2006 Fiscal Year Final Research Report Summary
Identification of immunosuppressant-sensitive genes: a study using model organisms
| Project/Area Number |
15209010
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General pharmacology
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| Research Institution | Kobe University |
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Principal Investigator |
KUNO Takayoshi Kobe University, Department of genome sciences, Professor, 大学院医学系研究科, 教授 (50144564)
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| Co-Investigator(Kenkyū-buntansha) |
SHUNTOH Hisato Kobe Gakuin University, Faculty of Rehabilitation, Professor, 総合リハビリテーション, 教授 (70206259)
SUGIURA Reiko Kinki University, School of Pharmacy, Proffessor, 薬学部, 教授 (90294206)
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| Project Period (FY) |
2003 – 2006
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| Keywords | calcineurin / immunosuppressant / farnesyltransferase / valproic acid / Rab / phospholipid / MAP kinase / リン脂質 |
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| Research Abstract |
1. In fission yeast, calcineurin dephosphorylates and activates the Przl transcription factor. Here, we identified the calcineurin-dependent response element (CDRE) in the promoter region of przl(+) gene and monitored the calcineurin activity in living cells using a destabilized luciferase reporter gene fused to three tandem repeats of CDRE. The Pck2 protein kinase C-Pmkl mitogen-activate protein kinase pathway was required for the stimulation of calcineurin via Yam8/Cchl-mediated Ca(2+) influx, but it was not required for the stimulation by elevated extracellular Ca(2+), suggesting two distinct pathways for calcineurin activation.
2. Here we isolated vic (viable in the presence of immunosuppressant and chloride ion) mutants. One of the mutants, vicl-1, carried a missense mutation in the cppl+ gene encoding a beta subunit of the protein farnesyltransferase. Analysis of the mutant strain revealed that Rho2 is a novel target of Cppl. Moreover, Cppl and Rho2 act upstream of Pck2-Pmk1 MAPK
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signaling pathway, thereby resulting in the vic phenotype upon their mutations.
3. To gain further insights into the molecular mechanisms of valproic acid action (VPA), a genetic screen for fission yeast mutants that show hypersensitivity to VPA was performed. One of the genes we identified was vps45+ which encodes a member of the Sec1/Munc18 family that is implicated in membrane trafficking. Our results suggest that VPA affects membrane trafficking, which leads to the enhanced sensitivity to cell wall damage in fission yeast.
4. In the same screen using the immunosuppressant drug FK506 in fission yeast, we isolated gdil-ill, a mutant allele of the essential gdil+ gene encoding Rab GDP-dissociation inhibitor. Notably, overexpression of spo20+, encoding a phosphatidylcholine/phosphatidylinositol transfer protein, rescued gdil-ill mutation. The gdil-ill and spo20-KC104 mutations are synthetically lethal. Our findings suggest that Spo20 modulates Gdil function via regulation of phospholipid metabolism of the membranes. Less
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