A Unique Type V CRISPR-Cas System Encoded by a Group of Thermus Viruses

CRISPR-Cas are widespread adaptive immune systems that protect bacteria and archaea from mobile genetic elements such as bacteriophages. Metagenomic sequencing identified CRISPR-Cas systems in phage genomes; however, their functions remain largely unknown. Here, we present Cas12r-CRISPR, a novel type V CRISPR-Cas system encoded by Lalka phages infecting thermophilic Thermus bacteria. We determined Cas12r-CRISPR PAM consensus sequence and crRNA structure and showed, that when provided with appropriate spacers and expressed in Thermus thermophilus , Cas12r-CRISPR efficiently interferes with plasmid transformation as well as infection by diverse Thermus phages. In the course of Lalka phage infection, the Cas12r-CRISPR locus is expressed with middle phage genes and its transcripts are among the most abundant phage RNAs. Notably, most Cas12r-CRISPR spacers target integrative mobile elements widespread in Thermus genomes. Both Lalka phages and targeted integrative mobile elements use host tRNA genes as attachment sites. We therefore propose that Cas12r-CRISPR participates in an inter-MGEs conflict.