USP14 promotes K63‐linked RIG‐I deubiquitination and suppresses antiviral immune responses

Retinoic acid‐inducible gene I (RIG‐I) is a critical RNA virus sensor that initiates antiviral immune response through K63‐linked ubiquitination. In this study, we demonstrated USP14, a deubiquitinating enzyme, as a negative regulator in antiviral responses by directly deubiquitinating K63‐linked RIG‐I. USP14 knockdown significantly enhanced RIG‐I‐triggered type I IFN signaling and inhibited vesicular stomatitis virus (VSV) replication both in mouse peritoneal macrophages and THP1 cells. USP14 overexpression in HeLa cells attenuated RIG‐I‐triggered IFN‐β expression and promoted VSV replication. Besides, USP14‐specific inhibitor, IU1, increased RIG‐I‐mediated type I IFN production and antiviral responses in vitro and in vivo. In addition, USP14 could interact with RIG‐I and remove RIG‐I K63‐linked polyubiquitination chains. This article is the first to report that USP14 acts as a negative regulator in antiviral response through deubiquitinating K63‐linked RIG‐I. These findings provide insights into a potential new therapy targeting USP14 for RNA virus‐related diseases.     

Tetraspanins CD9 and CD151 at the immune synapse support T‐cell integrin signaling

Understanding how the immune response is activated and amplified requires detailed knowledge of the stages in the formation of the immunological synapse (IS) between T lymphocytes and antigen‐presenting cells (APCs). We show that tetraspanins CD9 and CD151 congregate at the T‐cell side of the IS. Silencing of CD9 or CD151 blunts the IL‐2 secretion and expression of the activation marker CD69 by APC‐conjugated T lymphocytes, but does not affect the accumulation of CD3 or actin to the IS, or the translocation of the microtubule‐organizing center toward the T‐B contact area. CD9 or CD151 silencing diminishes the relocalization of α4β1 integrin to the IS and reduces the accumulation of high‐affinity β1 integrins at the cell–cell contact. These changes are accompanied by diminished phosphorylation of the integrin downstream targets FAK and ERK1/2. Our results suggest that CD9 and CD151 support integrin‐mediated signaling at the IS.     

Epstein–Barr virus‐induced gene 3 suppresses T helper type 1, type 17 and type 2 immune responses after Trypanosoma cruzi infection and inhibits parasite replication by interfering with alternative macrophage activation

The Epstein–Barr virus‐induced gene 3 (EBI3) is a member of the interleukin‐12 (IL)‐12) family structurally related to the subunit p40 of IL‐12 and forms a heterodimer either with the p28 subunit to build IL‐27 or with p35 to form IL‐35. Interleukin‐27 is secreted by antigen‐presenting cells whereas IL‐35 appears to be produced mainly by regulatory T cells and regulatory B cells but both cytokines negatively regulate inflammatory immune responses. We here analysed the function of EBI3 during infection with the intracellular parasite Trypanosoma cruzi. Compared with C57BL/6 wild‐type mice, EBI3‐deficient (EBI3^?/?) mice showed a higher parasitaemia associated with an increased mortality rate. The EBI3^?/? mice displayed an elevated inflammatory immune response with an increased production of T helper type 1 (Th1‐), Th2‐ and Th17‐derived cytokines. The increased Th2 immune response appears to have over‐ridden the otherwise protective Th1 and Th17 immune responses by the induction of arginase‐1‐expressing alternatively activated macrophages in these mice. Hence, neutralization of IL‐4 and arginase‐1 activity partially restored protective immune responses in EBI3^?/? mice. So far, our results demonstrate that EBI3 is an essential general regulator of inflammatory immune responses in experimental Chagas disease and is required for control of T. cruzi infection by inhibiting Th2‐dependent alternative macrophage activation. Further studies are needed to dissect the underlying mechanisms and clarify whether EBI3 association with IL‐27 or/and IL‐35 accounts for its anti‐inflammatory character in parasitic disease.     

