| 研究課題/領域番号 |
22K20651
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| 研究種目 |
研究活動スタート支援
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| 配分区分 | 基金 |
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| 審査区分 |
0702:細胞レベルから個体レベルの生物学およびその関連分野
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| 研究機関 | 名古屋大学 |
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研究代表者 |
Kozgunova Elena 名古屋大学, 高等研究院(理), 特任助教 (90786120)
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| 研究期間 (年度) |
2022-08-31 – 2025-03-31
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| 研究課題ステータス |
交付 (2023年度)
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| 配分額 *注記 |
2,860千円 (直接経費: 2,200千円、間接経費: 660千円)
2023年度: 1,430千円 (直接経費: 1,100千円、間接経費: 330千円)
2022年度: 1,430千円 (直接経費: 1,100千円、間接経費: 330千円)
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| キーワード | cell division / Physcomitrium patens / genetic screening / CRISPR/Cas9 / ヒメツリガネゴケ / CRISPR遺伝子編集 / 細胞分裂 / コケ / 植物 |
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| 研究開始時の研究の概要 |
運動性のある動物細胞とは異なり、多くの植物細胞は硬い細胞壁の中に籠もっており、細胞分裂の際に生じたエラーは、その後に修正することができず、植物の発達を妨げることになる。植物は動物とは異なる細胞分裂機構を発達させてきたというのが長年の仮説である。一方で、植物の細胞分裂に関する知識の多くは、動物の制御遺伝子のホモログの機能解析により得られている. そのため、植物特有の細胞分裂の仕組みについては、動物研究の進歩の範囲内での知識の拡大にとどまる。本研究の目的は、細胞分裂、特に紡錘体の形成と配置に関わる植物特異的な遺伝子を、独創的かつ革新的なスクリーニング手法で見つけ出し、その機能を解明することである。
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| 研究実績の概要 |
The core idea of this project is to combine CRISPR gene editing with the technological advantages of the model plant Physcomitrium patens (Physcomitrium patens) to achieve genetic screening with unprecedented speed and efficiency. This study focuses on genes that have not yet been functionally evaluated. We used transcriptome data from various cell cycles and developmental stages to create a library with genes that are thought to be involved in cell division. Then, about 800 genes were selected and genetic screening using CRISPR/Cas9 gene editing was carried out.
Based on the CRISPR/Cas9 screening in moss Physcomitrium patens, we identified three promising mutants, that are likely involved in the plant cell division and cytoskeleton organization. These genes have not been functionally analyzed previously, therefore we were able to identify novel players in plant cell division using CRISPR/Cas9 screening. Two of identified genes are conserved in bryophytes and ferns, but appear to be lost in angiosperms, suggesting that cell division machinery has evolved and lost certain components during plant evolution. Another gene is highly conserved across multiple plant species. Next, we plan to focus on detailed functional analysis of newly identified genes.
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
3: やや遅れている
理由
At present, we are carrying out functional analysis of genes identified through CRISPR/Cas9 screening. We perform localization check by expressing respective protein fusion with GFP and observing where does protein localize inside the cell. We found that one of the proteins localize to the chromosomes during cell division, another one to the spindle and phragmoplast midzone. Due to certain problems, such as low level of protein expression, we so far were unable to observe localization of the third protein and are currently working to resolve this problem by overexpressing the protein. In addition , we perform detailed imaging analysis of mutants to gain insights into each gene function. We are also currently preparing manuscript for publication based on these data.
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| 今後の研究の推進方策 |
In future, we will further investigate protein functions. In addition to localization analysis and mutant phenotype, we plan to perform in vitro assays. By purifying the protein and testing its interaction with DNA or microtubules, we can gain better understanding of its function. We also aim to reveal protein-protein interactions through co-immunoprecipitation to isolate protein complexes, followed by mass-spectrometry to identify binding partners.
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