Recurrent acquisition of nuclease-protease pairs in antiviral immunity

Abstract Antiviral immune systems diversify by integrating new genes into existing pathways, creating new mechanisms of viral resistance. We identified genes encoding a predicted nuclease paired with a trypsin-like protease repeatedly acquired by multiple, otherwise unrelated antiviral immune systems in bacteria. Cell-based and biochemical assays revealed the nuclease is a proenzyme that cleaves DNA only after activation by its partner protease. Phylogenetic analysis showed that two distinct immune systems, Hachiman and AVAST, use the same mechanism of proteolytic activation despite their independent evolutionary origins. Examination of nuclease-protease inheritance patterns identified caspase-nuclease (canu) genomic loci that confer antiviral defense in a pathway reminiscent of eukaryotic caspase activation. These results uncover the coordinated activities of pronucleases and their activating proteases within different immune systems and show how coevolution enables defense system innovation. Competing Interest Statement The Regents of the University of California have patents issued and pending for CRISPR technologies on which J.A.D. is an inventor. J.A.D. is a cofounder of Azalea Therapeutics, Caribou Biosciences, Editas Medicine, Evercrisp, Scribe Therapeutics, Isomorphic Labs, and Mammoth Biosciences. J.A.D. is a scientific advisory board member at BEVC Management, Evercrisp, Caribou Biosciences, Scribe Therapeutics, Mammoth Biosciences, The Column Group and Inari. She also is an advisor for Aditum Bio. J.A.D. is Chief Science Advisor to Sixth Street, a Director at Johnson & Johnson, Altos and Tempus, and has a research project sponsored by Apple Tree Partners. All other authors declare that they have no competing interests.