Structures of the CRISPR genome integration complex

Wright AV, Liu JJ, Knott GJ, Doxzen KW, Nogales E, Doudna JA.

Science. 2017; 357 (6356):1113-1118.

Speaker: Chia-Yu Kang (康家瑜)                                    Time: 15:00~16:00, Apr. 25, 2018

Commentator: Dr. Wen-Yih Jeng (鄭文義老師)         Place: Room 601

Abstract

  Bacterial and archaeal insert spacer sequence acquired from foreign DNA into CRISPR loci to generate bacterial adaptive immunity (1). The CRISPR-Cas systems depend on the Cas1-Cas2 integrase to capture and insert short foreign DNA fragment, enabling adaption to new viruses (2, 3). However, it is still not clear how Cas1-Cas2 complex recognizes the targeted sequence. Here, the authors present the structure of the Cas1-Cas2 integrase bound to both substrate and target DNA complexes, as well as a cryo-electron microscopic structure of the full CRIPSR locus integration complex with the accessory protein integration host factor (IHF). They found the Cas1-Cas2 complex from Escherichia coli shows unexpectedly indirect sequence recognition in identifying the site of integration. These processes rely mainly on active-site positioning and structural features of DNA. In addition, IHF sharply binds the target DNA that allows Cas1 to interact with an upstream recognition motif of integration site. Therefore, this mechanism contributes to increase both specificity and efficiency of integration between IHF and Cas1 to suppress off-target integration in CRISPR locus. Changes in CRISPR architecture between Cas1 active sites could reprogram the integrase to recognize different target sites. These results explain how Cas1-Cas2 integrase recognize a sequence-dependent DNA structure to ensure site-selective CRISPR array expansion during the initial step of bacterial adaptive immunity. Also, a structural context can be applied to genome tagging for genetic modification.

References

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