A large-scale type I CBASS antiphage screen identifies the phage prohead protease as a key determinant of immune activation and evasion.

Cyclic oligonucleotide-based signaling system (CBASS) is an antiviral system that protects bacteria from phage infection and is evolutionarily related to human cGAS-STING immunity. cGAS-STING signaling is initiated by the recognition of viral DNA, but the molecular cues activating CBASS are incompletely understood. Using a screen of 975 type I CBASS operon-phage challenges, we show that operons with distinct cGAS/DncV-like nucleotidyltransferases (CD-NTases) and CD-NTase-associated protein (Cap) effectors exhibit marked patterns of phage restriction.

eIF3d controls the persistent integrated stress response.

Cells respond to intrinsic and extrinsic stresses by reducing global protein synthesis and activating gene programs necessary for survival. Here, we show that the integrated stress response (ISR) is driven by the non-canonical cap-binding protein eIF3d that acts as a critical effector to control core stress response orchestrators, the translation factor eIF2α and the transcription factor ATF4. We find that during persistent stress, eIF3d activates the translation of the kinase GCN2, inducing eIF2α phosphorylation and inhibiting general protein synthesis.

Convergent mutations in phage virion assembly proteins enable evasion of Type I CBASS immunity.

CBASS is an anti-phage defense system that protects bacteria from phage infection and is evolutionarily related to human cGAS-STING immunity. cGAS-STING signaling is initiated by viral DNA but the stage of phage replication which activates bacterial CBASS remains unclear. Here we define the specificity of Type I CBASS immunity using a comprehensive analysis of 975 operon-phage pairings and show that Type I CBASS operons composed of distinct CD-NTases, and Cap effectors exhibit striking patterns of defense against dsDNA phages across five diverse viral families.

Moonlighting translation factors: multifunctionality drives diverse gene regulation.

Translation factors have traditionally been viewed as proteins that drive ribosome function and ensure accurate mRNA translation. Recent discoveries have highlighted that these factors can also moonlight in gene regulation, but through functions distinct from their canonical roles in protein synthesis. Notably, the additional functions that translation factors encode are diverse, ranging from transcriptional control and extracellular signaling to RNA binding, and are highly regulated in response to external cues and the intrinsic cellular state.

Eukaryotic Initiation Factor 5A2 Regulates Expression of Antiviral Genes.

Translation factors are essential for regulation of protein synthesis. The eukaryotic translation initiation factor 5A (eIF5A) family is made up of two paralogues - eIF5A1 and eIF5A2 - which display high sequence homology but distinct tissue tropism. While eIF5A1 directly binds to the ribosome and regulates translation initiation, elongation, and termination, the molecular function of eIF5A2 remains poorly understood.

cGAS-like receptors sense RNA and control 3'2'-cGAMP signalling in Drosophila.

Cyclic GMP-AMP synthase (cGAS) is a cytosolic DNA sensor that produces the second messenger cG[2'-5']pA[3'-5']p (2'3'-cGAMP) and controls activation of innate immunity in mammalian cells. Animal genomes typically encode multiple proteins with predicted homology to cGAS, but the function of these uncharacterized enzymes is unknown. Here we show that cGAS-like receptors (cGLRs) are innate immune sensors that are capable of recognizing divergent molecular patterns and catalysing synthesis of distinct nucleotide second messenger signals.

A phosphorylation-regulated eIF3d translation switch mediates cellular adaptation to metabolic stress.

Shutoff of global protein synthesis is a conserved response to cellular stresses. This general phenomenon is accompanied by the induction of distinct gene programs tailored to each stress. Although the mechanisms driving repression of general protein synthesis are well characterized, how cells reprogram the translation machinery for selective gene expression remains poorly understood.

CBASS Immunity Uses CARF-Related Effectors to Sense 3'-5'- and 2'-5'-Linked Cyclic Oligonucleotide Signals and Protect Bacteria from Phage Infection.

cGAS/DncV-like nucleotidyltransferase (CD-NTase) enzymes are immune sensors that synthesize nucleotide second messengers and initiate antiviral responses in bacterial and animal cells. Here, we discover Enterobacter cloacae CD-NTase-associated protein 4 (Cap4) as a founding member of a diverse family of >2,000 bacterial receptors that respond to CD-NTase signals. Structures of Cap4 reveal a promiscuous DNA endonuclease domain activated through ligand-induced oligomerization.

A molecular filter for the cnidarian stinging response.

All animals detect and integrate diverse environmental signals to mediate behavior. Cnidarians, including jellyfish and sea anemones, both detect and capture prey using stinging cells called nematocytes which fire a venom-covered barb via an unknown triggering mechanism. Here, we show that nematocytes from use a specialized voltage-gated calcium channel (nCa) to distinguish salient sensory cues and control the explosive discharge response.

