Biology subjects

Sohn, J.

Publications and source records attributed to Sohn, J..

7 recordsLinked to original sources

bioRxiv2026

Meso2EM: a cross-scale CLEM workflow linking mesoscale functional imaging to targeted electron microscopy

Meso2EM is a correlative light and electron microscopy workflow that transfers neurons selected from mesoscale functional images to targeted electron microscopy. We recorded Ca{superscript 2} signals from layer 2/3 neurons across a contiguous 3 x 3 mm cortical field in awake mice and reidentified a selected neuron after fixation and tangential sectioning. Lectin-labeled vascular architecture served as a shared landmark across in vivo two-photon imaging, confocal microscopy, laboratory micro-CT of resin-embedded tissue, and block-surface scanning electron microscopy, guiding focused-ion-beam scanning electron microscopy to the target cell body. The same progressive-targeting principle also supported serial ATUM-SEM reconstruction of an in vivo-tracked dendrite and serial transmission electron microscopy of optically selected dendrites from a patch-clamp-recorded Martinotti cell. Meso2EM therefore provides a practical route for preserving target identity across large changes in scale and specimen state while restricting electron-microscopy acquisition to a selected region.

neuroscience↗

bioRxiv2026

cGAS bends unpaired DNA to form an unconventional structure that hyperactivates the innate immune response

cGAS is a pattern-recognition receptor for dsDNA and forms 2cGAS:2DNA dimers followed by oligomerization into phase-separated condensates when fully-complementary DNA is studied. However, many DNAs are not fully complementary. We report that DNA with unpaired regions such as those found during transcription, recombination or replication (designated as bubble-DNA, Bu-DNA) causes cGAS hyper-activation. Hyperactivation is observed by Bu-DNA embedded in linear DNA, circular DNA, plasmid DNA and mitochondria DNA. Bu-DNA binds significantly more tightly to the cGAS catalytic domain than paired-DNA but suppresses condensation. Cryo-EM and single-molecule FRET reveal that cGAS forms 2cGAS:1DNA complexes by bending Bu-DNA into a V-shape using the unpaired region as a hinge, limiting its oligomeric state. This uncovers a novel mode of cGAS activation attributed to pattern diversity within pattern ligands.

immunology↗

bioRxiv2026

Protein-DNA mesh assembly drives dsDNA-specific and duplex length-dependent activation of cGAS

Cyclic G/AMP synthase (cGAS) forms condensates on dysregulated double-stranded (ds) DNA to trigger inflammatory responses. Currently, how it specifically recognizes dsDNA and why activation depends on duplex length remain poorly understood. Using cryo-electron microscopy, biochemical assays, and single-molecule methods, we show that full-length cGAS assembles a protein*dsDNA mesh by reiteratively propagating dimers-of-dimers, driving an associative phase transition. Previously uncharacterized N- and C-terminal interactions, together with inter-dimer junction-loops, track the B-form groove and bend dsDNA to build an extensive mesh network spanning multiple duplexes. These interactions are critical for cross-stabilizing active cGAS, revealing why higher-order assembly on long duplexes maximizes signaling activity. Remarkably, cGAS can build the protein*dsDNA mesh on a single contiguous duplex by entangling its oligomerization platform, resulting in a mechanically resilient and kinetically stable signaling platform. Together, our findings establish higher-order mesh assembly of cGAS as the foundation for dsDNA selectivity, duplex-length-dependent activation, and condensate formation.

biophysics↗

bioRxiv2025

The atypical filament assembly underpins the inflammasome-independent functions of IFI16.

