ERG Functionally Overlaps with Other Ets Proteins in Promoting TH9 Cell Expression of Il9 during Allergic Lung Inflammation.

CD4+ TH cells develop into subsets that are specialized in the secretion of particular cytokines to mediate restricted types of inflammation and immune responses. Among the subsets that promote development of allergic inflammatory responses, IL-9-producing TH9 cells are regulated by a number of transcription factors. We have previously shown that the E26 transformation-specific (Ets) family members PU.1 and Ets translocation variant 5 (ETV5) function in parallel to regulate IL-9. In this study we identified a third member of the Ets family of transcription factors, Ets-related gene (ERG), that mediates IL-9 production in TH9 cells in the absence of PU.1 and ETV5. Chromatin immunoprecipitation assays revealed that ERG interaction at the Il9 promoter region is restricted to the TH9 lineage and is sustained during murine TH9 polarization. Knockdown or knockout of ERG during murine or human TH9 polarization in vitro led to a decrease in IL-9 production in TH9 cells. Deletion of ERG in vivo had modest effects on IL-9 production in vitro or in vivo. However, in the absence of PU.1 and ETV5, ERG was required for residual IL-9 production in vitro and for IL-9 production by lung-derived CD4 T cells in a mouse model of chronic allergic airway disease. Thus, ERG contributes to IL-9 regulation in TH9 cells.

Comparative Analysis of Alternative Splicing Profiles in Th Cell Subsets Reveals Extensive Cell Type-Specific Effects Modulated by a Network of Transcription Factors and RNA-Binding Proteins.

Alternative splicing (AS) plays an important role in the development of many cell types; however, its contribution to Th subsets has been clearly defined. In this study, we compare mice naive CD4+ Th cells with Th1, Th2, Th17, and T regulatory cells and observed that the majority of AS events were retained intron, followed by skipped-exon events, with at least 1200 genes across cell types affected by AS events. A significant fraction of the AS events, especially retained intron events from the 72-h time point, were no longer observed 2 wk postdifferentiation, suggesting a role for AS in early activation and differentiation via preferential expression of specific isoforms required during T cell activation, but not for differentiation or effector function. Examining the protein consequence of the exon-skipping events revealed an abundance of structural proteins encoding for intrinsically unstructured peptide regions, followed by transmembrane helices, ß strands, and polypeptide turn motifs. Analyses of expression profiles of RNA-binding proteins (RBPs) and their cognate binding sites flanking the discovered AS events revealed an enrichment for specific RBP recognition sites in each of the Th subsets. Integration with publicly available chromatin immunoprecipitation sequencing datasets for transcription factors support a model wherein lineage-determining transcription factors impact the RBP profile within the differentiating cells, and this differential expression contributes to AS of the transcriptome via a cascade of cell type-specific posttranscriptional rewiring events.

Pancreas transplantation for cystic fibrosis: A frequently missed opportunity.

Cystic fibrosis (CF) is an inherited autosomal recessive disorder. Despite optimized therapy, the majority of affected individuals ultimately die of respiratory failure. As patients with CF are living longer, extra-pulmonary manifestations may develop including pancreatic failure, which manifests as exocrine insufficiency, and CF-related diabetes (CFRD). Both of these can be managed through pancreas transplantation. Pancreas transplantation is usually performed in combination with another organ, most often with a kidney transplant for end-stage diabetic nephropathy. In the CF patient population, the two settings where inclusion of a pancreas transplant should be considered would be in combination with a lung transplant for CF pulmonary disease, or in combination with a liver for CF-related liver disease with cirrhosis. This report will discuss this topic in detail, including a review of the literature regarding combinations of lung/pancreas and liver/pancreas transplant.

STAT5 promotes accessibility and is required for BATF-mediated plasticity at the Il9 locus.

T helper cell differentiation requires lineage-defining transcription factors and factors that have shared expression among multiple subsets. BATF is required for development of multiple Th subsets but functions in a lineage-specific manner. BATF is required for IL-9 production in Th9 cells but in contrast to its function as a pioneer factor in Th17 cells, BATF is neither sufficient nor required for accessibility at the Il9 locus. Here we show that STAT5 is the earliest factor binding and remodeling the Il9 locus to allow BATF binding in both mouse and human Th9 cultures. The ability of STAT5 to mediate accessibility for BATF is observed in other Th lineages and allows acquisition of the IL-9-secreting phenotype. STAT5 and BATF convert Th17 cells into cells that mediate IL-9-dependent effects in allergic airway inflammation and anti-tumor immunity. Thus, BATF requires the STAT5 signal to mediate plasticity at the Il9 locus.

