Aug 2026· International Journal of Molecular Sciences· Vol 27 16, pp. 7478· 0 citations· 181 references
Medicine
TL;DR
Competition with adenosine deaminase acting on RNA 1 (ADAR1), melanoma differentiation-associated protein 5 (MDA5), double-stranded RNA-dependent protein kinase (PKR), the cyclic guanosine monophosphate–adenosine monophosphate–adenosine monophosphate synthase–stimulator of interferon genes (cGAS–STING) pathway, and other nucleic-acid-sensing
Abstract
In contrast to foreign nucleic acids, some of our own endogenously synthesized nucleic acids may become immunologically active without being considered "foreign". For example, abnormalities in chromatin organization, transcription termination, ribonucleic acid (RNA) splicing, and RNA editing, together with damage to mitochondrial integrity, may render normally functional deoxyribonucleic acid (DNA) and RNA persistently available and aberrantly structured ligands for innate immunity. Z-DNA-binding protein 1 (ZBP1), recently identified as an important component of this innate immune system, recognizes both left-handed DNA (Z-DNA) and left-handed RNA (Z-RNA) using its tandem Z-alpha (Zα) domains and couples recognition of these conformational states to receptor-interacting serine/threonine-protein kinase 1 (RIPK1)-, receptor-interacting serine/threonine-protein kinase 3 (RIPK3)-, and mixed-lineage kinase domain-like pseudokinase (MLKL)-dependent inflammatory and cell-death pathways. More recent studies have also shown that ZBP1 plays a role in recognizing damaged self-nucleic acids associated with tauopathies, Alzheimer's disease (AD), traumatic brain injury (TBI), and amyloid-associated neuroinflammation. The nucleic-acid forms associated with these conditions include transposable-element activation, extended repeat-containing transcripts, RNA-RNA duplexes or RNA:DNA hybrids, oxidized mitochondrial DNA (mtDNA), and intercellularly transferred nucleic acids, all of which may exhibit substrate structures compatible with Z-form formation. Signaling by ZBP1 does not occur simply based upon nucleic-acid abundance; rather, signaling occurs after prolonged exposure to a nucleic acid when it persists in a structurally competent state, sufficient receptors are present to bind its exposed regions, the receptor proteoforms are competent to participate in signaling, receptor-interacting protein homotypic interaction motif (RHIM)-dependent assembly occurs, and the appropriate adaptor molecules are present. Furthermore, the identity of the cell type expressing ZBP1 determines whether the response produces RIPK3-MLKL-dependent neuronal injury, microglia-mediated inflammation, apoptosis, or mixed cell death. Finally, competition with adenosine deaminase acting on RNA 1 (ADAR1), melanoma differentiation-associated protein 5 (MDA5), double-stranded RNA-dependent protein kinase (PKR), the cyclic guanosine monophosphate-adenosine monophosphate synthase-stimulator of interferon genes (cGAS-STING) pathway, and other nucleic-acid-sensing proteins divides the available pool of endogenous nucleic acids among the outcomes of immune tolerance, type I interferon (IFN-I) signaling, translational inhibition, neuroinflammation, and necroptosis.
