CREB-binding protein, also known as CREBBP or CBP or KAT3A, (where CREB is cAMP response element-binding protein) is a coactivator encoded by the CREBBP gene in humans, located on chromosome 16p13.3. CBP has intrinsic acetyltransferase functions; it is able to add acetyl groups to both transcription factors as well as histone lysines, the latter of which has been shown to alter chromatin structure making genes more accessible for transcription. This relatively unique acetyltransferase activity is also seen in another transcription enzyme, EP300 (p300). Together, they are known as the p300-CBP coactivator family and are known to associate with more than 16,000 genes in humans; however, while these proteins share many structural features, emerging evidence suggests that these two co-activators may promote transcription of genes with different biological functions.
For example, CBP alone has been implicated in a wide variety of pathophysiologies including colorectal cancer as well as head and neck squamous cell carcinoma. In these diseases, association of CBP with β-catenin has been shown to promote cancer cell proliferation and disease aggressiveness, whereas p300/ β-catenin leads to cell differentiation and/ or apoptosis. CBP has also been shown to help modulate liver function via maintenance of energy homeostasis in response to changes in cell nutrition conditions by regulating the activity of transcription factors and genes responsible for lipogenesis and gluconeogenesis. CBP is also implicated in the etiologies of several other diseases including hematologic malignancies and other solid tumors, diabetes, schizophrenia, Alzheimer's disease, depression, and many other neurological conditions.
Structure The functional CBP is approximately 7362 nucleotides long and encodes 2,441 amino acids. CBP does not directly interact with promoter elements; it is brought to the site via protein-protein interactions that are the result of its different structural domains complexing with other transcriptional co-activators.
Transitional adapter zinc finger domains
CBP has two transitional adapter zinc finger (TAZ) domains that each consist of four alpha helices stabilized by zinc ions. Both TAZ1 and TAZ2 domains favor the hydrophobic residues within amphipathic amino acid sequences, and their binding affinity is enhanced by the interaction of acidic residues with positive side chains. Factors that bind to TAZ2 are also potentially regulated via acetylation due to its proximity to the acetyltransferase domain (KAT).
Cysteine-histidine rich regions Although there are three cysteine-histidine rich domains identified within CBP, only domains 1 and 3 (CH1, CH3) have had their structural functions resolved. A number of factors that associate with CBP bind to either the CH1 or CH3 domain, or to both, despite their locations at opposing ends of the protein. To date, little is known about the interaction between these two domains. The primary structure of CH1 and CH3 has been shown to break down into consensus sequences that are capable of chelating zinc ions (Zn2+). Experiments performed also showed that the residues in these sequences are obligatory for transcriptional coactivation to occur by either CH1 or CH3. The CH2 region, located in the middle of the protein, in its acetyltransferase domain, does not contain this consensus sequence, and has not been conclusively shown to bind zinc ions.
Kinase-inducible domain interacting domain Aka KIX domain, CREB binding domain, MYB interaction domain
The kinase-inducible domain (KID) interacting domain (KIX) also is the protein domain on CBP and p300 where heterodimers form from the interaction of CBP (or p300) with other transcription factors and coactivators. It consists of three alpha and two 310 helices that have high affinity for amphipathic protein sequences. Interestingly, these helices can fold into a number of different conformers which enables the domain to maintain both a level of promiscuity while also exerting regulatory control. The KIX domain controls the rate of transcription and has been shown to be critical for hematopoietic differentiation. Some of the proteins that bind to this domain have been shown to bind competitively—such as CREB and Myb—whereas others bind via allosteric cooperation as in the case of MLL and Myb.
Bromodomain
Bromodomains (BRD) consist of approximately 110 amino acids, that function to recognize acetylated lysine molecules. They consist of four left-handed alpha helices connected by loops which form hydrophobic binding pockets. The bromodomain of CBP binds to regions of the genome are rich with acetylated lysine residues, meaning they have lost their positive charge, decreasing histone affinity for DNA, which makes the region more open and accessible for transcription. Acetylated p53 and STAT3 have been shown to bind to CBP's bromodomain.
Lysine acetyltransferase domain
The 380 protein residue lysine acetyltransferase (KAT) domain is arguably one of the most important and identifying structural components of CBP. Its activity is regulated via phosphorylation of CBP. What's interesting is that it is able to acetylate not just histones but other proteins as well. Currently there are over 100 known substrates for CBP's KAT domain including proteins such as p53, E2F-1-3, GATA-1, MyoD and CREB. The ability of CBP to acetylate p53 via its KAT domain, which then increases p53's affinity for CBP's bromodomain showcases an interesting mechanism by which this unique protein can self-servingly regulate gene transcription.
Nuclear receptor coactivator binding domain Aka interferon response factor binding domain (IBiD)
The nuclear receptor coactivator binding domain (NCBD) located at the c-terminus of CBP, in its unbound state, fluctuates between multiple conformations. Binding of a target protein to NCBD will cause it to fold into three helices specifically functioned to associated with the disordered domains of its binding partners. Among the proteins known to bind to this region include ACTR (p160)--a coactivator for the thyroid and retinoid receptors, its homolog SRC-1, p53, and SMAD.
