RESUMO
Prokaryotes have developed numerous defense strategies to combat the constant threat posed by the diverse genetic parasites that endanger them. Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas loci guard their hosts with an adaptive immune system against foreign nucleic acids. Protection starts with an immunization phase, in which short pieces of the invader's genome, known as spacers, are captured and integrated into the CRISPR locus after infection. Next, during the targeting phase, spacers are transcribed into CRISPR RNAs (crRNAs) that guide CRISPR-associated (Cas) nucleases to destroy the invader's DNA or RNA. Here we describe the many different molecular mechanisms of CRISPR targeting and how they are interconnected with the immunization phase through a third phase of the CRISPR-Cas immune response: primed spacer acquisition. In this phase, Cas proteins direct the crRNA-guided acquisition of additional spacers to achieve a more rapid and robust immunization of the population.
Assuntos
Bactérias/genética , Sistemas CRISPR-Cas/genética , Imunidade/genética , Animais , DNA/genética , RNA/genéticaRESUMO
Type II CRISPR-Cas systems defend prokaryotes from bacteriophage infection through the acquisition of short viral DNA sequences known as spacers, which are transcribed into short RNA guides to specify the targets of the Cas9 nuclease. To counter the potentially devastating propagation of escaper phages with mutations in the target sequences, the host population acquires many different spacers. Whether and how pre-existing spacers in type II systems affect the acquisition of new ones is unknown. Here, we demonstrate that previously acquired spacers promote additional spacer acquisition from the vicinity of the target DNA site cleaved by Cas9. Therefore, CRISPR immune cells acquire additional spacers at the same time as they destroy the infecting virus. This anticipates the rise of escapers or related viruses that could escape targeting by the first spacer acquired. Our results thus reveal Cas9's role in the generation of immunological memories.
Assuntos
Sistemas CRISPR-Cas/genética , DNA Intergênico/genética , DNA Viral/metabolismo , RNA Guia de Cinetoplastídeos/genética , Staphylococcus aureus/genética , Streptococcus thermophilus/genética , Bacteriófagos/genética , Proteína 9 Associada à CRISPR/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Staphylococcus aureus/imunologia , Staphylococcus aureus/virologia , Streptococcus thermophilus/imunologia , Streptococcus thermophilus/virologiaRESUMO
Viruses have evolved inhibitors to counteract the CRISPR immune response, but they are not fully potent and need some time to be expressed after the beginning of infection. In this issue of Cell, Borges et al. and Landsberger et al. show that sequential infection gradually immunosuppresses the host to allow effective CRISPR inhibition.
Assuntos
Bacteriófagos/genética , Vírus/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente EspaçadasRESUMO
Antibiotics target conserved bacterial cellular pathways or growth functions and therefore cannot selectively kill specific members of a complex microbial population. Here, we develop programmable, sequence-specific antimicrobials using the RNA-guided nuclease Cas9 (refs.1,2) delivered by a bacteriophage. We show that Cas9, reprogrammed to target virulence genes, kills virulent, but not avirulent, Staphylococcus aureus. Reprogramming the nuclease to target antibiotic resistance genes destroys staphylococcal plasmids that harbor antibiotic resistance genes and immunizes avirulent staphylococci to prevent the spread of plasmid-borne resistance genes. We also show that CRISPR-Cas9 antimicrobials function in vivo to kill S. aureus in a mouse skin colonization model. This technology creates opportunities to manipulate complex bacterial populations in a sequence-specific manner.
