Regulation of the Activity in the p53 Family Depends on the Organization of the Transactivation Domain

Abstract

Despite high sequence homology among the p53 family members, the regulation of their transactivation potential is based on strikingly different mechanisms. Previous studies revealed that the activity of TAp63α is regulated via an autoinhibitory mechanism that keeps inactive TAp63α in a dimeric conformation. While all p73 isoforms are constitutive tetramers, their basal activity is much lower compared with tetrameric TAp63. We show that the dimeric state of TAp63α not only reduces DNA binding affinity, but also suppresses interaction with the acetyltransferase p300. Exchange of the transactivation domains is sufficient to transfer the regulatory characteristics between p63 and p73. Structure determination of the transactivation domains of p63 and p73 in complex with the p300 Taz2 domain further revealed that, in contrast to p53 and p73, p63 has a single transactivation domain. Sequences essential for stabilizing the closed dimer of TAp63α have evolved into a second transactivation domain in p73 and p53.

TA*p63 and GTAp63 achieve tighter transcriptional regulation in quality control by converting an inhibitory element into an additional transactivation domain

“…To experimentally investigate these predictions we mutated the residues most likely important for transactivation to alanines and investigated their influence on the transactivation potential of TAp63γ, TA*p63γ, and GTAp63γ, each carrying the FWL mutations. As expected TAp63γ FWL is inactive due to the importance of these three aa for interaction with p300 16 , while the two other FWL mutants still show distinct induction of the promotor. Only simultaneous mutation of the additional two motifs predicted in the TA*/GTA and GTA* peptides leads to a completely inactive protein (Fig.…”

“…In earlier studies we had shown that inhibition of the transcriptional activity of TAp63α is mechanistically based on the formation of a closed and only dimeric state with decreased affinity to both the DNA 6,8 and coactivators such as p300 16 . To investigate if TA*p63α and GTAp63α adopt a closed, dimeric conformation as well, we expressed both isoforms and TAp63α as control in H1299 cells and analyzed their oligomeric state by Blue-Native (BN)-PAGE.…”

“…In p63, the TAD consists of a single helix 16 that is critical for stabilization of the dimeric state 9 . Mutating the three aa F16A, W20A, and L23A (FWL mutant) in TAp63α disrupts the closed state and triggers the formation of an open tetrameric conformation 8 .…”

C-terminal α Domain of p63 Binds to p300 to Coactivate β-Catenin

“…The coactivation mode of p63 (ΔNp63α) was distinguished from the well-described transactivation mode of p53 and TA-p63, because the coactivation required stronger induction of β-catenin and functional WREs to which TCF/LEF proteins bind. The TA domain of TA-p63 isoforms interacts with p300 with a stretch conserved among p53 family proteins, (Figure 3 D ) for transcriptional activation [7]. In the case of TAp63α, interestingly, the ‘transcription inhibitory’ domain (TID) at the extremely C-terminus of the α-domain directly binds to the TA domain to block the transactivating function [32], [33], [34].…”

“…The TA domain of p63 has a single p300-binding site, whose sequences are conserved among p53 family proteins, to transactivate target genes [7]. In contrast, ΔN-p63 proteins lack the entire TA domain and instead have ΔN-specific sequences of only 14 amino acid residues.…”