1995 Fiscal Year Final Research Report Summary
Analysis of the androgen response element of mouse EGF gene by use of the androgen receptor expressed in Echerichia coli
| Project/Area Number |
06807144
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Functional basic dentistry
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| Research Institution | Iwate Medical University |
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Principal Investigator |
NEMOTO Takayuki Iwate Med.Univ., Sch.Dent., Lecturer, 歯学部, 講師 (90164665)
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| Co-Investigator(Kenkyū-buntansha) |
KYAKUMOTO Seiko Iwate Med.Univ., Sch.Dent., Assistant, 歯学部, 助手 (90118274)
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| Project Period (FY) |
1994 – 1995
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| Keywords | submandibular gland / androgen receptor / androgen response element / EGF / HSP90 / down-regulation / dimerization |
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| Research Abstract |
We studied the action mechanism of androgen in the mouse submandibular gland that shows the typical sexual dimorphism. To prepare the adsorbent specifically interacting with potential androgen response element, the DNA-domain of human androgen receptor (ARDBD) was expressed in E.coli. ARDBD fused to glutathione S-transferase (GST) existed as a dimer via GST-GST interaction and had a high affinity for mouse mammary tumor virus DNA.However, elimination of GST mojety by thrombin cleavage caused the reduction of the binding affinity, indicating that the dimerization of ARDBD is essential for the DNA binding of high affinity. The affinity column with GST-ARDBD was incubated with DNA fragments derived from 5'-upstream region of mouse EGF gene promoter. The column specifically trapped the 130 bp fragment located at -727- -598b of EGF gene. Comparison of the sequence with those of androgen-dependent androgen genes revealed the segment homologous among these genes.
The effect of testosterone on the expression of AR mRNA in mouse submandibular glands was examined. The AR mRNA level was higher in females than in males. Castration resulted in elevation of the level, while testosterone injection to female mice caused the reduction. Thses results show that the AR mRNA level is down-regulated by testosterone.
We finally investigated the mechanism of dimerization of the 90-kDa heat shock protein (HSP90), a binding component of steroid receptors. The dimeric structure of HSP90alpha is mediated by the C-terminal 191 amino acids and consists of duplicate interactions of the C-terminal region (Met628/Ala629-Asp732) of one subunit and the adjacent more N-terminal region (Val542-Tyr627/Met628) of the other subunit. The amino acids 290-732 were sufficient for the interaction with steroid receptors.
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[Publications] Nemoto, T., Matsusaka, T., Ota, M., Ohara-Nemoto, Y., Kaneko, M., Horigome, T.and Inano, K.: "Kinship between self-oligomerization and the estrogen receptor-interacting activities of the 90-kDa heat shock protein" Steroid Biochem. (Life Sci.Adv.). (in press).
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「研究成果報告書概要(欧文)」より
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[Publications] Ema M., Matsushita N., Sogawa K., Ariyama T., Inagawa J., Nemoto T., Ota, M., Oshimura, M., Fujii-Kuriyama, Y.: "Human arylhydrocarbon receptor : functional expression and chromosomal assignment to 7p21" J.Biochem.116. 845-851 (1994)
Description
「研究成果報告書概要(欧文)」より
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