Fine mapping and regulation of the association of p53 with p34cdc2

Abstract

In vivo p53 is multiply phosphorylated by dierent protein kinases suggesting a central role for phosphorylation in modulating p53 function. In addition, p53 was found to be associated with two protein kinases, p34 cdc2 and protein kinase CK2. Here we report the precise mapping of the interaction sites of p53 ± p34 cdc2 complexes. The p34 cdc2 binding site on human p53 maps to one distinct C-terminal site LQIRGRERFE (aa 330 ± 339) close to the corresponding phosphorylation site at serine 315. In order to test whether phosphorylation of p53 might in¯uence the binding of p53 to p34 cdc2 phosphorylation mutants of the C-terminus of p53, which mimick permanent phosphorylation, were tested on their ability to bind to p34 cdc2 in vitro. Substitution of serine 315 (the p34 cdc2 phosphorylation site) with aspartic acid had only little eect on complex formation whereas an exchange of serine 392 (the protein kinase CK2 phosphorylation site) to aspartic acid resulted in a signi®cant reduced relative binding anity of p53 to p34 cdc2 . The same result was obtained when the Cterminus of p53 was phosphorylated by puri®ed protein kinase CK2 prior to examination of complex formation. In addition, the speci®city of the complex formation has been checked by competition experiments with full length p53 proteins and the in¯uence of cyclin B on complex formation was examined.

Characterization of the oligomerization defects of two p53 mutants found in families with Li–Fraumeni and Li–Fraumeni-like syndrome

“…For example, Lomax et al (1998) have shown that both L344P and R337C have reduced ability to bind to MDM2 which forms a negative feedback regulatory loop with p53. Other proteins such as tms1 (Wagner et al, 1995) and p34 cdc2 (Wagner et al, 1998) interact with residues in the tet domain spanning the R337C mutation site. The occurrence of these two mutations in familial cancers underlines the important role the tet domain plays in p53 function.…”

Phosphorylation of the CCAAT Displacement Protein (CDP)/Cux Transcription Factor by Cyclin A-Cdk1 Modulates Its DNA Binding Activity in G2

“…In agreement with this, a Cdk2 binding domain was mapped between residues 45 and 60 of p21 WAF1/CIP1 , a region that is fully conserved in the p27Kip1 inhibitor (63). Moreover, a distinct Cdk1 binding domain was identified within the carboxylterminal domain of p53 (48,64).…”

The Conserved CPH Domains of Cul7 and PARC Are Protein-Protein Interaction Modules That Bind the Tetramerization Domain of p53

“…In addition, these residues are hydrophobic (Fig. 5, C and D) and have been proposed to be a potential site for interaction with other proteins (33,34). Residues Gly 334 and Arg 335 are also surface-exposed, and they belong to the turn that links the ␣-helices to the ␤-sheets.…”