| Co-Investigator(Kenkyū-buntansha) |
TSUNOO Hajime Meiji Insutitute of Health Science, Director & Scientists, 所長, 研究者
TAMURA Shigehiko Kyushu Univ., Faculty of Science, Research associate, 理学部, 助手 (90236753)
HARANO Tomoyuki Kyushu Univ., Faculty of Science, Research associate, 理学部, 助手 (80037275)
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| Budget Amount *help |
¥16,700,000 (Direct Cost: ¥16,700,000)
Fiscal Year 1998: ¥4,300,000 (Direct Cost: ¥4,300,000)
Fiscal Year 1997: ¥5,800,000 (Direct Cost: ¥5,800,000)
Fiscal Year 1996: ¥6,600,000 (Direct Cost: ¥6,600,000)
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| Research Abstract |
With the support of this Grants-in Aid for Scientific Research 0 8 5 5 7 0 1 1 we obtained two major types of findings :
I) Isolation, characterization, and complementation group analysis of novel CHO cell mutants defective in peroxisome biogenesis
In addition to the previously isolated, three complementation groups (CGs) of peroxisome-deficient CHO cell mutants, ZP24, Z65, and ZP92, we isolated ZPlO7 (the same group as Z24), ZP1O5/ZP139, ZPIO9, ZP11O, ZP114, ZP119, ZP124, and ZP126, by the P9OH/UV method. CG analysis by PEX cDNA transfection and/or cell fusion with previously identified CGs of mutant cells, including 12 CGs of fibroblasts derived from patients with peroxisome biogenesis disorders (PBDs), revealed that ZP110, ZP114, and ZP126 are distinct from human CGs. Thus, it is evident that peroxisome assembly requires at least 15 gene-products.
II) Cloning of novel peroxin genes By genetic functional complementation assay of newly isolated CHO cell mutants, we cloned several peroxin cDNAs (PEXs), including PEX1, PEX12, and PEX19 for ZP1O7, ZP1O9, and ZP119, respectively. We then delineated that gene mutations in PEX1, PEX12, and PEX19 are responsible for PBDs of CG-I (E), CG-III, and CG-J, respectively. Moreover, we isolated PEX1O and PEX16 by expressed sequence tag (EST) DNA search using yeast genes and demonstrated that inactivation of PEX]O and PEX16 is the genetic cause of CG-VII (B) and CG-IX (D) PBDs, respectively. We also found two isoforms of the peroxin Pex5p (PTS 1 receptor) using CG-ll CHO mutants, ZP1O5 and ZP139. Impaired import of PTS1 proteins in the mutant cells was restored by both, shorter and longer, forms of Pex5p. More strikingly, the longer isoform of Pex5p was found to be involved in import of PTS2 proteins as well.
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