽  
SIMON RICHARDSON ◽  
LAURENCE HARBIGE ◽  
GIULIA GETTI
Keyword(s):  
Mammalian Cells ◽  
Molecular Mechanisms ◽  
Secondary Infection ◽  
Human Macrophages ◽  
Leishmania Aethiopica ◽  
Reliable Model ◽  
Infected Cells ◽  
Insight Into ◽  
Apoptotic Induction

SummaryLeishmaniasis develops after parasites establish themselves as amastigotes inside mammalian cells and start replicating. As relatively few parasites survive the innate immune defence, intracellular amastigotes spreading towards uninfected cells is instrumental to disease progression. Nevertheless the mechanism of Leishmania dissemination remains unclear, mostly due to the lack of a reliable model of infection spreading. Here, an in vitro model representing the dissemination of Leishmania amastigotes between human macrophages has been developed. Differentiated THP-1 macrophages were infected with GFP expressing Leishmania aethiopica and Leishmania mexicana. The percentage of infected cells was enriched via camptothecin treatment to achieve 64·1 ± 3% (L. aethiopica) and 92 ± 1·2% (L. mexicana) at 72 h, compared to 35 ± 4·2% (L. aethiopica) and 36·2 ± 2·4% (L. mexicana) in untreated population. Infected cells were co-cultured with a newly differentiated population of THP-1 macrophages. Spreading was detected after 12 h of co-culture. Live cell imaging showed inter-cellular extrusion of L. aethiopica and L. mexicana to recipient cells took place independently of host cell lysis. Establishment of secondary infection from Leishmania infected cells provided an insight into the cellular phenomena of parasite movement between human macrophages. Moreover, it supports further investigation into the molecular mechanisms of parasites spreading, which forms the basis of disease development.

scholarly journals Chaperone Control of the Activity and Specificity of the Histone H3 Acetyltransferase Rtt109

2008 ◽  
Vol 28 (13) ◽  
pp. 4342-4353 ◽  
Author(s):  
Jeffrey Fillingham ◽  
Judith Recht ◽  
Andrea C. Silva ◽  
Bernhard Suter ◽  
Andrew Emili ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Histone Acetyltransferase ◽  
Histone H3 ◽  
Histone Chaperone ◽  
Genetic Interactions ◽  
Whole Genome ◽  
Genome Screen ◽  
Associated Lesions

ABSTRACT Acetylation of Saccharomyces cerevisiae histone H3 on K56 by the histone acetyltransferase (HAT) Rtt109 is important for repairing replication-associated lesions. Rtt109 purifies from yeast in complex with the histone chaperone Vps75, which stabilizes the HAT in vivo. A whole-genome screen to identify genes whose deletions have synthetic genetic interactions with rtt109Δ suggests Rtt109 has functions in addition to DNA repair. We show that in addition to its known H3-K56 acetylation activity, Rtt109 is also an H3-K9 HAT, and we show that Rtt109 and Gcn5 are the only H3-K9 HATs in vivo. Rtt109's H3-K9 acetylation activity in vitro is enhanced strongly by Vps75. Another histone chaperone, Asf1, and Vps75 are both required for acetylation of lysine 9 on H3 (H3-K9ac) in vivo by Rtt109, whereas H3-K56ac in vivo requires only Asf1. Asf1 also physically interacts with the nuclear Hat1/Hat2/Hif1 complex that acetylates H4-K5 and H4-K12. We suggest Asf1 is capable of assembling into chromatin H3-H4 dimers diacetylated on both H4-K5/12 and H3-K9/56.

scholarly journals Involvement of Histone Lysine Methylation in p21 Gene Expression in Rat KidneyIn Vivoand Rat Mesangial CellsIn Vitrounder Diabetic Conditions

2016 ◽  
Vol 2016 ◽  
pp. 1-12 ◽  
Author(s):  
Xiangjun Li ◽  
Chaoyuan Li ◽  
Xiaoxia Li ◽  
Peihe Cui ◽  
Qifeng Li ◽  
...  
Keyword(s):  
Gene Expression ◽  
Molecular Mechanisms ◽  
Mesangial Cells ◽  
Histone H3 ◽  
Diabetic Rats ◽  
Lysine Methylation ◽  
Histone Lysine Methylation ◽  
Histone Lysine

Diabetic nephropathy (DN), a common complication associated with type 1 and type 2 diabetes mellitus (DM), characterized by glomerular mesangial expansion, inflammation, accumulation of extracellular matrix (ECM) protein, and hypertrophy, is the major cause of end-stage renal disease (ESRD). Increasing evidence suggested that p21-dependent glomerular and mesangial cell (MC) hypertrophy play key roles in the pathogenesis of DN. Recently, posttranscriptional modifications (PTMs) have uncovered novel molecular mechanisms involved in DN. However, precise regulatory mechanism of histone lysine methylation (HKme) mediating p21 related hypertrophy associated with DN is not clear. We evaluated the roles of HKme and histone methyltransferase (HMT) SET7/9 in p21 gene expression in glomeruli of diabetic rats and in high glucose- (HG-) treated rat mesangial cells (RMCs). p21 gene expression was upregulated in diabetic rats glomeruli; chromatin immunoprecipitation (ChIP) assays showed decreased histone H3-lysine9-dimethylation (H3K9me2) accompanied with enhanced histone H3-lysine4-methylation (H3K4me1/3) and SET7/9 occupancies at the p21 promoter. HG-treated RMCs exhibited increased p21 mRNA, H3K4me level, SET7/9 recruitment, and inverse H3K9me, which were reversed by TGF-β1 antibody. These data uncovered key roles of H3Kme and SET7/9 responsible for p21 gene expressionin vivoandin vitrounder diabetic conditions and confirmed preventive effect of TGF-β1 antibody on DN.

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