Anti-dimethyl (Lys4) dimethyl (Lys9) Histone H3

Anti-dimethyl (Lys4) dimethyl (Lys9) Histone H3

This antibody only detects Histone H3 when dimethylated on both Lysine 4 and Lys9.

~17 kDa

Dimethylated Lys4 & Lys9

Synthetic peptide corresponding to residues surrounding and including dimethlyated Lys 4 and dimethlyated Lys9 of Histone H3.

Methylation

Histone H3 is one of the five main histone proteins involved in the structure of chromatin in eukaryotic cells. Featuring a main globular domain and a long N-terminal tail, H3 is involved with the structure of the nucleosomes of the 'beads on a string' structure. The N-terminal tail of histone H3 protrudes from the globular nucleosome core and can undergo several different types of epigenetic modifications that influence cellular processes. These modifications include the covalent attachment of methyl or acetyl groups to lysine and arginine amino acids and the phosphorylation of serine or threonine. Acetylation of histone H3 occurs at several different lysine positions in the histone tail and is performed by a family of enzymes known as Histone Acetyl Transferases (HATs). Acetylation of lysine14 is commonly seen in genes that are being actively transcribed into RNA.

• Human
• Vertebrates

Human tested. Expected to react with most other species based on extreme conserved homology among species.

Chromatin Immunoprecipitation:
An independent lab has shown that this antibody performs in chromatin immunoprecipitation (ChIP).

HeLa nuclear extract

Evaluated by Western Blotting on HeLa acid extracted nuclear preps (positive) and recombinant Histone H3 (negative).

Western Blotting Analysis:
1:500 dilution of this antibody was used to dimethyl (Lys4) dimethyl (Lys9) Histone H3 in acid extracted HeLa nuclear extracts.

Affinity Purfied

Affinity purified rabbit polyclonal antibody in buffer containing 0.1 M Tris-Glycine (pH7.4), 150 mM NaCl with 0.05% sodium azide.

Stable for 1 year at 2-8ºC from date of receipt.

P84243

NM_003493

• H3.3A
• H3.3B
• H3F3

• H3K4me2K9me2
• Histone H3 (di methyl K4, di methyl K9)
• H3 histone, family 3A

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• Chromatin Immunoprecipitation (ChIP)
• Western Blotting

Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Nucleosomes consist of approximately 146 bp of DNA wrapped around a histone octamer composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin fiber is further compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures. This gene is intronless and encodes a member of the histone H3 family. Transcripts from this gene lack polyA tails; instead, they contain a palindromic termination element. This gene is located separately from the other H3 genes that are in the histone gene cluster on chromosome 6p22-p21.3.

FUNCTION:Variant histone H3 which replaces conventional H3 in a wide range of nucleosomes in active genes. Constitutes the predominant form of histone H3 in non-dividing cells and is incorporated into chromatin independently of DNA synthesis. Deposited at sites of nucleosomal displacement throughout transcribed genes, suggesting that it represents an epigenetic imprint of transcriptionally active chromatin. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling. Ref.14 Ref.18 Ref.22

SUBUNIT STRUCTURE: The nucleosome is a histone octamer containing two molecules each of H2A, H2B, H3 and H4 assembled in one H3-H4 heterotetramer and two H2A-H2B heterodimers. The octamer wraps approximately 147 bp of DNA. Interacts with HIRA, a chaperone required for its incorporation into nucleosomes. Ref.14

SUBCELLULAR LOCATION: Nucleus.

DEVELOPMENTAL STAGE: Expressed throughout the cell cycle independently of DNA synthesis.

PTM: Acetylation is generally linked to gene activation. Acetylation on Lys-10 impairs methylation at Arg-9. Acetylation on Lys-19 and Lys-24 favors methylation at Arg-18.

Citrullination at Arg-9 and/or Arg-18 by PADI4 impairs methylation and represses transcription.

Asymmetric dimethylation at Arg-18 by CARM1 is linked to gene activation. Symmetric dimethylation at Arg-9 by PRMT5 is linked to gene repression.

Specifically enriched in modifications associated with active chromatin such as methylation at Lys-5, Lys-37 and Lys-80. Methylation at Lys-5 facilitates subsequent acetylation of H3 and H4. Methylation at Lys-80 is associated with DNA double-strand break (DSB) responses and is a specific target for TP53BP1. Methylation at Lys-10 and Lys-28, which are linked to gene repression, are underrepresented. Methylation at Lys-10 is a specific target for HP1 proteins (CBX1, CBX3 and CBX5) and prevents subsequent phosphorylation at Ser-11 and acetylation of H3 and H4. Methylation at Lys-5 and Lys-80 require preliminary monoubiquitination of H2B at 'Lys-120'. Methylation at Lys-10 and Lys-28 are enriched in inactive X chromosome chromatin.

