Analysis of dsRNA mechanism of response in insect cell

• Project/Area Number: 23580077
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Applied entomology
• Research Institution: Kyushu University
• Principal Investigator: LEE JAEMAN 九州大学, (連合)農学研究科(研究院), 助教 (50404083)
• Co-Investigator(Kenkyū-buntansha): 日下部 宜宏 九州大学, 大学院農学研究院, 教授 (30253595)
• Project Period (FY): 2011 – 2013
• Project Status: Completed (Fiscal Year 2013)
• Budget Amount: ¥5,330,000 (Direct Cost: ¥4,100,000、Indirect Cost: ¥1,230,000); Fiscal Year 2013: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2012: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2011: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
• Keywords: RNA interference / 遺伝子機能阻害 / dsRNA応答機構 / BmAgo1 / BmAgo2 / BmMLV / RISC因子 / RNAi / RNAi誘導実験 / siRNA / miRNA / Soaking RNAi誘導 / 遺伝子機能解析 / sid-1 / BmN4細胞

Research Abstract

In order to establish efficient RNA interference induction system in insect cells, we introduced several membrane proteins, which have the ability to uptake dsRNA into the cells. Also the cellular responses against dsRNA uptake were analyzed using silkworm cells. As a result, we found that C. elegans Sid-1 membrane protein possessed an ability to uptake dsRNA efficiently. Using this protein, several transgenic insect cell lines were successfully established. Furthermore, almost all RNAi related components of silkworm were isolated and characterized.

