TY - JOUR
T1 - The Streptococcus thermophilus CRISPR/Cas system provides immunity in Escherichia coli
AU - Sapranauskas, Rimantas
AU - Gasiunas, Giedrius
AU - Fremaux, Christophe
AU - Barrangou, Rodolphe
AU - Horvath, Philippe
AU - Siksnys, Virginijus
N1 - Funding Information:
Programme Gilibert between the Lithuanian Ministry of Education and Science and by French Ministry of Foreign and European affairs (MAEE); French Ministry of Higher education and Research (MESR); grant #R100 (3rd priority Strengthening of Capacities of Researchers and Scientists of the 2007–2013 Operational Programme for Human Resources Development implemented by the Research Council of Lithuania). Funding for open access charge: Danisco.
PY - 2011/11
Y1 - 2011/11
N2 - The CRISPR/Cas adaptive immune system provides resistance against phages and plasmids in Archaea and Bacteria. CRISPR loci integrate short DNA sequences from invading genetic elements that provide small RNA-mediated interference in subsequent exposure to matching nucleic acids. In Streptococcus thermophilus, it was previously shown that the CRISPR1/Cas system can provide adaptive immunity against phages and plasmids by integrating novel spacers following exposure to these foreign genetic elements that subsequently direct the specific cleavage of invasive homologous DNA sequences. Here, we show that the S. thermophilus CRISPR3/Cas system can be transferred into Escherichia coli and provide heterologous protection against plasmid transformation and phage infection. We show that interference is sequence-specific, and that mutations in the vicinity or within the proto-spacer adjacent motif (PAM) allow plasmids to escape CRISPR-encoded immunity. We also establish that cas9 is the sole cas gene necessary for CRISPR-encoded interference. Furthermore, mutation analysis revealed that interference relies on the Cas9 McrA/HNH-and RuvC/RNaseH-motifs. Altogether, our results show that active CRISPR/Cas systems can be transferred across distant genera and provide heterologous interference against invasive nucleic acids. This can be leveraged to develop strains more robust against phage attack, and safer organisms less likely to uptake and disseminate plasmid-encoded undesirable genetic elements.
AB - The CRISPR/Cas adaptive immune system provides resistance against phages and plasmids in Archaea and Bacteria. CRISPR loci integrate short DNA sequences from invading genetic elements that provide small RNA-mediated interference in subsequent exposure to matching nucleic acids. In Streptococcus thermophilus, it was previously shown that the CRISPR1/Cas system can provide adaptive immunity against phages and plasmids by integrating novel spacers following exposure to these foreign genetic elements that subsequently direct the specific cleavage of invasive homologous DNA sequences. Here, we show that the S. thermophilus CRISPR3/Cas system can be transferred into Escherichia coli and provide heterologous protection against plasmid transformation and phage infection. We show that interference is sequence-specific, and that mutations in the vicinity or within the proto-spacer adjacent motif (PAM) allow plasmids to escape CRISPR-encoded immunity. We also establish that cas9 is the sole cas gene necessary for CRISPR-encoded interference. Furthermore, mutation analysis revealed that interference relies on the Cas9 McrA/HNH-and RuvC/RNaseH-motifs. Altogether, our results show that active CRISPR/Cas systems can be transferred across distant genera and provide heterologous interference against invasive nucleic acids. This can be leveraged to develop strains more robust against phage attack, and safer organisms less likely to uptake and disseminate plasmid-encoded undesirable genetic elements.
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U2 - 10.1093/nar/gkr606
DO - 10.1093/nar/gkr606
M3 - Article
C2 - 21813460
AN - SCOPUS:80755145195
SN - 0305-1048
VL - 39
SP - 9275
EP - 9282
JO - Nucleic acids research
JF - Nucleic acids research
IS - 21
ER -