Interactions Proteins shown to interact specifically with CBP Actr Ap-1 Apc5 Apc7 Atr BRCA1 C-FOS C-JUN C-MYB Chrebp CITED2 Ddb1 (p127) Ddb2 (p48) Ddx3 Dhx9 DNA2 helicase E1A E2F Elk1 ER Ets1 Fen1 Foxo1 Gal4 GATA1 GR hATF4 HNF-1 Hif-1α Ku70 MDM2 MLL MyoD NF-κB p53 p73 Parp1 pCREB pCAF PCNA Pit1 RAR RXR SCRAP SMAD Src1 Srebp1c STAT1 STAT2 STAT3 Sumo T3R TBP Tdg TFIIB TFIIE TFIIF Wrn Xpa Zta (BZLF1)
Function
This gene is ubiquitously expressed and is involved in the transcriptional coactivation of many different transcription factors. CBP has two critical mechanisms by which it is able to regulate gene expression: as an acetyltransferase, and as a protein scaffold that helps recruit and construct the complexes that are necessary for transcription or chromatin remodeling. Phosphorylation of CBP increases its acetyltransferase activity, a process hypothesized to be regulated in a cell cycle dependent manner. Recent results suggest that novel CBP-mediated post-translational N-glycosylation activity alters the conformation of CBP-interacting proteins, leading to regulation of gene expression, cell growth and differentiation,
Distinction from p300 Often in scientific papers (especially in less recent ones), CBP and p300 are used interchangeably as CBP/p300. This is a reasonable amalgamation given the sequence homology, structural similarity and binding behaviors of these two proteins. However, ongoing research shows that CBP and p300 maintain distinct biological functions and should therefore not be conflated.
It has been reported that despite having common histone substrates, when there are lower amounts of histone or acetyl-CoA available, CBP and p300 have different, preferred acetyltransferase substrates. In an experiment performed involving the Kaposi sarcoma-associated herpesvirus, the pathological protein (vIRF), was shown to be upregulated by CBP and repressed by p300. p300 homozygous knockout studies in mice were embryonic lethal, with improper neurulation and poor heart development occurring during their limited survival. Additionally, the fibroblasts isolated from these mice were unable to properly proliferate and were lacking the retinoic acid receptor. Transgenically altered mice homozygous for mutated copies of CBP (missing the KAT domain), were also embryonic lethal, however instead, these mice had poor vascular angiogenesis and abnormal hematopoiesis characterized by the absence of progenitor cell proliferation and an altered hematopoietic microenvironment. The fact that both CBP and p300 homozygous knockouts were embryonic lethal suggest that these factors play a critical role in embryogenesis. The differential phenotypes of these embryos also indicate that CBP and p300 regulate different aspects of embryological development.
Role in cell cycle regulation
Studies performed in the late 1990s demonstrated that peak CBP acetyltransferase activity occurs at the transition between the G1/S phase of the cell cycle checkpoint. Given the concentrations of CDK2 present in the cell at this stage of the cell cycle, it was hypothesized that CDK2 may be a key regulator of these post-translational modifications. It turned out that administration of cyclin E/CDK2 inhibitors did in fact inhibit the enzymatic activity of CBP's KAT domain. Other proteins that have been shown to phosphorylate CBP include MAP Kinase, PKA and CAMK4. Ser-133 has been shown to be an important residue that is phosphorylated by PKA to initiate CBP transcriptional activity.
The E2F family of transcription factors are critical for taking a cell from G1 to S phase of the cell cycle. They bind to a sequence consensus in the promoter region of genes involved in DNA replication. CBP (and p300) have been shown to interact with E2F proteins as both a coactivator, and as an acetyltransferase, the latter of which causes increased E2F DNA binding affinity. A knockout study that was published in 2000, which used a microinjection of an antibody against CBP/p300 significantly diminished the number of cells able to progress into S phase, further supporting the idea that CBP is essential in the transcription of factors necessary G1/S phase transition.
CBP is also believed to facilitate the process of DNA replication during S phase by acetylating histones around origins of replication. Acetylation of histones, specifically lysine residues in the histones, weakens the electric charge interaction between the histone and DNA, causing this area to become more open and accessible for the machinery required for DNA replication. Two histone acetylation markers that have been associated with active regions include histone 3 lysine 18 acetylation (H3K18ac) and histone 3 lysine 27 acetylation (H3K27ac). It has also been shown that CBP acetylates two endonucleases (FEN1, DNA2) that are involved in processing Okazaki fragments.
Another key component of the cell cycle that is regulated by CBP is the anaphase promoting complex/cyclosome (APC/C). This complex consists of numerous subunits that are grouped into two subdomains, the "Arc Lamp" and the "Platform," and functions as an E3 ubiquitin ligase that targets components related to the cell cycle such as cyclin B, securin, and PLK1 for proteasome degradation. Two subunits of the APC/C have been shown to directly interact with CBP: AP5, which is located in the Platform subdomain, and AP7, located in the Arc Lamp subdomain. Experiments performed using RNAi to completely suppress CBP and p300 showed significant increases in protein concentrations for those normally targeted by the APC/C, and caused a number of cells to become arrested in the mitotic phase of the cell cycle.