Self versus non-self discrimination during CRISPR RNA-directed immunity

Abstract

Bacteria and archaea that take up exogenous DNA are equipped with host immunity systems that can recognize and eliminate the foreign DNA. One such system is mediated by the CRISPR genes, which encode small RNAs (crRNAs). CRISPR loci consist of repeats and spacer sequences. When the crRNA spacer sequence pairs with complementary invading DNA, it is marked for elimination. Luciano Marraffini and Erik Sontheimer resolve how the spacer DNA within the CRISPR loci themselves is not recognized as foreign; sequences outside the spacer show perfect pairing to the crRNAs while invading DNA will have mismatches. Differential complementarity of this type may underlie many types of self/non-self recognition — a key function in all immune systems. Distinguishing self from non-self is a vital function for immune systems to repel invaders without inducing autoimmunity. One system, which protects bacteria and archaea from invasion by phage and plasmid DNA, involves clustered, regularly interspaced, short palindromic repeat (CRISPR) loci. Here, in Staphylococcus epidermidis, the mechanism of CRISPR self/non-self discrimination is defined. All immune systems must distinguish self from non-self to repel invaders without inducing autoimmunity. Clustered, regularly interspaced, short palindromic repeat (CRISPR) loci protect bacteria and archaea from invasion by phage and plasmid DNA through a genetic interference pathway1,2,3,4,5,6,7,8,9. CRISPR loci are present in ∼40% and ∼90% of sequenced bacterial and archaeal genomes, respectively10, and evolve rapidly, acquiring new spacer sequences to adapt to highly dynamic viral populations1,11,12,13. Immunity requires a sequence match between the invasive DNA and the spacers that lie between CRISPR repeats1,2,3,4,5,6,7,8,9. Each cluster is genetically linked to a subset of the cas (CRISPR-associated) genes14,15,16 that collectively encode >40 families of proteins involved in adaptation and interference. CRISPR loci encode small CRISPR RNAs (crRNAs) that contain a full spacer flanked by partial repeat sequences2,17,18,19. CrRNA spacers are thought to identify targets by direct Watson–Crick pairing with invasive ‘protospacer’ DNA2,3, but how they avoid targeting the spacer DNA within the encoding CRISPR locus itself is unknown. Here we have defined the mechanism of CRISPR self/non-self discrimination. In Staphylococcus epidermidis, target/crRNA mismatches at specific positions outside of the spacer sequence license foreign DNA for interference, whereas extended pairing between crRNA and CRISPR DNA repeats prevents autoimmunity. Hence, this CRISPR system uses the base-pairing potential of crRNAs not only to specify a target, but also to spare the bacterial chromosome from interference. Differential complementarity outside of the spacer sequence is a built-in feature of all CRISPR systems, indicating that this mechanism is a broadly applicable solution to the self/non-self dilemma that confronts all immune pathways.