Z-nucleic acid-binding protein 1 (ZBP1) is a cytosolic innate immune sensor that detects left-handed Z-DNA and Z-RNA, structures arising from endogenous retroelements, splicing stress, R-loops, viruses, or viral mimicry. Originally viewed as an antiviral receptor, ZBP1 has emerged as a sentinel of genomic and transcriptomic instability. Upon ligand binding, ZBP1 recruits receptor-interacting serine/threonine-protein kinase 1 (RIPK1) and receptor-interacting serine/threonine-protein kinase 3 (RIPK3) via RIP homotypic interaction motif domains; in humans, RIPK1 acts as an essential scaffold to form the necrosome, which phosphorylates mixed lineage kinase domain-like pseudokinase (MLKL) and triggers necroptosis, a lytic, immunogenic cell death. This pathway is intimately regulated by reactive oxygen species (ROS): oxidative stress promotes RIPK1 activation, and necroptosis further drives a mitochondrial ROS burst, establishing a feed-forward amplification loop. In cancer, therapeutic induction of viral mimicry (e.g., using curaxins or splicing inhibitors) combined with tumor-localized ROS generation activates ZBP1-driven necroptosis, leading to the release of damage-associated molecular patterns and tumor antigens. This process converts immunosuppressive “cold” tumors into inflamed “hot” tumors, enhancing dendritic cell maturation, CD8+ T cell infiltration, and sensitivity to immune checkpoint blockade. Integrating epigenetic priming, nanomedicine, and patient stratification according to ZBP1/RIPK3/MLKL pathway status holds promise for overcoming therapeutic resistance. Thus, ZBP1-mediated nucleic acid surveillance represents a central innate checkpoint bridging genomic stress to antitumor immunity.
Background and aim Ras GTPase-activating protein SH3 domain-binding protein 1 (G3BP1) has been shown to bind cytosolic nucleic acid sensors including retinoic acid-inducible gene I (RIG-I) and cyclic GMP-AMP synthase (cGAS) in innate immunity. As a scaffolding protein enhancing protein-protein and protein-RNA interactions during cellular stress response, we examine whether G3BP1 could bind additional innate immune signaling molecules and investigate the relevance and mechanisms underlying these interactions. Methods To define G3BP1 function in dsRNA-triggered anti-viral signaling and identify additional interacting molecules, we generated siRNA- and CRISPER-Cas9-mediated G3BP1-knockdown and knockout HEK293T cells, conducted biochemical experiments in human and murine cells including co-immunoprecipitation, domain mutagenesis and mapping, as well as LC-MS/MS analyses to study protein post-translational modifications. We also performed p(I:C)-stimulation assays to measure interferon production, pharmacological inhibition studies to determine enzyme-substrate specificity, as well as confocal immunofluorescence microscopy to examine protein subcellular localization. Results Our study reveals a role for G3BP1 in binding TBK1 and IRF3, two important molecules in innate immune signaling leading to the production of IFN-β. Deficiency in G3BP1 reduces TBK1–IRF3 complex formation and affects IRF3 phosphorylation, nuclear translocation, and the production of IFN-β during p(I:C) stimulation while the phosphorylation of TBK1 was largely preserved. We further show that G3BP1 constitutively associates with IRF3 and recruits TBK1 upon p(I:C) stimulation. These findings suggest that G3BP1 acts as a scaffold to recruit activated TBK1 and facilitates its activation of IRF3. Domain mapping experiments indicated that the C-terminal RGG region of G3BP1 is critical for binding IRF3 and TBK1. LC–MS/MS analysis revealed human G3BP1 to be arginine-methylated at R435 and R460 residues upon p(I:C)-stimulation, and mutagenesis experiments indicated that R460 is required for its efficient binding of TBK1. We further show that the arginine methyltransferase PRMT5 associates with and promotes the symmetric arginine-dimethylation of G3BP1 and pharmacological inhibition of PRMT5 impaired G3BP1–TBK1 interaction. Conclusions Together, our data indicate that G3BP1 could bind PRMT5, TBK1 and IRF3 and reveal G3BP1 as a signaling hub that constitutively binds IRF3 and is arginine-methylated by PRMT5 to recruit TBK1 for IRF3 phosphorylation and activation and the induction of interferon production during host antiviral response.
