| Project/Area Number |
21590146
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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 |
Environmental pharmacy
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| Research Institution | National Institute of Health Sciences |
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Principal Investigator |
KIKUCHI Yutaka 国立医薬品食品衛生研究所, 衛生微生物部, 室長 (10234197)
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| Project Period (FY) |
2009 – 2011
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| Project Status |
Completed (Fiscal Year 2011)
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| Budget Amount *help |
¥4,680,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥1,080,000)
Fiscal Year 2011: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000)
Fiscal Year 2010: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
Fiscal Year 2009: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
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| Keywords | 食品衛生学 / プリオン / スプライシング / GPIアンカー / 選択的スプライシング |
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| Research Abstract |
The human prion protein(PrP) is a glycoprotein with a glycosylphosphatidylinositol(GPI) anchor at its C-terminus. I previously reported alternative splicing within exon 2 of the PrP gene(PRNP) in the human glioblastoma cell line T98G. The open reading frame(ORF) of the alternatively spliced mRNA lacked the GPI anchor signal sequence and encoded a 230 amino acid polypeptide. Its product, GPI-anchorless PrP(GPI-PrPSV), was unglycosylated and soluble in non-ionic detergent, and was found in the cytosolic fraction. I also detected low levels of alternatively spliced mRNA in human brain and non-neuronal tissues. Here I report alternative splicing within exon 3 of the PrP gene in the bovine cornea cell line BCE C/D-1b and the ovis aries(sheep) brain cell line OA1.I reported an intronic sequence with the canonical dinucleotides for splicing(GT at the 5' end and AG at the 3' end) within PRNP exon 2.However, I identified the non-canonical GC-AG introns within exon 3 of bovine and ovis aries PrP genes. The ORFs of the alternatively spliced mRNA lacked the GPI anchor signal sequence, and the bovine and sheep PrPSV encoded a 260 and 252 amino acid polypeptide, respectively. Relative quantitative RT-PCR analysis, which measures PrP mRNA in total RNA, revealed that low levels of PrPSV mRNA are constitutively expressed in bovine and ovis aries brains. Variant forms arising from alternative mRNA splicing within exon 3 have not been reported in cattle and sheep. This study is the first report of an alternative splicing by intron retention within PrP gene exon 3.
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