Details

Autor(en) / Beteiligte

• Shiriaeva, Anna A
• Kuznedelov, Konstantin
• Fedorov, Ivan
• Musharova, Olga
• Khvostikov, Timofey
• Tsoy, Yuliya
• Kurilovich, Elena
• Smith, Gerald R
• Semenova, Ekaterina
• Severinov, Konstantin

Titel

Host nucleases generate prespacers for primed adaptation in the E. coli type I-E CRISPR-Cas system

Ist Teil von

• Science advances, 2022-11, Vol.8 (47), p.eabn8650-eabn8650

Ort / Verlag

United States: American Association for the Advancement of Science

Erscheinungsjahr

2022

Link zum Volltext

Quelle

Free E-Journal (出版社公開部分のみ）

Beschreibungen/Notizen

• CRISPR-Cas systems provide prokaryotes with adaptive immunity against foreign nucleic acids. In , immunity is acquired upon integration of 33-bp spacers into CRISPR arrays. DNA targets complementary to spacers get degraded and serve as a source of new spacers during a process called primed adaptation. Precursors of such spacers, prespacers, are ~33-bp double-stranded DNA fragments with a ~4-nt 3' overhang. The mechanism of prespacer generation is not clear. Here, we use FragSeq and biochemical approaches to determine enzymes involved in generation of defined prespacer ends. We demonstrate that RecJ is the main exonuclease trimming 5' ends of prespacer precursors, although its activity can be partially substituted by ExoVII. The RecBCD complex allows single strand-specific RecJ to process double-stranded regions flanking prespacers. Our results reveal intricate functional interactions of genome maintenance proteins with CRISPR interference and adaptation machineries during generation of prespacers capable of integration into CRISPR arrays.