histone modification in living cells
TRANSCRIPT
1Department of Plant Biotechnology03/11/2016
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Contents
Histones are a group of basic protein that associate with DNA and help the DNA to condense it into chromatin.
Histones contain a large proportion of the positively charged (basic) amino acids, lyseine and arginine in their structure.
DNA is negatively charged due to the phosphate groups on its backbone.
These result of these opposite charges is strong attraction and therefore high binding affinity between histones and DNA structure called nucleosome.
DNA wraps around histones, they also play a role in gene regulation.
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Luger et al., Nature 1997
Principles of Biochemistry, Lehninger 4th Edition.
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Introduction
Histones
The basic unit of chromatin is the nucleosome core particle, which contains 147 bp of DNA wrapped nearly twice around an octamer of the core histones.
Each nucleosome is separated by 10–60 bp of ‘linker’ DNA, and the resulting nucleosomal array constitutes a chromatin fiber of ~10 nm in diameter.
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Genome Organization
Mol. Bio of Gene, Watson et al., 7th edition
Luger et al., Nature 1997
Histone proteins are of two types:
Core Histones - H2A, H2B, H3, and H4 Linker Histones - H1
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The eight histones in the core are arranged into a (H3)2(H4)2 tetramer and a pair of H2A–H2B dimers.
The tetramer and dimers come together to form a left-handed superhelical ramp around which the DNA wraps.
Hydrogen bonds between the DNA backbone and the amide group on the main chain of histone proteins
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Types of Histones
Mol. Bio of Gene, Watson et al., 7th edition
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Histone H1 is not part of the nucleosome core particle. Instead, it binds to the linker DNA and is referred to as a linker histone.
H1 is half as abundant as the other histones, which is consistent with the finding that only one molecule of H1 can associate with a nucleosome.
Since H2A packages DNA molecules into chromatin, the packaging process will effect gene expression.
Refers to a variety of closely related proteins that vary often by only a few amino acids.
H2A plays a major role in determining the overall structure of chromatin. Inadvertently, H2A has been found to regulate gene expression.
H2B is also involved with the structure of the nucleosomes of the 'beads on a string' structure.
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H1
H2A
H2B
http://www.actrec.gov.in/histome/histone_main.php
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H4 is a structural component of the nucleosome, and is subject to covalent modification ,including acetylation and methylation, which may alter expression of genes located on DNA associated with its parent histone octamer.
Featuring a main globular domain and a long N-terminal tail.
Also an important protein in the emerging field of epigenetics, where its sequence variants and variable modification states are thought to play a role in the dynamic and long term regulation of genes.
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H4
H3
http://www.actrec.gov.in/histome/histone_main.php
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The core histones each have an amino-terminal extension, called a tail because it lacks a defined structure and is accessible within the intact nucleosome.
These tails are rich in number of lysine (K) and arginine (R) residues.
The C-terminal end is primarily responsible for histone-DNA and histone-histone interactions.
The N-terminal tails stand as targets of post-transational modifications (PTMs), which may modify the structure of chromatin play an essential role in regulating gene expression.
© J.H.Waterborg,1999
© Maacmillan pub.Luger et al., 199703/11/2016
(g/mol)
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N-terminal tails of histones are the most accessible regions of these peptide as they protrude from the nucleosome and possess no specific structure.
The major function of PTMs is to either create sites for the recruitment of specific factors or modify existing sites so as to abolish previous interactions.
Chromatin must be first made relaxed to allow access of cellular machineries to chromatin DNA.
The amino-terminal portion of the core histone proteins contains a flexible and highly basic tail region, which is conserved across various species and is subject to various PTM.
Chromatin can be highly packed or loosely packed, and correlated to the gene expression levels.
Post-translational modification (PTM) of histones is a crucial step in epigenetic regulation of a gene.
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N-termini of the core histones
©1999, J.H.Waterborg, UMKC
Zdenko Herceg1 and Rabih Murr1,2
Histone Modification
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Modifications in histone proteins affects the structure of chromatin.
