Invivo CRISPR screening directly targeting testicular cells

IR (HANDLE) Open Access
  • Noguchi, Yuki
    Graduate School of Biostudies, Kyoto University; Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University
  • Onodera, Yasuhito
    Global Center for Biomedical Science and Engineering, Faculty of Medicine, Hokkaido University
  • Miyamoto, Tatsuo
    Department of Molecular and Cellular Physiology, Yamaguchi University, Graduate School of Medicine
  • Maruoka, Masahiro
    Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University; Center for Integrated Biosystems, Institute of Biomedical Sciences, Academia Sinica
  • Kosako, Hidetaka
    Division of Cell Signaling, Fujii Memorial Institute of Medical Sciences, Institute of Advanced Medical Sciences, Tokushima University
  • Suzuki, Jun
    Graduate School of Biostudies, Kyoto University; Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University; Center for Integrated Biosystems, Institute of Biomedical Sciences, Academia Sinica; CREST, Japan Science and Technology Agency

Description

CRISPR-Cas9 short guide RNA (sgRNA) library screening is a powerful approach to understand the molecular mechanisms of biological phenomena. However, its in vivo application is currently limited. Here, we developed our previously established in vitro revival screening method into an in vivo one to identify factors involved in spermatogenesis integrity by utilizing sperm capacitation as an indicator. By introducing an sgRNA library into testicular cells, we successfully pinpointed the retinal degeneration 3 (Rd3) gene as a significant factor in spermatogenesis. Single-cell RNA sequencing (scRNA-seq) analysis highlighted the high expression of Rd3 in round spermatids, and proteomics analysis indicated that Rd3 interacts with mitochondria. To search for cell-type-specific signaling pathways based on scRNA-seq and proteomics analyses, we developed a computational tool, Hub-Explorer. Through this, we discovered that Rd3 modulates oxidative stress by regulating mitochondrial distribution upon ciliogenesis induction. Collectively, our screening system provides a valuable in vivo approach to decipher molecular mechanisms in biological processes.

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Details 詳細情報について

  • CRID
    1050018042603018112
  • ISSN
    2666979X
  • HANDLE
    2433/287358
  • Text Lang
    en
  • Article Type
    journal article
  • Data Source
    • IRDB

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