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.

scholarly journals Set2-Dependent K36 Methylation Is Regulated by Novel Intratail Interactions within H3

2009 ◽  
Vol 29 (24) ◽  
pp. 6413-6426 ◽  
Author(s):  
James N. Psathas ◽  
Suting Zheng ◽  
Song Tan ◽  
Joseph C. Reese
Keyword(s):  
Amino Acids ◽  
Catalytic Activity ◽  
Chromatin Remodeling ◽  
Posttranslational Modifications ◽  
Active Site ◽  
Transcription Initiation ◽  
Gene Activation ◽  
N Terminus

ABSTRACT Posttranslational modifications to histones have been studied extensively, but the requirement for the residues within the tails for different stages of transcription is less clear. Using RNR3 as a model, we found that the residues within the N terminus of H3 are predominantly required for steps after transcription initiation and chromatin remodeling. Specifically, deleting as few as 20 amino acids, or substituting glutamines for lysines in the tail, greatly impaired K36 methylation by Set2. The mutations to the tail described here preserve the residues predicted to fill the active site of Set2, and the deletion mimics the recently described cleavage of the H3 tail that occurs during gene activation. Importantly, maintaining the charge of the unmodified tail by arginine substitutions preserves Set2 function in vivo. The H3 tail is dispensable for Set2 recruitment to genes but is required for the catalytic activity of Set2 in vitro. We propose that Set2 activity is controlled by novel intratail interactions which can be influenced by modifications and changes to the structure of the H3 tail to control the dynamics and localization of methylation during elongation.

scholarly journals Analysis of temporal changes in the expression of estrogen-regulated genes in the uterus

2003 ◽  
Vol 30 (3) ◽  
pp. 347-358 ◽  
Author(s):  
H Watanabe ◽  
A Suzuki ◽  
M Kobayashi ◽  
E Takahashi ◽  
M Itamoto ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Expression Profiles ◽  
Gene Activation ◽  
Gene Expression Profiles ◽  
Ovariectomized Mice ◽  
Two Phases ◽  
Er Alpha ◽  
Inducible Genes

In order to understand early events caused by estrogen in vivo, temporal uterine gene expression profiles at early stages were examined using DNA microarray analysis. Ovariectomized mice were exposed to 17beta-estradiol and the temporal mRNA expression changes of ten thousand various genes were analyzed. Clustering analysis revealed that there are at least two phases of gene activation during the period up to six hours. One involved immediate-early genes, which included certain transcription factors and growth factors as well as oncogenes. The other involved early-late genes, which included genes related to RNA and protein synthesis. In clusters of down-regulated genes, transcription factors, proteases, apoptosis and cell cycle genes were found. These hormone-inducible genes were not induced in estrogen receptor (ER) alpha knockout mice. Although expression of ERbeta is known in the uterus, these findings indicate the importance of ERalpha in the changes in gene expression in the uterus.

scholarly journals A Molecular Genetic Dissection of the Evolutionarily Conserved N Terminus of Yeast Rad52

Genetics ◽  
2002 ◽  
Vol 161 (2) ◽  
pp. 549-562
Author(s):  
Uffe H Mortensen ◽  
Naz Erdeniz ◽  
Qi Feng ◽  
Rodney Rothstein
Keyword(s):  
Dna Damage ◽  
Amino Acid ◽  
Dna Binding ◽  
Molecular Genetic ◽  
Mitotic Recombination ◽  
Amino Acid Residues ◽  
Γ Irradiation ◽  
Evolutionarily Conserved ◽  
N Terminus ◽  
Γ Ray

Abstract Rad52 is a DNA-binding protein that stimulates the annealing of complementary single-stranded DNA. Only the N terminus of Rad52 is evolutionarily conserved; it contains the core activity of the protein, including its DNA-binding activity. To identify amino acid residues that are important for Rad52 function(s), we systematically replaced 76 of 165 amino acid residues in the N terminus with alanine. These substitutions were examined for their effects on the repair of γ-ray-induced DNA damage and on both interchromosomal and direct repeat heteroallelic recombination. This analysis identified five regions that are required for efficient γ-ray damage repair or mitotic recombination. Two regions, I and II, also contain the classic mutations, rad52-2 and rad52-1, respectively. Interestingly, four of the five regions contain mutations that impair the ability to repair γ-ray-induced DNA damage yet still allow mitotic recombinants to be produced at rates that are similar to or higher than those obtained with wild-type strains. In addition, a new class of separation-of-function mutation that is only partially deficient in the repair of γ-ray damage, but exhibits decreased mitotic recombination similar to rad52 null strains, was identified. These results suggest that Rad52 protein acts differently on lesions that occur spontaneously during the cell cycle than on those induced by γ-irradiation.

