Functional screening of a soil metagenomic library identifies prokaryotic DNA glycosylases involved in antiviral defense

Bacteria adapt to phage predation through the evolution of a vast assortment of defense systems. However, the relatively scant availability of prokaryotic sequencing data prevents full access to the diversity of bacterial immune mechanisms. To overcome this limitation, we infected E. coli carrying a soil metagenomic DNA library with the lytic coliphage T4 to isolate clones carrying protective genes. Following this approach, we identified Brig1, a DNA glycosylase that excises alpha-glucosyl-hydroxymethylcytosine nucleobases from the bacteriophage T4 genome to generate abasic sites and inhibit viral replication. Brig1 homologs that provide immunity against T-even phages are present in the genus Nocardioides and additional homologs are found in multiple anti-phage defense loci across distinct clades of bacteria. Our study highlights the benefits of screening unsequenced DNA and reveals prokaryotic DNA glycosylases as important players in the bacteria-phage arms race.