Toxoplasma gondii is an important neurotropic pathogen that establishes latent infections in humans that can cause toxoplasmosis in immunocompromised individuals. It replicates inside host cells and has developed several strategies to manipulate host immune responses. However, the cytoplasmic pathogen-sensing pathway that detects T. gondii is not well-characterized. Here, we found that cyclic GMP-AMP synthase (cGAS), a sensor of foreign dsDNA, is required for activation of anti-T. gondii immune signaling in a mouse model. We also found that mice deficient in STING (Sting gt/gt mice) are much more susceptible to T. gondii infection than WT mice. Of note, the induction of inflammatory cytokines, type I IFNs, and interferon-stimulated genes in the spleen from Sting gt/gt mice was significantly impaired. Sting gt/gt mice exhibited more severe symptoms than cGAS-deficient mice after T. gondii infection. Interestingly, we found that the dense granule protein GRA15 from T. gondii is secreted into the host cell cytoplasm and then localizes to the endoplasmic reticulum, mediated by the second transmembrane motif in GRA15, which is essential for activating STING and innate immune responses. Mechanistically, GRA15 promoted STING polyubiquitination at Lys-337 and STING oligomerization in a TRAF protein-dependent manner. Accordingly, GRA15-deficient T. gondii failed to elicit robust innate immune responses compared with WT T. gondii. Consequently , GRA15 ؊/؊ T. gondii was more virulent and caused higher mortality of WT mice but not Sting gt/gt mice upon infection. Together, T. gondii infection triggers cGAS/STING signaling, which is enhanced by GRA15 in a STING-and TRAF-dependent manner. The protozoan parasite Toxoplasma gondii can infect nearly all warm-blooded animals (1, 2). As for humans, nearly 30% of the world's population is infected with T. gondii (3). In healthy adults, T. gondii is controlled by the immune system and remains dormant in the brain. However, in immunocompromised individuals, a defect of the immune system leads to the reactivation of the T. gondii parasites and the development of toxoplasmosis. Reactivated parasite replication causes life-threatening brain damage with brain abscesses and necrotic areas (4). Thus, HIV/AIDS patients, cancer patients, and organ transplant recipients are highly susceptible to T. gondii infection. The infection of T. gondii parasites is recognized by pattern recognition receptors (PRRs). 4 Previous studies showed that TLR11 is the PRR of T. gondii in murine cells. TLR11 is able to detect the actin-binding protein Profilin, which is required for entry of T. gondii during infection. TLR11 and TLR12 form a heterodimer in murine dendritic cells (DC) after sensing Profilin and activate adaptor protein MyD88 to initiate downstream signaling for defense against T. gondii (5). Moreover, TLR7 and TLR9 are able to compensate for the loss of TLR11 by activating

Loss of function/dysregulation of inhibitor of growth 4 (ING4) and hyperactivation of NF-κB are frequent events in many types of human malignancies. However, the molecular mechanisms underlying these remarkable aberrations are not understood. Here, we report that ING4 is physically associated with JFK. We demonstrated that JFK targets ING4 for ubiquitination and degradation through assembly of an Skp1-Cul1-F-box (SCF) complex. We showed that JFK-mediated ING4 destabilization leads to the hyperactivation of the canonical NF-κB pathway and promotes angiogenesis and metastasis of breast cancer. Significantly, the expression of JFK is markedly up-regulated in breast cancer, and the level of JFK is negatively correlated with that of ING4 and positively correlated with an aggressive clinical behavior of breast carcinomas. Our study identified SCF JFK as a bona fide E3 ligase for ING4 and unraveled the JFK-ING4-NF-κB axis as an important player in the development and progression of breast cancer, supporting the pursuit of JFK as a potential target for breast cancer intervention.

IFN regulatory factor 3 (IRF3) is critical for the transcription of type I IFNs in defensing virus and promoting inflammatory responses. Although several kinds of posttranslational modifications have been identified to modulate the activity of IRF3, whether atypical ubiquitination participates in the function regulation, especially the DNA binding capacity of IRF3, is unknown. In this study, we found that the ovarian tumor domain containing deubiquitinase OTUD1 deubiquitinated IRF3 and attenuated its function. An atypical ubiquitination, K6-linked ubiquitination, was essential for the DNA binding capacity of IRF3 and subsequent induction of target genes. Mechanistically, OTUD1 cleaves the viral infection–induced K6-linked ubiquitination of IRF3, resulting in the disassociation of IRF3 from the promoter region of target genes, without affecting the protein stability, dimerization, and nuclear translocation of IRF3 after a viral infection. Otud1−/− cells as well as Otud1−/− mice produced more type I IFNs and proinflammatory cytokines after viral infection. Otud1−/− mice were more resistant to lethal HSV-1 and VSV infection. Consistent with the former investigations that IRF3 promoted inflammatory responses in LPS-induced sepsis, Otud1−/− mice were more susceptible to LPS stimulation. Taken together, our findings revealed that the DNA binding capacity of IRF3 in the innate immune signaling pathway was modulated by atypical K6-linked ubiquitination and deubiquitination process, which was regulated by the deubiquitinase OTUD1.

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