Analysis of wild-type and mutant p21WAF-1 gene activities.
The p21WAF-1 gene is positively regulated by the wild-type p53 protein. p21WAF-1 has been shown to interact with several cyclin-dependent kinase complexes and block the activity of G1 cyclin-dependent kinases (cdks). Mutational analysis with the p21WAF-1 gene localized a site, at amino acid residues 21 and 24 in the amino terminus of the protein, for p21WAF-1 binding to cyclins D and E. This region of the protein is conserved (residues 21 to 26) in other p21WAF-1 family members, p27kip-1 and p57kip-2. The same p21WAF-121,24 mutant also fails to bind to cyclin D1-cdk 4 or cyclin E-cdk 2 complexes in vitro, suggesting that amino acid residues 21 and 24 are important for p21WAF-1-cdk-cyclin trimeric complex interactions. The p21WAF-1 wild-type protein will suppress tumor cell growth in culture while p21WAF-1 mutant proteins with defects in residues 21 and 24 fail to suppress tumor cell growth. The overexpression of cyclin D or E in these cells will partially overcome the growth suppression of wild-type p21WAF-1 protein in cells. These results provide evidence that p21WAF-1 acts through cyclin D1-cdk4 and cyclin E-cdk2 complexes in vivo to induce the growth suppression. The p21WAF-1 binding sites for cyclins (residues 21 to 26), cdk2 (residues 49 to 71), and proliferating-cell nuclear antigen (residues 124 to 164) have all been mapped to discrete sites on the protein.
Lin
J
J
Department of Molecular Biology, Princeton University, New Jersey 08544, USA.
Reichner
C
C
Wu
X
X
Levine
A J
AJ
D000595:Amino Acid Sequence; D002455:Cell Division / Q000235:genetics; D056741:Cyclin D; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics* / Q000378:metabolism; D016158:Genes, p53; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
C493956:CDKN1A protein, human; D056741:Cyclin D; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D016159:Tumor Suppressor Protein p53