Interferon regulatory factor-4 activates IL-2 and IL-4 promoters in cooperation with c-Rel.

Interferon regulatory factor (IRF)-4 is a member of the IRF transcription factor family, whose expression is primarily restricted to lymphoid and myeloid cells. In T-cells, IRF-4 expression is induced by T-cell receptor (TCR) cross-linking or treatment with phorbol-12-myristate-13-acetate (PMA)/Ionomycin, and IRF-4 is thought to be a critical factor for various functions of T-cells. To elucidate the IRF-4 functions in human adult T-cell leukemia virus type 1 (HTLV-1)-infected T-cells, which constitutively express IRF-4, we isolated IRF-4-binding proteins from T-cells, using a tandem affinity purification (TAP)-mass spectrometry strategy. Fourteen proteins were identified in the IRF-4-binding complex, including endogenous IRF-4 and the nuclear factor-kappaB (NF-κB) family member, c-Rel. The specific association of IRF-4 with c-Rel was confirmed by immunoprecipitation experiments, and IRF-4 was shown to enhance the c-Rel-dependent binding and activation of the interleukin-4 (IL-4) promoter region. We also demonstrated that IL-2 production was also enhanced by exogenously-expressed IRF-4 and c-Rel in the presence of P/I, in T-cells, and that the optimal IL-2 and IL-4 productions in vivo was IRF-4-dependent using IRF-4-/- mice. These data provide molecular evidence to support the clinical observation that elevated expression of c-Rel and IRF-4 is associated with the prognosis in adult T-cell leukemia/lymphoma (ATLL) patients, and present possible targets for future gene therapy

Designer probiotics for the prevention and treatment of human diseases

10.1016/j.cbpa.2017.04.011CURRENT OPINION IN CHEMICAL BIOLOGY408-1

Serum Starvation Activates NF-κB Through G Protein β2 Subunit-Mediated Signal

Several cell stresses induce nuclear factor-kappaB (NF-κB) activation, which include irradiation, oxidation, and UV. Interestingly, serum-starving stress-induced NF-κB activation in COS cells, but not in COS-A717 cells. COSA717 is a mutant cell line of COS cells that is defective of the NF-κB signaling pathway. We isolated genes with compensating activity for the NF-κB pathway and one gene encoded the G protein b2 (Gb2). Gb2 is one of the G rotein-coupled receptor signaling effectors. In COS-A717 cells, Gb2 expression is significantly reduced. In Gb2 cDNA-transfected COS-A717 cells, the NF-κB activity was increased along with the recovery of Gb2 expression. Furthermore, serum-starving stress induced the NF-κB activity in Gb2-transfected COS-A717 cells. Consistently, the serum-starved COS cells with siRNA-reduced Gb2 protein expression showed decreased NF-κB activity. These results indicate that Gb2 is required for starvation-induced NF-κB activation and constitutive NF-kB activity. We propose that serum contains some molecule(s) that strongly inhibits NF-κB activation mediated through Gβ2 signaling

Engineering Escherichia coli for diagnosis and management of hyperuricemia

10.3389/fbioe.2023.1191162Frontiers in Bioengineering and Biotechnology1

Development of a Proline-Based Selection System for Reliable Genetic Engineering in Chinese Hamster Ovary Cells

10.1021/acssynbio.0c00221ACS SYNTHETIC BIOLOGY971864-187

An Engineered Probiotic Produces a Type III Interferon IFNL1 and Reduces Inflammations in <i>in vitro</i> Inflammatory Bowel Disease Models

The etiology of inflammatory bowel diseases (IBDs) frequently results in the uncontrolled inflammation of intestinal epithelial linings and the local environment. Here, we hypothesized that interferon-driven immunomodulation could promote anti-inflammatory effects. To test this hypothesis, we engineered probiotic Escherichia coli Nissle 1917 (EcN) to produce and secrete a type III interferon, interferon lambda 1 (IFNL1), in response to nitric oxide (NO), a well-known colorectal inflammation marker. We then validated the anti-inflammatory effects of the engineered EcN strains in two in vitro models: a Caco-2/Jurkat T cell coculture model and a scaffold-based 3D coculture IBD model that comprises intestinal epithelial cells, myofibroblasts, and T cells. The IFNL1-expressing EcN strains upregulated Foxp3 expression in T cells and thereafter reduced the production of pro-inflammatory cytokines such as IL-13 and -33, significantly ameliorating inflammation. The engineered strains also rescued the integrity of the inflamed epithelial cell monolayer, protecting epithelial barrier integrity even under inflammation. In the 3D coculture model, IFNL1-expressing EcN treatment enhanced the population of regulatory T cells and increased anti-inflammatory cytokine IL-10. Taken together, our study showed the anti-inflammatory effects of IFNL1-expressing probiotics in two in vitro IBD models, demonstrating their potential as live biotherapeutics for IBD immunotherapy

Engineered commensal microbes for diet-mediated colorectal-cancer chemoprevention

10.1038/s41551-017-0181-yNATURE BIOMEDICAL ENGINEERING2127-3

Author Correction: Engineered commensal microbes for diet-mediated colorectal-cancer chemoprevention (vol 2, pg 27, 2018)

10.1038/s41551-020-0580-3NATURE BIOMEDICAL ENGINEERING47754-75

Characterization of dsRNA-induced pancreatitis model reveals the regulatory role of IFN regulatory factor 2 (Irf2) in trypsinogen5 gene transcription

Mice deficient for interferon regulatory factor (Irf)2 (Irf2−/− mice) exhibit immunological abnormalities and cannot survive lymphocytic choriomeningitis virus infection. The pancreas of these animals is highly inflamed, a phenotype replicated by treatment with poly(I:C), a synthetic double-stranded RNA. Trypsinogen5 mRNA was constitutively up-regulated about 1,000-fold in Irf2−/− mice compared with controls as assessed by quantitative RT-PCR. Further knockout of IFNα/β receptor 1(Ifnar1) abolished poly(I:C)-induced pancreatitis but had no effect on the constitutive up-regulation of trypsinogen5 gene, indicating crucial type I IFN signaling to elicit the inflammation. Analysis of Ifnar1−/− mice confirmed type I IFN-dependent transcriptional activation of dsRNA-sensing pattern recognition receptor genes MDA5, RIG-I, and TLR3, which induced poly(I:C)-dependent cell death in acinar cells in the absence of IRF2. We speculate that Trypsin5, the trypsinogen5 gene product, leaking from dead acinar cells triggers a chain reaction leading to lethal pancreatitis in Irf2−/− mice because it is resistant to a major endogenous trypsin inhibitor, Spink3