Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

SPT16 (D7I2K) Rabbit mAb #12191

Applications Reactivity Sensitivity MW (kDa) Isotype
W ChIP H M R Mk (Hm) (X) (Z) (B) (Dg) (GP) (Hr) Endogenous 140 Rabbit IgG

Applications Key:  W=Western Blotting  ChIP=Chromatin IP
Reactivity Key:  H=Human  M=Mouse  R=Rat  Hm=Hamster  Mk=Monkey  X=Xenopus  Z=Zebrafish  B=Bovine  Dg=Dog  GP=Guinea Pig  Hr=Horse
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

SPT16 (D7I2K) Rabbit mAb recognizes endogenous levels of total SPT16 protein.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu662 of human SPT16 protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using SPT16 (D7I2K) Rabbit mAb.

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HCT 116 cells starved for 48 hr then serum stimulated with 10% FBS for 15 min and either 10 μl of SPT16 (D7I2K) 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 SimpleChIP® Human c-Fos Promoter Primers #4663, SimpleChIP® Human c-Fos Exon 3 Primers #12010, SimpleChIP® Human EGR1 Promoter Primers #5549, and SimpleChIP® Human EGR1 Intron 3 Primers #11953. 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

Supressor of Ty-16 (SPT16) and structure-specific recognition protein-1 (SSRP1) are subunits of the facilitates chromatin transcription (FACT) complex that is essential for transcription elongation (1,2). FACT facilitates RNA polymerase-dependent transcription of chromatin templates by destabilizing the nucleosomes within the open reading frames of active genes (3-5). FACT destabilizes the nucleosomes, which would otherwise act as barriers to RNA polymerase transcription activity, by disrupting histone-histone and histone-DNA contacts that lead to the eviction of the histone H2A-H2B dimer (2,3,6). FACT may also function as a histone chaperone to reassemble nucleosomes after RNA polymerase passage (7). In addition to transcription, FACT activity has been shown to have a role in DNA replication in yeast and in DNA repair by contributing to the activation of p53 by CK2 and by facilitating histone H2AX-H2B exchange upon DNA damage (8-10).

  1. Winkler, D.D. and Luger, K. (2011) J Biol Chem 286, 18369-74.
  2. Orphanides, G. et al. (1999) Nature 400, 284-8.
  3. Orphanides, G. et al. (1998) Cell 92, 105-16.
  4. Birch, J.L. et al. (2009) EMBO J 28, 854-65.
  5. Orphanides, G. and Reinberg, D. (2000) Nature 407, 471-5.
  6. Keller, D.M. and Lu, H. (2002) J Biol Chem 277, 50206-13.
  7. Belotserkovskaya, R. et al. (2003) Science 301, 1090-3.
  8. Schlesinger, M.B. and Formosa, T. (2000) Genetics 155, 1593-606.
  9. Keller, D.M. and Lu, H. (2002) J Biol Chem 277, 50206-13.
  10. Heo, K. et al. (2008) Mol Cell 30, 86-97.

Application References

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

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