Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

CBP (D6C5) Rabbit mAb #7389

Applications Reactivity Sensitivity MW (kDa) Isotype
W IP IF-IC ChIP H M R Mk Endogenous 300 Rabbit IgG

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IF-IC=Immunofluorescence (Immunocytochemistry)  ChIP=Chromatin IP
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

CBP (D6C5) Rabbit mAb recognizes endogenous levels of total CBP protein. This antibody does not cross-react with p300 protein.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a recombinant protein specific to the amino terminus of human CBP protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using CBP (D6C5) Rabbit mAb.

IF-IC

IF-IC

Confocal immunofluorescent analysis of HeLa cells using CBP (D6C5) Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 293 cells, treated with Forskolin #3828 (30 μM, 1h) and either 20 μl of CBP (D6C5) Rabbit mAb 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 human ALS2 exon 1 primers, SimpleChIP® Human NR4A3 Promoter Primers #4829, 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

CBP (CREB-binding protein) and p300 are highly conserved and functionally related transcriptional co-activators that associate with transcriptional regulators and signaling molecules, integrating multiple signal transduction pathways with the transcriptional machinery (1,2). CBP/p300 also contain histone acetyltransferase (HAT) activity, allowing them to acetylate histones and other proteins (2). Phosphorylation of p300 at Ser89 by PKC represses its transciptional acitivity, and phosphorylation at the same site by AMPK disrupts the association of p300 with nuclear receptors (3,4). Ser1834 phosphorylation of p300 by Akt disrupts its association with C/EBPβ (5). Growth factors induce phosphorylation of CBP at Ser437, which is required for CBP recruitment to the transcription complex (6). CaM kinase IV phosphorylates CBP at Ser302, which is required for CBP-dependent transcriptional activation in the CNS (7). The role of acetylation of CBP/p300 is of particular interest (2,8). Acetylation of p300 at Lys1499 has been demonstrated to enhance its HAT activity and affect a wide variety of signaling events (9).

  1. Goodman, R.H. and Smolik, S. (2000) Genes Dev 14, 1553-77.
  2. Chan, H.M. and La Thangue, N.B. (2001) J. Cell Sci. 114, 2363-2373.
  3. Yuan, L.W. and Gambee, J.E. (2000) J. Biol. Chem. 275, 40946-40951.
  4. Yang, W. et al. (2001) J. Biol. Chem. 276, 38341-38344.
  5. Guo, S. et al. (2001) J. Biol. Chem. 276, 8516-8523.
  6. Zanger, K. et al. (2001) Mol. Cell 7, 551-558.
  7. Impey, S. et al. (2002) Neuron 34, 235-244.
  8. Yuan, L.W. and Giordano, A. (2002) Oncogene 21, 2253-2260.
  9. Thompson, P.R. et al. (2004) Nat. Struct. Mol. Biol. 11, 308-315.

Application References

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For Research Use Only. Not For Use In Diagnostic Procedures.

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