自己組織化における外的制御の研究

Field: Biochemistry, Genetics and Molecular Biology (Life Sciences)

The ubiquitin system is one component of the biological regulatory mechanisms, and is related in a variety of cellular processes including the cell cycle control, DNA repair, stress response and transcriptional control. Ubiquitin, a highly conserved eukaryotic protein, can be found either free or bond to the various substrate proteins. The covalent attachment of ubiquitin to the substrate proteins is accomplished by the sequential reaction of ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzyme (E2) and ubiquitin ligase (E3). The ubiquitin-substrate conjugates are rapidly degraded by the 26S proteasome, a multisubunit complex of protease.<br /> To date, a number of different E2s have been characterized. All known E2s share a conserved domain of approximately 16kDa and a specific cystein residue which is capable of accepting the ubiquitin. It is thought that the diversity of E2s is required for the selective ubiquitination of the substrate proteins which mediate different cellular functions. <br /> To isolate E2s systematically, he has developed a new screening method which utilized a common activity of E2 enzymes. Consequently, he has been isolated four members of a family of E2s from the fission yeast Schizosaccharomyces pombe, and termed them ubcP1 - 4. Comparison of the deduced amino acid sequences of ubcP1 - 4 cDNA with all of so far known E2s from the various organisms revealed that ubcP4 is a novel member of a family of E2s. The deduced amino acid sequence of ubcP4 is 20 - 30% identical to a family of E2s and contains a specific cystein residue which is capable of accepting the ubiquitin. <br /> Disruption of the coding region of ubcP4 gene shows that ubcP4 is an essential gene for growth in the fission yeast. The repression of the UbcP4 synthesis caused the several phenotypes such as G2 arrest, metaphase arrest and cut (cell untimely torn). The detailed phenotype analysis revealed that ubcP4 is required for the both cell cycle transitions; from G2 to M phase and from metaphase to anaphase. Interestingly, the several phenotypes in metaphase/anaphase caused by the loss of ubcP4 function, are strikingly similar to those of cut9 ts mutants; cut9 is also required for the metaphase/anaphase transition and is a potential component of ubiquitin ligase (E3), termed anaphase-promoting complex (APC) or cyclosome. Therefore, he concludes from the phenotype similarities that ubcP4 may function in cooperation with APC at metaphase/anaphase. <br /> "Checkpoint controls" delay mitosis until DNA synthesis and repair of DNA damage are complete. The DNA damage-induced delay is mainly due to a temporary G2 arrest and provide the time necessary for the cell to complete DNA repair before mitosis. In this study, he showed that the amount of UbcP4 is decreased in response to ultraviolet irradiation, and the cells overexpressing ubcP4 are sensitive to ultraviolet irradiation. These data suggest that the decreased level of UbcP4 is required for the cells when DNA is damaged. Since the repression of ubcP4 synthesis caused the G2 arrest, he concludes that the specific ubiquitin-pathway involving ubcP4 could be required for the DNA damage-induced G2 arrest. <br /> In conclusion, he has isolated a novel member of a family of E2s, ubcP4, and showed that it is required for the both of G2/M transition and metaphase/anaphase transition, and possibly for the checkpoint control in response to DNA damage.