Phosphorylated at Thr-4 by GSG2/haspin during prophase and dephosphorylated during anaphase. At centromeres, specifically phosphorylated at Thr-12 from prophase to early anaphase, probably DAPK3. Phosphorylation at 'Ser-11' by AURKB is crucial for chromosome condensation and cell-cycle progression during mitosis and meiosis. In addition phosphorylation at 'Ser-11' by RPS6KA4 and RPS6KA5 is important during interphase because it enables the transcription of genes following external stimulation, like mitogens, stress, growth factors or UV irradiation and result in the activation of genes, such as c-fos and c-jun. Phosphorylation at Ser-11, which is linked to gene activation, prevents methylation at Lys-10 but facilitates acetylation of H3 and H4. Phosphorylation at Ser-11 by AURKB mediates the dissociation of HP1 proteins (CBX1, CBX3 and CBX5) from heterochromatin. Phosphorylation at 'Ser-11' is also an essential regulatory mechanism for neoplastic cell transformation. Phosphorylated at Ser-29 by MLTK isoform 1, RPS6KA5 or AURKB during mitosis or upon ultraviolet B irradiation. Phosphorylation on Ser-32 is specific to regions bordering centromeres in metaphase chromosomes.

Ubiquitinated By similarity.

SEQUENCE SIMILARITY: Belongs to the histone H3 family.

SEQUENCE CAUTION:The sequence CAH73371.1 differs from that shown. Reason: Erroneous gene model prediction.

Histones are highly conserved proteins that serve as the structural scaffold for the organization of nuclear DNA into chromatin. The histones have an amino terminal tail, a globular domain, and a carboxy-terminal tail. The four core histones, H2A, H2B, H3 and H4 assemble into the octamer (2 molecules of each). Subsequently, 146 base pairs of DNA are wrapped in a sequence-independent manner around the octamer, forming the basic subunit of chromatin, the nucleosome. The distance between nucleosomes varies from species to species but generally is between 180 and 200 base pairs for higher organisms. Histone H1, the most common form of linker histone, binds to nucleosomal DNA at the point from which the DNA exits the nucleosome, and is required for higher order packing of chromatin.

Histones are modified post-translationally by the actions of enzymes in both the nucleus and cytoplasm that deposit specific functional groups. These modifications help to regulate the processes that depend on DNA, such as transcription, DNA repair, recombination and replication. The most commonly studied and best understood modifications are acetylation, phosphorylation, methylation, and to a lesser extent ubiquitination. The modifications occur predominantly on the amino terminal tails that extend out beyond the nucleosome core particle, but certain modifications have also been identified on the C-terminal tails and globular domains of some histones. Acetylation occurs on the epsilon amino group of lysine residues of all four core histones, and increases in acetylation in are correlated strongly with increases in gene expression.

Indeed, many histone acetyltransfease enzymes (HATs) form the catalytic subunits of transcriptional activating protein complexes. Histone deacetylases (HDACs) remove acetyl marks, antagonizing the activity of HATs and lead to decreases in transcription (see above). Phosphorylation occurs on serine residues in the amino termini of all four core histones and in multiple regions of H1. Phosphorylation of serines 10 and 28 of H3 occur during chromosome condensation in mitosis, and antibodies to these sites are excellent mitotic markers. H2B is phosphorylated at lysines 14 and 32 in cells undergoing apoptosis, and the histone variant H2A.X is phosphorylated at serine 139 in response to DNA damage. Histone methylation has recently become a popular research topic, and occurs on both lysine and arginine residues. Arginine methylation appears to be associated predominantly with transcriptional activation, whereas two specific lysine methylation events on histone H3 are hallmarks of either active chromatin (euchromatin) or silenced chromatin (heterochromatin). Ubiquitination is the least understood of the histone modifications and occurs on the C-terminal tails of H2A and H2B, and in some cases is a necessary precursor to specific histone methylation events.

Using a technique known as chromatin immunoprecipitation (ChIP), it is possible to analyze the variety of histone modifications present within a given promoter region or even an entire gene locus. Antibodies specific to the modification of interest are used to enrich for regions of chromatin (sheared to a manageable size and harvested from cells) that contain the modification, and various detection methods (Southern blot, PCR, microarray) are employed to detect specific DNA sequences within the enriched chromatin. This data is very useful in analyzing the involvement of a modification in specific biological processes.

Anti-dimethyl (Lys4) dimethyl (Lys9) Histone H3

1mg/mL

Polyclonal Antibody

100 µL

Affinity Purified

Rabbit