1999 Fiscal Year Final Research Report Summary
Regulation of the functional activity of Anaphase Promoting Complex
| Project/Area Number |
10680676
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Cell biology
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| Research Institution | Tokyo University of Pharmacy and Life Science |
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Principal Investigator |
TANAKA Hirofumi Tokyo University of Pharmacy & Life Science, Life Science, Associated Professor, 生命科学部, 助教授 (30146899)
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| Project Period (FY) |
1998 – 1999
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| Keywords | mammalian cells / cell cycle / mitosis / ubiquitin / proteolysis / cyclin B |
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| Research Abstract |
Anaphase-promoting complex (APC) functions as a cell cycle-regulated ubiquitin ligase that mediates destruction of cell cycle-regulatory factors such as cyclin B and cut2 by proteasomes dirung mitosis. APC is activated at metaphase-anapgase transition and remains active until late G1 phase. APC consists of 12 subunits at least in mammalian cells, which were recently identified. The APC activity has been thought to be regulated by cell cycle-specific phosphorylation and dephosphorylation of ithe subunits, but the precise regulatory mechanism of APC activation has remained unclear. Furthermore, the binding of APC and its secific ubiquitin conjugating enzyme (UbcH10) has not been reported, and the mechanism of substrate recognition remains unresolved. We identified a novel protein H10BH(UbcH10-binding protein with a hect-like domain) by the yeast two-hybrid system with UbcH10 as a bait. H10BH binds with UbcH10 in vitro and in vivo. Furthermore, H10BH interacts with cyclin B and Cdc27, an APC substrate and an APC component, respectively. H10BH recognizes the cyclin-box in cyclin B by its C-terminal domain.. H10BH greatly enhanced the ubiquitination of cyclin B in the presence of HeLa cell lysates. When H10BH was incubated with ubiquitin, E1, UbcH10 and ubiquitin, the self-ubiquitination of H10BH was observed. H10BH also showed a weak activity of ubiquitin-ligase for cyclin B in the absence of APC. These activities were mediated by the C-terminal hect-like domain of H10BH. Homolgy search of H10BH revealed that Saccharomyces cerevisiae genome contains a homologue of H10BH. Desruption of this gene showed that it is a essential gene for the groth of budding yeasts. These results suggests that H10BH is a novel regulator of APC or a ubiquitin-ligase itself.
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