Phage homing endonuclease amplifies anti-defense genes to evade bacterial immunity

Bacteriophages must overcome diverse bacterial immune systems, yet the molecular principles enabling such escape remain poorly understood. Here, we show that the phage homing endonuclease SegB facilitates immune evasion by promoting the segmental amplification of anti-defense loci. The antiphage defense Septu inhibits phage T6 replication by cleaving the variable loop of tRNA^Tyr. We show that SegB enables immune evasion by amplifying a genomic segment that contains the full-length tRNA^Tyr gene. This repeat expansion increases tRNA^Tyr expression, allowing the phage to overcome Septu immunity. SegB also mediates in trans amplification of distinct anti-defense genes that counteract OLD and toxin-antitoxin ToxIN defense systems. Collectively, our findings demonstrate that SegB-mediated segmental amplification represents a versatile mechanism by which phages rapidly adapt to and circumvent diverse bacterial antiphage defenses.