Research Products

• [Journal Article] Co-expression of Escherichia coli RNase III in silkworm cells improves the Efficiency of RNA interference induced by long hairpin dsRNAs (2013)
  • Author(s): JaeMan Lee, Yoshito Kojin, Hiroaki Mon, Tsuneyuki Tatsuke, Yoshitaka Miyagawa, Takahiro Kusakabe
  • Journal Title: Insect Sci Volume: 20 Pages: 69-77
• [Journal Article] A MC motif in silkworm Ago1 are indispensible for translational repression (2013)
  • Author(s): Li Zhu, Yuuki Masaki, Tsuneyuki Tatsuke, Hiroaki Mon, Zhiqing Li, Jian Xu, JaeMan Lee, Takahiro Kusakabe
  • Journal Title: Insect Mol. Biol Volume: 22 Pages: 320-330
• [Journal Article] Soaking RNAi-mediated modification of Sf9 cells for baculovirus expression system by ectopic expression of Caenorhabditis elegans SID-1 Appl (2013)
  • Author(s): Jian Xu, Yudai Nagata, Y., Hiroaki Mon, Zhiqing Li, Li Zhu, Kazuhiro Iiyama, Takahiro Kusakabe, JaeMan Lee
  • Journal Title: Microbiol. Biotech Volume: 97 Pages: 5921-5931
• [Journal Article] Genome-wide identification of Argonaute 1- and Argonaute 2-regulating genes revealed an inhibition of macula-like virus by RNAi pathway in the silkworm (2013)
  • Author(s): Li Zhu, Zhiqing Li, Tsuneyuki Tatsuke, Daojun Cheng, Jian Xu, Kaito Yoshimura, Hiroaki Mon, Kazuhiro Iiyama, JaeMan Lee, Qingyou Xia, Takahiro Kusakabe
  • Journal Title: J. Insect Biotechnol. Seric Volume: 82 Pages: 19-23
• [Journal Article] Characterization of Tudor-sn-containing granules in the silkworm, Bombyx mori (2013)
  • Author(s): Li Zhu, Tsuneyuki Tatsuke, Hiroaki Mon, Zhiqing Li, Jian Xu, JaeMan Lee, Takahiro Kusakabe
  • Journal Title: Insect Biochem. Mol. Biol Volume: 43 Pages: 664-674
• [Journal Article] Characterization of Tudor-sn-containing granules in the silkworm, Bombyx mori (2013)
  • Author(s): Li Zhu, Tsuneyuki Tatsuke, Hiroaki Mon, Zhiqing Li, Jian Xu, Jae Man Lee, Takahiro Kusakabe
  • Journal Title: Insect Biochemistry and Molecular Biology Volume: 43 Issue: 8 Pages: 664-674
  • DOI: 10.1016/j.ibmb.2013.04.004
  • Peer Reviewed
• [Journal Article] Establishment of a soaking RNA interference and Bombyx mori nucleopolyhedrovirus (BmNPV)-hypersensitive cell line using Bme21 cell (2013)
  • Author(s): Jian Xu, Hiroaki Mon, Takahiro Kusakabe, Zhiqing Li, Li Zhu, Kazuhiro Iiyama, Atsushi Masuda, Takumi Mitsudome, Jae Man Lee
  • Journal Title: Applied Microbiology and Biotechnology Volume: 97 Issue: 24 Pages: 10435-10444
  • DOI: 10.1007/s00253-013-5279-x
  • Peer Reviewed
• [Journal Article] Coexpression of Escherichia coli RNase III in silkworm cells improves the efficiency of RNA interference induced by long hairpin dsRNAs (2013)
  • Author(s): Lee, J.M., Kojin, Y., Tatsuke, T., Mon, H., Miyagawa, Y. and Kusakabe, T.
  • Journal Title: Insect Science Volume: 20 Issue: 1 Pages: 69-77
  • DOI: 10.1111/j.1744-7917.2012.01569.x
  • Peer Reviewed
• [Journal Article] Effective RNA interference in cultured silkworm cells mediated by overexpression of Caenorhabditis elegans SID-1 (2012)
  • Author(s): Hiroaki Mon, Isao Kobayashi, Shinji Ohkubo, Shuichiro Tomita, JaeMan Lee, Hideki Sezutsu, Toshiki Tamura, Takahiro Kusakabe
  • Journal Title: RNA Biol Volume: 9 Pages: 40-46
• [Journal Article] RNA interference induction by long hairpin dsRNAs expressed from chromosomal DNA of Bombyx mori cells (2012)
  • Author(s): JaeMan Lee, Yoshito Kojin, Hiroaki Mon, Tsuneyuki Tatsuke, Yoshitaka Miyagawa, Takahiro Kusakabe
  • Journal Title: J. Fac. Agr Volume: (in press)
  • NAID: 120004756198
