The Ppl protein senses 3'-hydroxyl DNA overhangs and NTP depletion to halt phage infection

The continuous battle between hosts and viruses leads to the development of numerous antiviral defense systems. The recent discovery of previously unrecognized defense modules has greatly expanded our understanding of the interaction between hosts and viruses. The Ppl defense system of Escherichia coli is a single-protein module that provides antiphage immunity through an unknown mechanism. Here, we show that upon phage infection, the vigorous phage replication consumes cellular nucleotide triphosphate (NTP), eliminating the inhibitory effect of the C-terminal NTPase on the N-terminal polymerase and histidinol phosphatase (PHP) exonuclease domain. Nevertheless, the 3'-5' exonuclease activity of the PHP domain remains harmless to the cells until it detects 3'-hydroxyl (-OH) overhangs in DNA caused by phage-encoded homing endonucleases, such as Gp5.3, thereby triggering "abortive infection" (Abi). The activation of the Ppl defense system necessitates the concurrent presence of two signals, highlighting a sophisticated mechanism to inhibit autoimmunity.