| Project/Area Number |
07044249
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| Research Category |
Grant-in-Aid for international Scientific Research
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| Allocation Type | Single-year Grants |
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| Section | Joint Research |
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
SHIMIZU Akira Center for Molecular Biology and Genetics, Kyoto University, 遺伝子実験施設, 教授 (00162694)
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| Co-Investigator(Kenkyū-buntansha) |
STORB Ursula Beate Department of Molecular Genetics and Cell Biology, The University of Chicago, 分子遺伝学・細胞生物学科, 教授
MARTIN Teren シカゴ大学, 分子遺伝学・細胞生物学科, 教授
AGATA Yasutoshi Center for Molecular Biology and Genetics, Kyoto University, 遺伝子実験施設, 助手 (60263141)
MARTIN Terence Edwin Department of Molecular Genetics and Cell Biology, The University of Chicago
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| Project Period (FY) |
1995 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| Budget Amount *help |
¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 1996: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1995: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Keywords | Ig class Switching / Ig gene / S region / Gene rearrangement / germ-line transcript / trans-splicing / trans-mRNA / Multiple isotype expression / アレルギー / CD40リガンド |
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| Research Abstract |
We cloned the constant (C) region gene of chicken immunoglobulin (Ig)mu chain. Nucleotide sequence analysis revealed that there are two types of tandem repetitive sequences apanning up to a few kilobases locating just upstream of the first exon of C_<mu> gene. Both of the repeats consist of unit sequences of 5 base long, resembling to the S_<mu> region of mammalian gene. One of the two types, closer to the C_<mu> gene consists of tandem repeats of (C/T) (C/A) CAG complementary to the mammalian S_<mu> region. These findings suggest the DNA recombination during the class switching in chicken, and importance of the repetitive nature of S regions.
We next tested whether the Ig trans-mRNAs, which consist of a variable (V) region of a transgene and the C regions of endogenous isotypes, can be expressed from a transgene lacking S region which is necessary for the DNA recombination of class switching. Spleen cells of C line mouse, a transgenic mouse carrying a rearranged mouse gamma2b gene from which the S_<mu> and S_<gamma2b> regions were completely deleted during the construction the transgene, were stimulated with LPS and IL-4 to induce the trans-mRNA expression. Upon the stimulation, specific induction of gamma1 and epsilon trans-mRNAs were detected by RT-PCR using specific primers for transgenic V region and endogenous C region genes. Structures of the detected PCR products were confirmed by nested PCRs with specific, inner primers and their direct sequencing using an automated sequencer. Our data indicate that the trans-mRNA expression is independent of S region mediated mechanism, i.e., DNA (S-S) recombination. This also supports our previous hypothesis that Ig trans-mRNAs are expressed by RNA mediated mechanism, i.e., trans-splicing, and thus multiple isotypes of Ig can be expressed through this mechanism.
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