title: Identification of an additional negative regulatory region for p53 sequence-specific DNA binding.
abstract: The DNA binding activity of p53 is crucial for its tumor suppressor function and is subject to tight regulation. Previous studies revealed that the inhibitory function of the p53 C terminus is implicated in the latent, low affinity sequence-specific DNA binding activity of p53 in the uninduced state. Sequence-specific DNA binding of p53 has been shown to be activated by several posttranslational modifications and interacting proteins that target predominantly the C terminus. Moreover, several authors have shown that synthetic peptides corresponding to p53 C-terminal sequences activate p53 sequence-specific DNA binding. In an effort to identify the interaction site of p53 with these activating peptides we assessed complex formation between p53 deletion constructs and C-terminal activating peptides by peptide affinity precipitation. This study revealed that two distal regions of the p53 molecule contribute synergistically to the interaction with activating C-terminal peptides: amino acids 80-93 and 364-393. The C-terminal residues 364-393 are already well characterized as having negative regulatory function. DNA binding analyses with these deletion constructs reveal a comparable negative regulatory activity for residues 80-93, defining this region as a previously unidentified negative regulatory domain of p53. Furthermore, synthetic peptides spanning this newly identified proline-rich negative regulatory region (residues 80-93) are able to activate p53 sequence-specific DNA binding in vitro. We suggest that both negative regulatory regions, residues 80-93 and 364-393, contribute cooperatively to the maintenance of the latent, low-affinity DNA binding conformation of p53.
authors.affiliation: Institute for Bone and Joint Disorders and Cancer, Bayer Corporation, Pharmaceutical Division, 400 Morgan Lane, West Haven, CT 06516, USA. beate.muellertiemann@schering.de
authors.forename: B F
authors.initials: BF
authors.lastname: Müller-Tiemann
authors.forename: T D
authors.initials: TD
authors.lastname: Halazonetis
authors.forename: J J
authors.initials: JJ
authors.lastname: Elting
mesh_terms: D001665:Binding Sites / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli; D016158:Genes, p53; D011485:Protein Binding / Q000235:genetics; D017421:Sequence Analysis; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
chemical_list: D004269:DNA, Bacterial; D016159:Tumor Suppressor Protein p53