Identifying phage proteins that activate the bacterial innate immune system

Aaron Whiteley — Wed, 02 Jul 2025 15:00:00 +0000

Identifying phage proteins that activate the bacterial innate immune system Aaron Whiteley Wed, 07/02/2025 - 09:00

Toni Nagy , Gina Gersabeck , Amy Conte , Aaron Whiteley

BioRxiv (2025). PubMed PMID: 40631175; PubMed Central PMCID: PMC12236752

Abstract

Bacteria have evolved sophisticated antiphage systems that halt phage replication upon detecting specific phage triggers. Identifying phage triggers is crucial to our understanding of immune signaling, however, they are challenging to predict. Here we used an expansive plasmid library that expressed 400 phage protein-coding genes from 6 different phages to identify novel triggers of known and undiscovered antiphage systems. We transformed our library into 72 diverse strains of E. coli . Each strain natively harbors a different suite of antiphage systems whose activation typically inhibits growth. By tracking plasmids that were selectively depleted, we identified over 100 candidate phage trigger- E. coli pairs. Two phage trigger proteins were investigated in detail, revealing a novel antiphage system that detects multiple phage tail fiber proteins and identifying major capsid protein as the activating ligand of the antiphage system Avs8. These experiments provide a unique dataset for continued definition of the molecular details of the bacterial immune system.

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Nagy TA, Gersabeck GW, Conte AN, Whiteley AT. Identifying phage proteins that activate the bacterial innate immune system. bioRxiv [Preprint]. 2025 Jul 2:2025.07.02.662641. doi: 10.1101/2025.07.02.662641. PMID: 40631175; PMCID: PMC12236752.

Nagy TA, Gersabeck GW, Conte AN, ➤Whiteley AT | BioRxiv 2025

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DnaJ mediates phage sensing by the bacterial NLR-related protein bNACHT25

Aaron Whiteley — Fri, 30 May 2025 15:00:00 +0000

DnaJ mediates phage sensing by the bacterial NLR-related protein bNACHT25 Aaron Whiteley Fri, 05/30/2025 - 09:00

Amy Conte , Madison Ruchel , Samantha Ridgeway , Emily Kibby , Toni Nagy , Aaron Whiteley

PLoS Biology (2025). PubMed PMID: 40446071; PubMed Central PMCID: PMC12169576

Abstract

Bacteria encode a wide range of antiphage systems and a subset of these proteins are homologous to components of the human innate immune system. Mammalian nucleotide-binding and leucine-rich repeat containing proteins (NLRs) and bacterial NLR-related proteins use a central NACHT domain to link detection of infection with initiation of an antimicrobial response. Bacterial NACHT proteins provide defense against both DNA and RNA phages. Here we investigate the mechanism of phage detection by the bacterial NLR-related protein bNACHT25 in E. coli. bNACHT25 was specifically activated by Emesvirus ssRNA phages and analysis of MS2 phage escaper mutants that evaded detection revealed a critical role for Coat Protein (CP). A genetic assay showed CP was sufficient to activate bNACHT25 but the two proteins did not directly interact. Instead, we found bNACHT25 requires the host chaperone DnaJ to detect CP and protect against phage. Our data support a model in which bNACHT25 detects a wide range of phages using an indirect mechanism that may involve guarding a host cell process rather than binding a specific phage-derived molecule.

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Citation

Conte AN, Ruchel ME, Ridgeway SM, Kibby EM, Nagy TA, Whiteley AT. DnaJ mediates phage sensing by the bacterial NLR-related protein bNACHT25. PLoS Biol. 2025 May 30;23(5):e3003203. doi: 10.1371/journal.pbio.3003203. [Epub ahead of print] PubMed PMID: 40446071.

Conte AN, Ruchel ME, Ridgeway SM, Kibby EM, Nagy TA, ➤Whiteley, AT | PLoS Biology 2025

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Identifying phage proteins that activate the bacterial innate immune system

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DnaJ mediates phage sensing by the bacterial NLR-related protein bNACHT25

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