2017 Fiscal Year Annual Research Report
RNA干渉におけるRISC形成への核内転写因子の関与
| Project/Area Number |
16H04740
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| Research Institution | University of Tsukuba |
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Principal Investigator |
Liu Qinghua 筑波大学, 国際統合睡眠医科学研究機構, 教授 (90723792)
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| Project Period (FY) |
2016-04-01 – 2019-03-31
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| Keywords | 核酸 / RNA / RISC |
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| Outline of Annual Research Achievements |
We purified histone H1 from nuclear extracts of Drosophila S2 cells. We identified HP1BP3, one of 13 human H1-like chromatin proteins, as a pri-miRNA binding protein that specifically associates with the Microprocessor. Knockdown of HP1BP3, but not other H1 variants, compromised pri-miRNA processing by resulting in the premature release of pri- miRNA transcripts from the chromatin. ChIP studies reveal genome-wide colocalization of HP1BP3 & Drosha and HP1BP3-dependent Drosha binding to actively transcribed miRNA loci. HP1BP3 exhibits a novel pri-miRNA binding activity and promotes the Drosha-pri-miRNA association in vivo. Together, these studies suggest that HP1BP3 promotes co-transcriptional miRNA processing through chromatin retention of nascent pri-miRNA transcripts.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
We have an excellent research team and received help from outstanding collaborators.
1. We collaborated with Dr. Mikiko Siomi at University of Tokyo, who is a world’s leader in small RNA research, to con duct co-localization studies of GFP-TAF11 and Dicer-2/R2D2, using excellent anti-Dicer-2 and anti-R2D2 monoclonal antibo dies that were previously generated in the Siomi lab.
2. Our lab has strong expertise in classical biochemistry. We are good at generating highly purified recombinant Dicer-2 /R2D2 and TAF11 proteins using insect cell-expression system. We developed all of the in vitro assays for analysis of RL C and RISC assembly.
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| Strategy for Future Research Activity |
The recent revolutionary progress in high-resolution imaging by cryoEM and near-atomic resolution structures from heterogeneous datasets made it necessary for us to aim for high-resolution structures of the C3PO/RNA complexes. We are currently using cryoEM to reconstruct 3D structure of the Drosophila RLC complex using recombinant Dicer-2-R2D2, TAF11 and duplex siRNA.
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Research Products
(5 results)
