TOK-1, a novel p21Cip1-binding protein that cooperatively enhances p21-dependent inhibitory activity toward CDK2 kinase.
A p21(Cip1/Waf1/Sdi1) is known to act as a negative cell-cycle regulator by inhibiting kinase activity of a variety of cyclin-dependent kinases. In addition to binding of the cyclin-dependent kinase to the N-terminal region of p21, p21 is also bound at its C-terminal region by proliferating cell nuclear antigen (PCNA), SET/TAF1, ... and calmodulin, indicating the versatile function of p21. In this study, we cloned cDNA encoding a novel protein named TOK-1 as a p21 C-terminal-binding protein by a two-hybrid system. Two splicing isoforms of TOK-1, TOK-1alpha and TOK-1beta, comprising 322 and 314 amino acids, respectively, were co-localized with p21 in nuclei and showed a similar expression profile to that of p21 in human tissues. TOK-1alpha, but not TOK-1beta, directly bound to the C-terminal proximal region of p21, and both were expressed at the G(1)/S boundary of the cell cycle. TOK-1alpha also preferentially bound to an active form of cyclin-dependent kinase 2 (CDK2) via p21, and these made a ternary complex in human cells. Furthermore, the results of three different types of experiments showed that TOK-1alpha enhanced the inhibitory activity of p21 toward histone H1 kinase activity of CDK2. TOK-1alpha is thus thought to be a new type of CDK2 modulator.
Mesh Terms:
Amino Acid Sequence, Animals, Base Sequence, Blotting, Northern, CDC2-CDC28 Kinases, COS Cells, Calcium-Binding Proteins, Carrier Proteins, Cell Cycle, Cell Cycle Proteins, Cell Line, Cell Nucleus, Cloning, Molecular, Codon, Terminator, Cyclin-Dependent Kinase 2, Cyclin-Dependent Kinase Inhibitor p21, Cyclin-Dependent Kinases, Cyclins, DNA, Complementary, Fluorescent Antibody Technique, Indirect, Glutathione Transferase, Hela Cells, Humans, Models, Genetic, Molecular Sequence Data, Mutagenesis, Site-Directed, Nuclear Proteins, Phosphotransferases, Plasmids, Proliferating Cell Nuclear Antigen, Protein Binding, Protein Isoforms, Protein Structure, Tertiary, Protein-Serine-Threonine Kinases, Recombinant Proteins, Time Factors, Tissue Distribution, Two-Hybrid System Techniques
Amino Acid Sequence, Animals, Base Sequence, Blotting, Northern, CDC2-CDC28 Kinases, COS Cells, Calcium-Binding Proteins, Carrier Proteins, Cell Cycle, Cell Cycle Proteins, Cell Line, Cell Nucleus, Cloning, Molecular, Codon, Terminator, Cyclin-Dependent Kinase 2, Cyclin-Dependent Kinase Inhibitor p21, Cyclin-Dependent Kinases, Cyclins, DNA, Complementary, Fluorescent Antibody Technique, Indirect, Glutathione Transferase, Hela Cells, Humans, Models, Genetic, Molecular Sequence Data, Mutagenesis, Site-Directed, Nuclear Proteins, Phosphotransferases, Plasmids, Proliferating Cell Nuclear Antigen, Protein Binding, Protein Isoforms, Protein Structure, Tertiary, Protein-Serine-Threonine Kinases, Recombinant Proteins, Time Factors, Tissue Distribution, Two-Hybrid System Techniques
J. Biol. Chem.
Date: Oct. 06, 2000
PubMed ID: 10878006
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