Reconstruction of in vitro piRNA biogenesis using silkworm germ line BmN4 cells

• Project/Area Number: 25840041
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Functional biochemistry
• Research Institution: The University of Tokyo
• Principal Investigator: NISHIDA Kazumichi 東京大学, 理学(系)研究科(研究院), 助教 (10598436)
• Project Period (FY): 2013-04-01 – 2015-03-31
• Project Status: Completed (Fiscal Year 2014)
• Budget Amount: ¥4,420,000 (Direct Cost: ¥3,400,000、Indirect Cost: ¥1,020,000); Fiscal Year 2014: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000); Fiscal Year 2013: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
• Keywords: piRNA生合成 / PIWIタンパク質 / RNAヘリカーゼタンパク質 / 生殖細胞 / カイコ / piRNA / トランスポゾン / BmVasa

Outline of Final Research Achievements

piRNAs associate with PIWI proteins and silence selfish transposable elements. piRNA biogenesis consists of two sequential steps; primary piRNA processing and the Ping-Pong cycle. However, piRNA biogenesis remains elusive. We aimed to elucidate about piRNA pathway.
A Bombyx mori germ cell line, BmN4, is an excellent tool to investigate the molecular pathways of piRNA biogenesis. BmN4 expresses two PIWI proteins (Siwi and BmAgo3). We show that Siwi binds with BmVasa. BmVasa is DEAD-box RNA helicase protein. A lack of BmVasa abolishes the Ping-Pong cycle. Also, Siwi is associated with BmSpn-E. BmSpn-E, unlike BmVasa, is necessary for primary piRNA production. We propose a new model for piRNA biogenesis, where the BmSpn-E function in primary piRNA processing, whereas BmVasa is important for the Ping-Pong cycle.

Research Products

• [Journal Article] Respective functions of two distinct Siwi complexes assembled during piRNA biogenesis in insect germ cells. (2015)
  • Author(s): Nishida, KM., Iwasaki, YW., Murota, Y., Nagao, A., Mannen, T., Kato, Y., Siomi, H., and Siomi, MC.
  • Journal Title: Cell Reports Volume: 10 Issue: 2 Pages: 193-203
  • DOI: 10.1016/j.celrep.2014.12.013
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Presentation] Respective functions of two distinct Siwi, a silkworm PIWI, complexes assembled during piRNA biogenesis in insect germ cells (2014)
  • Author(s): Kazumichi M. Nishida, Yuka W. Iwasaki, Yukiko Murota, Taro Mannen, Haruhiko Siomi and Mikiko C. Siomi
  • Organizer: 日本分子生物学会
  • Place of Presentation: パシフィコ横浜 (横浜)
  • Year and Date: 2014-11-25 – 2014-11-27
• [Presentation] Respective functions of two distinct Siwi, a silkworm PIWI, complexes assembled during piRNA biogenesis in insect germ cells (2014)
  • Author(s): Kazumichi M. Nishida, Yuka W. Iwasaki, Yukiko Murota, Taro Mannen, Haruhiko Siomi and Mikiko C. Siomi
  • Organizer: joint Australia and Japan RNA
  • Place of Presentation: University o Technology Sydney (Australia)
  • Year and Date: 2014-11-02 – 2014-11-05
• [Presentation] カイコpiRNA生合成経路におけるBmQinの機能解析 (2013)
  • Author(s): 西田 知訓
  • Organizer: 第15回日本RNA学会年会
  • Place of Presentation: ひめぎんホール(愛媛県)
• [Presentation] BmVasa involve discharge of cleavage product from piRISC in silkworm piRNA biogenesis
  • Author(s): 西田 知訓
  • Organizer: 第14回 東京大学生命科学シンポジウム
  • Place of Presentation: 東京都