CRISPRi-mediated repression of three repressors induces the expression of three related bacteriophages.

The FA1090 isolate encodes nine prophage islands (Ngoɸ1-9). Ngoɸ1-3 contain genes consistent with a -dsDNA bacteriophage (phage). Saturating transposon-sequencing screens using two different isolates predicted that multiple prophage genes were essential, including three putative transcriptional repressors: (present in Ngoɸ1), (present in Ngoɸ2), and (present in Ngoɸ3). All three genes display homology to the Lambda phage , a regulator important for maintaining the lysogenic state and inhibiting lytic induction, but these proteins are not close paralogs. Using a -derived Type I-C CRISPR-interference system, we show that these orthologs are essential, as the knockdown of each gene results in bacterial death. We determined that the repression of the three orthologs resulted in the significant induction of phage gene expression. Finally, we detected -like phage particles released from following repression of , , or . We hypothesize that these orthologs are critical for preventing phage lytic infection and cell death and allow to benefit from the carriage and expression of prophage genes.IMPORTANCEBacteriophage, or phage, are bacteria-infecting viruses and are the most abundant natural entities in the world. Here, we report that 's three most complete double-stranded DNA prophage islands each encode essential and related transcriptional repressors. CRISPRi-mediated repression of these transcriptional repressors leads to a significant increase in prophage gene expression and phage induction. This study marks an important initial step in studying the interaction between and its resident phage.