Co-Investigator(Kenkyū-buntansha) |
KOHN Leonald d. NIDDK,Cell Regulation Section, Chief, Chief
SUGAWA Hideo KYOTO UNIVERSITY,Lecturer, 医学研究科, 講師 (70162857)
AKAMIZU Takashi KYOTO UNIVERSITY,Assistant, 医学研究科, 助手 (20231813)
MORI Toru KYOTO UNIVERSITY,Professor, 医学研究科, 教授 (40026894)
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Research Abstract |
We investigated signal transduction mechanism of the TSH receptor by site-directed mutagenesis of the three cytoplasmic loops and C-terminal tail and following observations were obtained. For Gs interaction, the middle portion of the second cytoplasmic loop is most important. For Gq interaction, widely distributed regions such as entire region of the first cytoplasmic loop, the middle portion of the second cytoplasmic loop, the N-terminal and C-terminal portions of the third cytoplasmic loop and N-terminal one third of the C-terminal tail are crucial. Substitution mutants at residues 617-620 in the third cytoplasmic loop showed constitutive activation of the cAMP and inositol phosphate levels. This is the first example of constitutively activated glycoprotein hormone receptor mutations. After that, TSH receptor activating mutations in hyperfunctioning thyroid adenomas were reported. We also showed that TSH receptor mutations Asp^<633>*Glu/Tyr, Thr^<622>*Ile, and Phe^<631>*Cys found in hyperfunctioning thyroid adenomas were constitutively activated by expression experiments and proved that those were direct cause of the disease. All activating glycoprotein hormone receptor mutation so far reported were located in the transmembrane region. We showed that a deletion mutant in the extracellular domain was also constitutively activated. Further, exoplasmic loop was shown to be related signal transduction also. The letter two observations suggest that interaction between the extracellular region and the transmembrane region is important event for signal transdution. As for LH receptor, we also found new activation mutations in familial and sporadic male precocious puberty and showed that those were the direct cause of the disease by expression experiments. We also investigated the mechanism of activation of the LH receptor by in vitro mutagenesis and found that the importance of the side chain of the Asp^<578> for maintaining the inactive state of the receptor.
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