Repression of ferritin light chain translation by human eIF3.

A central problem in human biology remains the discovery of causal molecular links between mutations identified in genome-wide association studies (GWAS) and their corresponding disease traits. This challenge is magnified for variants residing in non-coding regions of the genome. Single-nucleotide polymorphisms (SNPs) in the 5' untranslated region (5'-UTR) of the ferritin light chain () gene that cause hyperferritinemia are reported to disrupt translation repression by altering iron regulatory protein (IRP) interactions with the mRNA 5'-UTR.

A complex suite of loci and elements in eukaryotic type II topoisomerases determine selective sensitivity to distinct poisoning agents.

Type II topoisomerases catalyze essential DNA transactions and are proven drug targets. Drug discrimination by prokaryotic and eukaryotic topoisomerases is vital to therapeutic utility, but is poorly understood. We developed a next-generation sequencing (NGS) approach to identify drug-resistance mutations in eukaryotic topoisomerases.

Modular Architecture of the STING C-Terminal Tail Allows Interferon and NF-κB Signaling Adaptation.

Stimulator of interferon genes (STING) is a key regulator of type I interferon and pro-inflammatory responses during infection, cellular stress, and cancer. Here, we reveal a mechanism for how STING balances activation of IRF3- and NF-κB-dependent transcription and discover that acquisition of discrete signaling modules in the vertebrate STING C-terminal tail (CTT) shapes downstream immunity. As a defining example, we identify a motif appended to the CTT of zebrafish STING that inverts the typical vertebrate signaling response and results in dramatic NF-κB activation and weak IRF3-interferon signaling.

Bacterial cGAS-like enzymes synthesize diverse nucleotide signals.

Cyclic dinucleotides (CDNs) have central roles in bacterial homeostasis and virulence by acting as nucleotide second messengers. Bacterial CDNs also elicit immune responses during infection when they are detected by pattern-recognition receptors in animal cells. Here we perform a systematic biochemical screen for bacterial signalling nucleotides and discover a large family of cGAS/DncV-like nucleotidyltransferases (CD-NTases) that use both purine and pyrimidine nucleotides to synthesize a diverse range of CDNs.

Ancient Origin of cGAS-STING Reveals Mechanism of Universal 2',3' cGAMP Signaling.

In humans, the cGAS-STING immunity pathway signals in response to cytosolic DNA via 2',3' cGAMP, a cyclic dinucleotide (CDN) second messenger containing mixed 2'-5' and 3'-5' phosphodiester bonds. Prokaryotes also produce CDNs, but these are exclusively 3' linked, and thus the evolutionary origins of human 2',3' cGAMP signaling are unknown. Here we illuminate the ancient origins of human cGAMP signaling by discovery of a functional cGAS-STING pathway in Nematostella vectensis, an anemone species >500 million years diverged from humans.

eIF3 targets cell-proliferation messenger RNAs for translational activation or repression.

Regulation of protein synthesis is fundamental for all aspects of eukaryotic biology by controlling development, homeostasis and stress responses. The 13-subunit, 800-kilodalton eukaryotic initiation factor 3 (eIF3) organizes initiation factor and ribosome interactions required for productive translation. However, current understanding of eIF3 function does not explain genetic evidence correlating eIF3 deregulation with tissue-specific cancers and developmental defects.

Integrase-mediated spacer acquisition during CRISPR-Cas adaptive immunity.

Bacteria and archaea insert spacer sequences acquired from foreign DNAs into CRISPR loci to generate immunological memory. The Escherichia coli Cas1-Cas2 complex mediates spacer acquisition in vivo, but the molecular mechanism of this process is unknown. Here we show that the purified Cas1-Cas2 complex integrates oligonucleotide DNA substrates into acceptor DNA to yield products similar to those generated by retroviral integrases and transposases.

Structure-guided reprogramming of human cGAS dinucleotide linkage specificity.

Cyclic dinucleotides (CDNs) play central roles in bacterial pathogenesis and innate immunity. The mammalian enzyme cGAS synthesizes a unique cyclic dinucleotide (cGAMP) containing a 2'-5' phosphodiester linkage essential for optimal immune stimulation, but the molecular basis for linkage specificity is unknown. Here, we show that the Vibrio cholerae pathogenicity factor DncV is a prokaryotic cGAS-like enzyme whose activity provides a mechanistic rationale for the unique ability of cGAS to produce 2'-5' cGAMP.

Peroxisomes are signaling platforms for antiviral innate immunity.

Peroxisomes have long been established to play a central role in regulating various metabolic activities in mammalian cells. These organelles act in concert with mitochondria to control the metabolism of lipids and reactive oxygen species. However, while mitochondria have emerged as an important site of antiviral signal transduction, a role for peroxisomes in immune defense is unknown.