2001 Fiscal Year Final Research Report Summary
In situ visualization of the signal proteins in the nervous system of the ascidian embryo
| Project/Area Number |
11680663
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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 |
Biophysics
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| Research Institution | MURORAN INSTITUTE OF TECHNOLOGY (2000-2001)
Himeji Institute of Technology (1999) |
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Principal Investigator |
IWASA Tatsuo MURORAN INSTITUTE OF TECHNOLOGY, Professor, 工学部, 教授 (00133926)
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| Project Period (FY) |
1999 – 2000
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| Keywords | G protein / Signal transduction / Molecular cloning / Microinjection / Protochordata / GFP / Embryogenesis / Regulation of gene expression |
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| Research Abstract |
In order to investigate G protein-coupling signaling system of neural system of ascidian embryo, we have isolated CDNA clones of G protein alpha and beta subunit. Five different CDNA clones of G protein alpha subunit were isolated from a CDNA library of ascidian larvae. The deduced amino acid sequence of three of them (HrGi, HrGq and HrGs) showed high homology to those of human Gi, Gq and Gs, respectively. The other two (HrGn and HrGx) were supposed to define a novel sub family within the Gi abd Gq families, respectively.
Whole-mount in situ hybridization and northern blot analysis revealed that the messages of all these G protein alpha subunits were found during embryogenesis and in adult organs with distinct spatial and temporal expression patterns.
We cloned 5'-franking sequence of HrGn and constructed HrGn/GFP fusion gene. The microinjection of the construct into fertilized eggs of ascidian revealed that the genomic fragment containing 1.4 Kb upstream of the putative translation start site contains the promoter region responsible for the expression pattern characteristic for HrGn.
Then, we constructed 1.4-HrGn/HrGn-coding/GFP fusion gene, which has the full length coding region of HrGn alpha subunit and GFP. Microinjection experiment of this construct is the final purpose of this project and now on going.
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