scholarly journals Long-Range Nucleosome Ordering Is Associated with Gene Silencing in Drosophila melanogaster Pericentric Heterochromatin

2001 ◽  
Vol 21 (8) ◽  
pp. 2867-2879 ◽  
Author(s):  
Fang-Lin Sun ◽  
Matthew H. Cuaycong ◽  
Sarah C. R. Elgin
Keyword(s):  
Drosophila Melanogaster ◽  
Gene Silencing ◽  
Long Range ◽  
Chromatin Structure ◽  
Regulatory Region ◽  
Pericentric Heterochromatin ◽  
Dnase I ◽  
Micrococcal Nuclease ◽  
Hypersensitive Sites ◽  
Nucleosome Array

ABSTRACT We have used line HS-2 of Drosophila melanogaster, carrying a silenced transgene in the pericentric heterochromatin, to investigate in detail the chromatin structure imposed by this environment. Digestion of the chromatin with micrococcal nuclease (MNase) shows a nucleosome array with extensive long-range order, indicating regular spacing, and with well-defined MNase cleavage fragments, indicating a smaller MNase target in the linker region. The repeating unit is ca. 10 bp larger than that observed for bulkDrosophila chromatin. The silenced transgene shows both a loss of DNase I-hypersensitive sites and decreased sensitivity to DNase I digestion within an array of nucleosomes lacking such sites; within such an array, sensitivity to digestion by MNase is unchanged. The ordered nucleosome array extends across the regulatory region of the transgene, a shift that could explain the loss of transgene expression in heterochromatin. Highly regular nucleosome arrays are observed over several endogenous heterochromatic sequences, indicating that this is a general feature of heterochromatin. However, genes normally active within heterochromatin (rolled and light) do not show this pattern, suggesting that the altered chromatin structure observed is associated with regions that are silent, rather than being a property of the domain as a whole. The results indicate that long-range nucleosomal ordering is linked with the heterochromatic packaging that imposes gene silencing.

Chromatin structure of a P-element-transduced hsp-28 gene in Drosophila melanogaster

1986 ◽  
Vol 6 (11) ◽  
pp. 4126-4129
Author(s):  
J C Eissenberg ◽  
S C Elgin
Keyword(s):  
Gene Expression ◽  
Drosophila Melanogaster ◽  
Chromatin Structure ◽  
Dnase I ◽  
P Element ◽  
Direct Selection ◽  
Wild Type ◽  
Dnase I Hypersensitive Sites ◽  
Hypersensitive Sites ◽  
Selection For

The Drosophila hsp-28 gene was heat inducible when transduced to novel chromosomal sites even when no direct selection for transduced gene expression was imposed. The pattern of DNase I-hypersensitive sites 5' to the wild type and transduced copy of hsp-28 was similar. In addition, DNase I-hypersensitive sites occurred within the P-element sequences flanking transduced loci.

scholarly journals GAGA Factor and the TFIID Complex Collaborate in Generating an Open Chromatin Structure at the Drosophila melanogaster hsp26 Promoter

2002 ◽  
Vol 22 (17) ◽  
pp. 6148-6157 ◽  
Author(s):  
Boris A. Leibovitch ◽  
Quinn Lu ◽  
Lawrence R. Benjamin ◽  
Yingyun Liu ◽  
David S. Gilmour ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Heat Shock ◽  
Chromatin Structure ◽  
Random Sequence ◽  
Regulatory Region ◽  
Open Chromatin ◽  
Gaga Factor ◽  
Factor Binding ◽  
Hypersensitive Sites ◽  
Tfiid Complex

ABSTRACT The upstream regulatory region of the Drosophila melanogaster hsp26 gene includes two DNase I-hypersensitive sites (DH sites) that encompass the critical heat shock elements. This chromatin structure is required for heat shock-inducible expression and depends on two (CT) n •(GA) n elements bound by GAGA factor. To determine whether GAGA factor alone is sufficient to drive formation of the DH sites, we have created flies with an hsp26/lacZ transgene wherein the entire DNA segment known to interact with the TFIID complex has been replaced by a random sequence. The replacement results in a loss of heat shock-inducible hsp26 expression and drastically diminishes nuclease accessibility in the chromatin of the regulatory region. Chromatin immunoprecipitation experiments show that the decrease in TFIID binding does not reduce GAGA factor binding. In contrast, the loss of GAGA factor binding resulting from (CT) n mutations decreases TFIID binding. These data suggest that both GAGA factor and TFIID are necessary for formation of the appropriate chromatin structure at the hsp26 promoter and predict a regulatory mechanism in which GAGA factor binding precedes and contributes to the recruitment of TFIID.

scholarly journals Chromatin structure of a P-element-transduced hsp-28 gene in Drosophila melanogaster.

