2015 Fiscal Year Annual Research Report
| Project/Area Number |
15F15780
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| Research Institution | Institute of Physical and Chemical Research |
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Principal Investigator |
Carninci Piero 国立研究開発法人理化学研究所, ライフサイエンス技術基盤研究センター, 副センター長 (10333296)
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| Co-Investigator(Kenkyū-buntansha) |
BUDIC MARUSKA 国立研究開発法人理化学研究所, ライフサイエンス技術基盤研究センター, 外国人特別研究員
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| Project Period (FY) |
2015-11-09 – 2018-03-31
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| Keywords | SINEUP / upregulation / transcription factor / iPS / reprogramming |
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| Outline of Annual Research Achievements |
In order to better understand the upregulation of protein translation with SINEUPs, we analyzed mouse SINEUPs and determined their activity by measuring translation activation of GFP (green fluorescent protein) mRNA in human cell lines after transfection. The results indicate that the SINEB2 element is sufficient for SINEUP activity. We also determined the rules to create the most effective sense-antisense overlap in the specificity domain in SINEUPs. We found that the 44 nt antisense specificity domain is sufficient for upregulation of protein translation.
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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
To get better insight into the upregulation of protein translation with SINEUPs, we analyzed mouse SINEUPs and determined their activity by measuring the translation activation of GFP mRNA, using engineered SINEUPs that overlap the 5’ end of reporter mRNA. We tested the activity by overexpression in human HEK293T and HeLa cell lines. We characterized the functional components of SINEUPs by analyzing natural SINEUPs having different SINE elements. We determined the rules to create the most effective sense-antisense overlap in the specificity domain in SINEUPs.
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| Strategy for Future Research Activity |
We analyzed and optimized functional domains of SINEUPs in vitro. Based on these results we will design SINEUPs for reprogramming human iPS cells (hiPSCs) into neuron cells. Conversion of hiPSCs into neurons allows examination of healthy human neurons and of neurons derived from patients with variety of neurological diseases. Derivations of neurons from hiPSCs may allow us to examine specific subtypes of neurons, to generate human neurons for regenerative medicine, and to investigate changes in human neurons in neuropsychiatric disorders. One of the major limitations of deriving neurons from hiPSCs is cumbersome, variable and slow procedure needed for deriving neurons with functional properties from hiPSCs. We will develop a method for rapid conversion of hiPSCs into neurons. First, we will perform bioinformatic analysis of neuron differentiation time course data from the international research consortium FANTOM5 (http://fantom.gsc.riken.jp/5/). Next, we will identify and select specific transcription factors (TF) expressed in differentiated neurons, and engineer synthetic SINEUPs that will overlap the 5’ ends of the selected TF mRNAs. We will optimize the synthetic SINEUPs for reprogramming hiPSCs into neurons.
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