| Abstract
| - The newly discovered oncogenic protein gankyrin, which contains six ankyrin repeats, hasbeen reported to be involved in the phosphorylation and degradation of the retinoblastoma gene product,Rb. Using in vitro systems, we have identified a peptide fragment of gankyrin, 176LHLACDEERN185,which is responsible for binding of gankyrin to Rb. We further demonstrated a different mechanism forgankyrin to facilitate the phosphorylation of Rb, by binding with cyclin-dependent kinase 4 (CDK4).This binding does not inhibit the Rb-phosphorylating kinase activity of CDK4, but it competes with p16binding to CDK4 and counteracts the inhibitory function of p16. We then showed that binding of gankyrinto CDK4 and the consequent counter action of p16 function were not affected by the Rb-binding peptide176LHLACDEERN185, indicating that the two mechanisms are independent. They also involve differentstructural regions of gankyrin. While the Rb-binding motif is located at the fifth ankyrin repeat, truncationmutants of gankyrin, with the first three or four ankyrin repeats remaining, are sufficient for binding toCDK4 and for counteracting the inhibitory function of p16. These results demonstrate the potentialimportance of gankyrin in cell cycle control and tumorigenesis and suggest an expanded INK4-CDK4/6-Rb pathway.
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