Gene regulation
DNA damage and repair
Chromosome condensation
Heterochromatin
-Highly packed form- Low gene expression
Euchromatin
- Loosely packed form- High gene expression
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N-terminal tails of all histones are particularly of interest since they protrude out of the compact structure. These N-terminal tails are often subjected to a variety of post-translational modifications such as,
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Methyl Acetyl Phospho Ubiquitin SUMO
Bhaumik, Smith, and Shilatifard, 2007.
It has been proposed that these modifications result in a ‘code’ which can be read by proteins involved in gene expression and other DNA translations
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Types of Histone Modification
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Amino acid Residue
Modification Type
Modiying Enzyme
Lysine AcetylationDeacetylation
HAT HDAC
Lysine MethylationDemethylation
HMT HDM
Lysine UbiquitylationDeubiquitylation
Ub ligaseUb protease
Serine/ThreoninePhosphorylationDephosphorylation
KinasePhosphatase
Arginine MethylationDemethylation
PRMTDeiminase/Demethylase
Some Examples of Histone Modification and Modifiers
Others: Sumoylation (Lysine), ADP Ribosylation (Glutamate)
Post-translational Modification of Histone N-terminal Tails
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N-term tails reversible acetylated in Lys, particularly in H3+H4
While the globular core is involved in histone-histone packing and DNA-contact, the N-terminal tails point outwards and is available for interaction.
Acetylation also provides binding sites for a number of proteins with an approximately 100-amino-acid sequence motif called a bromodomain.
++++ +
Luger et al., 1997
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1. Acetylation
P.J. Barnes et al., Eur Respir J 2005
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Histone acetylation Histone deacetylation
Enzyme Histone acetyl transferases (HATs)
Histone deacetylases (HDACs)
Group Adds acetyl groups to histone tails
Removes acetyl groups from histone tails
Interacion with DNA
Reduces positive charge and weakens interaction of histones with DNA
Increases interaction of DNA and histones
©2006 Prous Science
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Acetylation Deacetylation
Role in Gene regulation
Allows transcription Repress transcription
Acetyl co.A
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Modification Histone Site Possible function
Acetylation H2A K4, K5,K7 Transcriptional activation
H2BK5, K11, K12, K15, K16 , K20 Transcriptional activation
H3 K4, K9, K23, K27 Transcriptional activation
K14Transcriptional activation, DNA repair, Transcription elongation
K18 Transcriptional activation, DNA repair
H4 K5, K12, K16 Transcriptional activation, DNA repair
K8 Transcriptional activation
Sites of histone acetylation with their function-
03/11/2016 P.J. Barnes et al., Eur Respir J 2005
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Role of Acetylation in Gene Expression
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It is the introduction of an Methyl functional group to only on Lysine or Arginine of the histone tail.
These reactions are catalyzed by enzymes with "histone methyltransferases (HMTs)”
Histone lysine methyl transferases (HKMTs) Methylate lysine (K) residues
Protein argenin methyl transferases (PRMTs) Methylate arginine (R) residues
A role in both activation and repression
Arginines can be mono- or dimethylated whereas lysines can be mono-, di- or trimethylated .
Arg
Lys
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2. Methylation
Eric J. Richards., et al., 2002
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Methylation can result in activation or repression of expression . Activation (H3K4, H3K36, H3K79) Trimethylation of histone H3 at lysine 4 (H3K4) is an universal active mark for transcription.
Repression (H3K9, H3K27, H4K20) Dimethylation of histone H3 at lysine 9 (H3K9) and at 27 (H3K27) are the universal signal for
transcriptional silencing.
Modification Histone Site Possible Function
Methylation H3 K4Permissive euchromatin (di-Me), Active euchromatin (tri-Me)
Transcriptional activation
K9Transcriptional silencing (tri-Me), DNA methylation (tri-Me), Heterochromatin formation
R17 Transcriptional activation
K27 Transcriptional silencing, X inactivation (tri-Me)
K36 Transcriptional elongation
K79 Euchromatin, Transcriptional elongation
H4 R3 Transcriptional activation
K20Transcriptional silencing (mono-Me), Heterochromatin (tri-Me).
K59 Transcriptional silencing
Sites of histone methylation with their function
03/11/2016http://www.actrec.gov.in/histome/histones.php?histone=H3
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Model for Role of Methylation in Heterochromatin Formation
Condensation assisted by recruitment of HMT (histone methyltransferase), where HP1 (heterochromatin protein 1) binds to H3K9-Me3 which methylates adjacent H3K9.
Chromatin condensed until a boundary element is reached and turned into heterochromatin
Methylation of histone tails long lasting compared to acetylation.
Can be Inherited by daughter cells: Responsible for X-inactivation in female.
Phillips, T. (2008) Nature Education
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Phosphorylation is the addition of a phosphate group (PO43−) to a molecule.
Phosphorylation is catalyzed by various specific protein kinases, whereas phosphatases mediate removal of the phosphate group.
Histones can also get phosphorylated and the most studied sites of histone phosphorylation are the serine 10 of histone H3 (H3S10) that is deposited by the Aurora-B kinase during mitosis.
Phosphorylation of histones, in particular phosphorylation of H2AX, has a role in DNA damage response and DNA repair.
Rapid phosphorylation of H2AX, at serine 129 (H2AX) by the PI3K kinases at double strand break (DSB) sites, is one of the first and most easily detectable DNA damage signaling post-translational events.
Role of histone phosphorylation in DNA repair
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3. Phosphorylation
Dorine Rossetto, et. al., Epigenetics 7:10 2012
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Modification Histone Site Possible Function
.Phosphorylation H2A S1
Mitosis, Transcriptional repression, Chromatin assembly
T119 MitosisS129 (S. cerevisiae) DNA repairS139 (mammalian H2AX) DNA repair
H2BS14 (vertebrates) Apoptosis
S33 (D. melanogaster) Transcriptional activation
H3 T3 Mitosis
S10Mitosis, meiosis, Transcriptional activation
T11 (mammals) MitosisS28 (mammals) Mitosis
H4 S1 Mitosis
Role of histone phosphorylation in transcription regulation
The phosphorylation of H3S10 (H3S10P) was initially linked to chromosome relaxation and segregation during mitosis and meiosis.
The role of H3S10P in chromatin condensation suggests that it should be involved in transcriptional activation.
Transient derepression by histone H3 phosphorylation. Mitogen-activated protein kinases (MAPK)
Signal molecule
absent
Signal molecule present
Sites of histone phosphorylation with their function
03/11/2016 Anna Sawicka et al., Biochimie 94 (2012)
Ubiquitination (or ubiquitylation) refers to the post-translational modification of the amino group of a lysine residue by the covalent attachment of one (monoubiquitination) or more (polyubiquitination) ubiquitin monomers.
Ubiquitin is a 76 amino acid protein highly conserved in eukaryotes.
Histone ubiquitination alters chromatin structure and allows the access of enzymes involved in transcription
Ubiquitination is carried out in three main steps: activation, conjugation, and ligation, performed by ubiquitin-activating enzymes (E1s), ubiquitin-conjugating enzymes (E2s), and ubiquitin ligases (E3s), respectively.
22Department of Plant BiotechnologyJian Cao et.al.,Frontiers in oncology
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4. Ubiquitination
Modification Histone Site Possible Function
Ubiquitylation H2AK119 (mammals) Spermatogenesis
H2BK120 (mammals) Meiosis
K123 (S. cerevisiae)
Transcriptional activation
Sites of histone ubiquitination with their function
Small Ubiquitin-like Modifier (or SUMO) proteins are a family of small proteins that are attached to and detached from other proteins in cell to modify their function.
Sumoylation consists in the addition of a “Small Ubiquitin-related MOdifier protein” (SUMO) of ~100 amino acids.
Histone sumoylation was first reported in 2003, when Shiio et al. found that H4 can be modified by SUMO and they suggested that this modification leads to the repression of transcriptional activity through the recruitment of HDACs and HP1 proteins
The putative sumoylation sites were identified as K6/7 and to a lesser extent K16/17 of H2B, K126 of H2A, and all four lysines in the N-terminal tail of H4.
Histone sumoylation has a role in transcription repression by opposing other active marks such as acetylation, methylation, ubiquitination etc.
