Discovery of genes essential for male fertility using CRISPR/Cas9-based genome editing technology

• Project/Area Number: 18K16735
• Research Category: Grant-in-Aid for Early-Career Scientists
• Allocation Type: Multi-year Fund
• Review Section: Basic Section 56030:Urology-related
• Research Institution: Osaka University
• Principal Investigator: Lu Yonggang 大阪大学, 免疫学フロンティア研究センター, 特任助教(常勤) (00817033)
• Project Period (FY): 2018-04-01 – 2022-03-31
• Project Status: Completed (Fiscal Year 2021)
• Budget Amount: ¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000); Fiscal Year 2021: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000); Fiscal Year 2020: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000); Fiscal Year 2019: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000); Fiscal Year 2018: ¥780,000 (Direct Cost: ¥600,000、Indirect Cost: ¥180,000)
• Keywords: Male reproduction / Sperm-egg fusion / Fertilization / CRISPR/Cas9 / Knockout / Mouse model / Male contraception / Spermatology / male reproduction / male contraception / sperm / sperm-egg fusion / sperm function / sperm-oocyte fusion / Sperm / Spermatogenesis / Testis / Genome editing / Male fertility / Reproductive biology / Male infertility / Gene knockout

Outline of Final Research Achievements

During the four-year funding period, the applicant has generated multiple knockout mouse lines lacking testis-specific genes and analyzed the resultant phenotype. This study demonstrates the feasibility and efficiency of using the CRISPR/Cas9-mediated knockout strategy to identify genes essential for male reproduction. Through the large-scale in vivo screening, we have discovered essential genes Lrrc23, C2cd6, Sof1, Tmem95, and Fimp, which play roles in regulating sperm motility and sperm-egg fusion. The relevant research outcomes have been published in international peer-reviewed journals.

Academic Significance and Societal Importance of the Research Achievements

My work provides explanations for idiopathic male infertility. The identified molecules may serve as biomarkers for male infertility and potential targets for non-hormonal male contraception.