Inflammasomes trigger cell death upon sensing various intracellular maladies. These supra-structures relay upstream signals by sequentially assembling architecturally congruent filaments via their pyrin domains (PYDs). Interferon Inducible Protein 16 (IFI16) is an innate immune sensor that detects dysregulated nucleic acids. Once presumed as an inflammasome receptor due to its PYD, the role of IFI16 has been much more appreciated in other innate immune pathways such as regulating interferon production and viral replication restriction. Here, a cryo-EM structure of the filament assembled by the PYD of IFI16(PYD) shows a helical architecture distinct from inflammasome PYD filaments. In silico Rosetta interaction energy calculations suggest that the helical architecture of the IFI16PYD filament is incompatible with those assembled by the central inflammasome adaptor ASC and its interacting partners. Cellular experiments further support that IFI16 fails to interact with ASC. Together, we provide the structural basis for the inflammasome-independent functions of IFI16.

biophysics↗

bioRxiv2024

Dimerization-dependent gel-like condensation with dsDNA underpins the activation of human cGAS

Cyclic G/AMP Synthase (cGAS) initiates inflammatory responses against pathogenic double-stranded (ds)DNA. Although it is well established that cGAS forms phase-separated condensates with dsDNA, their function remains poorly defined. We report here that the dimerization of cGAS on dsDNA creates a mesh-like network, leading to gel-like condensate formation. Although cGAS binds to and forms condensates with various nucleic acids, only dsDNA permits the dimerization necessary for activation and gelation. cGAS co-condenses dsDNA and other nucleic acids but retains a distinct dsDNA-mediated gel-like substate, which single-stranded RNA can dissolve and deactivate the enzyme. Moreover, gel-like, but not liquid-like, condensation not only protects bound dsDNA from exonucleases, but also limits the mobility of NTPs and the dinucleotide intermediate for efficient cGAMP synthesis. Together, our results show that enzymes can finetune surrounding microenvironments to regulate their signaling activities.

biochemistry↗

bioRxiv2022

Design Principles for Inflammasome Inhibition by Pyrin-Only-Proteins

Inflammasomes are filamentous signaling platforms essential for host defense against various intracellular calamities such as pathogen invasion and genotoxic stresses. However, dysregulated inflammasomes cause an array of human diseases including autoinflammatory disorders and cancer. It was recently identified that endogenous pyrin-only-proteins (POPs) regulate inflammasomes by directly inhibiting their filament assembly. Here, by combining Rosetta in silico, in vitro, and in cellulo methods, we investigate the target specificity and inhibition mechanisms of POPs. In contrast to a previous report, we find that POP1 is a poor inhibitor of the central inflammasome adaptor ASC. Instead, POP1 inhibits the assembly of upstream receptor PYD filaments such as those of AIM2, IFI16, NLRP3, and NLRP6. Moreover, not only does POP2 directly suppress the nucleation of ASC, but it can also inhibit the elongation of receptor filaments. In addition to inhibiting the elongation of AIM2 and NLRP6 filaments, POP3 potently suppresses the nucleation of ASC. Our Rosetta analyses and biochemical experiments consistently suggest that a combination of favorable and unfavorable interactions between POPs and PYDs is necessary for effective recognition and inhibition. Together, we reveal the intrinsic target redundancy of POPs and their inhibitory mechanisms.

biochemistry↗

bioRxiv2021

The DNA sensors AIM2 and IFI16 are NET-binding SLE autoantigens

Nucleic acid binding proteins are frequently targeted as autoantigens in systemic lupus erythematosus (SLE) and other interferon (IFN)-linked rheumatic diseases. The AIM-like receptors (ALRs) are IFN-inducible innate sensors that form supramolecular assemblies along double-stranded DNA of various origins. Here, we identify the ALR Absent in melanoma 2 (AIM2) as a novel autoantigen in SLE, with similar properties to the established ALR autoantigen interferon-inducible protein 16 (IFI16). Our SLE cohort revealed a frequent co-occurrence of anti-AIM2, anti-IFI16 and anti-DNA antibodies, and higher clinical measures of disease activity in patients positive for antibodies against these ALRs. We examined neutrophil extracellular traps (NETs) as DNA scaffolds on which these antigens might interact in a pro-immune context, finding that both ALRs bind NETs in vitro and in SLE renal tissues. We demonstrate that ALR binding causes NETs to resist degradation by DNase I, suggesting a mechanism whereby extracellular ALR-NET interactions may promote sustained IFN signaling. Our work suggests that extracellular ALRs bind NETs, leading to DNase resistant nucleoprotein fibers that are targeted as autoantigens in SLE.

immunology↗