Cell Signaling Technology
XP Monoclonal Antibody

Product Pathways - Development

RBPSUH (D10A4) XP® Rabbit mAb #5313

Applications Reactivity Sensitivity MW (kDa) Isotype
W ChIP H M R Mk Endogenous 61 Rabbit IgG

Applications Key:  W=Western Blotting  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

RBPSUH (D10A4) XP® Rabbit mAb recognizes endogenous levels of total RBPSUH protein.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Gln110 of human RBPSUH protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using RBPSUH (D10A4) XP® Rabbit mAb.

Chromatin IP

Chromatin IP

CUTLL1 cells were cultured in media with γ-secretase inhibitor (1 μM, 3 d) and then either harvested immediately (left panel) or washed and cultured in fresh media for 3 h (right panel). Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 cells and 10 µl of RBPSUH (D10A4) XP® 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 HES1 promoter primers, SimpleChIP® Human HES4 Promoter Primers #7273, 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

RBPSUH (Recombining Binding Protein, SUppressor of Hairless), also termed RBP-J or CSL, is the DNA-binding component of the transcription complex regulated by canonical Notch signaling. In the absence of Notch activation, RBPSUH suppresses target gene expression through interactions with a co-repressor complex containing histone deacetylase. Upon activation of Notch receptors, the Notch intracellular domain (NICD) translocates to the nucleus and binds to RBPSUH. This displaces the co-repressor complex and replaces it with a transcription activation complex that includes Mastermind-like (MAML) proteins and histone acetylase p300, leading to transcriptional activation of Notch target genes (1-3). RBPSUH is also the DNA-binding partner for Epstein-Barr virus (EBV) nuclear antigen 2 (EBNA2), a protein critical for latent viral transcription and immortalization of EBV-infected B cells (4,5).

  1. Ehebauer, M. et al. (2006) Sci STKE 2006, cm7.
  2. Borggrefe, T. and Oswald, F. (2009) Cell Mol Life Sci 66, 1631-46.
  3. Kopan, R. and Ilagan, M.X. (2009) Cell 137, 216-33.
  4. Henkel, T. et al. (1994) Science 265, 92-5.
  5. Strobl, L.J. et al. (1997) Immunobiology 198, 299-306.

Application References

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Companion Products

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

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