Molecular genetic analysis of myoC, a Dictyostelium myosin I

1995 ◽  
Vol 108 (3) ◽  
pp. 1093-1103 ◽  
Author(s):  
M.D. Peterson ◽  
K.D. Novak ◽  
M.C. Reedy ◽  
J.I. Ruman ◽  
M.A. Titus
Keyword(s):  
Molecular Genetic ◽  
Molecular Genetic Analysis ◽  
Ultrastructural Organization ◽  
Apparent Lack ◽  
Myosin I ◽  
Family Analysis ◽  
Mutant Cells ◽  
Single Knockout

The protozoan myosin Is are widely expressed actin-based motors, yet their in vivo roles remain poorly understood. Molecular genetic studies have been carried out to determine their in vivo function in the simple eukaryote Dictyostelium, an organism that contains a family of four myosin Is. Here we report the characterization of myoC, a gene that encodes a fifth member of this family. Analysis of the deduced amino acid sequence reveals that the myoC gene encodes a myosin that is homologous to the well-described Acanthamoeba myosin Is as well as to Dictyostelium myoB and -D. The expression pattern of the myoC mRNA is similar to that of myoB and myoD, with a peak of expression at times of maximal cell migration, around 6 hours development. Deletion of the myoB gene has been previously shown to result in mutant cells that are defective in pseudopod extension and phagocytosis. However, no obvious differences in cell growth, development, phagocytosis or motility were detected in cells in which the myoC gene had been disrupted by homologous recombination. F-actin localization and ultrastructural organization also appeared unperturbed in myoC- cells. This apparent ‘lack’ of phenotype in a myosin I single knockout cannot be simply explained by redundancy of function. Our results rather suggest that the present means of assessing myosin I function in vivo are insufficient to identify the unique roles of these actin-based motors.

scholarly journals Direct p53 Transcriptional Repression: In Vivo Analysis of CCAAT-Containing G2/M Promoters

2005 ◽  
Vol 25 (9) ◽  
pp. 3737-3751 ◽  
Author(s):  
Carol Imbriano ◽  
Aymone Gurtner ◽  
Fabienne Cocchiarella ◽  
Silvia Di Agostino ◽  
Valentina Basile ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Transcriptional Repression ◽  
Histone Deacetylases ◽  
Specific Binding ◽  
Cell Cycle Control ◽  
Tetramerization Domain ◽  
Before And After

ABSTRACT In response to DNA damage, p53 activates G1/S blocking and apoptotic genes through sequence-specific binding. p53 also represses genes with no target site, such as those for Cdc2 and cyclin B, key regulators of the G2/M transition. Like most G2/M promoters, they rely on multiple CCAAT boxes activated by NF-Y, whose binding to DNA is temporally regulated during the cell cycle. NF-Y associates with p53 in vitro and in vivo through the αC helix of NF-YC (a subunit of NF-Y) and a region close to the tetramerization domain of p53. Chromatin immunoprecipitation experiments indicated that p53 is associated with cyclin B2, CDC25C, and Cdc2 promoters in vivo before and after DNA damage, requiring DNA-bound NF-Y. Following DNA damage, p53 is rapidly acetylated at K320 and K373 to K382, histones are deacetylated, and the release of PCAF and p300 correlates with the recruitment of histone deacetylases (HDACs)—HDAC1 before HDAC4 and HDAC5—and promoter repression. HDAC recruitment requires intact NF-Y binding sites. In transfection assays, PCAF represses cyclin B2, and a nonacetylated p53 mutant shows a complete loss of repression potential, despite its abilities to bind NF-Y and to be recruited on G2/M promoters. These data (i) detail a strategy of direct p53 repression through associations with multiple NF-Y trimers that is independent of sequence-specific binding of p53 and that requires C-terminal acetylation, (ii) suggest that p53 is a DNA damage sentinel of the G2/M transition, and (iii) delineate a new role for PCAF in cell cycle control.

scholarly journals The role of the polycomb complex in silencing α-globin gene expression in nonerythroid cells

