CRISPR-Cas: Adapting to change

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Science  07 Apr 2017:
Vol. 356, Issue 6333, eaal5056
DOI: 10.1126/science.aal5056

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Variation in prokaryote adaptive immunity

To repel infection by phage and mobile genetic elements, prokaryotes have a form of adaptive immune response and memory invested in clustered regularly interspaced short palindromic repeats and associated proteins (CRISPR-Cas). This molecular machinery can recognize and remember foreign nucleic acids by capturing and retaining small nucleotide sequences. On subsequent encounters, the cognate CRISPR-Cas marshals enzymatic defenses to destroy infecting elements that contain the same sequences. Jackson et al. review the molecular mechanisms by which diverse CRISPR-Cas systems adapt and anticipate novel threats and evasive countermeasures from mobile genetic elements.

Science, this issue p. eaal5056

Structured Abstract


The arms race between prokaryotes and their perpetually evolving predators has fueled the evolution of a defense arsenal. The so-called CRISPR-Cas systems—clustered regularly interspaced short palindromic repeats and associated proteins—are adaptive immune defense systems found in bacteria and archaea. The recent exponential growth of research in the CRISPR field has led to the discovery of a diverse range of CRISPR-Cas systems and insight into their defense functions. These systems are divided into two major classes and six types. Each system consists of two components: a locus for memory storage (the CRISPR array) and cas genes that encode the machinery driving immunity. Information stored within CRISPR arrays is used to direct the sequence-specific destruction of invading genetic elements, including viruses and plasmids. As such, all CRISPR-Cas immune systems are reliant on the formation of CRISPR memories, known as spacers, to facilitate future defense. To form these memories, small fragments of invader nucleic acids are added as spacers to the CRISPR memory banks in a process termed CRISPR adaptation. The genetic basis of immunity means that CRISPR adaptation provides heritable benefits, an attribute that is unparalleled in eukaryotic immune systems. There is widespread evidence of highly active CRISPR adaptation in nature, and it is clear that these systems play important roles in shaping microbial evolution and global ecological networks.


CRISPR adaptation requires several processes, including selection and processing of spacer precursors and their subsequent localization to, and integration into, the CRISPR loci. Although our understanding of all facets of the CRISPR adaptation pathway is not yet complete, considerable progress has been made in the past few years. At the heart of CRISPR adaptation is a protein complex, the Cas1-Cas2 “workhorse,” which catalyzes the addition of new spacers to CRISPR memory banks. A combination of functional assays and high-resolution structures of Cas1-Cas2 complexes has recently led to major advances. There is now a sound understanding of how foreign DNA is converted to prespacer substrates and captured by the Cas1-Cas2 complex. After this, Cas1-Cas2 locates the genomic CRISPR locus and docks in the appropriate position for insertion of the new spacer into the CRISPR array, while duplicating a CRISPR repeat. The cues directing the docking of substrate-laden Cas1-Cas2 differ between systems, with some relying on intrinsic sequence specificity and others assisted by host proteins.

Before integration, accurate processing of the spacer precursors is required to ensure that the new spacers are compatible with the protein machinery in order to elicit CRISPR-Cas defense. For a given CRISPR-Cas system, spacers must typically be of a certain length and be inserted into the CRISPR in a specific orientation. It is becoming increasingly apparent that Cas1-Cas2 complexes from diverse systems are capable of ensuring that these system-specific factors are met with high fidelity.

New findings also account for the ordering of stored memories: Typically, the insertion of new spacers is directed to one end of CRISPR arrays, and it has been shown that this enhances immunity against recently encountered invaders. The chronological ordering of new spacers has enabled insights into the temporal dynamics of interactions between hosts and invaders that are constantly changing. Some CRISPR-Cas systems use existing spacers to recognize previously encountered elements and promote the formation of new CRISPR memories, a process known as primed CRISPR adaptation. Viruses and plasmids that have escaped previous CRISPR-Cas defenses through genetic mutations trigger primed CRISPR adaptation. Several recent studies have revealed that primed CRISPR adaptation is also strongly promoted by recurrent invaders, even in the absence of escape mutations. This has led to previously separate paradigms of invader destruction and primed CRISPR adaptation beginning to converge into a unified model.


CRISPR adaptation is crucial for ensuring both population-level protection through spacer diversity and protection of the host through invader clearance. Although many studies have explored CRISPR adaptation in a broad range of host-specific and metagenomic contexts, much of the mechanistic detail has been gleaned from studying a relatively small subset of systems. Thus, despite the relative wealth of mechanistic information about CRISPR adaptation in a few specific types, work in other systems continues to reveal distinct modes of operation for spacer acquisition. Therefore, studies of CRISPR adaptation in alternative systems are necessary to determine which processes are conserved and which are system-specific. An important remaining question is why the enhanced primed CRISPR adaptation commonly found in type I systems has not yet been observed in other types. Do other systems possess analogous mechanisms that have yet to be discovered, or does the absence of priming in these systems explain the prevalence of type I systems in nature? Future expansion of our understanding of how CRISPR adaptation is carried out in the diverse repertoire of CRISPR-Cas systems is vital for maximizing the potential for repurposing the spacer acquisition machinery in biotechnological applications. Commandeering CRISPR adaptation for on-demand memory formation will usher in a new era of biological information storage, with many applications that await discovery.

Many hues of the CRISPR-Cas adaptation machinery.

The complex of the Cas1 and Cas2 proteins, which is the workhorse of CRISPR adaptation in diverse CRISPR-Cas prokaryotic immune systems, is depicted with a DNA substrate. Despite the near-ubiquitous Cas1-Cas2 molecular machinery, type-specific differences in the insertion of new information into CRISPR memory banks are beginning to come to light.


Bacteria and archaea are engaged in a constant arms race to defend against the ever-present threats of viruses and invasion by mobile genetic elements. The most flexible weapons in the prokaryotic defense arsenal are the CRISPR-Cas adaptive immune systems. These systems are capable of selective identification and neutralization of foreign DNA and/or RNA. CRISPR-Cas systems rely on stored genetic memories to facilitate target recognition. Thus, to keep pace with a changing pool of hostile invaders, the CRISPR memory banks must be regularly updated with new information through a process termed CRISPR adaptation. In this Review, we outline the recent advances in our understanding of the molecular mechanisms governing CRISPR adaptation. Specifically, the conserved protein machinery Cas1-Cas2 is the cornerstone of adaptive immunity in a range of diverse CRISPR-Cas systems.

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