| Project/Area Number |
10680589
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Structural biochemistry
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| Research Institution | IWATE UNIVERSITY (1999)
Tokai University (1998) |
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Principal Investigator |
SAITOH Yasushi Faculty of Agriculture, Associate Professor, 農学部, 助教授 (70287100)
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| Co-Investigator(Kenkyū-buntansha) |
HADANO Shinji Institute of Medical Science, Tokai University, Assistant Professor, 総合医学研究所, 助手 (60281375)
IKEDA Joh-E Institute of Medical Science, Tokai University, Professor, 総合医学研究所, 教授 (50266467)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1998: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Keywords | ubiquitin / huntingtin / deubiquitinating enzyme / repetitive sequence / megasatellite / 4p15 / antisense RNA / アンチセンスRNA / huntingtin / CAGリピート / ポリグルタミン |
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| Research Abstract |
The nuclear inclusion body formation of ubiquitin-huntingtin conjugate must be controlled by ubiquitin conjugating enzymes and deubiquitinating enzymes. One ubiquitin-conjugating enzyme was reported to bind huntingtin. In contrast, the function of deubiqitinating enzyme is not clear yet.
We found a novel deubiquitinating enzyme gene named RS447-dUb in the RS447 megasatellite repeat. We examined the expression of the RS447-dUb in human tissues. Northern blot analysis using an antisense ORF probe identified 2.4 Kb transcript in heart, brain, liver and skeletal muscle. To confirm the presence of poly(A) containing RS447 RNA, cDNA clones that encodes a sense transcript of the RS447 were isolated by using a PCR-based extension method in conjunction with 5' and 3'-RACE. We also identified a functional promoter activity in upstream sequence of the RS447 ORF.
We also transfected a cosmid clone containing 7 tandem copies of RS447 into mouse A9 cells and confirmed the expression of RS447 deubiquitinating enzyme.
These results indicate that the RS447-repeating unit on this megasatellite repeat codes and actively expresses a functional deubiquitinating enzyme.
Although it is expressed ubiquitously in human tissues, RS447-dUb exhibited a unique expression pattern in that its complementary strand is transcribed as an antisense transcript predominantly in brain. This could modulate the level of RS447-dUb expression in the human brain. Further characterizations of the expression of sense and antisense transcripts of the RS447, and relationship with ubiquitin-huntingtin nuclear inclusion body formation are currently underway.
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