| 研究課題/領域番号 |
18J11869
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| 研究種目 |
特別研究員奨励費
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| 配分区分 | 補助金 |
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| 応募区分 | 国内 |
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| 研究分野 |
細胞生物学
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| 研究機関 | 筑波大学 |
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研究代表者 |
河村 有美 筑波大学, グローバル教育院, 特別研究員(DC2)
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| 研究期間 (年度) |
2018-04-25 – 2020-03-31
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| 研究課題ステータス |
交付 (2019年度)
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| 配分額 *注記 |
1,500千円 (直接経費: 1,500千円)
2019年度: 700千円 (直接経費: 700千円)
2018年度: 800千円 (直接経費: 800千円)
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| キーワード | extracellular vesicle / retrotransposon / horizontal gene transfer |
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| 研究実績の概要 |
Long interspersed element-1 (LINE-1 or L1) retrotransposons replicate through a copy-and-paste mechanism using an RNA intermediate. However, little is known about the physical transmission of retrotransposon RNA between cells. To examine the horizontal transfer of an active human L1 retrotransposon mediated by extracellular vesicles (EVs), human cancer cells were transfected with an expression construct containing a retrotransposition-competent human L1 tagged with a reporter gene. Using this model, active retrotransposition events were detected by screening for the expression of the reporter gene inserted into the host genome by retrotransposition. EVs including exosomes and microvesicles were isolated from cells by differential centrifugation. The enrichment of L1-derived reporter RNA transcripts were detected in EVs isolated from cells expressing active L1 retrotransposition. The delivery of reporter RNA was confirmed in recipient cells, and reporter genes were detected in the genome of recipient cells. Additionally, employing qRT-PCR, we found that host-encoded factors are activated in response to increased exposure to L1-derived RNA transcripts in recipient cells. Our results suggest that the horizontal transfer of retrotransposons can occur through the incorporation of RNA intermediates delivered via EVs and may have important implications for the intercellular regulation of gene expression and gene function.
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
2: おおむね順調に進展している
理由
To examine the horizontal transfer of an active human L1 retrotransposon mediated by extracellular vesicles (EVs), human cancer cells were transfected with an expression construct containing a retrotransposition-competent human L1 tagged with a reporter gene. Using this model, active retrotransposition events were detected by screening for the expression of the reporter gene inserted into the host genome by retrotransposition. The enrichment of L1-derived reporter RNA transcripts were detected in EVs isolated from cells expressing active L1 retrotransposition. The delivery of reporter RNA was confirmed in recipient cells, and reporter genes were detected in the genome of recipient cells.
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| 今後の研究の推進方策 |
EVs are packaged with abundant cellular information that has been shown to reflect the status of the cell, however, the mechanisms that regulate EV cargo selection is still largely unknown. Therefore, the biogenesis and cargo selection of EVs will be investigated to elucidate some of the biological processes that are regulated by EVs. The use of EVs as biomarkers for disease in liquid biopsy is a promising field, and overcoming the technical difficulties involved in the isolation and detection of EVs from biological samples would be beneficial in monitoring health and detecting diseases at an early stage. For this reason, new technologies in the isolation and detection of EVs from biological samles will be assessed, focusing on blood-derived EVs by microfluidics and the characterization of EV components.
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