Induction of p21(WAF1/CIP1) and inhibition of Cdk2 mediated by the tumor suppressor p16(INK4a).
The tumor suppressor p16(INK4a) inhibits cyclin-dependent kinases 4 and 6. This activates the retinoblastoma protein (pRB) and, through incompletely understood events, arrests the cell division cycle. To permit biochemical analysis of the arrest, we generated U2-OS osteogenic sarcoma cell clones in which p16 transcription could be induced. In these clones, binding of p16 to cdk4 and cdk6 abrogated binding of cyclin D1, p27(KIP1), and p21(WAF1/CIP1). Concomitantly, the total cellular level of p21 increased severalfold via a posttranscriptional mechanism. Most cyclin E-cdk2 complexes associated with p21 and became inactive, expression of cyclin A was curtailed, and DNA synthesis was strongly inhibited. Induction of p21 alone, in a sibling clone, to the level observed during p16 induction substantially reproduced these effects. Overexpression of either cyclin E or A prevented p16 from mediating arrest. We then extended these studies to HCT 116 colorectal carcinoma cells and a p21-null clone derived by homologous recombination. In the parental cells, p16 expression also augmented total cellular and cdk2-bound p21. Moreover, p16 strongly inhibited DNA synthesis in the parental cells but not in the p21-null derivative. These findings indicate that p21-mediated inhibition of cdk2 contributes to the cell cycle arrest imposed by p16 and is a potential point of cooperation between the p16/pRB and p14(ARF)/p53 tumor suppressor pathways.
Mitra
J
J
Departments of Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA.
Dai
C Y
CY
Somasundaram
K
K
El-Deiry
W S
WS
Satyamoorthy
K
K
Herlyn
M
M
Enders
G H
GH
D017209:Apoptosis / Q000235:genetics*; D042846:CDC2-CDC28 Kinases; D018797:Cell Cycle Proteins; D051357:Cyclin-Dependent Kinase 2; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000378:metabolism*; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000378:metabolism*; D004261:DNA Replication; D005786:Gene Expression Regulation; D006801:Humans; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D011132:Polyribosomes; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D016160:Retinoblastoma Protein / Q000378:metabolism; D015398:Signal Transduction; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D025521:Tumor Suppressor Proteins
C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D019941:Cyclin-Dependent Kinase Inhibitor p16; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D008869:Microtubule-Associated Proteins; D016160:Retinoblastoma Protein; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases