| Project/Area Number |
11670161
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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 |
Pathological medical chemistry
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| Research Institution | Kagoshima University (2000-2001)
National Cardiovascular Center Research Institute (1999) |
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Principal Investigator |
MIYATA Atsuro Kagoshima University, Faculty of Medicine, Dept. of Pharmacology, Professor, 医学部, 教授 (60183969)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 2000: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1999: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Keywords | PACAP / mouse / gene expression / neuropeptide / cardiovascular regulator / neurotrophic factor / FISH / Inr / 循環調節因子 / 神経選択的サイレンサー / PC12 / 遺伝子 / 染色体 |
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| Research Abstract |
Pituitary adenylate cyclase activating polypeptide (PACAP) is a pleitropic neuropeptide which belongs to secretin/glucagon family. It should be interesting that PACAP elicits both of neurotrophic action and cardiovascular regulation. In this project, we have cloned and characterized mouse PACAP gene, and further performed functional analysis of its 5' upstream regulatory region in order to clarify the regulatory mechanism of PACAP gene expression.
The gene encoding mouse PACAP was isolated from a mouse genomic library using mouse PACAP cDNA as a probe. Nucleotide sequencing revealed that PACAP gene spans 6.5 kb and consists of 5 exons and 4 introns. The 3 kb of 5' -flanking region sequence contained potential binding site for several transcription factor (CBP, AP-1, GATA, GHF-1). Fluorescence in situ hybridization study revealed that the mouse gene for PACAP is localized to band E5 of chromosome 17. A tissue-distribution study by RT-PCR in rat demonstrated that PACAP mRNA was widely exp
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ressed and particularly abundant in the thymus, the adrenal gland and the testis beside of the brain.
5' RACE and RNase protection assay demonstrated multiple transcription start sites. The most upstream region of transcriptional initiation site contains two TATA like motifs, the distal one is well-conserved between human and mouse, and functionally more potent than proximal one in terms of basal transcription. In other two regions there were found several Inr-like elements instead of TATA box. These findings indicate that the transcription of the single copy of PACAP gene is regulated by three independent promoters, together with different splicing of the transcript (at least 5 types), giving rise to mature PACAP mRNA which has the same open reading frame, but different 5' untranslated region. So it was suggested that the expression of PACAP gene is regulated in dual mode, induciblly and constitutively. Furthermore deletion analysis using a luciferase reproter gene construct demonstrated the presence of several enhancer or supressor elements. Less
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