23Department of Plant BiotechnologyShiio et. al., PNAS, 2003
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5. Sumoylation
http://www.pnas.org/content/100/23/13118/F1.expansion.html
ADP-ribosylation is the addition of an ADP-ribose moiety onto a protein using NAD+ as a substrate.
Mono(ADP-ribosyl)ation is mediated by ADP ribosyl transferases (ART) and the enzymes responsible for the PARation (Poly-ADP-ribosylation) are the poly(ADP-ribose) polymerases (PARPs).
PARP1 prefers to linker histone H1 while PARP2 prefers core histones.
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Zdenko Herceg & Rabih MurrSascha Beneke* www.frontiersin.org, 2012
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6. ADP-Ribosylation
FIGURE 1 | Poly(ADP-ribose) polymerase1 in DNA repair
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H1 to nucleosomes increases chromatin compaction, ADP-ribosylation of H1 is suggested to alter the chromatin structure and possibly the chromatin composition.
Messner and Michael , Trends in Cell Biology September 201103/11/2016
1
2
3
ADP-ribosylation cycle of histones in chromatin
Suggested biological consequences of mono- or poly-ADP-ribosylated histones in chromatin.
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The histone code is a hypothesis that the transcription of genetic information encoded in DNA is in part regulated by chemical modifications to histone proteins, primarily on their unstructured ends .
This hypothesis predicts that-
•Distinct modifications of the histone tails will induce interaction affinities for chromatin-associated proteins.
•Modifications may be interdependent generate various combinations
•Local concentrations and combinations of differently modified nucleosomes determine qualities of higher order chromatin.
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Histone code
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Readers-
Chromodomain•CH3(Methyl)- recognition domain•HP1 has a chromodomain•Targets to Me-lys or H3K9me•Promote packed “CLOSED” chromatin
Demethylation of Lys 9 in H3 tailfacilitates phosphorylation (P) of Ser 10Acetylation (Ac) of Lys 9 and 14leads to “OPEN” chromatin
BromodomainBinds to acetylated lysines “OPEN”
Wide range of histone modifications >>>
Writers: enzymes that add a mark
Readers: proteins that bind to and “interpret” the mark
Erasers: enzymes that remove a mark
Tarakhovsky, A., Nature Immunology, 2010.
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Reading / translating the histone code
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The core histones are among the most conserved eukaryotic proteins; therefore, the nucleosomes formed by these proteins are very similar in all eukaryotes. But there are numerous histone variants found in eukaryotic cells.
Such unorthodox histones can replace one of the four standard histones to form alternate nucleosomes and may serve to demarcate particular regions of chromosomes
Examples-
Moggs and Orphanides, oxicological Sciences, 2004.
Play role in DNA damage repair
When chromosomal DNA is broken, H2A.X located adjacent to the break is phosphorylated at a serine residue that is not present in H2A.
PhosphorylatedH2A.X is specifically recognized by DNA repair enzymes leading to their localization at the site of DNA damage.
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Histone Varients
1) H2A.X is a variant of H2A
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In the centromeric region of chromosome , CENP-A replaces the histone H3 subunits in nucleosomes.
These nucleosomes are incorporated into the kinetochore that mediates attachment of the chromosome to the mitotic spindle core structure of the nucleosome.
Consistent with this interaction being critical for kinetochore formation, loss of CENP-A interferes with the association of kinetochore components with centromeric DNA.
Alteration of chromatin by incorporation of histone variants. incorporation of CENP-A in place of histone H3 is proposed to act as a binding site for one or more protein components of the kinetochore. Mol. Bio of Gene, Watson et al., 7th edition
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2) CENP-A varient histone H3
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The decreased HAT activity of CBP is a key contributor to the RSTS phenotype.
Features- Broad thumbs and toes, facial abnormalities, congenital heart defects, and increased risk of tumor formation.
Frequency- This condition is uncommon; it occurs in an estimated 1 in 100,000 to 125,000 newborns.
Additional features of the disorder can include eye abnormalities, heart and kidney defects, dental problems, and obesity.
Infants born with this severe form of the disorder usually survive only into early childhood.