1986 ◽  
Vol 6 (11) ◽  
pp. 4126-4129 ◽  
Author(s):  
J C Eissenberg ◽  
S C Elgin
Keyword(s):  
Gene Expression ◽  
Drosophila Melanogaster ◽  
Chromatin Structure ◽  
Dnase I ◽  
P Element ◽  
Direct Selection ◽  
Wild Type ◽  
Dnase I Hypersensitive Sites ◽  
Hypersensitive Sites ◽  
Selection For

The Drosophila hsp-28 gene was heat inducible when transduced to novel chromosomal sites even when no direct selection for transduced gene expression was imposed. The pattern of DNase I-hypersensitive sites 5' to the wild type and transduced copy of hsp-28 was similar. In addition, DNase I-hypersensitive sites occurred within the P-element sequences flanking transduced loci.

Major histocompatibility complex class I genes in murine fibrosarcoma IC9 are down regulated at the level of the chromatin structure

1989 ◽  
Vol 9 (7) ◽  
pp. 3136-3142
Author(s):  
U Maschek ◽  
W Pülm ◽  
S Segal ◽  
G J Hämmerling
Keyword(s):  
Major Histocompatibility Complex ◽  
Major Histocompatibility Complex Class ◽  
Chromatin Structure ◽  
Dnase I ◽  
Class I ◽  
Micrococcal Nuclease ◽  
Complex Class ◽  
Major Histocompatibility ◽  
Histocompatibility Complex ◽  
Nuclear Extracts

The fibrosarcoma IC9 is deficient in the expression of the major histocompatibility complex class I genes Kb, Kk, and Dk and expresses only the Db molecule. Because class I deficiency may enable tumor cells to escape the immune response by cytotoxic T lymphocytes, we investigated why the class I genes are not expressed. Expression of the silent class I genes could not be induced, but all known DNA-binding factors specific for class I genes could be detected in nuclear extracts of IC9 cells. After cloning of the silent Kb gene from the IC9 cells and subsequent transfection of this cloned Kb gene into LTK- and IC9 cells, normal Kb antigens were expressed on the cell surface of both cell lines. Digestion of the chromatin of IC9 cells with micrococcal nuclease and DNase I showed a decreased nuclease sensitivity of the silent class I genes in comparison with active genes and the absence of DNase I hypersensitive sites in the promoter region of the silent Dk gene. These findings demonstrate that class I expression is turned off by a cis-acting regulatory mechanism at the level of the chromatin structure.

The chromatin structure of Saccharomyces cerevisiae autonomously replicating sequences changes during the cell division cycle

1991 ◽  
Vol 11 (10) ◽  
pp. 5301-5311
Author(s):  
J A Brown ◽  
S G Holmes ◽  
M M Smith
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Division ◽  
Chromatin Structure ◽  
Dnase I ◽  
S Phase ◽  
Cell Division Cycle ◽  
Characteristic Change ◽  
The Core ◽  
Dnase I Hypersensitive Sites ◽  
Hypersensitive Sites

The chromatin structures of two well-characterized autonomously replicating sequence (ARS) elements were examined at their chromosomal sites during the cell division cycle in Saccharomyces cerevisiae. The H4 ARS is located near one of the duplicate nonallelic histone H4 genes, while ARS1 is present near the TRP1 gene. Cells blocked in G1 either by alpha-factor arrest or by nitrogen starvation had two DNase I-hypersensitive sites of about equal intensity in the ARS element. This pattern of DNase I-hypersensitive sites was altered in synchronous cultures allowed to proceed into S phase. In addition to a general increase in DNase I sensitivity around the core consensus sequence, the DNase I-hypersensitive site closest to the core consensus became more nuclease sensitive than the distal site. This change in chromatin structure was restricted to the ARS region and depended on replication since cdc7 cells blocked near the time of replication initiation did not undergo the transition. Subsequent release of arrested cdc7 cells restored entry into S phase and was accompanied by the characteristic change in ARS chromatin structure.

scholarly journals Formation of stable chromatin structures on the histone H4 gene during differentiation in Tetrahymena thermophila.

1986 ◽  
Vol 6 (8) ◽  
pp. 3014-3017 ◽  
Author(s):  
D S Pederson ◽  
K Shupe ◽  
G A Bannon ◽  
M A Gorovsky
Keyword(s):  
Chromatin Structure ◽  
Transcriptional Control ◽  
Tetrahymena Thermophila ◽  
Micrococcal Nuclease ◽  
Histone H4 ◽  
Hypersensitive Sites ◽  
Nuclear Differentiation ◽  
Histone Gene Transcription ◽  
The Relationship ◽  
I Gene

The relationship between chromatin structure and the transcriptional activity of the histone H4-I gene of Tetrahymena thermophila was explored. Indirect end-labeling studies demonstrated that major DNase I- and micrococcal nuclease-hypersensitive sites flank the active macronuclear genes but not the inactive micronuclear genes. Runon transcription experiments with isolated macronuclei indicated that histone gene transcription rates decreased when cells were starved. However, macronuclear nuclease-hypersensitive sites persisted upon starvation. Thus, one level of transcriptional control of the H4-I gene results in altered chromatin structure and is established during nuclear differentiation. The rate of transcription is also controlled, but not through hypersensitive site-associated structures.

