Interleukin-4 induced leukocyte differentiation

Monocytes, macrophages and dendritic cells (DCs) are important mediators of innate immune system, whereas T lymphocytes are the effector cells of adaptive immune responses. DCs play a crucial role in bridging innate and adaptive immunity. Naïve CD4+ Th progenitors (Thp) differentiate to functionally distinct effector T cell subsets including Th1, Th2 and Th17 cells, which while being responsible for specific immune functions have also been implicated in pathological responses, such as autoimmunity, asthma and allergy. The main objective of this thesis is to dissect the signalling networks involved in the IL-4 induced differentiation of two important leukocyte subtypes, Th2 cells and DCs. Gene expression profiling lead to identification of over 200 genes which are differentially expressed during cytokine induced differentiation of human monocytes to DCs or macrophages and which are likely to be essential for the proper biological functions of these cell types. Transcriptome analysis demonstrated the dynamic regulation of gene expression by IL-12 and IL-4 during the initiation of Th cell differentiation, which was partly counteracted by an immunosuppressive cytokine, TGFβ, present in the culture media. Results from RNAi mediated gene knockdown experiments and global gene expression analysis elucidated that SATB1 regulates multiple genes important for Th cell polarization or function as well as may compete with GATA3 for the reciprocal regulation of IL-5 transcription. In conclusion, the results obtained have extended our system-level understanding of the immune cell differentiation processes and provide an excellent basis for the further functional studies which could lead to development of improved therapeutic approaches for a range of immunological conditions.Siirretty Doriast

Activating transcription factor 3 is a positive regulator of human IFNG gene expression

IL-12 and IL-18 are essential for Th1 differentiation, whereas the role of IFN-α in Th1 development is less understood. In this microarray-based study, we searched for genes that are regulated by IFN-α, IL-12, or the combination of IL-12 plus IL-18 during the early differentiation of human umbilical cord blood CD4+ Th cells. Twenty-six genes were similarly regulated in response to treatment with IL-12, IFN-α, or the combination of IL-12 plus IL-18. These genes could therefore play a role in Th1 lineage decision. Transcription factor activating transcription factor (ATF) 3 was upregulated by these cytokines and selected for further study. Ectopic expression of ATF3 in CD4+ T cells enhanced the production of IFN-α, the hallmark cytokine of Th1 cells, whereas small interfering RNA knockdown of ATF3 reduced IFN-γ production. Furthermore, ATF3 formed an endogenous complex with JUN in CD4+ T cells induced to Th1. Chromatin immunoprecipitation and luciferase reporter assays showed that both ATF3 and JUN are recruited to and transactivate the IFNG promoter during early Th1 differentiation. Collectively, these data indicate that ATF3 promotes human Th1 differentiation

Aryl hydrocarbon receptor is required for optimal B-cell proliferation

The aryl hydrocarbon receptor (AhR), a transcription factor known for mediating xenobiotic toxicity, is expressed in B cells, which are known targets for environmental pollutants. However, it is unclear what the physiological functions of AhR in B cells are. We show here that expression of Ahr in B cells is up‐regulated upon B‐cell receptor (BCR) engagement and IL‐4 treatment. Addition of a natural ligand of AhR, FICZ, induces AhR translocation to the nucleus and transcription of the AhR target gene Cyp1a1, showing that the AhR pathway is functional in B cells. AhR‐deficient (Ahr (−/−)) B cells proliferate less than AhR‐sufficient (Ahr (+/+)) cells following in vitro BCR stimulation and in vivo adoptive transfer models confirmed that Ahr (−/−) B cells are outcompeted by Ahr (+/+) cells. Transcriptome comparison of AhR‐deficient and AhR‐sufficient B cells identified cyclin O (Ccno), a direct target of AhR, as a top candidate affected by AhR deficiency

Activation of the aryl hydrocarbon receptor dampens the severity of inflammatory skin conditions

Environmental stimuli are known to contribute to psoriasis pathogenesis and that of other autoimmune diseases, but the mechanisms are largely unknown. Here we show that the aryl hydrocarbon receptor (AhR), a transcription factor that senses environmental stimuli, modulates pathology in psoriasis. AhR-activating ligands reduced inflammation in the lesional skin of psoriasis patients, whereas AhR antagonists increased inflammation. Similarly, AhR signaling via the endogenous ligand FICZ reduced the inflammatory response in the imiquimod-induced model of skin inflammation and AhR-deficient mice exhibited a substantial exacerbation of the disease, compared to AhR-sufficient controls. Nonhematopoietic cells, in particular keratinocytes, were responsible for this hyperinflammatory response, which involved upregulation of AP-1 family members of transcription factors. Thus, our data suggest a critical role for AhR in the regulation of inflammatory responses and open the possibility for novel therapeutic strategies in chronic inflammatory disorders

The RNA-binding protein TTP is a global post-transcriptional regulator of feedback control in inflammation.

RNA-binding proteins (RBPs) facilitate post-transcriptional control of eukaryotic gene expression at multiple levels. The RBP tristetraprolin (TTP/Zfp36) is a signal-induced phosphorylated anti-inflammatory protein guiding unstable mRNAs of pro-inflammatory proteins for degradation and preventing translation. Using iCLIP, we have identified numerous mRNA targets bound by wild-type TTP and by a non-MK2-phosphorylatable TTP mutant (TTP-AA) in 1 h LPS-stimulated macrophages and correlated their interaction with TTP to changes at the level of mRNA abundance and translation in a transcriptome-wide manner. The close similarity of the transcriptomes of TTP-deficient and TTP-expressing macrophages upon short LPS stimulation suggested an effective inactivation of TTP by MK2, whereas retained RNA-binding capacity of TTP-AA to 3'UTRs caused profound changes in the transcriptome and translatome, altered NF-κB-activation and induced cell death. Increased TTP binding to the 3'UTR of feedback inhibitor mRNAs, such as Ier3, Dusp1 or Tnfaip3, in the absence of MK2-dependent TTP neutralization resulted in a strong reduction of their protein synthesis contributing to the deregulation of the NF-κB-signaling pathway. Taken together, our study uncovers a role of TTP as a suppressor of feedback inhibitors of inflammation and highlights the importance of fine-tuned TTP activity-regulation by MK2 in order to control the pro-inflammatory response

RNA-binding proteins ZFP36L1 and ZFP36L2 promote cell quiescence.

