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TitleMolecular mechanism of Eco8-mediated anti-phage defense.
Journal, issue, pagesMol Cell, Vol. 85, Issue 22, Page 4229-44242.e4, Year 2025
Publish dateNov 20, 2025
AuthorsLinggang Yuan / Liqiao Xu / Bing Wu / Qingyang Liu / Yue Yao / Xiaoting Hua / Yu Feng /
PubMed AbstractEscherichia coli Eco8 is an anti-phage defense system consisting of a reverse transcriptase, a class 3 overcoming lysogenization defect (OLD) nuclease, and a DNA-RNA chimera called multi-copy single- ...Escherichia coli Eco8 is an anti-phage defense system consisting of a reverse transcriptase, a class 3 overcoming lysogenization defect (OLD) nuclease, and a DNA-RNA chimera called multi-copy single-stranded DNA (msDNA). Genetic and biochemical data suggest that Eco8-mediated anti-phage defense is triggered by the phage single-stranded DNA (ssDNA)-binding proteins, but the underlying structural basis remains unknown. Here, we demonstrate that the DNA cleavage and ATP hydrolysis activities of the OLD nuclease are critical for Eco8-mediated anti-phage defense. We also determine the cryoelectron microscopy (cryo-EM) structures of Eco8 alone and in complex with the T7 phage ssDNA-binding protein. Structural analysis reveals that the reverse transcriptase, msDNA, and OLD nuclease form a megacomplex with a 4:4:4 stoichiometry. The T7 phage ssDNA-binding protein unwinds the msDNA and transforms Eco8 into an ATP-dependent DNA-degrading machinery. This study not only elucidates the molecular mechanism of Eco8-mediated anti-phage defense but also validates that msDNA serves as a sensor of phage DNA-modifying/binding proteins.
External linksMol Cell / PubMed:41172990
MethodsEM (single particle)
Resolution3.2 - 3.3 Å
Structure data

63268

9lp9

EMDB-63268, PDB-9lp9:
The cryo-EM structure of retron Eco8 in a standby state
Method: EM (single particle) / Resolution: 3.2 Å

63269

9lpa

EMDB-63269, PDB-9lpa:
The cryo-EM structure of retron Eco8 in an active state
Method: EM (single particle) / Resolution: 3.3 Å

Chemicals

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

Source
  • escherichia coli (E. coli)
KeywordsANTITOXIN/DNA/RNA / retron / multi-copy single-stranded DNA / Eco8 / anti-phage defense / OLD-family endonuclease / reverse transcriptase / ANTITOXIN-DNA-RNA complex

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