Bcl6 and Blimp1 reciprocally regulate ST2+ Treg-cell development in the context of allergic airway inflammation.

BACKGROUND: Bcl6 is required for the development of T follicular helper cells and T follicular regulatory (Tfr) cells that regulate germinal center responses. Bcl6 also affects the function of regulatory T (Treg) cells. OBJECTIVE: The goal of this study was to define the functions of Bcl6 in Treg cells, including Tfr cells, in the context of allergic airway inflammation. METHODS: We used a model of house dust mite sensitization to challenge wild-type, Bcl6fl/fl Foxp3-Cre, and Prdm1 (Blimp1)fl/fl Foxp3-Cre mice to study the reciprocal roles of Bcl6 and Blimp1 in allergic airway inflammation. RESULTS: In the house dust mite model, Tfr cells repress the production of IgE and Bcl6+ Treg cells suppress the generation of type 2 cytokine-producing cells in the lungs. In mice with Bcl6-deficient Treg cells, twice as many ST2+ (IL-33R+) Treg cells develop as are observed in wild-type mice. ST2+ Treg cells in the context of allergic airway inflammation are Blimp1 dependent, express type 2 cytokines, and share features of visceral adipose tissue Treg cells. Bcl6-deficient Treg cells are more susceptible, and Blimp1-deficient Treg cells are resistant, to acquiring the ST2+ Treg-cell phenotype in vitro and in vivo in response to IL-33. Bcl6-deficient ST2+ Treg cells, but not Bcl6-deficient ST2+ conventional T cells, strongly promote allergic airway inflammation when transferred into recipient mice. Lastly, ST2 is required for the exacerbated allergic airway inflammation in Bcl6fl/fl Foxp3-Cre mice. CONCLUSIONS: During allergic airway inflammation, Bcl6 and Blimp1 play dual roles in regulating Tfr-cell activity in the germinal center and in the development of ST2+ Treg cells that promote type 2 cytokine responses.

A conserved enhancer regulates Il9 expression in multiple lineages.

Cytokine genes are regulated by multiple regulatory elements that confer tissue-specific and activation-dependent expression. The cis-regulatory elements of the gene encoding IL-9, a cytokine that promotes allergy, autoimmune inflammation and tumor immunity, have not been defined. Here we identify an enhancer (CNS-25) upstream of the Il9 gene that binds most transcription factors (TFs) that promote Il9 gene expression. Deletion of the enhancer in the mouse germline alters transcription factor binding to the remaining Il9 regulatory elements, and results in diminished IL-9 production in multiple cell types including Th9 cells, and attenuates IL-9-dependent immune responses. Moreover, deletion of the homologous enhancer (CNS-18) in primary human Th9 cultures results in significant decrease of IL-9 production. Thus, Il9 CNS-25/IL9 CNS-18 is a critical and conserved regulatory element for IL-9 production.

STAT3 Activation Impairs the Stability of Th9 Cells.

Th9 cells regulate multiple immune responses, including immunity to pathogens and tumors, allergic inflammation, and autoimmunity. Despite ongoing research into Th9 development and function, little is known about the stability of the Th9 phenotype. In this study, we demonstrate that IL-9 production is progressively lost in Th9 cultures during several rounds of differentiation. The loss of IL-9 is not due to an outgrowth of cells that do not secrete IL-9, as purified IL-9 secretors demonstrate the same loss of IL-9 in subsequent rounds of differentiation. The loss of IL-9 production correlates with increases in phospho-STAT3 levels within the cell, as well as the production of IL-10. STAT3-deficient Th9 cells have increased IL-9 production that is maintained for longer in culture than IL-9 in control cultures. IL-10 is responsible for STAT3 activation during the first round of differentiation, and it contributes to instability in subsequent rounds of culture. Taken together, our results indicate that environmental cues dictate the instability of the Th9 phenotype, and they suggest approaches to enhance Th9 activity in beneficial immune responses.