Progression through the stages of lymphocyte development requires coordination of the cell cycle. Such coordination ensures genomic integrity while cells somatically rearrange their antigen receptor genes [in a process called variable-diversity-joining (VDJ) recombination] and, upon successful rearrangement, expands the pools of progenitor lymphocytes. Here we show that in developing B lymphocytes, the RNA-binding proteins (RBPs) ZFP36L1 and ZFP36L2 are critical for maintaining quiescence before precursor B cell receptor (pre-BCR) expression and for reestablishing quiescence after pre-BCR-induced expansion. These RBPs suppress an evolutionarily conserved posttranscriptional regulon consisting of messenger RNAs whose protein products cooperatively promote transition into the S phase of the cell cycle. This mechanism promotes VDJ recombination and effective selection of cells expressing immunoglobulin-μ at the pre-BCR checkpoint.This work was funded by the Biotechnology and Biological Sciences Research Council, a Medical Research Council CASE studentship with GSK, an MRC centenary award (A.G) and project grants from Bloodwise. DJH was supported by a Medical Research Council Clinician Scientist FellowshipThis is the author accepted manuscript. The final version is available from the American Association for the Advancement of Science via http://dx.doi.org/10.1126/science.aad597

SATB1 dictates expression of multiple genes including IL-5 involved in human T helper cell differentiation

Special AT-rich binding protein 1 (SATB1) is a global chromatin organizer and a transcription factor regulated by interleukin-4 (IL-4) during the early T helper 2 (Th2) cell differentiation. Here we show that SATB1 controls multiple IL-4 target genes involved in human Th cell polarization or function. Among the genes regulated by SATB1 is that encoding the cytokine IL-5, which is predominantly produced by Th2 cells and plays a key role in the development of eosinophilia in asthma. We demonstrate that, during the early Th2 cell differentiation, IL-5 expression is repressed through direct binding of SATB1 to the IL-5 promoter. Furthermore, SATB1 knockdown-induced upregulation of IL-5 is partly counteracted by down-regulating GATA3 expression using RNAi in polarizing Th2 cells. Our results suggest that a competitive mechanism involving SATB1 and GATA3 regulates IL-5 transcription, and provide new mechanistic insights into the stringent regulation of IL-5 expression during human Th2 cell differentiation. (Blood. 2010;116(9):1443-1453

Comparative analysis of human and mouse transcriptomes of Th17 cell priming

Uncontrolled Th17 cell activity is associated with cancer and autoimmune and inflammatory diseases. To validate the potential relevance of mouse models of targeting the Th17 pathway in human diseases we used RNA sequencing to compare the expression of coding and non-coding transcripts during the priming of Th17 cell differentiation in both human and mouse. In addition to already known targets, several transcripts not previously linked to Th17 cell polarization were found in both species. Moreover, a considerable number of human-specific long non-coding RNAs were identified that responded to cytokines stimulating Th17 cell differentiation. We integrated our transcriptomics data with known disease-associated polymorphisms and show that conserved regulation pinpoints genes that are relevant to Th17 cell-mediated human diseases and that can be modelled in mouse. Substantial differences observed in non-coding transcriptomes between the two species as well as increased overlap between Th17 cell-specific gene expression and disease-associated polymorphisms underline the need of parallel analysis of human and mouse models. Comprehensive analysis of genes regulated during Th17 cell priming and their classification to conserved and non-conserved between human and mouse facilitates translational research, pointing out which candidate targets identified in human are worth studying by using in vivo mouse models

Rhinoviruses in infancy and risk of immunoglobulin E sensitization

Previous data about the role of viruses in the development of allergic immunoglobulin E (IgE) sensitization are contradictory. The aim of this study was to determine the possible associations between exposure to different viruses (rhinovirus, enterovirus, norovirus, and parechovirus) during the first year of life and IgE sensitization. Viruses were analyzed from stool samples collected monthly from infants participating in a prospective birth cohort study. From that study, 244 IgE sensitized case children and 244 nonsensitized control children were identified based on their allergen-specific IgE antibody levels at the age of 6, 18, and 36 months. Stool samples (n = 4576) from the case and control children were screened for the presence of rhinovirus, enterovirus, norovirus, and parechovirus RNA by reverse transcription quantitative polymerase chain reaction. The study showed that rhinovirus was the most prevalent virus detected, present in 921 (20%) samples. None of the viruses were associated with IgE sensitization in the full cohort but after stratifying by sex, the number of rhinovirus positive samples was inversely associated with IgE sensitization in boys (odds ratio [OR]: 0.81; 95% confidence interval [CI]: 0.69-0.94; P = 0.006). There was also a temporal relation between rhinoviruses and IgE sensitization, as rhinovirus exposure during the first 6 months of life was associated with a reduced risk of subsequent IgE sensitization in boys (OR: 0.76; 95% CI: 0.6-0.94; P = 0.016). In conclusion, early exposure to rhinoviruses was inversely associated with IgE sensitization but this protective association was restricted to boys.Peer reviewe