| Co-Investigator(Kenkyū-buntansha) |
KAMBE Fukushi Nagoya Univ., Res.Inst.Environ., Med., Assoc.Prof., 環境医学研究所, 助教授 (00211871)
MURATA Yoshiharu Nagoya Univ., Res.Inst.Environ., Med., Prof., 環境医学研究所, 教授 (80174308)
KOMATSU Yukio Nagoya Univ., Res.Inst.Environ., Med., Prof., 環境医学研究所, 教授 (90135343)
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| Research Abstract |
We identified a thyroid hormone (T3)-responsive gene, ZAKI-4 in cultured human skin fibroblasts. It belongs to a family of genes that encode proteins containing a conserved motif. The motif binds to and inhibits calcineurin (CN). In this study, we identified three different ZAKI-4 transcripts, α, β1 and β2 in human brain by 5'-and 3'-RACE. The a transcript was identical to that we originally cloned and the other two were novel. The three transcripts are generated by alternative initiation and splicing from a single gene on the short arm of chromosome 6. It is predicted that β1 and β2 encode an identical protein product β, which differs from a in its N-terminus. Both isoforms associate with CN and inhibit its activity through an identical C-terminal region. An examination of expression profile of the three transcripts revealed that a transcript is expressed exclusively in brain, while b transcripts are expressed ubiquitously, most abundantly in brain, heart, skeletal muscle and kidney.
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It was also demonstrated that human skin fibroblasts express both α and β transcripts, raising a question which transcript is upregulated by T3. It was revealed that T3 markedly induced the expression of a isoform but not of β. This T3-mediated increase in the a isoform was associated with a significant decrease in endogenous CN activity. We further explored how T3 regulates the expression of a isoform in human skin fibroblasts. First, effect of calcineurin inhibitor FK506 and its analog rapamycin on T3-mdeiated regulation of ZAKI-4α was studied, because activation of CN was reported to cause an increased expression of a ZAKI-4 family gene, DSCR1. It was demonstrated that T3-mediated increase in ZAKI-4 expression was not inhibited by FK506 but completely abrogated by rapamycin. Since rapamycin is a specific inhibitor of mammalian target of rapamycin (mTOR), we investigated the involvement of mTOR in T3 action. It was demonstrated that T3 activates mTOR kinase through non-genomic action. These data for the first time demonstrated that T3 signaling cascade through mTOR to calcineurin inhibitor ZAKI-4α. Less
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