Dendritic cell homeostasis is maintained by nonhematopoietic and T‐cell‐produced Flt3‐ligand in steady state and during immune responses

Lymphoid‐tissue dendritic cells (DCs) are short‐lived and need to be continuously replenished from bone marrow‐derived DC progenitor cells. Fms‐related tyrosine kinase 3 is expressed during cellular development from hematopoietic progenitors to lymphoid‐tissue DCs. Fms‐related tyrosine kinase 3 ligand (Flt3L) is an essential, nonredundant cytokine for DC progenitor to lymphoid tissue DC differentiation and maintenance. However, which cells contribute to Flt3L production and how Flt3L cytokine levels are regulated in steady state and during immune reactions remains to be determined. Here we demonstrate that besides nonhematopoietic cells, WT T cells produce Flt3L and contribute to the generation of both classical DCs (cDCs) and plasmacytoid DCs in Flt3L^?/? mice. Upon stimulation in vitro, CD4⁺ T cells produce more Flt3L than CD8⁺ T cells. Moreover, in vivo stimulation of naïve OT‐II CD4⁺ T cells with OVA leads to increase of pre‐cDCs and cDCs in draining lymph nodes of Flt3L^?/? mice in a partially Flt3L‐dependent manner. Thus, Flt3L‐mediated lymphoid tissue DC homeostasis is regulated by steady‐state T cells as well as by proliferative T cells, fostering local development of lymphoid organ resident DCs.     

Obesity promotes prolonged ovalbumin‐induced airway inflammation modulating T helper type 1 (Th1), Th2 and Th17 immune responses in BALB/c mice

Clinical and epidemiological studies indicate that obesity affects the development and phenotype of asthma by inducing inflammatory mechanisms in addition to eosinophilic inflammation. The aim of this study was to assess the effect of obesity on allergic airway inflammation and T helper type 2 (Th2) immune responses using an experimental model of asthma in BALB/c mice. Mice fed a high‐fat diet (HFD) for 10 weeks were sensitized and challenged with ovalbumin (OVA), and analyses were performed at 24 and 48 h after the last OVA challenge. Obesity induced an increase of inducible nitric oxide synthase (iNOS)‐expressing macrophages and neutrophils which peaked at 48 h after the last OVA challenge, and was associated with higher levels of interleukin (IL)‐4, IL‐9, IL‐17A, leptin and interferon (IFN)‐γ in the lungs. Higher goblet cell hyperplasia was associated with elevated mast cell influx into the lungs and trachea in the obese allergic mice. In contrast, early eosinophil influx and lower levels of IL‐25, thymic stromal lymphopoietin (TSLP), CCL11 and OVA‐specific immunoglobulin (IgE) were observed in the obese allergic mice in comparison to non‐obese allergic mice. Moreover, obese mice showed higher numbers of mast cells regardless of OVA challenge. These results indicate that obesity affects allergic airway inflammation through mechanisms involving mast cell influx and the release of TSLP and IL‐25, which favoured a delayed immune response with an exacerbated Th1, Th2 and Th17 profile. In this scenario, an intense mixed inflammatory granulocyte influx, classically activated macrophage accumulation and intense mucus production may contribute to a refractory therapeutic response and exacerbate asthma severity.     

Vasostatin‐1 antigenic epitope mapping for induction of cellular and humoral immune responses to chromogranin A autoantigen in NOD mice

The chromogranin A (ChgA) 29–42 sequence is the antigenic epitope for the BDC2.5 CD4⁺ T‐cell receptor in NOD mice (H‐2^g7). We have now characterized the binding register of the ChgA 29–42 peptide for the I‐A^g7 molecule. Truncation of the peptide demonstrated that the KCVLEVISD sequence 34–42 is the binding register and extension of this sequence by flanking residues increased its binding affinity and antigenic capacity. We employed anti‐ChgA peptide antibodies generated against different fragments of ChgA for immunostaining of pancreatic islet sections from NOD mice. A strong immuno‐staining pattern was observed for the ChgA 17–38 peptide antibodies that overlap with the ChgA 29–42 sequence. Moreover, sera from diabetic NOD mice showed elevated titers of autoantibodies to the ChgA 29–42 peptide. These findings indicate that peptides from the N‐terminal region of ChgA are able to induce cellular and humoral immune responses in NOD mice.