Linking functions: an additional role for an intrinsically disordered linker domain in the transcriptional coactivator CBP

Abstract

The multi-domain transcriptional coactivators CBP/p300 integrate a multitude of signaling inputs, interacting with more than 400 proteins via one or more of their globular domains. While CBP/p300 function is typically considered in terms of these structured domains, about half of the protein consists of intrinsically disordered regions (IDRs) of varying length. However, these IDRs have only been thought of as linkers that allow flexible spatial arrangement of the structured domains, but recent studies have shown that similar IDRs mediate specific and critical interactions in other proteins. To examine the roles of IDRs in CBP, we performed yeast-two-hybrid screenings of placenta and lung cancer cDNA libraries, which demonstrated that the long IDR linking the KIX domain and bromodomain of CBP (termed ID3) can potentially bind to several proteins. The RNA-binding Zinc-finger protein 106 (ZFP106) detected in both libraries was identified as a novel substrate for CBP-mediated acetylation. Nuclear magnetic resonance (NMR) spectroscopy combined with cross-linking experiments and competition-binding assays showed that the fully disordered isolated ID3 transiently interacts with an IDR of ZFP106 in a fashion that disorder of both regions is maintained. These findings demonstrate that beside the linking function, ID3 can also interact with acetylation substrates of CBP.

Response to Comment on “Innovative scattering analysis shows that hydrophobic disordered proteins are expanded in water”

“…This discrepancy is not limited to the specific proteins selected by us or by Hofmann et al (10). We reanalyzed available SAXS data for other IDPs with sequences typical of folded proteins (11)(12)(13)(14)(15)(16)(17) using our new MFF procedure and compared these results to n extracted from available FRET studies of disordered proteins with foldable sequences ( Fig. 2A).…”

Proline Fingerprint in Intrinsically Disordered Proteins

“…Carbon‐13 direct‐detection NMR has proven to be very valuable for studying IDPs/IDRs . In particular, the detection of carbonyl carbon atoms (C′) constitutes a particularly useful tool for detection of proline residues .…”

“…In this context, long linkers of large proteins are paradigmatic. As one example, the CREB‐binding protein (CBP) is a large (2442 residues) protein in which half of the residues are predicted to form five long intrinsically disordered linkers connecting globular domains . Proline residues make up 11.3 % of the total amino acids, and 15.5 % of the amino acids in the classified disordered regions .…”

NMR Characterization of Long‐Range Contacts in Intrinsically Disordered Proteins from Paramagnetic Relaxation Enhancement in 13C Direct‐Detection Experiments

“…Certain residue types, such as glycine, serine, and threonine, have strong water–amide exchange broadening under physiological conditions, which causes a poor signal‐to‐noise ratio in 1 H detection experiments and prevents the determination of PREs. However, these residues, in addition to prolines, are highly abundant in IDPs and their role in local dynamics and phosphorylation events is key for the function of many IDPs . Therefore, the possibility of monitoring PRE effects for these residues is highly valuable.…”