RSTS - Facial features (A), left hand and feet showing broad thumb and big toes (B, C) and X-ray of both hands showing short broad thumbs (D). (Limb Malformations & Skeletal Dysplasia)
Huda Y. Zoghbi and Arthur L. Beaudet Cold Spring Harbor Laboratory Press, 2016
Mutations in the CREBBP gene, the gene provides instructions for making CREB binding protein, plays an essential role in controlling cell growth and division and prompting cells to mature
This mutation abolishes the histone acetyltransferase (HAT) activity of CBEP (Murata et al. 2001).
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Diseses associated with histone modification
1. Rubinstein–Taybi Syndrome (RSTS)
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The syndrome is caused by mutations in the RPS6KA3 gene (histone phosphorylation) and is located on the short arm of the X chromosome. Males are usually more severely affected than females.
Ribosomal protein S6 kinase, 90kDa, polypeptide 3, also known as RPS6KA3, is an enzyme that in humans is encoded by the RPS6KA3 gene.
The protein RSK2 which is encoded by the RPS6KA3 gene is a kinase which phosphorylates some substrates like CREB and histone H3.
A rare genetic disorder characterized by mental retardation and abnormalities of the head and facial and other areas.
Cardiac abnormalities affect 15% of the patients.
Coffin-Lowry syndrome 03/11/2016
2. Coffin-Lowry syndrome
CLSF- Coffin-Lowry Syndrome Foundation
Created in 1991 by Mary Hoffman in France.
CLS a general forum in which to exchange information, ideas and advice and a great resource for families affected by CLS.
Huda Y. Zoghbi and Arthur L. Beaudet Cold Spring Harbor Laboratory Press, 2016
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Techniques to Study Histone Modification
2) Mass Spectrometry (MS)
Other genome-wide techniques combined with ChIPs
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ChIP is a technique whereby a protein of interest is selectively immunoprecipitated from a chromatin preparation to determine the DNA sequences associated with it.
Chromatin immunoprecipitation (ChIP) has become the technique of choice to investigate protein–DNA interactions inside the cell.
ChIP has been used for mapping the localization of post-translationally modified histones and histone variants in the genome, and for mapping DNA target sites for transcription factors and other chromosome-associated proteins.
There are mainly two types of ChIP, primarily differing in the starting chromatin preparation.
Philippe Collas, Mol Biotechnol (2010) 45:87–100Thomas A. Methods in Molecular Biology, vol. 538
1. Cross-linked ChIP (XChIP)
Uses reversibly cross-linked chromatin as a starting material then sheared by sonication.
Mainly suited for mapping the DNA target of transcription factors or other chromatin-associated proteins,
2. Native ChIP (NChIP)
Chromatin sheared by micrococcal nuclease digestion.
Mainly suited for mapping the DNA
target of histone modifiers.
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1. Chromatin Immunoprecipitation (ChIP)
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1. DNA and associated proteins reversibly cross-linked with formaldehyde followed by cell lysis.
2. The DNA-protein complexes (chromatin-protein) are then sheared into ~500 bp DNA fragments by sonication or nuclease digestion.
3. Immunoprecipitation (IP) by using an appropriate protein-specific antibody.
4. Reverse cross linking and sequence determination
5. Enrichment of specific DNA sequences represents regions on the genome that the protein of interest is associated with in vivo.
Chromatin immunoprecipitation assay (ChIP) and various methods of analysis
Philippe Collas, Mol Biotechnol (2010) 45:87–10003/11/2016
Major Steps Involved in ChIP Assay
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Chromatin Immunoprecipitation (ChIP) – Seq Assay
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ChIP-on-chip (also known as ChIP-chip) is a technology that combines chromatin immunoprecipitaion ('ChIP') with DNA microarray ("chip").
Development of genomic DNA microarrays (chips) has, when combined with ChIP assays, enabled the mapping of transcription factor binding sites and of histone modifications.
The most prominent representatives of this class are transcription factors, replication -related proteins, like Origin Recognition Complex Protein (ORC), histones, their variants, and histone modifications.