Wen Song, Susana Soo-Yeon Kim, Candice Yam et al.· Frontiers in Immunology· 0 citations
RNA-binding proteins (RBPs) are key regulators of gene expression that shape cellular function in health and disease. However, the roles of RBPs in immune cells within the central nervous system (CNS) remain poorly understood. Here, we identify ARID5A as an RBP highly expressed in microglia and uncover its RNA-mediated regulatory functions using integrated multi-omics analyses of its RNA, DNA, and protein interactions. ARID5A regulates the splicing and translation of its RNA targets, many of which are integral to lysosomal, immune, and iron metabolism pathways. We confirm the functional relevance of this ARID5A-dependent RNA regulatory network by demonstrating that ARID5A modulates lysosomal activity, cytokine secretion, iron accumulation, and ferroptosis in iPSC-derived microglia. We further demonstrate that knockdown of microglial ARID5A reduces neuronal ferroptosis in co-cultures, underscoring the interconnected nature of these pathways. Moreover, in microglia harboring the TREM2-T66M mutation, ARID5A depletion restores dysregulated lysosomal and metabolic functions. Our results highlight the importance of protein-RNA interactions in regulating microglial cell biology.
Danielle M. Schafer, S. Blue, Shuhao Xu et al.· Nature Communications· 0 citations
Tolerance of endogenous nucleic acids is crucial for cellular homeostasis and when perturbed can lead to immune dysregulation. RNA-binding proteins (RBPs) bind, modify, and compartmentalize endogenous RNAs to prevent their recognition by the innate immune system. We found that loss of a highly abundant RBP, the cold shock domain containing Y-box binding protein 1 (YBX1), results in the spontaneous induction of an IL6/STAT3 inflammatory response and the upregulation of interferon-stimulated genes. A meta-analysis of other studies reveals that the depletion of YBX1 orthologs and paralogs results in a similar proinflammatory gene signature, suggesting a conserved role of Y-box proteins in maintaining innate immune homeostasis. Among RNAs that interact with YBX1, we tested whether endogenous RNA polymerase III (RNAP III) transcripts stimulate innate immune signaling when YBX1 is depleted. We demonstrate that the inflammatory responses induced by the loss of YBX1 are abrogated by inhibiting RNAP III transcription and RIG-I signaling. Hence, we hypothesize that by regulating RNAP III transcripts, YBX1 prevents their recognition by the innate immune system, uncovering a role of this RBP in maintaining cellular homeostasis and tolerance to endogenous RNAs. The conservation of this phenotype across YBX1 orthologs and paralogs argues that RNA shielding by cold shock domain proteins represents an evolutionarily conserved solution to the problem of endogenous RNA immunogenicity.
Tania Strilets, Sarah Dremel, Mariano A. Garcia-Blanco· Proceedings of the National...· 0 citations
Deletion of MEX3B inhibited caspase-4 and gasdermin D activation, pyroptosis, and secretion of inflammasome-dependent inflammatory cytokines in human cell lines and murine primary macrophages and suggested that MEX3B is a pan-inflammasome regulator targeting inflammatory caspases.
Penghua Wang, Jason G. Cahoon, Duomeng Yang et al.· Journal of Immunology· 0 citations
Cytosolic DNA and RNA sensing is crucial for innate immunity, playing essential roles in pathogen defense and autoinflammation induction. We reported the endogenous metabolite flavin adenine dinucleotide (FAD) as a molecular brake restraining both cytosolic DNA and RNA sensing. It bound directly to cytosolic nucleic acid sensors cyclic GMP-AMP synthase (cGAS) and retinoic acid-inducible gene I (RIG-I), occupying catalytic pockets to suppress their activity and downstream immune responses. Physiologically, FAD prevented self-nucleic acid-induced sterile inflammation and maintained immune homeostasis. FAD deficiency due to FAD synthase (FLAD1) ablation exacerbated auto-inflammation and cellular senescence. Upon viral infection, reduced FLAD1 activity lowered FAD amounts, which removed its inhibitory control over DNA and RNA sensors, thus facilitating extensive interferon-I (IFN-I) signaling activation. Consequently, FLAD1 depletion strengthened the innate immune response and protected mice from viral infection. Our findings identify FAD as a natural suppressor of both cytosolic DNA and RNA sensing, offering therapeutic potential for inflammatory diseases.
Yao Wang, Yanyan Shen, Jiayu Liu et al.· Immunity· 0 citations
