{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,8,17]],"date-time":"2026-08-17T18:31:18Z","timestamp":1786991478511,"version":"build-2736575974"},"posted":{"date-parts":[[2026,8,13]]},"group-title":"Microbiology","reference-count":57,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,8,13]],"date-time":"2026-08-13T00:00:00Z","timestamp":1786579200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.biorxiv.org\/about\/FAQ#license"}],"funder":[{"name":"NIH","award":["GM145888"],"award-info":[{"award-number":["GM145888"]}]},{"name":"NIH","award":["GM145060"],"award-info":[{"award-number":["GM145060"]}]},{"name":"Maloris Foundation"},{"name":"Memorial Sloan Kettering Cancer Center Core Grant","award":["P30-CA008748"],"award-info":[{"award-number":["P30-CA008748"]}]},{"name":"NIH Common Fund Transformative High Resolution Cryo-Electron Microscopy Program","award":["U24 GM129539"],"award-info":[{"award-number":["U24 GM129539"]}]},{"DOI":"10.13039\/100000893","name":"Simons Foundation","doi-asserted-by":"crossref","award":["SF349247"],"award-info":[{"award-number":["SF349247"]}],"id":[{"id":"10.13039\/100000893","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,8,13]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Bacteria use nucleotide second messengers to couple phage recognition to antiviral effector activation. Here, we identify Pucsar, an ofshoot of the Pycsar-related antiphage family, whose cyclases generate 3\u2019,5\u2019-cGMP from GTP, that are associated with Toll\/Interleukin-1 receptor (TIR) and transmembrane (TM) effectors. In the PucsarTIR system, cGMP produced by cyclase PucC, selectively activates the cognate PucTIR NADase. X-ray crystallography shows that PucC adopts a dimeric cyclase scaffold with a guanine-compatible base-recognition pocket. Cryo-EM and mutational analyses revealed that apo-PucTIR, composed of cyclic nucleotide binding and TIR domains, is self-sequestered as an inactive tetramer, in which TIR domains are proximal but misoriented. Sequential cGMP binding remodels this assembly through partially occupied intermediates and stabilizes a fully cGMP-bound dimer, that polymerizes into a zig-zag aligned filament, creating composite NAD\n                  <jats:sup>+<\/jats:sup>\n                  -binding pockets required for efficient NADase activity. These findings establish 3\u2032,5\u2032-cGMP as an antiphage second messenger and reveal how cyclic purine signaling converts self-sequestered TIR effectors into a NADase-competent filament.\n                <\/jats:p>","DOI":"10.64898\/2026.08.12.744502","type":"posted-content","created":{"date-parts":[[2026,8,14]],"date-time":"2026-08-14T04:50:20Z","timestamp":1786683020000},"source":"Crossref","is-referenced-by-count":0,"title":["Cyclase-generated 3\u2019,5\u2019-cGMP activates PucsarTIR antiphage defense through a stepwise zig-zag aligned filament assembly"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0009-0009-3267-8450","authenticated-orcid":false,"given":"Arpita","family":"Chakravarti","sequence":"first","affiliation":[{"name":"Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5965-0399","authenticated-orcid":false,"given":"Yi","family":"Wu","sequence":"additional","affiliation":[{"name":"School of Biological Sciences, University of Southampton, Southampton, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2229-8771","authenticated-orcid":false,"given":"A. 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H.","family":"Huang","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, University of Illinois, Urbana-Champaign, Urbana, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0771-253X","authenticated-orcid":false,"given":"L.","family":"Aravind","sequence":"additional","affiliation":[{"name":"Division of Intramural Research, National Library of Medicine, National Institutes of Health, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8238-1083","authenticated-orcid":false,"given":"Franklin L.","family":"Nobrega","sequence":"additional","affiliation":[{"name":"School of Biological Sciences, University of Southampton, Southampton, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9779-7778","authenticated-orcid":false,"given":"Dinshaw J.","family":"Patel","sequence":"additional","affiliation":[{"name":"Structural 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