Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

Acetyl-Histone H4 (Lys5) Antibody #9672

Applications Reactivity Sensitivity MW (kDa) Source
W IP IHC-P IF-IC ChIP H M R Mk (C) (Dm) (X) (Z) (B) (Pg) (Hr) (Ce) Endogenous 11 Rabbit

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IF-IC=Immunofluorescence (Immunocytochemistry)  ChIP=Chromatin IP
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  C=Chicken  Dm=D. melanogaster  X=Xenopus  Z=Zebrafish  B=Bovine  Pg=Pig  Hr=Horse  Ce=C. elegans
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Acetyl-Histone H4 (Lys5) Antibody detects endogenous levels of histone H4 only when acetylated on Lys5. This antibody does not cross-react with histone H4 acetylated on lysines 8, 12, or 16.

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to the amino terminus of histone H4 in which Lys5 is acetylated. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Acetyl-Histone H4 (Lys5) Antibody.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded mouse kidney using Acetyl-Histone H4 (Lys5) Antibody.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Acetyl-Histone H4 (Lys5) Antibody.


IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human colon carcinoma using Acetyl-Histone H4 (Lys5) Antibody.

IF-IC

IF-IC

Immunofluorescent analysis of HeLa cells, untreated (left) or treated with Trichostatin A (TSA) #9950 (right), using Acetyl-Histone H4 (Lys5) Antibody (green) and Golgin-97 Antibody (red). Actin filaments were labeled with a dye conjugated phalloidin (pseudocolored blue).

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells and either 20 μl of Acetyl-Histone H4 (Lys5) Antibody or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human GAPDH Exon 1 Primers #5516, SimpleChIP® Human RPL30 Exon 3 Primers #7014, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.


Background

The nucleosome, made up of four core histone proteins (H2A, H2B, H3, and H4), is the primary building block of chromatin. Originally thought to function as a static scaffold for DNA packaging, histones have now been shown to be dynamic proteins, undergoing multiple types of post-translational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (1,2). Histone acetylation occurs mainly on the amino-terminal tail domains of histones H2A (Lys5), H2B (Lys5, 12, 15, and 20), H3 (Lys9, 14, 18, 23, 27, 36 and 56), and H4 (Lys5, 8, 12, and 16) and is important for the regulation of histone deposition, transcriptional activation, DNA replication, recombination, and DNA repair (1-3). Hyper-acetylation of the histone tails neutralizes the positive charge of these domains and is believed to weaken histone-DNA and nucleosome-nucleosome interactions, thereby destabilizing chromatin structure and increasing the accessibility of DNA to various DNA-binding proteins (4,5). In addition, acetylation of specific lysine residues creates docking sites for a protein module called the bromodomain, which binds to acetylated lysine residues (6). Many transcription and chromatin regulatory proteins contain bromodomains and may be recruited to gene promoters, in part, through binding of acetylated histone tails. Histone acetylation is mediated by histone acetyltransferases (HATs), such as CBP/p300, GCN5L2, PCAF, and Tip60, which are recruited to genes by DNA-bound protein factors to facilitate transcriptional activation (3). Deacetylation, which is mediated by histone deacetylases (HDAC and sirtuin proteins), reverses the effects of acetylation and generally facilitates transcriptional repression (7,8).

Histone H4 lysine 5 is acetylated by multiple HAT proteins. Acetylation by Esa1p in yeast, or Tip60 in mammalian cells, may contribute to both transcriptional activation and DNA repair, including non-homologous end joining and replication-coupled repair (9-12). Histone H4 lysine 5 is also acetylated by CBP/p300, a family of HAT proteins that function as transcriptional co-activators for a large number of transcription factors (13).

  1. Peterson, C.L. and Laniel, M.A. (2004) Curr Biol 14, R546-51.
  2. Jaskelioff, M. and Peterson, C.L. (2003) Nat Cell Biol 5, 395-9.
  3. Roth, S.Y. et al. (2001) Annu Rev Biochem 70, 81-120.
  4. Workman, J.L. and Kingston, R.E. (1998) Annu Rev Biochem 67, 545-79.
  5. Hansen, J.C. et al. (1998) Biochemistry 37, 17637-41.
  6. Yang, X.J. (2004) Bioessays 26, 1076-87.
  7. Haberland, M. et al. (2009) Nat Rev Genet 10, 32-42.
  8. Haigis, M.C. and Sinclair, D.A. (2010) Annu Rev Pathol 5, 253-95.
  9. Clarke, A.S. et al. (1999) Mol Cell Biol 19, 2515-26.
  10. Kimura, A. and Horikoshi, M. (1998) Genes Cells 3, 789-800.
  11. Bird, A.W. et al. (2002) Nature 419, 411-5.
  12. Ikura, T. et al. (2000) Cell 102, 463-73.
  13. Schiltz, R.L. et al. (1999) J Biol Chem 274, 1189-92.

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