• [Journal Article] Molecular cloning of BmTUDOR-SN and analysis of its role in RNAi pathway in the silkworm, Bombyx mori (Lepidoptera : Bombycidae) (2012)
  • Author(s): Li Zhu, Tsuneyuki Tatsuke, Zhiqing Li, Hiroaki Mon, Jian Xu, JaeMan Lee, Takahiro Kusakabe
  • Journal Title: Appl. Entomol. Zool Volume: 47 Pages: 207-215
• [Journal Article] Molecular cloning of BmTUDOR-SN and analysis of its role in the RNAi pathway in the silkworm, Bombyx mori (Lepidoptera: Bombycidae) (2012)
  • Author(s): Zhu, L., Tatsuke, T., Li, Z., Mon, H., Xu, J., Lee, J.M. and Kusakabe, T.
  • Journal Title: Applied entomology and zoology Volume: 47 Pages: 207-215
  • Peer Reviewed
• [Journal Article] RNA Interference Induction by Long Hairpin dsRNAs Expressed from Chromosomal DNA of Bombyx mori Cells (2012)
  • Author(s): Lee, J.M., Kojin, Y., Tatsuke, T., Mon, H., Miyagawa, Y. and Kusakabe, T.
  • Journal Title: Journal of the Faculty of Agriculture, Kyushu University Volume: 57 Pages: 441-445
  • NAID: 120004756198
• [Journal Article] Effective RNA interference in cultured silkworm cells mediated by over expression of Caenorhabditis elegans SID-1 (2012)
  • Author(s): Mon H, Kobayashi I, Ohkubo S, Tomita S, Lee JM, Sezutsu H, Tamura T, Kusakabe T
  • Journal Title: RNA Biol Volume: 9 Issue: 1 Pages: 1-8
  • DOI: 10.4161/rna.9.1.18084
  • Peer Reviewed
• [Presentation] Role of silkworm RISC components in the basal defense against Bombyx mori nucleopolyhedrovirus (BmNPV) (2014)
  • Author(s): 祝 力・徐 剣・李 志清・門 宏明・ 李 在萬・日下部宜宏
  • Organizer: 日本蚕糸学会
  • Place of Presentation: 日本大学生物資源科学部
• [Presentation] Soaking RNAi and its application on insect sciences (2013)
  • Author(s): Takahiro Kusakabe, Hiroaki Mon, JaeMan Lee, Xu Jian, Li Zhiqing
  • Organizer: 韓国蚕糸学会本会
  • Place of Presentation: 大韓民国平昌郡
• [Presentation] カイコにおけるRNAi経路の解析 (2012)
  • Author(s): 吉末真吾、田附常幸、阪下浩介、門宏明、李在萬、日下部宜宏
  • Organizer: 日本蚕糸学会第82回大会蚕糸・昆虫機能利用学術講演会
  • Place of Presentation: 福岡市
• [Presentation] BmTudor-sn interact with BmAgo1 and BmAgo2 not to be involved in RNAi pathway, while to participate in stress granule formation in BmN4 cells (2012)
  • Author(s): Li Zhu, Tuneyuki Tatsuke, Hiroaki Mon, JaeMan Lee, Takahiro Kusakabe
  • Organizer: 日本蚕糸学会第82回大会蚕糸・昆虫機能利用学術講演会
  • Place of Presentation: 福岡市
• [Presentation] カイコ培養細胞NHEJにおける修復関連遺伝子ノックダウンの影響 (2012)
  • Author(s): 藤井美江、門宏明、高橋将晃、光延仁志、李在萬、日下部宜宏
  • Organizer: 日本蚕糸学会第82回大会蚕糸・昆虫機能利用学術講演会
  • Place of Presentation: 福岡市
• [Presentation] カイコ培養細胞におけるN-グリコシル化経路の改変とタンパク質発現系への応用 (2012)
  • Author(s): 永田祐大、副嶋泰彦、門宏明・李在萬、日下部宜宏
  • Organizer: 日本蚕糸学会第82回大会蚕糸・昆虫機能利用学術講演会
  • Place of Presentation: 福岡市
• [Presentation] カイコ培養細胞による多分岐型N-グリカンを持つ糖タンパク質の生産 (2012)
  • Author(s): 永田祐大、門宏明、李 在萬、日下部宜宏
  • Organizer: 平成２４年日本分子生物学会本会
  • Place of Presentation: 福岡国際会議場・マリンメッセ福岡
• [Presentation] カイコ培養細胞NHEJにおける修復関連遺伝子ノックダウンの影響 (2012)
  • Author(s): 藤井美江、門宏明、高橋将晃、光延仁志、李 在萬、日下部宜宏
  • Organizer: 平成２４年日本分子生物学会本会
  • Place of Presentation: 福岡国際会議場・マリンメッセ福岡
• [Presentation] カイコにおける RNAi 経路の解析 (2012)
  • Author(s): 吉末真吾 田附常幸 阪下浩介 門 宏明 李 在萬 日下部宜宏
  • Organizer: 平成24年度 蚕糸・昆虫機能利用学術講演会
  • Place of Presentation: 九州大学箱崎キャンパス文系地区 福岡
• [Presentation] Analysis of P-bodies and stress granules in silkworm BmN4 cells (2011)
  • Author(s): 祝力、田附常幸、李志清、門宏明、李在萬、日下部宜宏
  • Organizer: 第34回日本分子生物学会
  • Place of Presentation: 横浜市
• [Presentation] カイコβ-N-acetylglucosaminidase (BmFDL)のノックダウンがN-グリカン形成に与える影響/Effect of RNAi for BmFDL in the N-glycosylation pathway (2011)
  • Author(s): 永田祐大、副嶋泰彦、門宏明、李在萬、日下部宜宏
  • Organizer: 第34回日本分子生物学会
  • Place of Presentation: 横浜市