Philippe Collas, Mol Biotechnol (2010) 45:87–10003/11/2016
ii) ChIP–on-chip
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Chromatin immunoprecipitation followed by sequencing (ChIP–seq) is a technique for genome-wide profiling of DNA-binding proteins, histone modifications or nucleosomes.
1. Crosslinking
2. Chromatin is sheared by sonication (200–600 bp )
3. Immunoprecipitation
4. Reverse crosslinking DNA is assayed to determine the sequences bound by the protein.
5. Construction of a sequencing library,
6. Sequencing of DNA.
Overview of a ChiP–seq experiment.NATuRe RevIewS | GeneticsPeter J. Park 200903/11/2016
ii) ChIP–seq
Major Steps
In ChIP–seq, the DNA fragments of interest are sequenced directly instead of being hybridized on an array.
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Illumina Sequencing
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Chromatin Immunoprecipitation (ChIP) coupled with quantitative PCR can be used to investigate protein-DNA interaction at known genomic binding sites.
This technique is now used in a variety of life science disciplines including cellular differentiation, tumor suppressor gene silencing, and the effect of histone modifications on gene expression.
1. Cell fixation (cross-linking)
2. Chromatin shearing: by Sonication (100-500 bp)
3. Chromatin IP: Using specific ChIP-grade antibodies
4. Reverse cross-linking and DNA purification.
5. qPCR and analysis
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Major Steps
iii) ChIP-qPCR
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Revolution in Immunoassays
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Mass Spectrometry is an analytic technique that utilizes the degree of deflection of charged particles by a magnetic field to find the relative masses of molecular ions and fragments.
A mass spectrum is a plot of the ion signal as a function of the mass-to-charge ratio (m/z)
The two primary methods for ionization of whole proteins,
i). Electrospray ionization(ESI)
ii). Matrix-assisted laser desorption/ionization (MALDI).
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2. Mass Spectrometry (MS)
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1.Extraction of histones.
2. Further separation by SDS-PAGE
3. Protease digestion
4. Trypsin is used to cleave proteins
5. Reversed phase chromatography.
6. Mass spectrometer analysis
Quantification and identification of the example peptide H3 27-40 by mass spectrometry
A. The areas of peptides with different modification states are highlighted in grey. The presence of fragmentation spectra for the monomethylated peptide is also displayed.
B. MS/MS spectrum -fragment ions b2+ and b3+ clearly demonstrate methylation in K27 while y6+-y9+ show only propionylation in positions K36 and K37.
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Major Steps
Ignasi Forné et.al., CIPSM,2012
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Recent Review on Histone Modification
A genetically encoded probe for live-cell imaging of H4K20 monomethylation
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H4K20me1-Mintbody : A new probe to track histone modifications in living cells
Modification specific intracellular antibody
Genetically encoded single chain variable fragment of antibody (scFv) tagged with green fluorescent protein (GFP) recognizes and binds to histone H4 post-translationally modified at K20 position (H4K20me1).
Scientists at Tokyo Institute of Technology have developed a sensitive fluorescent antibody probe(Mintbody) that specifically detects monomethylation of lysine 20 in histone H4 in living cells.
To elucidate the relationship between histone modifications and cellular functions, it is important to monitor the dynamics of modifications in single living cells.
© 2016 The Author(s). Published by Elsevier Ltd.
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Mintbody
Two mintbodies have been developed to date1.H3K9Ac2.H3K20me1
Yuko Sato, et. al., JMB, 2016
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(a) Schematic structure of H4K20-mintbody.
(b)The localization of H4K20me1-mintbodies in living cells. H4K20me1-mintbody was concentrated in nuclei during interphase and bound to chromosomes during mitosis.
VH- Blue colour, LH- Red colour, The amino acids that differ in 12C8 are indicated in yellow. .Space filling models of two important residues (I95 and A99; letters shown in red) are blown-up to the side. The oxygen and nitrogen atoms arecolored in red and dark blue. Bars, 10 μm.
(c) The crystal structure of 15F11-scFv at 1.94 Å resolution.
Yuko Sato, et. al., JMB, 2016
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Conventional techniques used to study regulation by histone modifications are limited to fixed (dead) cells, thus preventing assessment of histone modification in single, living cells. e.g. ChIP-chip, ChIP-seq , ChIP-qPCR etc.
