Definition of the p53 functional domains necessary for inducing apoptosis

The p53 protein contains several functional domains necessary for inducing cell cycle arrest and apoptosis. The C-terminal basic domain within residue s 364-393 and the proline-rich domain within residues 64-91 are required fo r apoptotic activity. In addition, activation domain 2 within residues 43-6 3 is necessary for apoptotic activity when the N-terminal activation domain 1 within residues 1-42 is deleted (Delta AD1) or mutated (AD1(-)). Here we have discovered that an activation domain 2 mutation at residues 53-54 (AD 2(-)) abrogates the apoptotic activity but has no significant effect on cel l cycle arrest. We have also found that p53-(Delta AD2), which lacks activa tion domain 2, is inert in inducing apoptosis. p53-(AD2(-)Delta BD), which is defective in activation domain 2 and lacks the C-terminal basic domain, p53-(Delta AD2 Delta BD), which lacks both activation domain 2 and the C-te rminal basic domain, and p53-(Delta PRD Delta BD), which lacks both the pro line-rich domain and the C-terminal basic domain, are also inert in inducin g apoptosis. All four mutants are still capable of inducing cell cycle arre st, albeit to a lesser extent than wild-type p53. Interestingly, we have fo und that deletion of the N-terminal activation domain 1 alleviates the requ irement of the C-terminal basic domain for apoptotic activity. Thus, we hav e generated a small but potent p53-(Delta AD1 Delta BD) molecule. Furthermo re, we have determined that at least two of the three domains (activation d omain 1, activation domain 2, and the proline-rich domain), are required fo r inducing cell cycle arrest. Taken together, our results suggest that acti vation domain 2 and the proline-rich domain form an activation domain for i nducing pro-apoptotic genes or inhibiting antiapoptotic genes. The C-termin al basic domain is required for maintaining this activation domain competen t for transactivation or transrepression.