TY  - JOUR
TI  - p65 Chromatin Immunoprecipitation Protocol
AU  - Dudgeon, Crissy R.
VL  - 3
IS  - 8
PY  - 2013
DA  - 2013/04/20
SP  - e683
C1  - Bio-protocol 2013;3:e683
DO  - 10.21769/BioProtoc.683
UR  - https://doi.org/10.21769/BioProtoc.683
AB  - Chromatin Immunoprecipitation (ChIP) is an important procedure that allows you to verify if a certain protein is physically located at a regulatory region.  This information, taken together with other procedures such as luciferase assays and EMSAs, will give definitive proof that the query protein is involved in the transcription of a protein.  This procedure for p65 ChIP can be adapted to investigate other proteins; just a change of the antibody will suffice. The transcription factor known as NF-κB is a homo- or hetero-dimer consisting of members of the Rel/NFKB family.  The most abundant NF-κB complexes are made of two different proteins, p65 (Rel-A) and p50 (NFKB1).  The NF-κB complex is initially inhibited by IκB by direct binding, thus trapping NF-κB in the cytoplasm. After a stimulatory signal, IκB kinase (IKK) phosphorylates IκB, allowing IκB to undergo proteasome-mediated degradation.  The degradation of IκB and phosphorylation of p65 by multiple kinases activates NF-κB, allowing it to transport to the nucleus and cause the transcriptional activation of many of its target genes containing κB sites (consensus sequence: gggRNNYYcc, R = purine Y = pyrimidine), such as PUMA, IL-6, and TNF.
KW  - P65
KW  - NFkB
KW  - ChIP
KW  - Promoter
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False