Summary Binding of particulate antigens by antigen presenting cells (APC) is a critical step in immune activation. Previously, we demonstrated that uric acid crystals are potent adjuvants, initiating a robust adaptive immune response. However, the mechanisms of activation are unknown. Using atomic force microscopy as a tool for real time single cell activation analysis, we report that uric acid crystals can directly engage cellular membranes, particularly the cholesterol components, with a force substantially stronger than protein based cellular contacts. Binding of particulate substances activates Syk kinase-dependent signaling in dendritic cells (DCs). These observations suggest a mechanism whereby immune cell activation can be triggered by solid structures via membrane lipid alteration without the requirement for specific cell surface receptors, and a testable hypothesis for crystal-associated arthropathies, inflammation and adjuvanticity.

Uric acid is released from injured cells and can act as an adjuvant signal to the immune system. Uric acid crystals invoke strong inflammatory responses in tissues. Although their biological effects are evident and the associated signaling mechanisms are becoming clear, it remains unexplained as to why uric acid precipitates rapidly in vivo, in sharp contrast to the minimal crystallization in vitro. We report in this study that a group of IgM Abs is able to bind to these crystals, which is interesting in light that B cell-deficient mice do not sense the proinflammatory adjuvant effect of uric acid. The titers of these Abs increase upon immunization with uric acid crystals. We have produced large quantities of such mAbs. The purified IgM Abs can significantly facilitate uric acid precipitation to form the inflammatory crystals in vitro. Infusion of these Abs into B cell-deficient mice significantly increases the basal level of inflammation in these recipients and restores the host’s ability to sense uric acid’s adjuvanticity. Therefore, we have identified a factor in determining uric acid precipitation and possibly its ability to function as an endogenous adjuvant. This finding suggests a new mechanism of the pathogenesis of gouty arthritis and uric acid-induced immune activation.

Cross-talk between distinct protein post-translational modifications is critical for an effective DNA damage response. Arginine methylation plays an important role in maintaining genome stability, but how this modification integrates with other enzymatic activities is largely unknown. Here, we identify the deubiquitylating enzyme USP11 as a previously uncharacterised PRMT1 substrate, and demonstrate that the methylation of USP11 promotes DNA end-resection and the repair of DNA double strand breaks (DSB) by homologous recombination (HR), an event that is independent from another USP11-HR activity, the deubiquitylation of PALB2. We also show that PRMT1 is a ubiquitylated protein that it is targeted for deubiquitylation by USP11, which regulates the ability of PRMT1 to bind to and methylate MRE11. Taken together, our findings reveal a specific role for USP11 during the early stages of DSB repair, which is mediated through its ability to regulate the activity of the PRMT1-MRE11 pathway.

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The binding of antigens to professional antigen-presenting cells (APCs), particularly dendritic cells (DCs), is essential for immune activation. Previously, it was shown that uric acid crystals, the causative agent of gout, were released from dying and injured cells as a "danger signal" to initiate a robust adaptive immune response. But how uric acid crystals lead to APC activation at a molecular level is unknown. Using atomic force microscopy (AFM) to analyze single cell activation in real time, we demonstrate that uric acid crystals directly bind cellular membranes, particularly cholesterol components, with very high affinity. Binding is dependent on intracellular Syk-kinase and PI3-kinase signaling and is independent of extracellular protein receptors. These findings suggest a mechanism that activates the immune system without specific cell surface protein receptors and presents a testable hypothesis for inflammation to materials without evolutionarily sensible receptors such as materials produced post-industrially. This research has applications in vaccine development, biomaterials, inflammation, and crystal-associated arthropathies.

Uric acid is a product of purine metabolism that is secreted from injured or dying cells and can act as an endogenous adjuvant. However, uric acid must crystallize to monosodium urate (MSU) before it can activate dendritic cells and cause gouty inflammation. At normal serum uric acid levels, crystallization of uric acid is not observed in vitro. The crucial question therefore is how the crystals form and cause inflammation. It has been reported that serum from rabbits immunized with MSU, increased uric acid precipitation rates. When C57BL6 or Balb/c mice were immunized with MSU, they produced serum factors which bind to MSU crystals, a phenomenon absent in mu chain deficient IgH (or muMT) mice. These observations show that antibodies may be involved in MSU crystallization. Antibodies produced via B cell hybridomas, were observed to precipitate uric acid in vitro, with the variable region acting as the binding domain. Most importantly, the presence of antibody was instrumental in significantly increasing the basal level of inflammation and mediating the adjuvant effect of uric acid in antigen dependent T cell cytotoxicity in B cell deficient mice. Therefore, we have identified a factor in determining uric acid precipitation and possibly its ability to function as an endogenous adjuvant. This finding suggests a new mechanism of the pathogenesis of gouty arthritis and uric acid induced immune activation.

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