Endogenous CRISPR-Cas System-Based Genome Editing and Antimicrobials: Review and Prospects.

Yingjun Li,Nan Peng

doi:10.3389/fmicb.2019.02471

Yingjun Li, Nan Peng

Open Access

https://doi.org/10.3389/fmicb.2019.02471

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Journal: Frontiers in microbiology	Publication Date: Oct 25, 2019
Citations: 40	License type: CC BY 4.0

Affiliation: Huazhong Agricultural University

Abstract

CRISPR-Cas systems adapt “memories” via spacers from viruses and plasmids to develop adaptive immunity against mobile genetic elements. Mature CRISPR RNAs guide CRISPR-associated nucleases to site-specifically cleave target DNA or RNA, providing an efficient genome engineering tool for organisms of all three kingdoms. Cas9, Cas12, and Cas13 are single proteins with multiple domains that are the most widely used CRISPR nucleases of the Class 2 system. However, these CRISPR endonucleases are large in size, leading to difficulty for manipulation and toxicity for cells. Most archaeal genomes and half of the bacterial genomes encode different types of CRISPR-Cas systems. Therefore, developing endogenous CRISPR-Cas systems-based genome editing will simplify manipulations and increase editing efficiency in prokaryotic cells. Here, we review the current applications and discuss the prospects of using endogenous CRISPR nucleases for genome engineering and CRISPR-based antimicrobials.

Highlights

Site-specific nucleases which can introduce targeted DNA double-strand break (DSB) are employed for genome editing, facilitating the identification, characterization, and modification of important genetic element in the study of biological processes
We introduce basic concept and principle of CRISPR-CRISPR associated (Cas) systems, and summarize pre-existing applications of the endogenous CRISPR-Cas systems for genome editing in bacteria and archaea
Type VI systems encode the single effector protein Cas13 that cleaves target ssRNA, and the target RNA complementary to the CRISPR RNA (crRNA) can activate Cas13 to degrade collateral ssRNAs, similar to the ssDNA cleavage activity of Cas10 in type III systems (Shmakov et al, 2015; Abudayyeh et al, 2016; East-Seletsky et al, 2016; Smargon et al, 2017; Liu et al, 2017a); this property of Cas13a has been used for pathogen detection (East-Seletsky et al, 2016; Khambhati et al, 2019)

Summary

INTRODUCTION

Site-specific nucleases which can introduce targeted DNA double-strand break (DSB) are employed for genome editing, facilitating the identification, characterization, and modification of important genetic element in the study of biological processes. During the final stage of CRISPR-Cas-meditated immunity, Cas proteins and mature CRISPR RNA (crRNA) form a ribonucleoprotein complex (RNP) that recognizes and cleaves invading foreign DNA (or RNA) via base pairing of crRNA and target nucleic acids (Figure 1, see details below) (Brouns et al, 2008). Type VI systems encode the single effector protein Cas that cleaves target ssRNA, and the target RNA complementary to the crRNA can activate Cas to degrade collateral ssRNAs, similar to the ssDNA cleavage activity of Cas in type III systems (Shmakov et al, 2015; Abudayyeh et al, 2016; East-Seletsky et al, 2016; Smargon et al, 2017; Liu et al, 2017a); this property of Cas13a has been used for pathogen detection (East-Seletsky et al, 2016; Khambhati et al, 2019). Cas12a and Cas12b RNP complexes can catalyze non-specific ssDNA cleavage when bound with a complementary target ssDNA as an activator (Figure 1; Chen et al, 2018; Li et al, 2018)

HARNESSING ENDOGENOUS CRISPRCAS SYSTEMS FOR GENOME EDITING

FINAL REMARKS AND PROSPECT