CRISPR interference: RNA-directed adaptive immunity in bacteria and archaea

Abstract

Sequence-directed genetic interference pathways control gene expression and preserve genome integrity in all kingdoms of life. The importance of such pathways is highlighted by the extensive study of RNA interference (RNAi) and related processes in eukaryotes. In many bacteria and most archaea, clustered, regularly interspaced short palindromic repeats (CRISPRs) are involved in a more recently discovered interference pathway that protects cells from bacteriophages and conjugative plasmids. CRISPR sequences provide an adaptive, heritable record of past infections and express CRISPR RNAs -small RNAs that target invasive nucleic acids. Here, we review the mechanisms of CRISPR interference and its roles in microbial physiology and evolution. We also discuss potential applications of this novel interference pathway.The acquisition of new genes that confer a selective advantage is an important factor in genome evolution. Considerable proportions of bacterial and archaeal genomes consist of genes derived from the exchange of genetic material among related or unrelated species 1 , which is known as horizontal gene transfer (HGT). HGT occurs by uptake of environmental DNA (transformation) or by the incorporation of heterologous DNA carried on mobile genetic elements, such as plasmids (conjugation) and bacteriophages (transduction) 2 .However, only a miniscule fraction of acquired genes confers an immediate selective advantage. Therefore bacteria and archaea have developed many mechanisms to prevent HGT, such as DNA restriction and surface exclusion 2 . Recently, arrays of clustered, regularly interspaced short palindromic repeats (CRISPRs) have been identified as determinants of a novel genetic interference pathway that limits at least two major routes of HGT -conjugation and transduction. Like eukaryotic RNA interference (RNAi) and Competing interests statementThe authors declare no competing financial interests. DATABASES NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript related pathways (with which CRISPR interference is analogous but not homologous), CRISPR interference provides the host with an efficient antiviral defence mechanism.In contrast to other gene transfer and phage defence mechanisms, CRISPR interference is an adaptive immune system that can be reprogrammed to reject invading DNA molecules that have not been previously encountered. CRISPRs are separated by short spacer sequences that match bacteriophage or plasmid sequences and specify the targets of interference. Upon phage infection, CRISPR arrays can acquire new repeat-spacer units that match the challenging phage. Cells with this extended CRISPR locus will survive phage infection and thrive. Therefore the spacer content of CRISPR arrays reflects the many different phages and plasmids that have been encountered by the host, and these spacers can be expanded rapidly in response to new invasions. Accordingly, CRISPRs constitute a 'genetic memory' that ensures the rejection of new, returning and ever-present invading DNA m...

Novel configurations of type I and II CRISPR–Cas systems in Corynebacterium diphtheriae

“…CRISPR arrays can rapidly expand in response to new invasions by different phages and plasmids (Marraffini & Sontheimer, 2010). Analysis of the spacer diversity among all of the CRISPR arrays found a total of 221 unique spacer sequences (Table S1).…”

“…All the strains analysed in this study were isolated from the Americas and Europe over a long period (1896-2009), including isolates from the pre-and post-vaccination era, and showed distinctive direct repeat-spacer patterns. CRISPR arrays represent hypervariable loci and spacer diversity potentially reflects a history of prior invasions by different phages and plasmids (Marraffini & Sontheimer, 2010), making them a useful target for outbreak characterization and epidemiological investigations (Mokrousov et al, 2007, Mokrousov 2013. Therefore, while CRISPR-based typing may not necessarily provide information on evolutionary relationships between different strains it may offer high levels of discrimination for strain subtyping to study local epidemiology.…”

The impact of CRISPR repeat sequence on structures of a Cas6 protein–RNA complex

“…The clustered regularly interspaced short palindromic repeats (CRISPRs) are found in many bacteria and archaea genomes and are recently demonstrated to confer small RNA-based adaptive immunity against invasive genetic elements. [1][2][3][4][5][6][7][8] CRISPR loci typically consist of several to hundreds of short ($30-40-nucleotide) repeats separated by short ($30-40-nucleotide) variable sequences (spacers) and are associated with one or more heterogenous families of cas genes that encode Cas proteins. 9,10 The spacer sequences are short DNA segments derived from past infection and they establish the specificity for obstruction of future invasion.…”

Advancing biotechnology with CRISPR/Cas9: recent applications and patent landscape

“…1a). Once guided to the target, Cas9 catalytically cleaves the DNA sequence 3 nt upstream the 5′-NGG PAM, resulting in the activation of endogenous repair mechanisms, such as homologous recombination (HR) or non-homologous end joining (NHEJ) [2,71] (Fig. 1b).…”