Blood ◽  
2008 ◽  
Vol 112 (9) ◽  
pp. 3889-3899 ◽  
Author(s):  
David Garrick ◽  
Marco De Gobbi ◽  
Vasiliki Samara ◽  
Michelle Rugless ◽  
Michelle Holland ◽  
...  
Keyword(s):  
Histone Deacetylases ◽  
Globin Gene ◽  
Cpg Islands ◽  
Gene Activation ◽  
Globin Genes ◽  
Erythroid Cells ◽  
Polycomb Complex ◽  
Globin Gene Expression

Although much is known about globin gene activation in erythroid cells, relatively little is known about how these genes are silenced in nonerythroid tissues. Here we show that the human α- and β-globin genes are silenced by fundamentally different mechanisms. The α-genes, which are surrounded by widely expressed genes in a gene dense region of the genome, are silenced very early in development via recruitment of the Polycomb (PcG) complex. By contrast, the β-globin genes, which lie in a relatively gene-poor chromosomal region, are not bound by this complex in nonerythroid cells. The PcG complex seems to be recruited to the α-cluster by sequences within the CpG islands associated with their promoters; the β-globin promoters do not lie within such islands. Chromatin associated with the α-globin cluster is modified by histone methylation (H3K27me3), and silencing in vivo is mediated by the localized activity of histone deacetylases (HDACs). The repressive (PcG/HDAC) machinery is removed as hematopoietic progenitors differentiate to form erythroid cells. The α- and β-globin genes thus illustrate important, contrasting mechanisms by which cell-specific hematopoietic genes (and tissue-specific genes in general) may be silenced.

scholarly journals IL-1α is a DNA damage sensor linking genotoxic stress signaling to sterile inflammation and innate immunity

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Idan Cohen ◽  
Peleg Rider ◽  
Elena Vornov ◽  
Martin Tomas ◽  
Cicerone Tudor ◽  
...  
Keyword(s):  
Dna Damage ◽  
Histone Deacetylases ◽  
Genotoxic Stress ◽  
Sterile Inflammation ◽  
Surrounding Tissue ◽  
Cell Integrity ◽  
Environmental Signals ◽  
Damage Sensing ◽  
Novel Concept

Abstract Environmental signals can be translated into chromatin changes, which alter gene expression. Here we report a novel concept that cells can signal chromatin damage from the nucleus back to the surrounding tissue through the cytokine interleukin-1alpha (IL-1α). Thus, in addition to its role as a danger signal, which occurs when the cytokine is passively released by cell necrosis, IL-1α could directly sense DNA damage and act as signal for genotoxic stress without loss of cell integrity. Here we demonstrate localization of the cytokine to DNA-damage sites and its subsequent secretion. Interestingly, its nucleo-cytosolic shuttling after DNA damage sensing is regulated by histone deacetylases (HDAC) and IL-1α acetylation. To demonstrate the physiological significance of this newly discovered mechanism, we used IL-1α knockout mice and show that IL-1α signaling after UV skin irradiation and DNA damage is important for triggering a sterile inflammatory cascade in vivo that contributes to efficient tissue repair and wound healing.

scholarly journals Molecular Analysis of Drosophilaeyes absentMutants Reveals Features of the Conserved Eya Domain

Genetics ◽  
2000 ◽  
Vol 155 (2) ◽  
pp. 709-720 ◽  
Author(s):  
Quang T Bui ◽  
John E Zimmerman ◽  
Haixi Liu ◽  
Nancy M Bonini
Keyword(s):  
Protein Sequence ◽  
Molecular Genetic ◽  
Molecular Genetic Analysis ◽  
Special Importance ◽  
Missense Mutations ◽  
Eyes Absent ◽  
Sine Oculis ◽  
Eye Formation

AbstractThe eyes absent (eya) gene is critical to eye formation in Drosophila; upon loss of eya function, eye progenitor cells die by programmed cell death. Moreover, ectopic eya expression directs eye formation, and eya functionally synergizes in vivo and physically interacts in vitro with two other genes of eye development, sine oculis and dachshund. The Eya protein sequence, while highly conserved to vertebrates, is novel. To define amino acids critical to the function of the Eya protein, we have sequenced eya alleles. These mutations have revealed that loss of the entire Eya Domain is null for eya activity, but that alleles with truncations within the Eya Domain display partial function. We then extended the molecular genetic analysis to interactions within the Eya Domain. This analysis has revealed regions of special importance to interaction with Sine Oculis or Dachshund. Select eya missense mutations within the Eya Domain diminished the interactions with Sine Oculis or Dachshund. Taken together, these data suggest that the conserved Eya Domain is critical for eya activity and may have functional subregions within it.

Sign in / Sign up

Export Citation Format

Share Document