Structures of KIX domain of CBP in complex with two FOXO3a transactivation domains reveal promiscuity and plasticity in coactivator recruitment

Abstract

Forkhead box class O 3a (FOXO3a) is a transcription factor and tumor suppressor linked to longevity that determines cell fate through activating transcription of cell differentiation, survival, and apoptotic genes. Recruitment of the coactivator CBP/p300 is a crucial step in transcription, and we revealed that in addition to conserved region 3 (CR3) of FOXO3a, the C-terminal segment of CR2 (CR2C) binds CBP/p300 and contributes to transcriptional activity. CR2C and CR3 of FOXO3a interact with the KIX domain of CBP/p300 at both "MLL" and "c-Myb" binding sites simultaneously. A FOXO3a CR2C-CR3 peptide in complex with KIX exists in equilibrium between two equally populated conformational states, one of which has CR2C bound to the MLL site and CR3 bound to the c-Myb site, whereas in the other, CR2C and CR3 bind the c-Myb and MLL sites, respectively. This promiscuous interaction between FOXO3a and CBP/p300 is further supported by additional binding sites on CBP/ p300, namely, the TAZ1 and TAZ2 domains. In functional studies, our structure-guided mutagenesis showed that both CR2C and CR3 are involved in the activation of certain endogenous FOXO3a target genes. Further, phosphorylation of S626, a known AMP-dependent protein kinase target in CR3, increased affinity for CBP/p300 and the phosphomimetic mutation enhanced transactivation of luciferase. These findings underscore the significance of promiscuous multivalent interactions and posttranslational modification in the recruitment of transcriptional coactivators, which may allow transcription factors to adapt to various gene-specific genomic and chromatin structures and respond to cell signals.NMR solution structure | intrinsic disorder | promiscuous binding

Recognition of the disordered p53 transactivation domain by the transcriptional adapter zinc finger domains of CREB-binding protein

“…The p53 TAD forms an ensemble of conformational states in complex with the KIX domain of CBP/p300, with each of the AD1 and AD2 helices binding simultaneously in two distinct orientations and in two distinct site on the KIX surface (53). Similar promiscuous interactions with both KIX sites have also been observed for the bipartite transactivation domain of the FOXO3a transcription factor and are required for full transcriptional activity (68).…”

“…The ternary complex between p53, MDM2, and CBP/p300 seems to play a central role in regulation of p53 stability, determining in a phosphorylation-dependent manner whether p53 becomes polyubiquitinated and degraded or is activated to initiate transcription of p53-regulated stress response genes (25,67). The ability of multipartite systems to promiscuously interact with binding partners has been shown with other CBP/p300 binding proteins and may represent a widely used mechanism by which proteins can use disparate motifs to regulate transcription (53,68,69).…”

Dissecting allosteric effects of activator–coactivator complexes using a covalent small molecule ligand

“…The KIX domain interacts with more than 10 distinct transcriptional activators (23) via two different binding sites, with the resulting complexes leading to regulation of key functions such as hematopoiesis, cell cycle progression, and memory formation (Fig. 1B) (12,13,(24)(25)(26). Many of the KIX-binding TADs from activators such as pKID, MLL, and c-Myb are intrinsically disordered proteins that assume a helical structure only upon binding to interaction partners such as KIX (25,27,28).…”

FOXO factors and breast cancer: outfoxing endocrine resistance

“…FOXOs participate in several known transcriptional complexes, such as: i) ER (Schuur et al 2001, Zhao et al 2001; ii) p53 (Wang et al 2008); and iii) CREB binding protein (CBP)/p300. In the case of CBP/p300, the phospho-Thr32/Ser253-isoform has been shown to have lost the ability to physically associate with these transcriptional co-activator proteins (Wang et al , 2012a, and thus, this modification negates the transcriptionally favorable chromatin remodeling that this protein association generally promotes. CBP/p300 also functions to acetylate the lysine residues within the FOX-domain, thus repressing FOXO DNA-binding activity (see 'FOXO acetylation' section).…”