Prokaryotes employ CRISPR-Cas (clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins) adaptive immune systems to protect against viral infection. The CRISPR-Cas systems are encoded by about 90% of archaea and 50% of bacteria. A typical CRISPR locus is composed of an array of short direct repeats and interspersed spacer sequences (short DNA sequences from invading viruses), which is flanked by diverse cas genes. The rapid progression of CRISPR-Cas technology, from the discovery of its DNA cutting-activity to its widespread adoption as a genome-editing tool, has been astonishing. The technology have transformed scientists' ability to manipulate genomes and study molecular networks, and concurrently opened novel avenues for the treatment of genetic diseases.
The CRISPR-Cas immunity response consists of three stages. During the adaptation stage, also known as spacer acquisition, the processed foreign DNA (known as the protospacer) is integrated into the host CRISPR locus, yielding a new spacer. The CRISPR RNA (crRNA) expression and processing stage involves transcription of the CRISPR locus into a single pre-crRNA and further processing into mature crRNAs before being incorporated into the CRISPR-Cas interference modules. In the interference stage, a single Cas protein (or complex) uses the crRNA as a guide to cleave phage nucleic acid or plasmid bearing a complementary sequence to the spacer sequence of the crRNA. In type Ⅱ CRISPR-Cas system, The sgRNA, which consists of crRNA- and tracrRNA-derived sequences, directs Cas9 nuclease cleavage of the corresponding target to initiate gene editing. All of the components of CRISPR play important roles in the adaptive immune processes.
Fig 1. The stages of CRISPR-Cas immunity.
Fig 2. Function of Anti-CRISPR (Acr) protein.
CRISPR-Cas systems are bacterial anti-viral systems. Prokaryotes utilize CRISPR-mediated adaptive immune systems to kill the invading phages and mobile genetic elements, and in turn, the viruses evolve diverse anti-CRISPR (Acr) proteins to evade that immunity. While Acrs are prevalent in phages capable of lying dormant in a CRISPR-carrying host, their orthologs have been observed only infrequently in virulent phages. The ﬁeld of Acrs has rapidly garnered interest, largely due to potential applications modulating the cleavage activity of various Cas9 proteins. Tight control over Cas9 could prevent off-target cleavage in genome-editing applications, or lock Cas9 into useful catalytically inactive states. Bioinformatic methodologies have uncovered a number of Acrs that interfere with different types of CRISPR-Cas systems in a variety of manners.
Up to now, a total of 23 distinct families of anti-CRISPR genes have been reported. The proteins encoded by these genes are entirely distinct, with low sequence similarity. Based on the classification of target CRISPR immunity systems, these proteins are divided into two classes, Class I anti-CRISPRs and Class II anti-CRISPRs. Because many human pathogens encode CRISPR-Cas systems, phages used for gene therapy could be outﬁtted with a variety of anti-CRISPRs to expand their host range and prevent the bacterial adaptive immune response from being mounted. The discovery of anti-CRISPR proteins has opened up a new area of phage research and has provided a valuable addition to the CRISPR toolbox. Although lots of anti-CRISPRs have been successfully identified, there is a long way for us to go to unveil the details of this evolutionary war.
1. AP Hynes et al. Widespread anti-CRISPR proteins in virulent bacteriophages inhibit a range of Cas9 proteins. Nature Communications. 2018;9:2919.
2. Zhu et al. Structural insights into the inactivation of CRISPR-Cas systems by diverse anti-CRISPR proteins. BMC Biology, (2018) 16:32.
3. Luciano A. Marraffini. CRISPR-Cas immunity in prokaryotes. NATURE. 1 OCTOBER 2015; 526:55-61.
4. Pawluk et al. Anti-CRISPR: discovery, mechanism and function. NATURE REVIEWS | MICROBIOLOGY. JANUARY 2018; VOLUME 16:12-16.