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Substrate Generation for Endonucleases of CRISPR/Cas Systems
Published on: September 8, 2012
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タイプIIIのCRISPR-Casシステムは,周期性オリゴアデニラート第2伝達物質を生成する
Ole Niewoehner1, Carmela Garcia-Doval1, Jakob T Rostøl2
1Department of Biochemistry, University of Zurich, Winterthurerstrasse 190, CH-8057 Zurich, Switzerland.
Nature
|July 20, 2017
まとめ
タイプIIIのCRISPR-Casシステムは,Csm6 RNaseを活性化するためにサイクルオリゴアデニラートセカンドメッセンジャーを使用する. この発見は生まれつきの免疫シグナル伝達に似た CRISPR 干渉の新たな制御メカニズムを明らかにしています
科学分野:
- 微生物学
- 分子生物学
- 免疫学
背景:
- タイプIIIのCRISPR-Casシステムは,侵襲的な遺伝要素に対するプロカリオート免疫を提供します.
- タイプIIIのCRISPR干渉において,Csm6タンパク質は単独のRNaseとして作用するが,その活性化メカニズムは不明であった.
研究 の 目的:
- タイプIIIのCRISPR-CasシステムでCsm6の活性が調節されるメカニズムを解明する.
- 侵入者検知とCsm6の活性化をつなぐ信号経路を特定する
主な方法:
- バイオケミカルアッセイとタイプIIIのCRISPR-Casコンポーネントの遺伝子操作を用いてCsm6の活性化を調査した.
- Cas10サブユニットとCRISPR関連ロスマン折り (CARF) ドメインのCsm6調節における役割を分析した.
- 特定の突然変異の機能的影響を評価するために in vivo 試験を用いる.
主要な成果:
- Csm6はタイプIII干渉複合体によって生成される周期性オリゴアデニラート二次メッセンジャーによって活性化されていることが示された.
- 標的RNAの結合時に,この第2のメッセンジャーを生成するCas10サブユニットを特定した.
- Csm6のCARFドメインが周期性オリゴアデニラートと結合し,アロステル活性化につながることが示された.
- Cas10とCsm6のCARFドメインの変異は,このシグナル伝達経路の重要性を確認した.
結論:
- CRISPRの干渉を制御する 前例のないメカニズムを発見しました
- Cas10によって生成される循環性オリゴアデニラートは,Csm6をアロステリックに活性化し,標的認識とRNAの分解を結びつける.
- このシグナル伝達経路は,哺乳類の先天免疫におけるオリゴアデニラートシグナル伝達と概念的に類似している.
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