Research Products

• [Journal Article] Sperm membrane proteins DCST1 and DCST2 are required for sperm-egg interaction in mice and fish (2022)
  • Author(s): Noda Taichi、Blaha Andreas、Fujihara Yoshitaka、Gert Krista R.、Emori Chihiro、Deneke Victoria E.、Oura Seiya、Panser Karin、Lu Yonggang、Berent Sara、Kodani Mayo、Cabrera-Quio Luis Enrique、Pauli Andrea、Ikawa Masahito
  • Journal Title: Communications Biology Volume: 5 Issue: 1 Pages: 1-11
  • DOI: 10.1038/s42003-022-03289-w
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] C2cd6-encoded CatSperτ targets sperm calcium channel to Ca2+ signaling domains in the flagellar membrane (2022)
  • Author(s): Hwang Jae Yeon、Wang Huafeng、Lu Yonggang、Ikawa Masahito、Chung Jean-Ju
  • Journal Title: Cell Reports Volume: 38 Issue: 3 Pages: 110226-110226
  • DOI: 10.1016/j.celrep.2021.110226
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Sperm IZUMO1 Is Required for Binding Preceding Fusion With Oolemma in Mice and Rats (2022)
  • Author(s): Matsumura Takafumi、Noda Taichi、Satouh Yuhkoh、Morohoshi Akane、Yuri Shunsuke、Ogawa Masaki、Lu Yonggang、Isotani Ayako、Ikawa Masahito
  • Journal Title: Frontiers in Cell and Developmental Biology Volume: 9 Pages: 1-10
  • DOI: 10.3389/fcell.2021.810118
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] LRRC23 is a conserved component of the radial spoke that is necessary for sperm motility and male fertility in mice (2021)
  • Author(s): Zhang Xin、Sun Jiang、Lu Yonggang、Zhang Jintao、Shimada Keisuke、Noda Taichi、Zhao Shuqin、Koyano Takayuki、Matsuyama Makoto、Zhou Shushu、Wu Jiayan、Ikawa Masahito、Liu Mingxi
  • Journal Title: Journal of Cell Science Volume: 134 Issue: 20 Pages: 1-14
  • DOI: 10.1242/jcs.259381
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Cooperation-based sperm clusters mediate sperm oviduct entry and fertilization (2021)
  • Author(s): Qu Yongcun、Chen Qi、Guo Shanshan、Ma Chiyuan、Lu Yonggang、Shi Junchao、Liu Shichao、Zhou Tong、Noda Taichi、Qian Jingjing、Zhang Liwen、Zhu Xili、Lei Xiaohua、Cao Yujing、Li Wei、Li Wei、Plachta Nicolas、Matzuk Martin M.、Ikawa Masahito、Duan Enkui、Zhang Ying、Wang Hongmei
  • Journal Title: Protein & Cell Volume: 12 Issue: 10 Pages: 810-817
  • DOI: 10.1007/s13238-021-00825-y
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] CRISPR/Cas9-based genome editing in mice uncovers 13 testis- or epididymis-enriched genes individually dispensable for male reproduction (2020)
  • Author(s): Sun Jiang、Lu Yonggang、Nozawa Kaori、Xu Zoulan、Morohoshi Akane、Castaneda Julio M、Noda Taichi、Miyata Haruhiko、Abbasi Ferheen、Shawki Hossam H、Takahashi Satoru、Devlin Darius J、Yu Zhifeng、Matzuk Ryan M、Garcia Thomas X、Matzuk Martin M、Ikawa Masahito
  • Journal Title: Biology of Reproduction Volume: N/A Issue: 2 Pages: 183-194
  • DOI: 10.1093/biolre/ioaa083
  • NAID: 120007190927
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Sperm proteins SOF1, TMEM95, and SPACA6 are required for sperm－oocyte fusion in mice (2020)
  • Author(s): Taichi Noda, Yonggang Lu, Yoshitaka Fujihara, View ORCID ProfileSeiya Oura, View ORCID ProfileTakayuki Koyano, Sumire Kobayashi, Martin M. Matzuk, and View ORCID ProfileMasahito Ikawa
  • Journal Title: Proceedings of the National Academy of Sciences Volume: 117 Issue: 21 Pages: 11493-11502
  • DOI: 10.1073/pnas.1922650117
  • NAID: 120006957465
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Spermatozoa lacking Fertilization Influencing Membrane Protein (FIMP) fail to fuse with oocytes in mice (2020)
  • Author(s): Fujihara Yoshitaka、Lu Yonggang、Noda Taichi、Oji Asami、Larasati Tamara、Kojima-Kita Kanako、Yu Zhifeng、Matzuk Ryan M.、Matzuk Martin M.、Ikawa Masahito
  • Journal Title: Proceedings of the National Academy of Sciences Volume: 117 Issue: 17 Pages: 9393-9400
  • DOI: 10.1073/pnas.1917060117
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Prss55 but not Prss51 is required for male fertility in mice† (2020)
  • Author(s): Kobayashi Kiyonori、Endo Tsutomu、Matsumura Takafumi、Lu Yonggang、Yu Zhifeng、Matzuk Martin M、Ikawa Masahito
  • Journal Title: Biology of Reproduction Volume: - Issue: 2 Pages: 223-234
  • DOI: 10.1093/biolre/ioaa041
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] CIB4 Is Essential for the Haploid Phase of Spermatogenesis in Mice (2020)
  • Author(s): Zoulan Xu, Haruhiko Miyata, Yuki Kaneda, Julio M Castaneda, Yonggang Lu, Akane Morohoshi, Zhifeng Yu, Martin M Matzuk, Masahito Ikawa
  • Journal Title: Biology of Reproduction Volume: 102 Issue: 2 Pages: 1-9
  • DOI: 10.1093/biolre/ioaa059
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] CRISPR/Cas9-mediated genome editing reveals 30 testis-enriched genes dispensable for male fertility in mice† (2019)
  • Author(s): Yonggang Lu, Oura Seiya, Matsumura Takafumi, et al.
  • Journal Title: Biology of Reproduction Volume: 101 Issue: 2 Pages: 501-511
  • DOI: 10.1093/biolre/ioz103
  • Peer Reviewed / Open Access / Int'l Joint Research