The specificity of the H4K20me1-mintbody in living cells was verified using yeast mutants and mammaliancells in which this target modification was diminished.
In a roundworm Caenorhabditis elegans model, the H4K20me1-mintbody could be used to monitor changes in H4K20me1 over the cell cycle and localization of dosage-compensated X chromosomes without disrupting cell function.
Expression of the H4K20me1-mintbody allowed us to monitor the oscillation of H4K20me1 levels during the cell cycle.
This research also identified key amino acids responsible for H4K20me1-mintbody conformational stability, solubility, and consequently, functional performance using X-ray crystallography and genetic analyses.
This research has future implications and can be used to monitor the dynamics of histone modifications and genome integrity in single living cells without disturbing cellular functions.
In the future, development of additional mintbodies specific to diverse post-translational histone modifications will facilitate the identification of regulatory mechanisms that control epigenetic modifications.
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Importance
Yuko Sato, et. al., JMB, 2016
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Histone proteins are most important for packaging and ordering of DNA. N-terminal tails of histones are the most accessible for post translational modificatins. Histone tail modification participate in regulation of many processes including,
– Transcription activation– Transcription repression– DNA damage and repair– Chromatin assembly – Domain bindings– Cell cycle – Long-range packaging (heterochromatin formation, silencing)– Chromosome condensation
Post-translational modification (PTM) of histones play an impotanat role in epigenetic regulation of a gene.
Strategy for localizing histone marks- ChIP-chip, ChIP –seq., ChIP-qPCR, Mass Spectromery etc.
The H4K20me1-Mintbody could be used to monitor changes in H4K20me1 in living cells.
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Conclusion
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References
Sato, Y., Kujirai, T., Arai, R., Asakawa, H., Ohtsuki, C., Horikoshi, N., Yamagata, K., Ueda, J., Nagase, T., Haraguchi, T., Hiraoka, Y., Kimura, A., Kurumizaka, H. & Kimura, H., A genetically encoded probe for livecell imaging of H4K20 monomethylation, Journal of Molecular Biology (2016).
Mechanisms of Histone Modifications, Zdenko Herceg and Rabih Murr (download.bioon.com.cn/upload/201105/30165853_5528.pdf )
Shelley L Berger , Histone modifications in transcriptional regulation. Current Opinion in Genetics & Development 2002, 12:142–148.
Sascha Beneke Regulation of chromatin structure by poly(ADP-ribosyl)ation . www.frontiersin.org REVIEW ARTICLE September2012|Volume3|Article169 |
Peter J. Park ChIP–seq: advantages and challenges of a maturing technologyNATUREReviews | Genetics vOlume 10 | OCTOBER 2009 |
Anna Sawicka, Christian Seiser. Histone H3 phosphorylation e A versatile chromatin modification for different occasions. Biochimie 94 (2012) 2193e2201
Eric J. Richards and Sarah C.R. Elgin Epigenetic Codes for Review Heterochromatin Formation and Silencing: Rounding up the Usual Suspects. Cell, Vol. 108, 489–500, February 22, 2002,
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EpiTect® ChIP PCR System For reliable analysis of protein—DNA interactions (www.sabiosciences.com/manuals/1073038_PP_GEF_Chip_0712_lr.pdf )
Huda Y. Zoghbi1,2 and Arthur L. Beaudet. Epigenetics and Human Disease Published by Cold Spring Harbor Laboratory Press at University of Hong Kong Libraries on February 1, 2016
Vikki M. Weake and Jerry L. Workman, Histone Ubiquitination: Triggering Gene Activity , Molecular Cell 29, March 28, 2008
Mol Biology of the Gene by Watson 7th Edition
Principles of Biochemistry by Lehninger 4th Edition
https://en.wikipedia.org/wiki/Histone http://www.abcam.com/epigenetics/histone-modifications-a-guide https://www.youtube.com/watch?v=fCd6B5HRaZ8 https://www.youtube.com/watch?v=4oFdS9EN9Pk https://www.youtube.com/watch?v=eYrQ0EhVCYA
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