2024年新着論文 18　細胞情報学分野から論文が発表されました

In vivo CRISPR screening directly targeting testicular cells

Cell Genom. 2024 Feb 27:100510. doi: 10.1016/j.xgen.2024.100510. Online ahead of print.

Authors

Yuki Noguchi¹, Yasuhito Onodera², Tatsuo Miyamoto³, Masahiro Maruoka⁴, Hidetaka Kosako⁵, Jun Suzuki⁶

Affiliations

¹ Graduate School of Biostudies, Kyoto University, Konoe-cho, Yoshida, Sakyoku, Kyoto 606-8501, Japan; Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University, Yoshida-Honmachi, Sakyoku, Kyoto 606-8501, Japan.
² Global Center for Biomedical Science and Engineering, Faculty of Medicine, Hokkaido University, N15W7 Kita-ku, Sapporo, Hokkaido 060-8638, Japan.
³ Department of Molecular and Cellular Physiology, Yamaguchi University, Graduate School of Medicine, 1-1-1 Minami-Kogushi, Ube, Yamaguchi 755-8505, Japan.
⁴ Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University, Yoshida-Honmachi, Sakyoku, Kyoto 606-8501, Japan; Center for Integrated Biosystems, Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan.
⁵ Division of Cell Signaling, Fujii Memorial Institute of Medical Sciences, Institute of Advanced Medical Sciences, Tokushima University, 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
⁶ Graduate School of Biostudies, Kyoto University, Konoe-cho, Yoshida, Sakyoku, Kyoto 606-8501, Japan; Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University, Yoshida-Honmachi, Sakyoku, Kyoto 606-8501, Japan; Center for Integrated Biosystems, Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan; CREST, Japan Science and Technology Agency, Kawaguchi, Saitama 332-0012, Japan. Electronic address: jsuzuki@icems.kyoto-u.ac.jp.

PMID: 38447574
DOI: 10.1016/j.xgen.2024.100510

Free article

Abstract

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.

Keywords: CRISPR screening; Hub-Explorer; in vivo genome-wide screening; multi-omics; spermatogenesis.

Conflict of interest statement

Declaration of interests J.S. and Y.N. are inventors on a patent application of the in vivo genome-wide screening method toward spermatogenesis.

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