Upstream activation sequence-dependent alteration of chromatin structure and transcription activation of the yeast GAL1-GAL10 genes

1989 ◽  
Vol 9 (4) ◽  
pp. 1721-1732
Author(s):  
M J Fedor ◽  
R D Kornberg
Keyword(s):  
Chromatin Structure ◽  
Promoter Region ◽  
Protein A ◽  
Transcription Activation ◽  
Micrococcal Nuclease ◽  
Promoter Sequences ◽  
Upstream Activation Sequence ◽  
Protein Particles ◽  
Nucleosome Array ◽  
Activation Sequence

Conversion of the positioned nucleosome array characteristic of the repressed GAL1-GAL10 promoter region to the more accessible conformation of the induced state was found to depend on the upstream activation sequence, GAL4 protein, a positive regulator of transcription, and galactose, the inducing agent. The effect of the GAL4 protein-upstream activation sequence complex on the structure of adjacent chromatin required no other promoter sequences. Although sequences protected by histones in the repressed state became more accessible to micrococcal nuclease and (methidiumpropyl-EDTA)iron(II) cleavage following induction of transcription, DNA-protein particles containing these sequences retained the electrophoretic mobility of nucleosomes, indicating that the promoter region can be associated with nucleosomes under conditions of transcription activation.

scholarly journals DNase I- and micrococcal nuclease-hypersensitive sites in the human apolipoprotein B gene are tissue specific.

1988 ◽  
Vol 8 (1) ◽  
pp. 71-80 ◽  
Author(s):  
B Levy-Wilson ◽  
C Fortier ◽  
B D Blackhart ◽  
B J McCarthy
Keyword(s):  
Hela Cells ◽  
Apolipoprotein B ◽  
Hepg2 Cells ◽  
Transcriptional Start Site ◽  
Dnase I ◽  
Micrococcal Nuclease ◽  
Apo B ◽  
Human Apolipoprotein ◽  
Hypersensitive Sites ◽  
High Degree

We have mapped the DNase I- and micrococcal nuclease-hypersensitive sites present in the 5' end of the human apolipoprotein B (apo-B) gene in nuclei from cells expressing or not expressing the gene. Four DNase I-hypersensitive sites were found in nuclei from liver-derived HepG2 cells and intestine-derived CaCo-2 cells, which express the apo-B gene, but not in HeLa cells, which do not. These sites are located near positions -120, -440, -700, and +760 base pairs relative to the transcriptional start site. Undifferentiated CaCo-2 cells exhibited another site, near position -540. Six micrococcal nuclease-hypersensitive sites were found in nuclei from HepG2 and CaCo-2 cells, but not in HeLa cells or free DNA. These sites are located near positions -120, -390, -530, -700, -850, and +210. HepG2 cells exhibited another site, near position +460. Comparison of the DNA sequence of the 5' flanking regions of the human and mouse apo-B genes revealed a high degree of evolutionary conservation of short stretches of sequences in the immediate vicinity of each of the DNase I- and most of the micrococcal nuclease-hypersensitive sites.

scholarly journals The Forkhead Transcription Factor FoxI1 Remains Bound to Condensed Mitotic Chromosomes and Stably Remodels Chromatin Structure

2006 ◽  
Vol 26 (1) ◽  
pp. 155-168 ◽  
Author(s):  
Jizhou Yan ◽  
Lisha Xu ◽  
Gregory Crawford ◽  
Zenfeng Wang ◽  
Shawn M. Burgess
Keyword(s):  
Chromatin Structure ◽  
Fluorescent Protein ◽  
Living Cells ◽  
Dnase I ◽  
Higher Order ◽  
Micrococcal Nuclease ◽  
Stable Cell Line ◽  
Order Structure ◽  
Dna Fragments ◽  
Mitotic Chromosomes

ABSTRACT All forkhead (Fox) proteins contain a highly conserved DNA binding domain whose structure is remarkably similar to the winged-helix structures of histones H1 and H5. Little is known about Fox protein binding in the context of higher-order chromatin structure in living cells. We created a stable cell line expressing FoxI1-green fluorescent protein (GFP) or FoxI1-V5 fusion proteins under control of the reverse tetracycline-controlled transactivator doxycycline inducible system and found that unlike most transcription factors, FoxI1 remains bound to the condensed chromosomes during mitosis. To isolate DNA fragments directly bound by the FoxI1 protein within living cells, we performed chromatin immunoprecipitation assays (ChIPs) with antibodies to either enhanced GFP or the V5 epitope and subcloned the FoxI1-enriched DNA fragments. Sequence analyses indicated that 88% (106/121) of ChIP sequences contain the consensus binding sites for all Fox proteins. Testing ChIP sequences with a quantitative DNase I hypersensitivity assay showed that FoxI1 created stable DNase I sensitivity changes in condensed chromosomes. The majority of ChIP targets and random targets increased in resistance to DNase I in FoxI1-expressing cells, but a small number of targets became more accessible to DNase I. Consistently, the accessibility of micrococcal nuclease to chromatin was generally inhibited. Micrococcal nuclease partial digestion generated a ladder in which all oligonucleosomes were slightly longer than those observed with the controls. On the basis of these findings, we propose that FoxI1 is capable of remodeling chromatin higher-order structure and can stably create site-specific changes in chromatin to either stably create or remove DNase I hypersensitive sites.

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