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シロイヌナズナ単離細胞の概日リズム特性の研究

中村, 駿志 京都大学 DOI:10.14989/doctor.k23742

2022.03.23

概要

地球の自転による周期的な環境変化である昼夜サイクルに適応するために、多くの生物が概日時計を有している。概日時計は約24時間の周期のリズム(概日リズム)を生成し、環境変動をあらかじめ予測することによって生体内の状態を最適化している。シロイヌナズナ植物においては、約20個の時計遺伝子の転写―翻訳フィードバックループによって概日リズムが生み出されている。植物全体の細胞で時計遺伝子が発現することから、植物の概日時計は細胞時計の集合体として捉えることができるが、全ての細胞が均質な概日時計を有しているわけではない。先行研究から植物は組織特異的な細胞概日時計の集合体であり、茎頂から根などの他の器官への時間情報伝達が示唆された他、細胞時計は周りの細胞からの影響を受けていると考えられている。つまり、植物は器官や組織によって異なる不均一な細胞時計間を統合することによって、植物個体として外部環境の変動や細胞ごとの時計のばらつきに適応している。しかし、組織からの影響を受けない細胞時計の特徴や役割は依然として不明のままであった。本研究では、細胞自身が持つ固有の概日時計の特徴を明らかにすることによって、植物が細胞時計を統合し、個体として環境に適応する仕組みを理解することを目指した。プロトプラストとして細胞を物理的に単離し、個々の細胞概日リズムを様々な条件で調べることで、細胞が本来持つ概日時計の特徴を明らかにした。
 第1章では、シロイヌナズナの葉から単離した細胞の発光概日リズムを長期的に測定できる条件を確立した。従来の方法では葉由来単離細胞集団の概日リズムは急激に減衰し、3日程度しか発光概日リズムを検出することができなかった。培地条件や細胞密度を検討することで単離細胞集団の概日リズムを11日以上測定することに成功し、様々な概日リズム解析を可能にする実験系を確立した。
 第2章では、単一細胞レベルでの単離細胞の発光概日リズム測定系を確立し、細胞時計の特徴を明らかにした。概日リズムの三大性質である「自由継続性」、「温度補償性」、「同調性」を単離細胞が有することを明らかにし、植物の細胞一つ一つの概日時計が細胞自律的に機能することを実証した。また、葉由来単離細胞に加えて根由来単離細胞の概日リズムを調べることによって器官特異的な細胞時計の存在を示した。
 第3章では、単離細胞の細胞間ではたらく作用によって単離細胞の概日リズムの安定性や明暗に対する細胞間での正確性を向上させていることを示した。葉由来単離細胞を高細胞密度下でみられるその作用は時刻情報とは直接関係のない細胞の分泌物質によって引き起こされていることを明らかにした。
 単離細胞の発光概日リズムを単一細胞レベルで解析することによって、組織からの影響を受けない植物細胞時計の特徴を明確に示した。単離細胞という人工的な条件ではあるが、植物の概日時計研究を発展させるための基本的な知見を見出したことに加え、葉と根の細胞時計が植物の地上部や地下部といった植物内での不均一な環境に適合した性質を持つとのコンセプトを提唱した。

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